book of abstracts - Phytochemical Society of Europe

Transcript

Facolta’ di Farmacia
Programme
Programme &
& Abstracts
Abstracts
INTERNATIONAL
INTERNATIONAL PSE
PSE SYMPOSIUM
SYMPOSIUM
PHYT
CHEMICALS
in Nutrition
and Health
27-30 September 2011
Giovinazzo (BARI), Italy
INTERNATIONAL PSE SYMPOSIUM
Phytochemicals in Nutrition and Health
Programme & Abstracts
27-30 September 2011
Giovinazzo (BARI), Italy
FACOLTÀ DI FARMACIA
Organizing Committee
Pinarosa Avato, Chairman (Bari, Italy)
Maria Pia Argentieri, Treasurer (Bari, Italy)
Francesco Racaniello, Secretariat (Bari, Italy)
Scientific Committee
Giovanni Appendino (Novara, Italy)
Randolph Arroo (Leicester, UK)
Pinarosa Avato (Bari, Italy)
Lars Bohlin (Uppsala, Sweden)
Veronique Cheynier (Montpellier, France)
Massimo Curini (Perugia, Italy)
Virginia Lanzotti (Naples, Italy)
Wieslaw Oleszek (Pulawy, Poland)
Vassilios Roussis (Athens, Greece)
Satya Sarker (Wolverhampton, UK)
Angela Stafford (Sheffield,UK)
http://www.phytochemicalsociety.org/bari
Under the High Patronage and with the financial support of
Università degli Studi di Bari Aldo Moro
Facoltà di Farmacia, Università degli Studi di Bari Aldo Moro
Senato della Repubblica Italiana
Ministero della Salute
Regione Puglia
Città di Bari
Comune di Bitonto – Assessorato al Turismo
Fondazione Cassa di Risparmio di Puglia
Federazione Ordini Farmacisti Italiani
Ordine dei Farmacisti Provincia di Bari
Ordine dei Farmacisti Provincia di Brindisi
Società Italiana di Fitochimica
Aboca S.p.A
Indena S.p.A
Apivita Europe
Thermo Scientific
Waters
Extrasynthese
Farmalabor – Farmacisti Associati
Amarelli
Confetteria Museo Mario Mucci
F. Divella S.p.A
Battista Luigi Caffè & Bristol S.r.l
Biscottificio Panificio Farinella
Programme
Salone delle Stelle
Grand Hotel Riva del Sole
Giovinazzo (Bari)
Programme
Tuesday –27 September
12.00-15.00
Registration
15.00
Opening and welcome address
Pinarosa Avato, Chairperson
Corrado Petrocelli, Rector of “Universitá degli Studi di Bari Aldo Moro”
Roberto Perrone, Dean of “Facoltá di Farmacia, Universitá degli Studi di Bari Aldo Moro”
Luigi d’Ambrosio Lettieri, Senator of the Italian Republic, Standing Committee for Health;
Vice - President of the Italian Federation of Pharmacists
Pier Giorgio Zambonin, President of “Societá Chimica Italiana – Sezione Puglia”
Luisa Pistelli, Executive Board of “Societá Italiana di Fitochimica”
Wieslaw Oleszek, President of “Phytochemical Society of Europe”
Chair: Anna-Marja Aura
15.30-16.10
PL1
Giovanni Appendino (University of Piemonte Orientale, Italy)
Food-based drug discovery: overcoming the nutrition-medicine divide
16.10-16.30
SL1
Paul Reddell (EcoBiotics Ltd, Australia)
Discovery of new nutraceuticals from Australia’s tropical rainforests
16.30-16.50
SL2
Jafar M.I. Alqudah (Dublin Institute of Technology, Ireland)
Screening of irish fruit and vegetable germplasm for novel anti-tumour
and pesticidal compounds
16.50-17.30
Coffee Break
Chair: Giovanni Appendino
17.30-18.10
PL2
Mercedes Muzquiz (SGIT-INIA, Spain)
Bioactive compounds in legumes: pronutritive and antinutritive actions.
Implications for nutrition and health
18.10-18.30
SL3
Miriana Durante (ISPA-CNR, Italy)
Durum wheat by-products as natural sources of valuable nutrients
18.30-18.50
SL4
Lindsay Brown (University of Southern Queensland, Australia)
Anthocyanin-enriched purple carrot juice with low carotenoids
attenuates cardiovascular and hepatic symptoms induced by high
carbohydrate, high fat-feeding in rats
21.00
“Italian Dragees”: After dinner by “Confetteria & Museo Mario Mucci”
Wednesday –28 September
Chair: Paul J. Russell
09.00-09.40
PL3
Wolfgang Gelbmann (European Food Safety Authority – EFSA)
EFSA’s Role in Novel Foods and Health Claims
09.40-10.00
SL5
Marcello Salvatore Lenucci (University of Salento, Italy)
Tomato organic lycopene: the way to obtain a unique healthy product
10.00-10.20
SL6
Franz Bucar (University of Graz, Austria)
Isoflavone composition of soy and red clover dietary supplements
10.20-10.40
SL7
Juan Parrado (University of Sevilla, Spain)
Antioxidant and anti-inflammatory properties of a new soluble extract
from grape pomace
10.40-11.10
Coffee Break
Chair: Jean-Luc Wolfender
11.10-11.50
PL4
Veronique Cheynier (INRA, France)
Phenolic compounds: from plants to foods
11.50-12.10
SL8
Ana A. Matias (University of Lisboa, Portugal)
Polyphenols-rich extracts obtained from Prunus avium and Opuntia ficus
indica as natural anti-inflammatory modulators for Inflammatory Bowel
Diseases.
12.10-12.30
SL9
Emilio Jirillo (University of Bari, Italy)
Polyphenols from red wine exert potential beneficial effects to human
health
12.30-12.50
SL10
Magdalini A. Papandreou (University of Athens, Greece)
Differential antioxidant & anti-amyloid effects of a wild bluberry
polyphenol-rich extract: in vitro & in vivo studies
13.00-14.00
Lunch Break
14.00-15.00
Free Poster Session
Chair: Franz Bucar
15.00-15.40
PL5
Jean-Luc Wolfender (University of Geneva, Switzerland)
New trends in the analysis of natural products for the detailed metabolite
profiling of crude extracts
15.40-16.00
SL11 Vera Schulzova (University of Prague, Czech Republic)
Metabolomic profiling and fingerprinting: new effective approach for
plant characterisation and authentication
16.00-16.20
SL12
Margherita Zotti (University of Foggia, Italy)
Estrous cycle in female rats influences the neurochemical and
neurobehavioural effects of carvacrol
16.20-17.00
Coffee Break
Chair: Mercedes Muzquiz
17.00-17.40
PL6
Arnaud Bovy (Plant Research International, The Netherlands)
Breeding for improved flavour and health
17.40-18.00
SL13 Jiri Gruz (Palacký University, Czech Republic)
Application of purine derivative LGR-1814 improves functional
properties of field-grown lettuce (Lactuca sativa)
18.00-18.20
SL14 Anna Szakiel (University of Warsaw, Poland)
Comparison of triterpenoid content of Vaccinium vitis-idaea berries
and leaves collected from natural forest habitats in Finland and Poland
18.30-20.00
Poster Session 1 Authors presenting posters with odd numbers should be
present at their posters
21.00
Visit to the medieval town of Bitonto
Thursday –29 September
Chair: Virginia Lanzotti
09.00-09.40
PL7
Søren Balling Engelsen (University of Copenhagen, Denmark)
NMR metabolomics: advantages and disadvantages for studying
bioactive components in food
09.40-10.00
SL15
Marlene Monschein (University of Graz, Austria)
Formation of astragaloside IV from acylated astragalosides during
extraction of Astragali radix
10.00-10.20
SL16
Wanda Kisiel (Polish Academy of Sciences, Poland)
Sesquiterpenoids from aerial parts of Lactuca aculeata
10.20-10.40
SL17
Christine Tschiggerl (University of Graz, Austria)
The volatile fraction of herbal teas
10.40-11.10
Coffee Break
Chair: Veronique Cheynier
11.10-11.50
PL8
Anna-Marja Aura (VTT, Finland)
Phenolic nutritional phytochemicals as biomarkers of plant food intake
11.50-12.10
SL18
Annalisa Tassoni (University of Bologna, Italy)
Elicitation, production and release of polyphenols in fed-batch
fermentation of grape cells
12.10-12.30
SL19
Susana M. Cardoso (CERNAS, Portugal)
Insights into olive mill wastewaters phenolics
12.30-12.50
SL20
Antonella Leone (ISPA-CNR, Italy)
Molecular mechanisms underlie the biological activities of
phytochemicals: searching the gap
13.00-14.00
Lunch Break
14.00-15.00 Free Poster Session
Chair: Giuseppe Dalessandro
15.00-15.40
PL9
Barbara Ann Halkier (University of Copenhagen, Denmark)
Bioengineering of glucosinolates in yeast
15.40-16.00
SL21
Gemma Farrè (University of Lleida, Spain)
Corn metabolic engineering by creating a metabolic sink by expressing
the Arabidopsis homologue of the Orange (Or) gene
16.00-16.20
SL22
Mrinal K. Maiti (Indian Institute of Technology, India)
Production of nutritionally desirable fatty acids in seed oil of Indian
mustard (Brassica juncea L.) by metabolic engineering
16.20-17.00
Coffee Break
Chair: Wieslaw Oleszek
17.00-17.40
PSE-APIVITA Award Ceremony with the lecture by Award Winner
17.40-18.00
SL23
18.00-18.20
SL24 Marie-Hélène Teiten (Hôpital Kirchberg, Grand Duchy of Luxembourg)
Proteomic identification of differentially expressed proteins in curcumintreated prostate cancer cells
18.30-20.00
Poster Session 2 Authors presenting posters with even numbers should be
present at their posters
20.30
Hemant Poudyal (University of Queensland, Australia)
D-Linolenic acid-rich oil from chia seed (Salvia hispanica L.) induces
lipid redistribution, heart and liver protection in diet-induced obese rats
Symposium Dinner
Friday –30 September
Chair: Arnaud Bovy
09.00-09.40
PL10
Virginia Lanzotti (University of Naples, Italy)
Bioactive natural compounds from food plants
09.40-10.00
SL25
Olaf-Georg Issinger (University of Southern Denmark, Denmark)
Natural compounds as selective inhibitors for protein kinases and their
potential application in cancer research
10.00-10.20
SL26
Mariangela Miele (University of Genova, Italy)
Hazel and other angiosperm plants as new sources of compounds with
taxane-like activity
10.20-10.50
Coffee Break
Chair: Lindsay Brown
10.50-11.30
PL11
Paul J. Russel (Unilever, UK)
Naturals for masses - Chemistry approaches for safe commercialisations
11.30-11.50
SL27
Caroline Mathon (Official Food Control Authority, Switzerland)
Targeted screening of botanicals in herbal dietary supplements by LCMS/MS with an Information Independent Acquisition approach
11.50-12.10
SL28
Angela Mari (University of Salerno, Italy)
LC-MS/MS determination of marker metabolites in the ethanolic extract
and in mother tinctures of Vitex agnus castus fruits
12.10-12.30
14.00
Closing and Farewell
Post Symposium guided sightseeing tour to the UNESCO World Heritage town of Alberobello.
Buffet with wine tasting.
Oral Presentations
Oral Presentations – International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September 2011
PL1
Food-based drug discovery: overcoming the nutrition-medicine
divide
Giovanni Appendino
Dipartimento di Scienze Chimiche, Alimentari, Farmaceutiche e Farmacologiche, Università del
Piemonte Orientale, Via Bovio 6, 28100 Novara, Italy
Statins, the commercially most successful drugs ever, are derivatives of biological analogues of
lovastatin, a constituent of the red yeast of rice, an ingredient of the Oriental cuisine, and foodplant derived constituents have been developed as mainstream drugs (L-DOPA, theophylline).
Yet, edible plants have not yet systematically considered as a source of drugs-leads and drugs.
The growing erosion of the drug-dietary supplement interface provides now new opportunities to
explore the role of food-derived compounds for the complementary management of various
pathologies, especially in the realm of chronic-degenerative diseases, either as a stand-alone
agent, or in combination with mainstream drugs.
After an overview of the chemical diversity of food plants-derived phytochemicals and their
biological targets, recent studies on the identification of the elusive pharmacophore of curcumin
and the clinical development of a phospholipid formulation of curcuminoids will be summarized
as exemplificative of how modern science can contribute to the advancement of our knowledge in
this field.
11
SL1
Discovery of new nutraceuticals from Australia’s tropical rainforests
Paul Reddella, Victoria Gordona, Peter Parsonsb, Gerald Muenchc, Craig Williamsd
a
EcoBiotics Ltd, PO Box 1, Yungaburra, Queensland, 4884, Australia
Queensland Institute of Medical Research, Royal Brisbane Hospital, Herston, Queensland, 4029,
Australia
c
School of Medicine, University of Western Sydney, Locked Bag 1797
Penrith South DC, NSW 1797, Australia
d
Department of Chemistry, University of Queensland, St Lucia, 4072, Australia
[email protected]
b
Australia’s tropical rainforests are amongst the most ancient and diverse in the world. They
have a flora characterised by high endemism at both genus and species level and comprise a wide
range of forest types from humid lowland rainforests through to palm forests, seasonally
deciduous rainforests and tropical mountain cloud forests. The combination of a long geographic
isolation, high phylogenetic diversity and a wide range of physical environments creates ideal
conditions to underpin high and novel chemical diversity as plants evolve new ‘chemical
solutions’ to survive and flourish in these complex and dynamic biological systems.
To date, very few of Australia’s rainforest flora have been examined for their potential
applications in the food, cosmetics or pharmaceutical industries. For example, of the more than
200 species (from more than 40 plant families) that are known to produce edible fruits, nuts,
tubers, flowers and/or leaves, only the macadamia nut (Macadamia integrifolia; family
Proteaceae) and lemon myrtle (Backhousia citrodora; family Myrtaceae) have been
commercialised as mainstream ingredients in food or cosmetics, while a small number of other
species are used as local food products.
This presentation outlines our approach to discovery and early development of new
nutraceutical products from this unique flora. We will (i) highlight how an understanding of
rainforest ecology can be used to successfully target and predict potential sources of new
nutraceuticals with specific bioactivities and (ii) discuss a specific example of assays, biomarkers
and veterinary models that we are using to evaluate potential new anti-inflammatory and
neuroprotectant nutraceuticals.
12
SL2
Screening of irish fruit and vegetable germplasm for novel antitumor and pesticidal compounds
Alqudah J.M.I.1, 2, Reilly K.2, Gaffney M.T.2, Martin-Diana A.B.3 & Barry-Ryan C.1
1
Dublin Institute of Technology, School of Food Science and Environmental Health, Cathal Brugha St,
Dublin 1, Ireland.
2
Teagasc R&D Centre, Horticulture Development Department, Kinsealy, Dublin 17, Ireland.
3
Instituto Technológico Agrario, Junta de Castilla y León, Consejería de Agricultura y Ganadería,
España.
Plant secondary metabolites are a source of potential novel therapeutic and insecticidal agents.
This project is focused on screening Irish grown fruit and vegetables for novel bioactive
compounds, for use in food production and biopesticides. Bioassays used include lethality to brine
shrimp (Artemia salina) larva, a general bioassay for insecticidal and anti-tumour compounds and
the inhibition of Agrobacterium tumefaciens crown gall tumours on potato disc slices. To date
aqueous extracts of 173 plant samples have been screened using the brine shrimp bioassay (BSB).
Six crops showed positive lethality to brine shrimp at a dilution <0.02 v/v, beetroot (root),
cranberry (fruit), garlic (bulb), ginger (rhizome), red onion (bulb) and rhubarb (petiole). These
samples also showed significant inhibition of crown gall tumours in the potato disc bioassay
(PDB) (>20% inhibition). Methanolic extracts of cranberry (fruit), ginger (rhizome) and rhubarb
(petiole) showed positive lethality to brine shrimp in the BSB at concentration <1000 ppm.
Methanolic extracts of cranberry (fruit) and rhubarb (petiole) showed significant inhibition of
crown gall tumours in the PDB (>20%). There was a significant effect shown between aqueous
and methanolic extracts in the BSB and PDB. Phenolic compounds in aqueous extracts of the six
positive crop tissues were separated and analysed using thin layer chromatography plates sprayed
with Folin Denis reagent.
Brine shrimp bioassays were carried out for both aqueous and methanolic extracts from
different tissues of known varieties of beetroot ( ‘Pablo’ and ‘Rocket’), red onion (‘Red Spark’,
‘Red Baron’ and ‘Lilia’) and rhubarb (‘Red Early Superb’, ‘Sutton’ and ‘Victoria’). Beetroot and
red onion varieties showed negative results in the BSB for both aqueous and methanolic extracts.
Aqueous and methanolic extracts of all rhubarb varieties (petiole and leaf) gave positive results in
the BSB. The aqueous extracts of petiole and leaf of rhubarb varieties showed significant
inhibition of crown gall tumours in the PDB (>20%) apart from ‘Sutton’ (petiole) which gave
11.5% inhibition. Further investigations will allow identification of useful bio-active compounds
of plant origin with potential application in the food, pharmaceutical and biopesticide industry.
13
PL2
Bioactive compounds in legumes: pronutritive and antinutritive
actions. Implications for nutrition and health
Mercedes Muzquiz
Dpto. de Tecnología de Alimentos. SGIT-INIA. 28080 Madrid, Spain
[email protected]
Legumes seeds are employed as a protein source for animal and human nutrition not only for
their nutritional value (high in protein, lipids and dietary fibre), but also their adaptability to
marginal soils and climates. Human consumption of legumes has been increased in recent years,
being regarded as beneficial food ingredients. In fact, they are recommended as staple food by
health organisations and dieticians and they can be considered as functional foods [1].
Legume seeds contain a great number of compounds which qualify as phytochemicals with
significant potentials benefits to human health (for example, as anticarcinogenic,
hypocholesterolemic or hypoglycemic agents). The term phytochemical has also been used and
emphasizes the plant source of the majority of these disease-preventing non-nutritive compounds
[2].
Antinutritional compounds vary considerably in their biochemistry. They can be proteins
(protease inhibitors, α-amylases, lectins), glycosides (α-galactosides, vicine and convicine),
tannins, saponins, alkaloids. Hence, methods for their extraction, determination and quantification
are specific of each compound. They do not appear equally distributed in all legumes, and their
physiological effects are diverse. Some of these compounds are important in plant defence
mechanisms against predators or environmental conditions. Others are reserve compounds,
accumulated in seeds as energy stores in readiness for germination [3].
Processing generally improves the nutrient profile of legume seed by increasing in vitro
digestibility of proteins and carbohydrates and at the same time there are reductions in some
antinutritional compounds [4]. Most antinutritional factors are heat-labile, such as protease
inhibitors and lectins, so thermal treatment would remove any potential negative effects from
consumption. On the other hand tannins, saponins and phytic acid are heat stable but can be
reduced by dehulling, soaking, germination and/or fermentation.
However, in order to exert an effect, either local or systemic, these substances have to survive
at least to some extent the digestive process within the gastrointestinal tract. New directions in nonutritional factors research in the last decade have led to major developments in our understanding
of their role in nutrition. The scientific interest in no-nutritional factors is now also turning to
studies of their possible useful and beneficial applications as gut, metabolic and hormonal
regulators and as probiotic/prebiotic agents [5].
>1@ de Roos, N. M. (2004) In: Arnoldi, A. (Ed.), Functional Foods: Cardiovascular Disease and Diabetes.Woodhead
Publishing Ltd., Cambridge, pp. 1-9.
[2] Hasler CM, Bloch AS, Thomson CA, Enrione E, Manning C (2004). Journal of American Dietetic Association
104: 814-826.
>3@ Roberts M (1998). In: M.F. Roberts, M. Wink (eds). Alkaloids. Biochemistry, Ecology, and Medicinal
Applications. Plenum Press. New York and London p. 109-146.
>4@ Hajos Gy, Osagie AU (2004). In: Muzquiz M., Hill G.D., Cuadrado C., Pedrosa M.M., Burbano, C. (eds.)
Proceedings of the fourth international workshop on antinutritional factors in legume seeds and oilseeds, EAAP
publication No. 110, Toledo, Spain, pp. 293-305.
>5@ Pusztai A.,Bardocz S., Martin-Cabrejas M.A. (2004) In: Muzquiz M., Hill G.D., Cuadrado C., Pedrosa M.M.,
Burbano, C. (eds.) Proceedings of the fourth international workshop on antinutritional factors in legume seeds
and oilseeds, EAAP publication No. 110, Toledo, Spain, pp.87-100.
14
SL3
Durum wheat by-products as natural sources of valuable nutrients
Miriana Durante1, Marcello Lenucci2, Leonardo Rescio3, Giovanni Mita1, Sofia Caretto1
1
Istituto di Scienze delle Produzioni Alimentari (ISPA), CNR , Via Prov. le Lecce-Monteroni,
73100 Lecce, Italy
2
Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali (Di.S.Te.B.A.), Università del
Salento, Via Prov. le Lecce-Monteroni, I-73100 Lecce, Italy
3
PIERRE S.r.l., s.s. 476 Km 17, 650 Zona Industriale, I-73013 Galatina (LE), Italy
[email protected]
Based on epidemiological studies, the human consumption of whole grains is associated with a
reduced risk of health disorders, such as cancer and diabetes [1]. However, the outer layers of the
wheat kernel (bran plus aleurone layer) together with the embryo (germ), which contain several
bioactive compounds, are usually removed during the early stages of milling to produce flour or
semolina. As a consequence, mostly of the wheat valuable healthy molecules end up in the byproducts. Due to the increasing research interest in establishing new production systems of natural
vitamin E [2], attention has been focused on wheat milling by-products, which represent one of
the richest known natural sources of vitamin E [3]. Being Apulia the leading Italian region in
durum wheat (Triticum durum, ) production, this study was carried out to evaluate the possibility
of extracting oil from an enriched germ containing by-product of durum wheat milling by
supercritical carbon dioxide (SC-CO2). The SC-CO2 technology gives totally solvent free extracts
improving product quality, safety and healthiness. Different operative parameters were tested to
optimize oil and vitamin E extraction yields. When 300 bar pressure, 50 °C temperature, 4 L/min
CO2 flow and 1 hour extraction time were used, an oil yield of ~6 wt %, containing ~4 mg total
tocopherols/g, was obtained. A slight improvement (~7 wt % oil; ~5 mg total tocopherols/g oil)
was reached when the starting material was oven-dried for 72 hours at 60°C (residual humidity ~3
% ) and reduced to a particle size of 30 mesh. The oil characterization was carried out by HPLC
and GC-MS. The -form was the most abundant tocopherol (2.33 mg/g oil), followed by β/γ–
tocopherols (1.99 mg/g oil). Among the identified fatty acids, linoleic was definitely the most
abundant (~50%), followed by moderate amounts of oleic (~27%) and palmitic (~19%) acids.
Finally, carotenoids were present in the SC-CO2 extracted oil as lutein, zeaxanthin and β-carotene.
Oil stability was also evaluated under different combinations of storage conditions (25 °C, 4 °C,
day-light, darkness) measuring changes in vitamin E content and hydroperoxide production.
Altogether these results highlight the importance of wheat milling industry by-products as rich
sources of valuable natural nutrients.
[1] Meyer K.A., Kushi L.H., Jacobs D.R. Jr, Slavin J., Sellers T.A., Folsom A.R. (2000). Carbohydrates, dietary fiber,
incident type 2 diabetes mellitus in older women. American Journal of Clinical Nutrition, 71: 921-930.
[2] Caretto S., Nisi R., Paradiso A., De Gara L. (2010). Tocopherol production in plant cell cultures.. Molecular
Nutrition and Food Research, 54 (5): 726-730.
[3] Atwell W.A. (2001). Wheat Flour. Eagan Press, St. Paul, Minnesota.
15
SL4
Anthocyanin-enriched purple carrot juice with low carotenoids
attenuates cardiovascular and hepatic symptoms induced by high
carbohydrate, high fat feeding in rats
Lindsay Browna and Hemant Poudyalb
a
Department of Biological and Physical Sciences, University of Southern Queensland,
Toowoomba 4350, AUSTRALIA; bSchool of Biomedical Sciences, The University of Queensland,
Brisbane, 4072, AUSTRALIA
[email protected]
Purple carrots (Dacus carota L.) are rich in anthocyanins such as cyanidin-3-(2ʹʹ-xylose-6ʹʹsinapoyl-glucose-galactoside) imparting the distinctive colour [1, 2]. In the current study, we have
investigated whether administration of purple carrot juice (5% of the diet; 15.2 mg/kg/d) for the
last 8 weeks of a 16 weeks protocol reverses the structural and functional changes in the heart and
liver in rats fed a high carbohydrate, high fat diet (HCHF) (*P<0.05; n=8-10/group). In this model
of diet-induced metabolic syndrome, HCHF-fed rats developed hypertension, infiltration of
inflammatory cells, cardiac fibrosis, increased cardiac stiffness, endothelial dysfunction, glucose
intolerance, increased abdominal fat deposition, altered plasma lipid profile, increased hepatic fat
deposition and fibrosis and increased plasma liver enzymes. Purple carrot juice (PC) decreased
percent body weight gain (HCHF 14.3±0.9, HCHF+PC 10.3±0.6*) and normalised glucose
tolerance, systolic blood pressure (HCHF 145±4.5, HCHF+PC 127.3±1.1* mmHg), left
ventricle+septum weight (HCHF 22.3±1.3, HCHF+PC 17.3±0.7* mg/mm tibial length),
abdominal circumference (HCHF 22.9±0.6, PC 20.2±0.4* cm), visceral fat pad deposition (HCHF
751±48, HCHF+PC 452±31* mg/mm tibial length), infiltration of inflammatory cells into the left
ventricle, collagen deposition in the left ventricle (HCHF 7.8±0.6, HCHF+PC 4.2±0.7* %surface
area), diastolic stiffness (HCHF 28.7±0.8, HCHF+PC 22.4±1.3*), acetylcholine-induced
vasodilatation, plasma CRP concentrations (HCHF 102.3±4.6, HCHF+PC 23.5±9.2 μmol/L),
plasma lipid profile and liver enzymes. Additionally, histological examination of the liver showed
reduced inflammatory cell infiltration, fibrosis and lipids deposition. Our data suggests that purple
carrot improves cardiovascular and hepatic structure and function through anti-inflammatory
effects mediated by its anthocyanin content.
[1] Sun, T., Simon, P.W.,Tanumihardjo, S.A. (2009) Antioxidant phytochemicals and antioxidant capacity of
biofortified carrots ( Daucus carota L.) of various colors. J Agric Food Chem, 57: 4142-4147.
[2] Charron, C.S., Kurilich, A.C., Clevidence, B.A., Simon, P.W., Harrison, D.J., Britz, S.J., Baer, D.J.,Novotny, J.A.
(2009) Bioavailability of anthocyanins from purple carrot juice: effects of acylation and plant matrix. J Agric
Food Chem, 57: 1226-30.
16
PL3
EFSA’s role in novel foods and claims
Wolfgang Gelbmann
Nutrition Unit European Food Safety Authority – EFSA Largo N. Palli 5/A, I-43121 Parma
[email protected]
G
Set up in 2002, the European food Safety Authority (EFSA) provide independent scientific
advice, opinions, information, and technical support for Community legislation and policies in all
fields which have a direct or indirect impact on food and feed safety - including animal health and
welfare and plant protection, and nutrition in relation to Community legislation.
EFSA’s risk assessments provide a sound scientific basis for defining policy-driven legislative
or regulatory measures required to ensure a high level of consumer protection with regards to food
safety. EFSA’s risk assessments are carried out by its Scientific Committee and Scientific Panels,
each composed of external experts and specialised in different aspects of food and feed safety.
The EFSA Scientific Panel on Dietetic Products, Nutrition and Allergies (NDA) is the Panel
dealing with questions on dietetic products, nutrition and allergies, and other associated subjects
such as Novel Food (NF) and Claims.
At the request of the European Commission, the NDA Panel carries out risk assessment of NF
(ingredients), defined as food (ingredients) that have not been used for human consumption to a
significant degree within the Community, further specified by the NF Regulation (EC) Nº 258/9.
Based on an Opinion of the former Scientific Committee for Food, the European Commission has
issued Recommendation 97/618/EC of 29 July 1997 concerning the scientific aspects and the
presentation of information necessary to support applications for the placing on the market of
novel foods and novel food ingredients and the preparation of initial assessment.
Information is required on the specification of the NF, effect of the production process,
history of the organism used as the source of the NF, anticipated intake and extent of use,
previous human exposure to the NF or its source, the nutritional information, microbiological
and toxicological information. The requirements for individual applicants however might differ
depending on the composition and the history of the NF. The assessment of NF concerns only
risk that might be associated with consumption and is not an assessment of the efficacy of the
NF with regard to any claimed benefit.
On the basis of the Novel Food Regulation and the European Commissions Recommendation,
the NDA Panel has so far delivered 42 scientific opinions on the safety assessment of a variety of
NF such as foodstuffs with added phytosterols, substances with antioxidative properties like
synthetic zeaxanthin and lycopene from various sources (Blakeslea trispora, tomato oleoresin,
synthetic), NF from third countries (Chia seeds, Allanblackia seed oil, Noni Juice, Noni leaves),
Enova oil and vegetable oils high in unsaponifiable matter, NF ingredients with a new or
intentionally modified molecular structure (α-cyclodextrin), extracts from Alfalfa, licorice,
fermented black beans, yeast and Lentinus edodes, and such to which a production process has
been applied (Ice structuring proteins).
Regulation (EC) No 1924/2006 harmonises the provisions that relate to nutrition and health
claims and establishes rules governing the Community authorisation of health claims made on
foods. This Regulation requires that scientific substantiation should be the main aspect to be taken
into account for the use of nutrition and health claims and the food business operators using
claims should justify them. A claim should be scientifically substantiated by taking into account
the totality of the available scientific data, and by weighing the evidence. In addition, in order to
bear claims, foods will have to have appropriate nutrient profiles which will be set. This will
enhance the consumers' ability to make informed and meaningful choice. In order to ensure
17
PL3
18
harmonised scientific assessment of these claims, the European Food Safety Authority (EFSA)
should carry out such assessments.
The substantiation of health claims includes the assessment of whether or not the
food/constituent is sufficiently defined and characterised, to assure EFSA that the food/constituent
for which the claim is made is the same as that for which the evidence on efficacy is provided.
Also the claimed effect should be defined and comprise a beneficial physiological effect
(‘beneficial to human health’). Finally, an assessment will be made of whether or not a cause and
effect relationship exists between the consumption of the food/constituent and the claimed effect.
In case the Panel considers that a cause-effect relationship is established, it needs to be assessed
whether the quantity of food/pattern of consumption required to obtain the claimed effect can be
consumed within a balanced diet, whether the proposed wording reflects the scientific evidence
(consumer understanding is not addressed by EFSA), and whether the proposed conditions of use
are appropriate.
In every case, in assessing each food/health relationship that forms the basis of a claim, all the
evidence from the pertinent studies is weighed with respect to its overall strength, consistency and
biological plausibility, taking into account the quality of individual studies and with particular
regard to the target population group and the conditions of use proposed for the claimed effect.
In this presentation an update on the EFSA’s work on claims will be given.
SL5
Tomato organic lycopene: the way to obtain a unique healthy
product
Marcello Salvatore Lenucci1, Leonardo Rescio2, Gabriella Piro1, Giuseppe Dalessandro1
1
Di.S.Te.B.A., Università del Salento, Via prov. Lecce-Monteroni, 73100 Lecce, Italy
2
Pierre S.r.l., S.P. 362 Km 17,650, 73013 Galatina (LE), Italy
[email protected]
Lycopene is a carotenoid found in a wide variety of vegetables and fruits. There are more than
six hundred natural carotenoids, but only twenty of those have been found in human blood and
tissues. Lycopene is the most abundant carotenoid in human blood, followed by β-carotene, lutein
and zeaxanthin [1]. The purpose of this work was to obtain a natural, solvent-free lycopene extract
from red-ripe tomato fruits. This project was developed in three different steps finalised to control
and optimise the entire productive cycle, from fresh tomatoes to the production of organic
lycopene. The first step of this work was the accurate selection, cultivation and production of
organic grade certified tomato cultivars characterised by high-lycopene content, followed by the
biochemical analyses of their antioxidant characteristics [2]. The second step was the set-up of a
process to obtain a freeze-dried tomato matrix suitable for the extraction by supercritical carbon
dioxide (SC-CO2) from the new selected high-lycopene cultivars [3]. The last step was the
optimization of SC-CO2 lycopene extraction from a defined blend of the tomato matrix with a
lipid co-matrix obtained from roughly crushed hazelnut seeds. This procedure allowed the
simultaneous extraction of lycopene from the tomato matrix and oil from hazelnut co-matrix
which improved the solubility of the pigment and increased the yield of the recovered endproduct, according to the Pierre S.r.l. patented co-extraction technology [3-5]. The extract was an
over-saturated solution of lycopene in hazelnut oil (oleoresin). This innovative procedure showed
high lycopene extraction yields (>80%). The extracted oleoresin represents a healthy and high
added-value source of lycopene, suitable for nutraceutical, cosmetic and pharmaceutical purposes.
It was registered as the first organic grade lycopene. This innovative product is characterized by:
1) the absence of toxic impurities such as solvents, pesticides, heavy metals and dioxins; 2) high
natural bioavailability, increased by the presence of the co-extracted natural components from
hazelnuts and tomatoes; 3) high antioxidant activity, due to the presence of tomato carotenoids
and hazelnut tocopherols and phytosterols.
[1] Rao A.V., Tomatoes, Lycopene & Human Health. Preventing Chronic Diseases. Caledonian Science Press,
Badalona (2006).
[2] Lenucci M.S., Cadinu D., Taurino M., Piro G., Dalessandro G. (2006) Antioxidant composition in cherry and
high-pigment tomato cultivars. Journal of Agricultural and Food Chemistry 54: 2606-2613 (2006).
[3] Lenucci M.S., Caccioppola A., Durante M., Serrone L., Leonardo R., Piro G., Dalessandro G. (2010) Optimisation
of biological and physical parameters for lycopene supercritical CO 2 extraction from ordinary and high-pigment
tomato cultivars. Journal of the Science of Food and Agriculture, 90: 1709-1718.
[4] Vasapollo G., Longo L., Rescio L., Ciurlia L. (2004). Innovative supercritical CO2 extraction of lycopene from
tomato in the presence of vegetable oil as co-solvent. Journal of Supercritical Fluids, 29: 87-96.
[5] Ciurlia L., Bleve M., Rescio L. (2009) Supercritical carbon dioxide co-extraction of tomatoes (Lycopersicum
esculentum L.) and hazelnuts (Corylus avellana L.): a new procedure in obtaining a source of natural lycopene.
Journal of Supercritical Fluids, 49: 338-344.
19
SL6
Isoflavone composition of soy and red clover dietary supplements
Franz Bucar, Elvira Knauder
Institute of Pharmaceutical Sciences, Department of Pharmacognosy, University of Graz,
Universitätsplatz 4, A-8010 Graz, Austria
[email protected]
Isoflavones from soybean and red clover are supposed to have beneficial effects by alleviating
postmenopausal symptoms or by reducing cancer risk of hormone dependent cancers. However,
their efficiency and risk-benefit balance is still discussed controversially [1,2].
Isoflavone preparations are usually marketed as nutraceuticals (dietary supplements). Previous
investigations have shown that the proportion of aglycones and glycosides can reveal a wide range
[3] but usually this information is not included in the declaration of the products. However, in
order to estimate the phytoestrogenic activity the amount of isoflavone aglycones (in free and
glycosidated form) in the applied dose should be known.
During the last years by our group a number of isoflavone dietary supplements have been
analysed for their isoflavonoid pattern using HPLC-PDA and LC-PDA-ESI-MS analysis.
Sixteen products contained soy extracts, 4 contained red clover isoflavone extracts and 4 were
manufactured from mixtures of soy and red clover extracts. Only one half of the products
contained the declared amounts ± 15 %. Red clover extracts generally showed a high proportion
of aglycones, i.e. 99.1 – 86.8 % of total amount were present as free aglycones, calculated in
molar concentrations. Biochanin A and formononetin were the major isoflavones in all
investigated red clover supplements. On the other hand, soy products varied to a much greater
extent (5.0 – 98.1 % of total amount as glycosides, mean ± S.D. 78.7 ± 29.8 %, calculated in
molar concentrations) with daidzin, glycitin and genistin as main glycosides whereas acetylated
glycosides usually were present only in minor quantities.
Due to the different absolute amounts and varying ratios of isoflavone aglycones and
glycosides in the investigated products, a declaration of the dose as “isoflavone aglycone
equivalents” seems to be advantageous.
R4
O
R3
R2
O
R1
Compound
R1
R2
R3
R4
Daidzein
OH
H
H
OH
Genistein
OH
OH
H
OH
Glycitein
OH
H
OCH3
OH
Biochanin A
OCH3
OH
H
OH
Formononetin
OCH3
H
H
OH
[1] Dixon RA. (2004) Phytoestrogens. Annu. Rev. Plant Biol.,. 55: 225-261
[2] Zhao E., Mu Q. (2011) Phytoestrogen biological actions on mammalian reproductive system and cancer growth.
Sci. Pharm. 79: 1-20
[3] Schwartz H., Sontag G. (2008) Determination of isoflavones in nutritional supplements by HPLC with
coulometric electrode array detection. Monatsh. Chem., 139: 865-872
20
SL7
Antioxidant and anti-inflammatory properties of a new soluble
extract from grape pomace
Candiracci M1., Castaño A1., SantaMaría C1., Revilla E1., Dominguez-Barragán M1.,
Rodríguez-Morgado B1., García-Antras D1., Carballo M3., Bautista J1. Herrera , M.D2., and
Parrado J1.
1
Departamento de Bioquímica, Biología Molecular. Facultad de Farmacia Universidad de Sevilla.
Departamento de Farmacología. Facultad de Farmacia Universidad de Sevilla.
C/Prof. García Gonzalez 2 , Sevilla 41012
3
CITIUS. Universidad de Sevilla
2
Grape pomace (GP) is a byproduct from wine industry consisting of skins, seeds, and stems. In
order to valorize the GP, we have developed an enzymatic process that give rise to a new product
consisting in a soluble extract enriched in polyphenols (GPE).
Moderate and regular consumption of red wine has been reported to reduce the risk of
cardiovascular diseases [1]. This beneficial effect is attributed, in part, to their high content in
polyphenolic compounds that prevent oxidation of low-density lipoproteins [2]. Thus, we have
tested antioxidative effect of GPE by three different techniques: Phospho molybdate, ORAC and
DDPH. Our results clearly show a dose-dependent antioxidative property of GPE.
On the other hand, it’s widely accepted that inflammation plays a pivotal role in
neurodegenerative process as Parkinson´s or Alzheimer disease. The hallmark of
neuroinflammation is the activation of microglia, but although microglial activation is the brain´s
major defense against immune challenge, activated microglia may also contribute to
neurodegeneration through the release of proinflammatory and/or cytotoxic factors as IL-1E,
TNF-D, NO and reactive oxygen intermediates among others. Thus it can be assumed that a
degree of brain inflammation is required for repair of damaged tissue but excesive inflammation
causes neuronal cells death [3,4]. In this sense, it will be interesting to search for molecules that
could help to control inflammation in the CNS.
Given that flavonoids and tocopherols inhibit release of proinflammatory cytokines after LPSactivation of microglia [5,6] we search whether the GPE exerts an antiinflammtory effect on N13
microglia cells stimulated with LPS. Interestingly our results show that GPE decreases not only
the level of mRNAs of IL-1E, TNF-D and iNOS, but also iNOS protein.
In conclusion our results deeply support that GPE exerts beneficial effect on pathological
conditions related to aged as cardiovascular and neurodegenerative processes.
[1] Renaud S. et al. Lancet 339 : 1523–6, 1992,
[2] Serafini M., et al. J. Nutr. Biochem. 11: 585–90, 2000.
[3] Block M.L. et al. Nature Review Neuroscience 8: 57- 69, 2007.
[4] Park K.W. et al. Exp Mol Med 39 (6) : 812-19, 2007.
[5] Lu X. et al. J. Neuroinflammation 7: 46-60, 2010,
[6] Ha S. K., et al. Neuroscience letters 485(3): 143-7, 2010.
21
PL4
Phenolic compounds : from plants to foods
Véronique Cheynier
INRA, UMR1083 Sciences pour l’Oenologie, place Viala, 34060 Montpellier, France.
Phenolic compounds are a large class of plant secondary metabolites, including phenolic acids,
stilbenes, a variety of flavonoids (e.g. flavonols, flavan-3-ols, anthocyanins), and polymeric
compounds, based on these three groups of compounds. They play a role in plant defence against
biotic and abiotic stress and in attraction processes involved in seed dissemination and pollination.
For example, anthocyanins are the pigments of most red and blue plant organs, while flavonols
protect plant tissues against UV irradiation and proanthocyanidins (PA, i.e. flavan-3-ol oligomers
and polymers, also called condensed tannins) are involved in defence against herbivores, fungi
and viruses. Phenolic compounds are equally important for the quality of plant based foods: they
are responsible for the colour of red fruits, juices and wines and substrates for enzymatic
browning, and are also involved in flavour and taste properties. In particular, astringency is
ascribed to precipitation of salivary proteins by tannins. In addition, phenolic compounds may
contribute to the health benefit associated to dietary consumption of fruits and vegetables.
During food processing and storage, plant phenolics are converted to a variety of derived
compounds, which also affect quality. Understanding the structure of the resulting compounds and
their particular properties, as well as the impact of processes on composition changes is essential
for controlling food quality. Extraction of phenolics into beverages such as juices, wines and teas,
depending on their structure but also on the characteristics of the plant organ, is another important
issue for plant transformation industry which has devoted much effort to the development of new
extraction processes.
While lower molecular weight compounds are quite easily analysed by HPLC hyphenated with
UV-visible spectrophotometry and mass spectrometry, analysis of polymeric compounds remains
a challenge. Indeed, strong interactions of polymeric phenolics with plant cell wall material limit
their extraction. Besides, their polydispersity results in poor resolution and detection, especially in
the case of derived structures. However, approaches combining depolymerisation and small angle
X-ray scattering (SAXS) provided new insights on the structure of PA oxidation products
(Poncet-Legrand et al., 2010).
Current knowledge on phenolic reactions and resulting structures and their impact on colour,
taste, and health properties will be presented. Changes taking place during transformation
processes will be illustrated in the case of wine-making, with special emphasis on extraction
technologies and oxidation reactions.
Poncet-Legrand C, Cabane B, Bautista-Ortın AB, Carrillo S, Fulcrand H, Perez J, Vernhet A (2010),
Biomacromolecules, 11, 2376–2386.
22
SL8
Polyphenols-rich extracts obtained from Prunus avium and opuntia
ficus indica as natural anti-inflammatory modulators for
inflammatory bowel diseases.
S. Nunes1, A.T. Serra1,2, M. R. Bronze1,2,3, Catarina M.M. Duarte1,2, A.A. Matias 1,2,3*
1
Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Avenida da República,
Estação Agronómica Nacional, 2780-157 Oeiras, Portugal;
2
Instituto de Biologia Experimental e Tecnológica, Avenida da República, Quinta-do-Marquês, Estação
Agronómica Nacional, Apartado 12, 2781-901 Oeiras, Portugal;
3
iMED, Faculdade de Farmácia da Universidade de Lisboa, Av. das Forças Armadas, 1649-019, Lisboa,
Portugal.
[email protected]
There are evidences that the regular consumption of polyphenols in early life could reduce or
delay Inflammatory Bowel Diseases (IBD) development in humans. However, IBD are usually
treated with pharmacological agents, which could produce secondary effects. It is therefore,
urgent to find new and safe compounds able to contribute to prevention or even to treatment of
these inflammatory diseases.
In the present study we investigated the preventive effectiveness of polyphenolic-rich extracts
obtained from Sweet cherries (Prunus avium) and cactus pear (Opuntia ficus indica) on
inflammatory expression in intestinal epithelial human cell lines (caco-2) induced with specific
stimuli (LPS and TNF-α combination).
The selected fruits were processed in order to prepare the polyphenols-rich extracts (PRE) and
further characterized in terms of polyphenolic compounds, namely anthocyanins and betalains
using colorimetric assays and chromatographic techniques.
The inflammatory response of the PRE’s were assessed using intestinal epithelial cells (seeded
in transwell™ plates) after pro-inflammatory stimulus (LPS and TNF-α cocktail). The secretion of
pro-inflammatory markers (NO excretion, IL-8 and IL-1β secretion) was assayed in the media
from both apical and basolateral compartments for inflamed cells as compared to the control.
The monolayer integrity and permeability were assessed by the determination of transepithelial
electrical resistance values.
When Caco-2 cells were simultaneously incubated with PREs and the inflammatory stimuli, no
significant change in cytotoxicity and TEER values was observed.
In inflamed cells, the incubation with Cherry extract promotes the maintenance of epithelium
barrier integrity of the inflamed cells and also reduced significantly the IL- 8 secretion and NO
excretion.
Cactus pear extract provoked a slightly decrease in the IL-8 over-secretion.
In conclusion, this study provides evidence that cherry and cactus pear extracts could
downregulate the inflammatory response in inflamed intestinal epithelial cells.
Further investigation on this subject are in progress.
[1] Kelley DS, Rasooly R, Jacob RA, Kader AA, Mackey BE. Consumption of Bing sweet cherries lowers circulating
concentrations of inflammation markers in healthy men and women. J Nutr. 2006, 136:981–6.
[2] Ferretti G, Bacchetti T, Belleggia A, Neri D, Cherry Antioxidants: From Farm to Table, Molecules 2010, 15,
6993-7005.
23
SL9
Polyphenols from red wine exert potential beneficial effects to human
health
Thea Magronea, Sergio Fontanab, Emilio Jirilloa
a
Department of Basic Medical Sciences, University of Bari, P.zza G. Cesare 11, 70125 Bari, Italy
Farmalabor Srl, via Oberdan 52, 70053 Canosa di Puglia, Italy
[email protected]
b
Polyphenols (flavonoids and resveratrol) are highly distributed in the plant kingdom and their
biological properties have been extensively studied [1].
Over the past few years, we have investigated the in vitro and in vivo effects of polyphenols
from red grapes. In particular, we have provided evidence that polyphenols isolated from Vitis
Vinifera Negroamaro are able to induce release of nitric oxide from human monocytes and to
modulate either innate or adaptive human immune response [2]. In this respect, these polyphenols
are able to maintain the immunological homeostasis, thus balancing the
inflammatory/antinflammatory pathway [3]. In addition, polyphenols interfere with the activation
of the NF-NB pathway induced by endotoxins, thus attenuating the proinflammatory wave of
cytokines in clinical settings, such as sepsis [4].
Similar effects have been obtained using fermented grape mark (FGM) from Vitis Vinifera
Negroamaro and Koshu. Quite interestingly, Koshu-FGM has the ability to attenuate murine
experimental colitis in vivo [5]. This finding is also supported by the evidence that Koshu-FGM
reduces the secretion of interleukin 1E and tumor necrosis factor D from colitis mice homogenised
colon [6]. These data are also supported by in vitro results which clearly demonstrated the ability
of both Koshu-FGM and Negroamaro-FGM to induce T regulatory cells (CD4+CD25+FoxP3+) in
the human system. Finally, both FGM are able to mitigate allergic reactions inhibiting mast cell
and basophil degranulation, release of IgE, infiltration of eosinophils in the respiratory system and
passive cutaneous anaphylaxis [7,8].
Clinical trials in patients with allergic disease and inflammatory bowel disease are in progress.
[1] Magrone T., Jirillo E. (2010) Polyphenols from red wine are potent modulators of innate and adaptive immune
responsiveness. Proc Nutr Soc, 69: 279-285
[2] Magrone T., Tafaro A., Jirillo F., Panaro MA., Cuzzuol P., Cuzzuol AC., Pugliese V., Amati L., Jirillo E., Covelli
V. (2007) Red wine consumption and prevention of atherosclerosis: an in vitro model using human peripheral
blood mononuclear cells. Curr Pharm Des, 13: 3718-3725
[3] Magrone T., Tafaro A., Jirillo F., Amati L., Jirillo E., Covelli V. (2008) Elicitation of immune responsiveness
against antigenic challenge in age-related diseases: effects of red wine polyphenols. Curr Pharm Des, 14: 27492757
[4] Magrone T., Panaro MA., Jirillo E., Covelli V. (2008) Molecular effects elicited in vitro by red wine on human
healthy peripheral blood mononuclear cells: potential therapeutical application of polyphenols to diet-related
chronic diseases. Curr Pharm Des, 14: 2758-2766
[5] Magrone T., Kumazawa Y., Jirillo E. (2011) Natural Antioxidants and their Derivatives: Biology and Clinical
Application. Curr Top Med Chem, [Epub ahead of print]
[6] Magrone T., Jirillo E. (2011) Potential Application of Dietary Polyphenols from Red Wine to Attaining Healthy
Ageing. Curr Top Med Chem, [Epub ahead of print]
[7] Tominaga T., Kawaguchi K., Kanesaka M., Kawauchi H., Jirillo E., Kumazawa Y. (2010) Suppression of type-I
allergic responses by oral administration of grape marc fermented with Lactobacillus plantarum.
Immunopharmacol Immunotoxicol, 32: 593-599
[8] Kaneko M., Kanesaka M., Yoneyama M., Tominaga T., Jirillo E., Kumazawa Y. (2010) Inhibitory effects of
fermented grape marc from Vitis vinifera Negroamaro on antigen-induced degranulation. Immunopharmacol
Immunotoxicol, 32: 454-461.
24
SL10
Differential antioxidant & anti-amyloid effects of a wild blueberry
polyphenol-rich extract: in vitro & in vivo studies
Magdalini A. Papandreou1, Maria Tsachaki2, Spiros Efthimiopoulos2, Dorothy KlimisZacas3, Marigoula Margarity1, Fotini N. Lamari4
1
Dept. Biol., Univ. Patras, 2Dept. Biol., Univ. Athens, 3Dept. Food Science & Human Nutrition, Univ.
Maine, Orono, ME 04469, USA, 4Lab. Pharmacogn. & Chem. Natural Products, Dept. Pharmacy, Univ.
Patras, Greece.
[email protected]
Blueberries, a rich source of natural anthocyanin antioxidants, have demonstrated a broad
spectrum of beneficial biomedical functions. In this study, the effects of a daily (7-day)
intraperitoneal (i.p.) administration of a polyphenol-rich extract (PrB) of Vaccinium angustifolium
(wild blueberries), on cognitive functions were examined in healthy adult (4 months old), male
Balb-c mice (n=8/group) by passive avoidance test. Brain oxidative markers, caspase-3 and
acetylcholinesterase (AChE) activity were determined in whole brain homogenates (minus
cerebellum). Results showed that PrB-treated mice exhibited a significant improvement in
learning and memory, accompanied by reduced lipid peroxidation products, higher total brain
antioxidant activity and reduced caspase-3 activity. Furthermore, salt- and detergent-soluble
AChE activity was significantly decreased. Thus, the significant cognitive enhancement observed
in adult mice after short-term i.p. supplementation with the blueberry extract, is closely related to
higher brain antioxidant properties and inhibition of AChE activity. To further delineate the
neuroprotective mechanisms, we investigated the antioxidant effects of PrB in three different cell
lines against H2O2-induced oxidative damage. PrB suppresses H2O2-initiated SH-SY5Y
intracellular cell death (MTT assay) and oxidation (DCF assay). Moderate effects were observed
on CHOAPP770 cells, while further damage on HEK293 cells was shown after co-treatment with
250 μΜ H2O2 and PrB. Since Amyloid Precursor Protein (APP) altered metabolism, Aβoverproduction/aggregation are key pathological hallmarks in AD, we further studied the effects
of PrB on Aβ-fibrillogenesis, in vitro, utilizing the thioflavine T assay, and on APP-metabolism in
CHOAPP770 cells. Blueberry polyphenols inhibited Aβ-aggregation in a time-dependent manner,
while in CHOAPP770 cells, no alterations in APP metabolism were observed as assessed by western
blot. Taken together, our results suggest that blueberry polyphenols exhibit antioxidant and/or
pro-oxidant properties according to the cellular environment inhibit Aβ-fibrillogenesis in vitro but
have no effect on APP metabolism.
25
PL5
New trends in the analysis of natural products for the detailed
metabolite profiling of crude extracts
Jean-Luc Wolfender
School of Pharmaceutical Sciences, EPGL, University of Geneva, University of Lausanne, 30, quai ErnestAnsermet, CH-1211 Geneva, Switzerland
Plant natural products (NPs) have provided the inspiration for a large number of the active
ingredients in medicine. Their success is probably due to the effects of evolutionary pressure to
create biologically active molecules. The large chemical space occupied by NPs is directly linked
to a high variability of their intrinsic physicochemical properties that render their separation,
detection and characterization challenging. In order to identify all these metabolites, crude plant
extract profiling is essential. This is a challenging task that requires methods providing high
chromatographic resolution for detailed profiling or high throughput for rapid quantification or
fingerprinting analysis. Furthermore these methods should give on-line spectroscopic information
for the identification of each individual metabolites for dereplication purposes. In this respect
hyphenated techniques such as LC-MS and LC-NMR have played a key role over the last three
decades. In phytochemical analysis, the recent introduction of Ultra High Pressure Liquid
Chromatography (UHPLC) systems operating at very high pressures and using sub-2 μm packing
columns have allowed a remarkable decrease in analysis time and increase in peak capacity,
sensitivity and reproducibility compared to conventional HPLC. In complement to this powerful
chromatographic method, the introduction of benchtop time–of-flight (TOF)-MS instruments
provide sensitive detection and high MS resolution. For de-novo identification of NPs on-line LCNMR, introduced in the early 90’s, has evolved towards sensitive at-line microflow NMR
approaches combining pre-concentration of the LC peaks of interest prior to NMR measurement
(LC-SPE-NMR, capillary NMR: CapNMR). With such methods structure determination of
targeted compounds at the low microgram level is possible and complement the search in MS and
chemotaxonomic data bases for dereplication. The potential and limitations as well as some new
trends in the development of UHPLC-MS and micro NMR will be discussed. In particular
examples related to metabolomics and de novo biomarker identification will be presented.
26
Oral Presentations – International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September
SL11
Metabolomic profiling and fingerprinting: new effective approach
for plant characterisation and authentication
Vera Schulzova, Hana Novotna, Lukas Vaclavik, Anna Hurajova, Jana Hajslova.
Institute of Chemical Technology, Department of Food Chemistry and Analysis, Technická 3, 16628,
Prague 6, Czech Republic
[email protected]
Food and medicinal plants are a good source of different phytonutrients and nutraceuticals.
Content of biological active compounds such as vitamins, carotenoids, phenols, flavonoids,
terpenoids etc. differs in individual varieties. The geographic origin and climatic conditions also
influence the content of phytonutriets significantly. Until now, a wide range of analytical
approaches has been tested for control of quality and authenticity. In our most recent research
projects, we have implemented a novel approach based on ambient mass spectrometry [1]. Using
the Direct Analysis in Real Time (DART) ion source hyphenated with a high resolution Time of
Flight Mass Spectrometer (TOFMS), metabolomic fingerprint (mass spectrum) can be obtained in
a few seconds. Metabolomics may be used either for ´fingerprinting´ of samples to perform
comparative analyses aimed at detection of differences or for ´profiling´ in which individual
sample components (both primary and secondary metabolites) are identified for further
chemometric assessment. Using relevant statistical tools, locality, crop years, varieties, and way
of farming could be separated. In any case, relevant databases of metabolomic profiles of well
characterised samples should be established to have a larger data set for statistical processing thus
obtaining recognition and prediction models.
In this study, DART-TOFMS analysis was used for characterisation and authentication of
different plant materials. The results of several case studies concerned with authentication of food
and medicinal plants (flax seeds, potato tubers, tomatoes, apples, peppers, rose hip, lemon balm,
peppermint, stinging nettle etc.) will be presented.
This study was carried out within the project MSM 6046137305 and within specific university
research MSMT no. 21/2010 supported by Ministry of Education, Youth and Sports of the Czech
Republic.
[1] Hajslova J., Cajka T., Vaclavik L. (2011) Challenging applications offered by direct analysis in real time (DART)
in food-quality and safety analysis. Trend Anal. Chem. 30, 204-218
27
SL12
Estrous cycle in female rats influences the neurochemical and
neurobehavioural effects of carvacrol
Margherita Zottia, Marilena Colaiannaa, Maria Grazia Morgesea, Lucia Ciuffredaa, Paolo
Tuccia, Pinarosa Avatob and Luigia Trabacea
a
Dipartimento di Scienze Biomediche, Università, V.le Pinto 1, 71121 Foggia, Italy
Dipartimento Farmaco-Chimico, Università, Via Orabona 4, 70125 Bari, Italy
[email protected]
b
Carvacrol, the major natural terpenoid constituent of oregano [1] and thyme essential oils is a
small and lipophilic compound likely to readily cross the blood-brain barrier. It has been approved
for food use and included in the chemical flavorings list, although little is known on its safety.
The use of plant extracts is relatively high among women, especially middle-aged, and even
more common among women with depression [2,3]. Some authors have reported antimicrobial,
anticancer and antioxidant properties for this terpene, hovewer its possible action in the
mammalian central nervous system has not been extensively investigated.
The aim of this study was to evaluate the neurochemical and behavioural effects of carvacrol
on physiological and endocrine profile of female rats in proestrus and diestrus phases,
characterized by high and low estrogen levels, respectively. In particular, serotonin and its
metabolite tissue content in prefrontal cortex and nucleus accumbens, 2h after carvacrol
administration (0.15 and 0.45 g/Kg p.o.), were measured. Effects were examined in behavioural
tests for alterations in motor activity and depression-related behaviours.
While in proestrus carvacrol reduced serotonin and related metabolite levels in both brain
areas, no effects were observed in diestrus phase. Only in proestrus phase, carvacrol induced a
depressive-like behaviour in the forced swimming test (FST), without changes in ambulation. The
improvement of performance in FST after subchronic treatment with fluoxetine (s.c., 20 mg/kg),
suggested the specific involvement of serotonergic system in the depressive-like effects of
carvacrol. Regarding endocrine profile, we found that in proestrus phase, carvacrol reduced
estradiol levels.
Taken together our data suggest that carvacrol has an estrous-stage specific effect on
depressive-behaviours and endocrine parameters involving serotonergic system.
[1] Morone-Fortunato, I., Avato P. (2008) Plant Cell Tiss Organ Cult 93, 139-149
[2] Brett, K. M., and Keenan, N. L. (2007). Menopause, New York
[3] Barnes, P. M., Powell-Griner, E., McFann, K., and Nahin, R. L. (2004). Advance data, United States
28
PL6
Breeding for improved flavour and health
Yury Tikunova,b, Sjaak van Heusdena,b, Jos Molthoffa, Henri van de Geestb, Ana-Rosa
Ballesterc, Irene Romeroa,b, Antonio Granelld, Ric de Vosa,b, Arnaud Bovya,b
a
Plant Research International, P.O. Box 16, 6700 AA, Wageningen, Netherlands
Centre for Biosystems Genomics, P.O. Box 98, 6700PB, Wageningen, Netherlands
c
Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas
(IATA-CSIC), Valencia, Spain
d
Instituto de Biología Molecular y Celular de Plantas (IBMCP-CSIC), Universidad Politécnica de
Valencia (UPV), Valencia, Spain
b
Traditionally the most important targets in fruit and vegetable breeding are producer traits,
such as yield, disease resistance and shelf life. In the past decade, however, the importance of
consumer-driven quality traits, such as flavour, nutritional value and appearance have been
increasingly recognised. This is not only due to growing consumer demands for more healthy
food with an excellent quality, but also to the availability of various omics tools, which make it
possible to study complex, multifactorial traits, such as flavour and health. These quality traits are
to a large extent determined by the phytochemical composition of our food products. In order to
improve the consumer-quality of food crops, we aim to explore and utilise the natural diversity for
flavour- and health-related metabolites. Our general strategy involves the elucidation of
biochemical pathways underlying important quality traits using a genetical genomics approach
and the isolation of the responsible genes underlying quality-related mQTLs. Tomato is one of the
most important vegetable crops worldwide and an important model for fleshy fruits. Cultivated
tomato has a very narrow genetic base, due to severe genetic bottlenecks experienced during
domestication. In contrast, a lot of genetic diversity can be found in wild relatives of tomato,
which are therefore used as novel sources of important agronomic and quality traits. In particular
the use of introgression line (IL) populations, in which defined chromosomal segments of wild
species have been introgressed into an elite tomato background has been very successful. We
analysed a diverse collection of both cultivated and wild tomato germplasm for variation in
colour-, health- and flavour-related metabolites, in order to elucidate the biochemical and
molecular mechanisms underlying these quality traits. In this lecture, several examples of this
approach will be discussed:
1. By screening a Solanum chmielewskii IL population we elucidated the biochemical and
molecular basis underlying pink-coloured tomatoes, the predominant tomato type in Asia.
2. This population also revealed a large variation in health-related metabolites, such as
carotenoids, flavonoids and alkaloids. We are currently using a candidate gene approach to
clone the underlying genes.
3. A set 94 commercial tomato cultivars was evaluated for taste using trained sensory panels
and metabolite profiling. We observed a close correlation between the presence of
phenylpropanoid-derived volatiles and a “smoky” flavour. The release of these volatiles is
determined by their storage form as glycoside conjugate. By a combined genetic,
metabolomic and transcriptomic approach the underlying glycosyltransferase gene was
isolated.
This research was funded by the Dutch NWO genomics initiative Centre for Biosystems Genomics and the
EU project EUSOL (PL-016214).
29
SL13
Application of purine derivative LGR-1814 improves functional
properties of field-grown lettuce (Lactuca sativa)
Jiri Gruza,b and Lukas Spichala,b
a
Laboratory of Growth Regulators, Institute of Experimental Botany AS CR, Slechtitelu 11, CZ 783 71
Olomouc, Czech Republic
b
Centre of the Region Hana for Biotechnological and Agricultural Research, Faculty of Science, Palacky
University, Slechtitelu 11, CZ 783 71 Olomouc, Czech Republic
Lettuce is one of the most consumed vegetables in the world. It is particularly popular in
Argentina, Australia, Canada, Chile, New Zealand, United States of America and Uruguay, where
the mean consumption is 30.6 g person-1 day-1 according to International Estimated Daily Intake
published by WHO in 2006. Considering the phytochemical potential of lettuce, the manipulation
of endogenous metabolite levels is an advisable strategy to improve its functional properties. The
increase in specific bioactive components is also a competitive factor which may increase sales
and performance of food companies. In our study, we have found that single application of purine
derivative LGR-1814 improved functional properties of field-grown lettuce (Lactuca sativa).
ORAC antioxidant capacities of the lettuce plants treated with 10 and 50 μM LGR-1814 were
significantly increased by 21 and 32%, respectively. A similar trend was observed when TPC was
determined. The increase in antioxidant capacity was correlated with the UPLC-MS/MS
determined content of endogenous hydroxycinnamic acids (HCA), including ferulic and 4coumaric acids, while the content of hydroxybenzoic acids (HBA) was not altered. The fact that
levels of only HCA but not HBA were increased in the treated plants is in accordance with the
observed induction of PAL activity. The application of LGR-1814 did not result into any
significant reduction in yield which is a common side effect of stress and stress-related
accumulation of phenolic compounds. In accordance, the levels of malondialdehyde remained
unaffected, confirming that lettuce plants were not stressed by the treatment. Our data indicate
that LGR-1814 could find interesting applications in improving functional properties, shelf life,
and stress protection of edible plants.
30
SL14
Comparison of triterpenoid content of Vaccinium vitis-idaea berries
and leaves collected from natural forest habitats in Finland and
Poland
Anna Szakiela, Cezary Pączkowskia, Heini Koivuniemib, Satu Huttunenb
a
Department of Plant Biochemistry, Faculty of Biology, University of Warsaw, ul. Miecznikowa 1, 02-096
Warszawa, Poland
b
Botany Division, Department of Biology, University of Oulu, P.O. Box 3000, 90014 Oulu, Finland
[email protected]
Vaccinium vitis-idaea (cowberry, lingonberry), known for its edible berries and leaves used in
herbal medicine, is a rich source of dietary nutraceutics, phenolics and triterpenoids.
Plant material (berries and two types of foliage: young, immature, current-year leaves and old,
previous-year leaves) was collected in natural forest habitats in Finland (65°066 N; 25°458 E) and
Poland (52°455 N, 21°332 E) in late August and December 2010. Samples were dried, powdered
and extracted in Soxhlet apparatus with diethyl ether. Extracts were fractionated by CC and TLC
chromatography on silica gel, and analysed by GC-MS/FID.
Main triterpenoid profile of V. vitis-idaea of both origin consisted of 12 compounds, namely:
α- and β-amyrins, betulinol, erythrodiol, fernenol, friedelin, lupeol, svertenol, taraxasterol, uvaol,
oleanolic and ursolic acids. Other notable constituents were β–sitosterol, cycloartanol and
stigmasta-3,5-dien-7-one. Within each class of triterpenes, compounds based on the ursan
skeleton predominated, with α–amyrin being more abundant than β–amyrin, and ursolic acid and
uvaol found at higher levels than oleanolic acid and erythrodiol. However, some remarkable
qualitative and quantitative differences between berries and leaves obtained from the two
locations were found.
The content of triterpene acids was higher in Finnish (approx. 9 mg·g-1 d.w.) than in Polish
berries (8 mg·g-1 d.w.) whereas triterpenols were more than twice more abundant in Polish than in
Finnish fruits (1 and 0.4 mg·g-1 d.w., respectively). The principal triterpenol in Finnish berries
was fernenol whereas in Polish berries α–amyrin was predominating within this class of
compounds. Additionally, betulinol and taraxasterol were found in small amounts in Polish
berries, whilst they were not detected in Finnish fruits. Berries of both origin contained
comparable amounts of free β-sitosterol (1.3 mg·g-1 d.w.) but its esterified form was twice more
abundant in Finnish berries. In turn, the amounts of cycloartanol and stigmastadienone were
markedly higher in Polish fruits.
In leaves, the total content and profile of triterpenoids varied significantly not only depending
on the place of plant origin, but also on leaf age. Generally, in August young leaves contained less
triterpenes (4.6 and 3 mg in Finnish and Polish leaves, respectively) than old leaves (6.2 and 6.1
mg). These differences were less significant in December. Triterpene acids were again the most
abundant constituents (2.4 and 4.4 mg in young and old Finnish leaves, 3 and 6.1 mg in Polish
leaves). The most remarkable difference in triterpene profile concerned the occurrence of fernenol
and taraxasterol, the first occurring as the main triterpenol in Finnish, the latter in Polish leaves.
Acknowledgement. This research was supported by POIG.02.02.00-14-024/08-00.
31
PL7
NMR metabolomics: advantages and disadvantages for studying
bioactive components in food
Søren Balling Engelsen and Francesco Savorani
Department of Food Science, Faculty of Life Sciences, University of Copenhagen, Rolighedsvej 30, DK1958 Frederiksberg C, Denmark
[email protected]
Metabolomics is crucial for understanding human health and nutrition. Many lifestyle related
problems are in reality metabolic issues: a mismatch between what is ingested and the needs of
the organism. Food health and safety are areas reaping the benefits of metabolomics. Food is
eaten as a whole and therefore the combined effect of components has to be taken into account
when assessing the health and safety aspect of food and food processes.
In humans and animals, biologically relevant samples can routinely be obtained from blood,
small tissue biopsies, saliva, urine and faeces. Metabolomic profiling seeks to determine as many
chemical (and physical) features as possible and to identify metabolites or patterns of metabolites
that discriminate samples from treated individuals and controls [1].
The two dominant analytical platforms in metabolomics are Liquid Chromatography-Mass
Spectrometry (LC-MS) and NMR spectroscopy, which have quite complementary properties.
While LC-MS has much higher sensitivity NMR is inherently quantitative, un-biased and nondestructive and has unique properties for measuring fx lipoproteins and chylomicrons in blood [2].
The bottleneck for exploiting the powerful new analytical technologies has become the
bioinformatics analysis of the overwhelming amount of data. The combination of NMR
spectroscopy with chemometrics has proven to have a tremendous potential for metabonomics [3].
In this approach the multivariate analytical results are compared across large cohorts, in for
example intervention studies, for metabolites and/or patterns of metabolites that are able to isolate
the effect of the intervention. Chemometrics have also been useful in finding solutions to
imperfect analytical methodology that are prone to alignment, normalization and other
standardization problems [4, 5] as well as for advanced analysis aimed at exploiting the
experimental design and the data structure [6, 7].
This paper tries to sum up the current status of NMR metabonomics for studying bioactive
substances in food and their effect in the human body.
[1] Fiehn O. (2009), Metabolomics - the link between genotypes and phenotypes, Plant Molecular Biology, 48: 155171.
[2] Savorani F., Kristensen M., Larsen F.H., Astrup A., Engelsen S.B. (2010), High throughput prediction of
chylomicron triglycerides in human plasma by nuclear magnetic resonance and chemometrics, Metabolism &
Nutrition, 7:43.
[3] Nicholson J.K., Lindon J.C., Holmes E. (1999), 'Metabonomics': understanding the metabolic responses of living
systems to pathophysiological stimuli via multivariate statistical analysis of biological NMR spectroscopic data,
Xenobiotica, 29: 1181-1189.
[4] Savorani F., Tomasi G., Engelsen S.B. (2010), icoshift: a versatile tool for the rapid alignment of 1D NMR
spectra, Journal of Magnetic Resonance, 202:190-202.
[5] Rasmussen L.G., Savorani F., Larsen T.M., Dragsted L.O., Astrup A., Engelsen S.B. (2011), Standardization of
factors that influence human urine metabolomics, Metabolomics (2011), 7:71-83.
[6] van Velzen E.J.J., Westerhuis J.A., van Duynhoven J.P.M., van Dorsten F.A., Hoefsloot H.C.J., Jacobs D.M.,
Smit S., Draijer R., Kroner C.I. , Smilde A.K. (2008), Multilevel data analysis of a crossover designed human
nutritional intervention study, Journal of Proteome Research, 7:4483-4491.
[7] Bro R., Viereck N., Toft M., Toft H., Hansen P.I., Engelsen S.B. (2010), Mathematical chromatography solves the
cocktail party effect in mixtures using 2D spectra and PARAFAC, Trends in Analytical Chemistry, 29: 281-284.
32
SL15
Formation of astragaloside IV from acylated astragalosides during
extraction of Astragali radix
Monschein Ma, Ardjomand-Woelkart Ka, Wolf Ia, Rieder Jb, Bauer Ra
a
Institute of Pharmaceutical Sciences, Department of Pharmacognosy, Karl-Franzens-University Graz,
Universitaetsplatz 4, 8010 Graz, Austria
b
Bavarian State Research Center for Agriculture, Dept. AQU2, Lange Point 6, 85354 Freising, Germany
[email protected]
Astragaloside IV (AGS-IV), a cycloartane-type triterpene glycoside, is used as an active marker
for the quality control of Astragali Radix (Huangqi; Astragalus mongholicus Bunge,
Leguminosae) in Chinese Medicine [1]. Recently, when astragaloside malonates have been
identified, it has already been argued, that qualitative and quantitative analysis of astragaloside IV
may not give the correct information about the saponin profile of Astragalus roots [2]. We now
have confirmed by HPLC-ELSD (HPLC-Evaporative light scattering detection) analysis that
AGS-IV is formed in Astragalus extracts like astragaloside I and II by hydrolysis during
extraction. Therefore, for consistent assay results fully implemented hydrolysis with Ammonia R
is essential, and the extraction methods of the pharmacopoeias [3] have to be re-evaluated and
optimized.
Astragaloside IV
[1] The Chinese Pharmacopoeia (Beijing, China) Vol. 1 (2005) 249.
[2] Chu C, et al., J Sep Sci. 33(4-5): 570-81 (2010).
[3] European Pharmacopoeia Ph.Eur. 7.0
33
SL16
Sesquiterpenoids from aerial parts of Lactuca aculeata
Klaudia Michalska, Wanda Kisiel
Department of Phytochemistry, Institute of Pharmacology, Polish Academy of Sciences, 31-343 Krakow,
Poland
[email protected]
The wild lettuce Lactuca aculeata Boiss. (Asteraceae, tribe Cichorieae) is distributed in the
Near East and the Anatolian plateau. This species can be easily hybridized with Lactuca sativa L.
and thus occupy the primary gene pool for cultivated lettuce improvement. Sesquiterpene
lactones, including the guaianolide lactucin and its derivatives as well as the germacranolide
lactuside A, are particularly characteristic constituents of Lactuca species and their metabolic
function has been strongly implicated in defense against diseases and herbivorous insects. We
have recently confirmed the reputed analgesic and sedative activities of lactucin and its
derivatives in mice [1]. Moreover, biological studies have shown that 8-deoxylactucin downregulated cyclooxygenase-2 protein expression and nitric oxide production, and inhibited DNA
binding of the central transcription factor NFκB, promoting the expression of multiple genes in
response to inflammatory stimulators [2,3]. These data support some Lactuca species as sources
of functional food ingredients with anti-inflammatory properties.
Our previous phytochemical study (by HPLC) on variation of seven sesquiterpene lactones in
L. aculeata natural populations has shown that this plant is rich in 8-deoxylactucin (up to 1% of
dry wt.) [4].
The present study deals with the isolation of ten known sesquiterpene lactones from aerial parts
of L. aculeata. These compounds were isolated by silica gel column chromatography followed by
preparative TLC and/or semipreparative HPLC, and were characterized by spectroscopic methods
as 8-deoxylactucin, jacquinelin, 9α-hydroxyzaluzanin C and its 11β, 13-dihydroderivative,
vernoflexuoside and its 11β,13-dihydroderivative, ixerin F, macrocliniside A, crepidiaside B, and
lactuside A. In addition, a new lactucin-type guaianolide was isolated and its structure was
determined as 11β-hydroxycrepidiaside B on the basis of HR ESIMS, 1H and 13C NMR spectra
and 2D NMR techniques.
[1] Wesolowska A, Nikiforuk A, Michalska K, Kisiel W, Chojnacka-Wojcik E (2006) Analgesic and sedative
activities of lactucin and some lactucin-like guaianolides in mice. J. Ethnopharmacol. 107: 254-258
[2] Cavin C, Delannoy M, Malnoe A, Debefve E, Touche A, Courtois D, Schilter B (2005) Inhibition of the
expression and activity of cyclooxygenase-2 by chicory extract. Biochem. Biophys. Res. Commun. 327: 742- 749
[3] Ripoll C, Schmidt BM, Ilic N, Poulev A, Dey M, Kurmukov AG, Raskin I (2007) Anti-inflammatory effects
of a sesquiterpene lactone extract from chicory (Cichorium intybus L.) roots. Nat. Prod. Commun. 2:717-722
[4] Beharav A, Ben-David R, Malarz J, Stojakowska A, Michalska K, Dolezalowa I, Lebeda A, Kisiel W (2010)
Variation of sesquiterpene lactones in Lactuca aculeata natural populations from Israel, Jordan and Turkey.
Biochem. Syst. Ecol. 38: 602-611.
34
SL17
The volatile fraction of herbal teas
Christine Tschiggerl and Franz Bucar
Institute of Pharmaceutical Sciences, Department of Pharmacognosy, Karl-Franzens University Graz,
Universitätsplatz 4/1, 8010 Graz, Austria
[email protected], [email protected]
Herbal teas or tisanes are among the most commonly consumed beverages all over the world.
Often the beneficial health effects of these teas are mainly attributed to their volatiles. Results on
the quantitative and qualitative changes of the volatiles during the tea brewing process are scarce
[1-3]. Therefore the volatile fraction of rosemary- (Rosmarinus officinalis L., Lamiaceae),
lavender- (Lavendula angustifolia Mill., Lamiaceae), fennel- (Foeniculum vulgare Mill. subsp.
vulgare var. vulgare (Mill.) Thellung, Apiaceae), chamomile- (Matricaria recutita L., Asteraceae)
and thyme infusion (Thymus vulgaris L., Lamiaceae) has been investigated [4-6]. Additionally,
the influence of saponin plants (Primula veris L./P. elatior L. and Glycyrrhiza glabra L.) on the
volatile fraction in thyme herbal teas has been analyzed [6]. The genuine essential oils of the
relevant plant material were compared with the volatile fraction of their infusion extract. The
volatiles were obtained by two different methods, hydrodistillation and solid phase extraction.
Qualitative and semiquantitative analyses were performed by GC-MS. Some individual
compounds of the extracts obtained by SPE were quantified by internal standard method.
Volumetric quantification of the essential oil yield of the plant materials was performed by
hydrodistillation according to the European Pharmacopoeia. Analogically, the amount of volatile
fraction of the infusions was determined.
The results of our investigations can be summarized as follows:
1. Comparison of the chemical classes (hydrocarbons, oxides, alcohols/ethers,
aldehydes/ketones, esters) of the essential oils and of the volatile fraction of the infusion extracts
resulted in a different profile of chemical classes, which could be explained by the octanol-water
partition coefficient and the boiling point of the individual compounds. In all tea samples
significant losses of hydrocarbons could be recorded.
2. Physicochemical interactions between saponins and volatiles (thymol) exist. An influence on
the percentage distribution of chemical classes in the volatile fraction of the combined infusion
extracts (thyme and cowslip/liquorice) was observed.
3. About 15% to almost 40% of the initial essential oil could be extracted by infusions,
depending on the physicochemical nature of the constituents of the essential oil.
4. Hydrodistillation and solid phase extraction are in most cases comparable methods for
determining volatiles in the infusion extracts. Exceptions are herbs where reactions of the volatile
compounds occur during hydrodistillation processes as in case of chamomile.
5. Semiquantitative peak area% method is in most cases adequate to make conclusions about
transfer rates of individual compounds and the composition of the volatile fraction of tisanes.
[1] Carnat A., Carnat AP., Fraisse D., Lamaison JL. (1999) The aromatic and polyphenolic composition of lemon verbena tea.
Fitoterapia, 70: 44-49
[2] Carnat A., Carnat AP., Fraisse D., Ricoux L., Lamaison JL. (2004) The aromatic and polyphenolic composition of roman
camomile tea. Fitoterapia, 75: 32-38
[3] Bilia AR., Flamini G., Taglioli V., Morelli I., Vincieri FF. (2002) GC-MS analysis of essential oil of some commercial fennel
teas. Food Chem, 76: 307-310
[4] Tschiggerl C., Bucar F. (2010) Investigation of the volatile fraction of rosemary infusion extracts. Sci Pharm, 78: 483-492
[5] Tschiggerl C., Bucar F. (2010) Volatile fraction of lavender and bitter fennel infusion extracts. Nat Prod Commun, 9: 14311436.
35
PL8
Phenolic nutritional phytochemicals as biomarkers of plant food
intake
Anna-Marja Aura
VTT Technical Research Centre of Finland, P.O.Box 1000, Tietotie 2, Espoo, FI-02044 VTT, Finland.
Dietary recommendations define a healthy diet as a one containing substantial amounts of plant
foods. Plant foods are rich in dietary fibre and its co-passengers, polyphenols. Dietary fibre is by
definition non-digestible fraction of plant foods and is transported by intestinal peristalsis through
ileum to the colon, in which the fibre matrix is degraded and the polyphenols are released to
undergo larger structural changes by the microbiota than those apparent by the liver and upper
intestine. Polyphenols can be divided to six major groups: flavonoids, isoflavonoids, stillbenes,
lignans, phenolic acids and tannins: ellagitannins and proanthocyanins, which can be connected
with their colonic microbial metabolite profiles. Even more challenges in diversity are apparent as
the metabolical processes of plant foods are described by metabolomic profiling techniques. The
identified novel metabolites can be found also from the body fluids (plasma and urine) of the
volunteers consuming the same plant foods.
Dietary phenolic compounds undergo intestinal and hepatic metabolism, the products of which
are also shown in the plasma and urine after consumption of plant foods. Most of the complex
colonic metabolites and some phenolic acid metabolites return to the liver and undergo in the
enterohepatic-circulation. For this reason the residence time of colonic metabolites has been
shown to be over 35 hours before returning to baseline after a single dose. If a person eats or
drinks regularly polyphenol-rich foodstuffs, a prediction can be made that the phenolic metabolite
level can be fairly high and constant and a connection with systemic health benefits can be made.
The mechanisms, however, are under investigation. At the moment we can confirm that the
consumption of dietary polyphenols and the circulating metabolites correlate with the plant food
intake and serve as biomarkers of intake.
The work presented here is supported by project ETHERPATHS (EC Contract no: FP7-KBBE222639).
References
Aura, A-M. 2008. Microbial metabolism of dietary phenolic compounds in the colon. Phytochem Rev. 7 (2008): 407429.
36
SL18
Elicitation, production and release of polyphenols in fed-batch
fermentation of grape cells
Tassoni Annalisa and Ferri Maura
Department of Experimental Evolutionary Biology, University of Bologna, Via Irnerio 42, 40126, Bologna,
Italy; [email protected]
Flavonoids and stilbenes are naturally occurring polyphenols, found in several fruits,
vegetables and beverages. In the last years, the potential protective effects of many of these
compounds against cardiovascular and neurodegenerative diseases, as well as the
chemopreventive properties against cancer, have been largely investigated. Cell suspension
cultures have been obtained from Vitis vinifera cv. Barbera petioles and berries and treated with
several different elicitors in order to increase the in vitro production of stilbenes and flavonoids
[1, 2, 3].
Recently, the optimal conditions of grape cell cultures in batch and fed batch fermentations [4],
have been studied with the aim to specifically improve the production of mono-glucosylated
stilbenes, which are resveratrol derivatives. These compounds are physiologically as active as free
resveratrol in cardio- and chemoprotection but more stable and bioavailable after ingestion
through diet. Polyphenols were extracted from biomass and culture broths and stilbenes and
catechins were analyzed by reverse-phase HPLC-DAD separation [2]. In fed batch conditions the
production of mono-glucosides was largely increased together with that of free resveratrol (which
was 32-fold higher in bioreactor with respect to jar cultures). The elicitor chitosan [2] was tested
in a bioreactor system, demonstrating its efficacy in inducing stilbenes production (23-fold and
103-fold increase of mono-glucosides and free resveratrol respectively compared to untreated fed
batch culture). The bioreactor culture conditions also allowed the accumulation of other
polyphenols, such as catechins and in particular the most abundant was epigallocatechin-gallate
(EGCG, 90% of total catechins). The vast majority of the produced compounds was released in the
culture media, a result that represents a relevant advantage for the employment of grape cell
cultures in fed batch fermentation for the industrial production of plant polyphenols, also
considering the necessity of developing sustainable processes.
[1] Tassoni A. et al. (2005). Jasmonates and Na-orthovanadate promote resveratrol production in Vitis vinifera L. cv.
Barbera cell cultures. New Phytologist, 166: 895-905
[2] Ferri M. et al. (2009) Chitosan treatment induces changes of protein expression profile and stilbene distribution in
Vitis vinifera cell suspensions. Proteomics, 9: 610-624
[3] Ferri M. et al. (2011). Increasing sucrose concentrations promote phenylpropanoid biosynthesis in grapevine cell
cultures. Journal of Plant Physiology, 168: 189-195
[4] Ferri M. et al. (2011). Chitosan elicits mono-glucosylated stilbene production and release in fed-batch bioreactor
cultures of grape cells. Food Chemistry, 124: 1473-1479
37
SL19
Insights into olive mill wastewaters phenolics
Susana M. Cardosoa,b*, Soraia I. Falcãoa, António M. Peresb,c, Olivia R. Pereirad, Maria
R.M. Dominguese
a
CERNAS - Escola Superior Agrária, Instituto Politécnico de Coimbra, Bencanta, 3040-316 Coimbra,
Portugal;
b
CIMO, LSRE - Escola Superior Agrária, Instituto Politécnico de Bragança, Campus de Sta. Apolónia,
5301-855 Bragança, Portugal;
c
Escola Superior de Saúde, Instituto Politécnico de Bragança, Av. D. Afonso V, 5300-121 Bragança,
Portugal;
d
Centro de Espectrometria de Massa, Departamento de Química, Universidade de Aveiro, 3810-193
Aveiro, Portugal
[email protected]
Olive mill wastewaters (OMW) have high contents of phenolic compounds, which possess a
wide range of biological activities. Therefore, OMW are regarded as a potent source of biophenols
for food and pharmaceutical industries. Important biophenols occurring in OMW include tyrosol,
oleuropein and hydroxytyrosol, with the latter being commercially exploited as a novel ingredient
with a broad use in dietary supplements, functional foods and natural cosmetics [1,2].
Nevertheless, the phenolic profile of OMW is extremely complex and many compounds remain
unknown. The elucidation of new phenolic compounds in this matrix is an important task to
encourage the search of more bioactive compounds. Indeed, hydroxytyrosyl
acyclodihydroelenolate and p-coumaroyl-6’- secologanoside were recently identified in OMW
and proved to have greater antioxidant scavenging activity than hydroxytyrosol and oleuropein
[3].
The present study evaluates the recovery of some biophenols from three Portuguese OMW. For
that an ethyl acetate liquid-liquid extraction procedure, which has been previously optimized for
hydroxytyrosol extraction [4], has been used. On the other hand, this work also contributes to the
knowledge of OMW phenolic composition, through the elucidation of major phenolic constituents
in purified methanol (PME) or aqueous acetone extracts (PAAE).
Data showed that the ethyl acetate extracts mostly contained hydroxytyrosl, although its
content was much variable for the three OMW samples (0.25 – 1.91 g L-1). Oleuropein, tyrosol
and luteolin-7-O-glucoside were found in the range of 0.02 – 0.23 g L-1, 0.08 – 0.35 g L-1 and 0.3
– 0.75 g L-1, respectively. New phenolic compounds identified in PME comprised oleuropein and
ligstroside isomers which contain the glucose unit linked to the aromatic moiety, as well as some
di-glucoside derivatives of those two compounds. In addition, polymeric compounds derived from
ligstroside glucoside isomer or composed of oleuropein monomers were identified as major
components of the PAAE. Future studies focusing the abundance of these new phenolic
compounds in OMW, as well as their bioactivities, are now needed for determining their possible
industrial exploitation.
[1] Takaç S., Karakaya A. (2009). Recovery of Phenolic Antioxidants from Olive Mill Wastewater. Rec Pat Chem
Eng, 2: 230-237
[2] Soni MG., Burdock GA., Christian MS., Bitler CM., Crea R. (2006) Safety assessment of aqueous olive pulp
extract as an antioxidant or antimicrobial agent in foods. Food Chem Toxicol, 44: 903-915
[3] Obied H K., Karuso P., Prenzler PD., Robards K. (2007) Novel Secoiridoids with Antioxidant Activity from
Australian Olive Mill Waste. JAFC, 55: 2848-2853
[4] De Marco E., Savarese M., Paduano A., Sacchi R. (2007). Characterization and fractionation of phenolic
compounds extracted from olive oil mill wastewaters. Food Chem, 104: 858-867
38
SL20
Molecular mechanisms underlie
phytochemicals: searching the gap
the
biological
activities
of
Cristiano Longoa, Roberto Zefferinob, Antonella Leonea
a
Istituto di Scienze delle Produzioni Alimentari, Consglio Nazionale delle Ricerche (ISPA-CNR),
via Prov.le Lecce Monteroni, 73100, Lecce, Italy
b
Dipartimento di Scienze Mediche e del Lavoro, Università di Foggia, Foggia, Italy
[email protected]
The health benefit of plant-derived foods is established and new researches are identifying
active plant derived molecules among the thousands phytochemicals. Throughout life, the
exposure to specific phytochemicals can affect gene expression via reversible epigenetic
mechanisms. This has recently launched re-exploration of nutritional, botanical or
phytopharmaceutical compounds for their epigenetic effects in order to identify promising
nutraceuticals or cosmeceuticals [1].
Polyphenols from grape seed extracts (GSE) possess a broad spectrum of chemo-protective
properties, and lycopene, found in tomatoes and other vegetables, have been proposed to modulate
hormonal and immune systems, metabolic pathways, and gap junction intercellular
communication (GJIC). In animal cells, GJIC has been implicated in the cell growth control via
adaptive responses: differentiation, proliferation and apoptosis. GJIC is deficient in many human
tumors and its restoration or upregulation is associated with decreased cell proliferation. In the
carcinogenesis process, the reversible inhibition of GJIC has been hypothesized to be involved in
the tumor promotion phase.
Some innovative, safe and environmentally friendly processes such as supercritical carbon
dioxide (S-CO2) extraction are developing for the antioxidant production from plant matrices.
GSE and new S-CO2-extracted oleoresins obtained from tomato and tomato added with grape
seeds were analyzed for antioxidant activities and biological properties on human keratinocytes
[2] and human breast cancer cell (MCF-7) cultures. Plant extracts showed the ability to enhance
the GJIC and connexin 43 expression in human cell cultures, and were able to completely
overcome the GJIC inhibition induced by the tumor promoter HgCl2.
The knowledge of molecular mechanisms underlies the phytochemical effects and the
availability of environmentally friendly methods to produce plant-derived molecules with
potentially healthy properties, from natural sources (e.g., tomato and waste winery by-products),
could lay new bases for preventive nutrition.
[1] vel Szic KS., Ndlovu MN., Haegeman G., Berghe WV. (2010) Nature or nurture: Let food be your epigenetic
medicine in chronic inflammatory disorders. Biochemical Pharmacology, 80: 1816–183.
[2] Leone A., Zefferino R., Longo C., Leo L., Zacheo G. (2010) Supercritical CO 2-extracted tomato oleoresins
enhance gap junction intercellular communications and recover from mercury chloride inhibition in keratinocytes.
Journal of Agricultural Food Chemistry, 58: 4769-4778.
39
PL9
Bioengineering of glucosinolates production in yeast
B.A. Halkier1*, U.H. Mortensen2, L. Albertsen2, B. Salomonsen1, M. Mikkelsen1, J.K. Vester,
F. Geu-Flores1
1
VKR Centre for Pro-Active Plants, LIFE-KU, 1871 Frederiksberg C
Centre for Microbial Technology, DTU, 2800 Lyngby
[email protected]
2
Epidemiological studies have demonstrated reduced risk of developing cancer upon consumption
of diets rich in cruciferous vegetables. Key players in this chemoprevention are the natural
products glucosinolates, in particular the methionine-derived glucoraphanin which is highly
abundant in broccoli. Improved nutrition by functional foods or health-promoting dietary
supplements is an attractive means for prevention of lifestyle-based diseases. Towards this goal,
we have engineered the production of both simple and complex glucosinolates into tobacco (1,2)
transferred the entire glucoraphanin biosynthetic pathway consisting of thirteen genes from
Arabidopsis into the non-cruciferous tobacco by transient expression. The engineering involves
the chloroplast-localized chain elongation machinery (5 genes) that converts methionine to
dihomomethionine, and the cytosolic, ER-anchored core structure pathway (8 genes) that converts
dihomomethionine to the glucoraphanin. More recently, we have embarked on development of a
technology platform for engineering plant pathways exemplified by glucosinolates into yeast. Our
progress in this respect will be discussed.
[1] Geu-Flores et al. (2009) Engineering the production of benzylglucosinolate identifies an uncharacterized
biosynthetic enzyme. Nature Chem. Biol., 5, 575.
[2] Mikkelsen et al. (2010) Reconstitution of the glucoraphanin biosynthetic pathway. Mol.Plant. 3, 751.
40
SL21
The Arabidopsis Orange (Or) gene generates a metabolic sink and
enhances accumulation of carotenoids in cereal cells and plants
Gemma Farre1, Sol Rivera1, Chao Bai1, Ramon Canela1, Gerhard Sandmann2, Changfu
Zhu1, Teresa Capell1, Paul Christou1,3
1
Departament de Producció Vegetal i Ciencia Forestal, Universitat de Lleida, Lleida, Spain
Molecular Biosciences, J. W Goethe Universitaet, Frankfurt, Germany
3
Institucio Catalana de Recerca I Estudis Avancats, Barcelona, Spain
2
Carotenoids accumulate in chromoplasts, which arise directly from proplastids in dividing
tissues and from other non-photosynthetic plastids, such as leucoplasts and amyloplasts [1]. In all
cases, chromoplasts accumulate large amounts of carotenoids in specialized lipoproteinsequestering structures [2]. A spontaneous mutation in cauliflower lead to the discovery of the
Orange (Or) gene, which showed a profound effect on patterns of carotenoid accumulation in
cauliflower plants [3]. In plants cells, chromoplasts contain specialized structures that function as
a metabolic sink to sequester large amounts of carotenoids [4]. The orange (or) gene is
responsible for the accumulation of high levels of β-carotene in various tissues normally devoid of
carotenoids. A functional role for or involves the differentiation of non-colored plastids into
chromoplasts, thus generating a sink for carotenoid accumulation [5]. Lopez et al. [4] were able to
increase carotenoid content in potato tubers by expressing a cauliflower or gene under the control
of a potato granule-bound starch synthase (GBSS) promoter. The total carotenoid levels in the Orexpresing transgenic lines were up to 6-fold higher than in the wild-type tubers. This demonstrates
that creating a metabolic sink has a positive effect on carotenoid accumulation.
We have introduced the Arabidopsis or gene into white corn, which normally accumulates only
trace amounts of carotenoids (lutein and zeaxanthin). Expression of or was controlled by the
endosperm specific wheat LMW glutenin promoter. We report molecular and expression analyses
of T1 seeds from independent transgenic plants as a prelude to a more in depth analyses at the
metabolite level. Transgenic T1 seeds exhibited a color phenotype and even though carotenoid
levels have not been measured yet in these plants, our preliminary data provide exciting opportunities
to enhance carotenoid levels in corn. In order to investigate the mechanisms of carotenoid
accumulation and enhance carotenoid content in rice endosperm, we genetically engineered rice
(Oryza sativa) with the Arabidopsis thaliana orange gene (Ator), Z. mays phytoene synthase 1
(Zmpsy1), and Pantoea ananatis phytoene desaturase (PacrtI). Here we report preliminary results
in rice callus tissue which demonstrate that the Ator gene enhances carotenoid accumulation in
rice dedifferentiated tissues. We are in the process of regenerating plants from colored rice tissue
to ascertain whether carotenoid levels will also be increased in rice seeds.
We acknowledge financial support from Ministry of Science and Innovation-MICINN, Spain (Grant BFU200761413) and European Research Council Advanced Grant BIOFORCE.
[1] Marano et al. (1993); [2] Bartley and Scolnik (1995); [3] Crisp et al. (1975); [4] Lopez et al. (2008); [5] Lu et al.
(2006)
41
SL22
Production of nutritionally desirable fatty acids in seed oil of Indian
mustard (Brassica juncea L.) by metabolic engineering
Surajit Bhattacharyaa, Saheli Sinhab, Prabuddha Deyb, Natasha Dasa, Mrinal K. Maitia,b
a
Department of Biotechnology,
Adv. Lab. for Plant Genetic Engineering, Advanced Technology Development Center,Indian
Institute of Technology Kharagpur, Kharagpur 721302, India
[email protected]
b
Development of a designer oilseed crop with the improved yield attributes and enhanced
nutritional quality for the benefits of mankind and animal husbandry is now achievable with the
combination of genetic engineering and plant breeding. In spite of their immense importance, the
fatty acid profiles of most oilseed crops are imbalanced that necessitate the use metabolic
engineering strategies to overcome the various shortfalls in order to improve the nutritional
quality of these edible oils.
Indian mustard (Brassica juncea L.), being one of the important oilseed crops in Indian
subcontinent naturally contains ~50% nutritionally undesirable very long chain unsaturated fatty
acids (VLCUFAs), e.g. erucic acid (C22:1). In order to divert the carbon flux from the VLCUFAs
to other health beneficial fatty acids, a hairpin RNA-mediated gene silencing construct under the
seed specific napin promoter was used to down-regulate the endogenous fatty acid elongase [1].
The mature seeds of the transgenic B. juncea plant lines developed by Agrobacterium-mediated
transformation showed significant decrease (80-82%) in the VLCUFAs along with noteworthy
increase of oleic acid (C18:1), and displayed a few potentially yield attributing traits.
Further attempts have been initiated for seed-specific down-regulation of the endogenous
stearoyl-acyl carrier protein (ACP) desaturase by similar RNAi approach and over-expression of a
heterologous stearoyl-ACP thioesterase of FatB type [2]. For these purposes, two different genetic
constructs have been prepared for B. juncea transformation using Agrobacterium-based binary
plasmid. Transgenic lines are expected to have increased amount of stearic acid (C18:0), a
nutritionally desirable saturated fatty acid in B. juncea seed oil.
[1] Sinha S., Jha JK., Maiti MK., Basu A., Mukhopadhyay UK., Sen SK. (2007) Metabolic engineering of fatty acid
biosynthesis in Indian mustard (Brassica juncea) improves nutritional quality of seed oil. Plant Biotechnol Rep, 1:
185-197.
[2] Ghosh SK., Bhattacharjee A., Jha JK., Mondal AK., Maiti MK., Basu A., Ghosh D., Ghosh S., Sen SK. (2007)
Characterization and cloning of a stearoyl/oleoyl specific fatty acyl-acyl carrier protein thioesterase from the seeds
of Madhuca longifolia (latifolia). Plant Physiol. Biochem., 45: 887-897.
42
SL23
α-Linolenic acid-rich oil from chia seed (Salvia hispanica L.) induces
lipid redistribution, heart and liver protection in diet-induced obese
rats
Hemant Poudyala and Lindsay Brownb
a
School of Biomedical Sciences, The University of Queensland, Brisbane, 4072, AUSTRALIA; bDepartment
of Biological and Physical Sciences, University of Southern Queensland, Toowoomba 4350, AUSTRALIA.
[email protected]
Chia oil contains the essential n-3 fatty acid, D-linolenic acid (~58% ALA) [1, 2]. This study
has assessed whether chia oil attenuates the metabolic, cardiovascular and hepatic signs in rats fed
a high carbohydrate, high fat (H) diet (carbohydrates 52% (w/w), fat 24% (w/w) with 25% (w/v)
fructose in drinking water). Diets of the treatment groups were supplemented with 3% chia oil
after 8 weeks on H diet for a further 8 weeks. Compared to the H rats, chia oil-supplemented Hrats had improved insulin sensitivity and glucose tolerance, reduced visceral adiposity, decreased
hepatic steatosis, reduced cardiac and hepatic inflammation and fibrosis without changes in
plasma lipids. Chia oil induced lipid redistribution with lipid trafficking away from the visceral fat
and liver with increased accumulation in the heart. The stearoyl-CoA desaturase-1 substrate
(saturated fatty acids) concentrations were increased in the heart, liver and the adipose tissue of
chia oil supplemented rats with a decrease in the products (monounsaturated fatty acids). Transvaccenic acid was preferentially stored in the adipose tissue; the relatively inert oleic acid was
stored in sensitive organs such as liver and heart and linoleic acid, the precursor of arachidonic
acid, the pro-inflammatory fatty acid, was preferentially metabolised. These results strongly
suggest ALA-induced lipid redistribution underlies the heart and liver protection in diet-induced
obese rats.
[1]Alvarez-Chavez, L.M., Valdivia-Lopez, M.D., Aburto-Juarez, M.D.,Tecante, A. (2008) Chemical characterization
of the lipid fraction of Mexican chia seed (Salvia hispanica L.). International Journal of Food Properties, 11:
687-697.
[2] Ayerza, R. (2009) The seed's protein and oil content, fatty acid composition, and growing cycle length of a single
genotype of chia (Salvia hispanica L.) as affected by environmental factors. Journal of Oleo Science, 58: 347-354.
43
SL24
Proteomic identification of differentially expressed proteins in
curcumin-treated prostate cancer cells
Marie-Hélène Teiten a, Anthoula Gaigneaux a, Sébastien Chateauvieuxa, Billing AMb,
Renaut Jc, Mario Dicato a, Marc Diederich a
a
Laboratoire de Biologie Moléculaire et Cellulaire du Cancer, Hôpital Kirchberg, 9 rue Edward Steichen,
L-2540 Luxembourg, Grand Duchy of Luxembourg
b
Laboratoire National de la santé, Institute of Immunology, CRP-Sante, L-1950 Luxembourg.
c
CRP-Gabriel Lippmann, Department Environment and Agro-biotechnologies, L-4422 Belvaux,
Luxembourg
Prostate cancer is the most common cancer in men in the western world. Lifestyle, diet and
environmental as well as genetic factors (e.g. deregulations of the Wingless (Wnt) signaling
pathway) promote the malignant transformation of healthy prostatic epithelium. Due to the high
prevalence and the slow progressive development of prostate cancer, primary prevention appears
as an attractive strategy to eradicate prostate cancer. During the last decades, curcumin
(diferuloylmethane), a natural compound issued from the roots of turmeric plant Curcuma Longa,
was described as a potent chemopreventive agent as it exhibits anti-inflammatory, anticarcinogenic, anti-proliferative, anti-angiogenic and anti-oxidant properties in various cancer
cells. The present study was designed to identify proteins involved in curcumin anti-cancer
activity in androgen dependent and independent human prostate cancer cells using a twodimensional electrophoresis–based proteomic analysis (2D-DIGE). Our approach permitted to
revealed 425 proteins differentially expressed after curcumin treatment. Identification of selected
proteins by MALDI-TOF-MS led to the conclusion that curcumin modulated proteins implicated
in protein folding (Hsps family members, PP2R1A), RNA splicing (RBM17, DDX39), cell death
(HMGB1, NPM1), androgen receptor signaling (NPM1, FKBP4/FKBP52) and that this natural
compound has an impact on miR183 and miR141 expression. Taken altogether these data
reinforced the chemopreventive potential of curcumin by showing that curcumin modulates the
expression of proteins that potentially contribute to prostate carcinogenesis.
44
PL10
Bioactive natural compounds from food plants
Lanzotti V.
Dipartimento di Scienza degli Alimenti, Università di Napoli Federico II, via Università 100, 80055
Portici, Napoli, Italy.
[email protected]
Bioactive natural compounds from food plants have been the focus of researches for decades,
firstly due to their pharmacological effects, and secondly due to their defence properties against
plant diseases. Recent years have seen a renewed interest of such a research because of the
development of analytical techniques allowing the structure elucidation of compounds isolated in
low amounts and the application of genetic engineering and combinatorial chemistry to
phytochemistry.
Our research in this field started twenty years ago and was mainly addressed to the
phytochemical investigation of food plants belonging to Allium genus.1-3
In particular, we isolated and characterized of a large number of bioactive plant metabolites
including apolar compounds such as tiosulfinate and sulphur compounds but also polar
compounds such as flavonoid glycosides and saponins1-3 from A. porrum, A. ascalonicum, A.
triquetrum, A. cepa var. Tropea, A. elburzense, A. hirthifolium, A. atroviolaceum, and very
recently from A. cepa4 and A. sativum. 5
The isolated compounds subjected to a biological screening showed cytotoxicity, and
antiproliferative, antispasmodic and antifungal properties.
In addition, the isolation of a high number of structurally-related analogues allowed to perform
Structure Activity Relationship studies without any chemical modification which gave
information on the key pharmacophoric elements of these new class of potential drugs.
[1] Lanzotti, V. (2005) Bioactive Saponins from Allium and Aster plants, Phytochemistry Rev., 4: 95-110.
[2] Lanzotti, V. (2006) The analysis of onion and garlic, J. Chromat. A, 1112: 3-22.
[3] Benkemblia, N. and Lanzotti, V. (2007) Allium Thiosulfinates: Chemistry, Biological Properties and their
Potential Utilization in Food Preservation, Food, 1, 193-201.
[4] Lanzotti, V, Romano, A., Lanzuise S., Bonanomi G., Scala F. (2011) Antifungal saponins from white onion,
Allium cepa L., in preparation.
[5]Lanzotti, V., Barile E., Antignani E., Bonanomi G., Scala F. (2011) Bioactive Compounds from Allium sativum
L., in preparation
45
SL25
Natural compounds as selective inhibitors for protein kinases and
their potential application in cancer research
Johanne Hadsbjerga, Sarah Willkomma, Katharina Nöskea, Tine D Rasmussena, Elena
Brunsteinb, Karsten Niefindb, Olaf-Georg Issingera
a
Institut for Biokemi og Molekylærbiologi, Syddansk Universitet, Campusvej 55, 5230 Odense Denmark
Institut für Biochemie, Universität zu Köln, Zülpicher St. 47, 50674 Köln, Germany
[email protected]
b
Protein kinases have become the pharmaceutical industry’s most studied class of targets and
some 10 protein kinase inhibitors have so far been approved for the treatment of cancer. Recently,
protein kinase CK2 has been considered a druggable kinase, and increased efforts are undertaken
to find potent and selective inhibitors also for this kinase [1].
A Natural Compound Library from the "The NCI/DTP Open Chemical Repository" was
screened for inhibitors of protein kinase CK2. The screening was performed using the catalytic
subunit and the heterotetrameric holoenzyme using a luminometric test platform as described [2].
Several potential compounds were identified as CK2 inhibitors. We have focused on the
characterization of resorufin and hypericin by measuring IC50, Ki, Vmax and Km. Moreover, we
have determined the selectivity of the compounds towards a panel of 60 protein kinases and
investigated their effect on selected human prostate and kidney tumor cell lines.
When WiT49 and CAL-54 cell lines were treated with 50 μM hypericin 55% and 40%
inhibition of protein kinase CK2 activity was observed, respectively as compared to untreated
controls.
Special focus was on the activity and expression status of selected signaling molecules of key
signaling pathways, e.g. PI3K, AKT/PKB; p38, ERK, JNK, AMPK.
Treatment with 50 uM hypericin caused a reduction in S473 phosphorylation in AKT1/PKB
and an increase in the phosphorylation of T180/Y182 in p38.
How these alterations relate to the concomitant inhibition of CK2 activity is a matter of
discussion
[1] Guerra, B., & Issinger, O. (2008). Protein kinase CK2 in human diseases. Current medicinal chemistry, 15(19),
1870-1886.
[2] Boldyreff, B., Rasmussen, T. L., Jensen, H. H., Cloutier, A., Beaudet, L., Roby, P., & Issinger, O. (2008).
Expression and purification of PI3 kinase alpha and development of an ATP depletion and an alphascreen PI3
kinase activity assay. Journal of biomolecular screening:13(10),1035-1040.
46
SL26
Hazel and other angiosperm plants as new sources of compounds
with taxane-like activity
Mariangela Mielea,b, Anna Maria Mumota, Achille Zappab,c, Paolo Romanob, Laura
Ottaggiob
a
Department of Pharmaceutical Sciences, University of Genova, V.le Benedetto XV, 3, 16132, Genova,
Italy
b
Department of Advanced Diagnostic Technologies, National Institute for Cancer Research, Largo R.
Benzi, 10, 16132, Genova, Italy
c
Department of Informatics, Systems and Telematics, University of Genoa, Via Opera Pia 13, 16132,
Genova, Italy
[email protected]
Taxol is an effective antineoplastic drug, originally extracted from the bark of Taxus brevifolia,
acting on microtubule dynamics and affecting cell cycle progression[1]. The yield of Taxol and its
precursors from Taxus species is very low and it is not sufficient to satisfy the commercial
requirements. For this reason, most of the drug for clinical use is currently produced by semisynthesis, starting from a natural precursor, the taxane 10-deacetylbaccatin III [2]. The finding of
taxanes in differentiated and undifferentiated tissue of Corylus avellana [3,4], suggested that the
production of these compounds is not a peculiarity of Taxus genera, and that, developing
appropriate methodologies, other species could be used for the future production of Taxol and
taxanes.
The aim of the present work is to identify species other than the slow growing Taxus to be
employed as commercial source of Taxol and its precursors.
With this purpose, extracts from differentiated tissues of several plant species were analysed by
ELISA and HPLC. Methanolic extracts obtained from hazel and other angiosperms have been
shown to contain Taxol and other taxanes. Some of these extracts have also been shown to
possess antimitotic activity on cultured cancer cell lines. Our finding suggested that the
biosynthesis of taxanes occur in hazel as well as in other Angiosperms. Thus, other species could
be employed for the commercial production of Taxol and other antineoplastic compounds. Studies
are in progress to verify whether the enzyme involved in taxane biosynthesis, only reported for
Taxus species, are also present in different species producing taxanes. With this purpose a
comparison of involved genes is being carried out through specialized databanks and software.
This work was partially supported by grant of Compagnia di San Paolo.
[1] Wani M et al. (1971) Plant antitumor agents. VI. The isolation and structure of taxol, a novel antileukemic and
antitumor agent from Taxus brevifolia J Am Chem Soc, 93: 2325-2327.
[2] Holton RA et al. (1995) Taxol Science and Applications, Suffness M. CRC Press, Boca Raton 97-12.
[3] Bestoso F et al. (2006) In vitro cell cultures obtained from different explants of Corylus avellana produce Taxol
and taxanes BMC Biotechnology, 6: 45.
[4] Ottaggio L et al. (2008) Taxanes from Shells and Leaves of Corylus avellana JNP, 71: 58-60.
47
PL11
Naturals for the masses
safecommercialisation
–
Chemistry
approaches
for
Russell, P.J.
Safety & Environmental Assurance Centre, Unilever, Colworth Science Park, Sharnbrook, Bedford, UK
[email protected]
There is a growing consumer interest in food and cosmetic products which contain botanicals
as an ingredient with an established or perceived functional benefit which is often intrinsically
linked to traditional medicine cultures throughout the world. The challenges in safely
commercialising truly efficacious plant derived materials which are substantiated by strong
functional claims are great due to their inherent chemical complexity.
Traditional medicine usage is well documented for many natural products but can only be used
to support development programmes where both the indication and extract composition are the
same as historically used. This should be confirmed through raw material authentication and
comprehensive holistic fingerprinting approaches, which we define as a ‘unique visual pattern
representing the presence of known and/or unknown characteristic chemical components’. The
reductive nature of pharmacopeia marker approaches do not provide sufficient insight into the
composition of the complex material as a whole and the concept of a holistic analytical
representation also retains respect for the synergistic philosophy of completeness advocated in
many traditional therapies.
Partially or heavily refined extracts and those which are no longer being used for the traditional
indication are often desired for commercialisation to increase efficacy and provide protectable
intellectual property. The approach to risk assessment therefore requires a combination of
fingerprinting techniques and comprehensive individual chemical component investigation which
can be resource consuming. In addition to these challenges there appears to be a lack of definition
from international regulators as to the requirements for the safety assessment of natural products,
placing the emphasis on industry to design and provide robust supporting data packages.
Here we present a framework approach to define the analytical requirements to support the
development of natural products through a defined safety programme using risk based
approaches.
48
SL27
Targeted screening of botanicals in herbal dietary supplements by
LC-MS/MS with an information dependent acquisition approach
Mathon C.a,b, Duret M.a, Edder P. a, Bieri S.a, Christen P.b
a
Official Food Control Authority of Geneva, Quai Ernest-Ansermet 22, 1211 Geneva 4, Switzerland
School of Pharmaceutical Sciences, EPGL, University of Geneva, University of Lausanne, Quai ErnestAnsermet 30, CH-1211 Geneva 4, Switzerland
[email protected]
b
Herbal Dietary Supplements (HDS) are more popular now than ever before. The commercial
success of botanical ingredients is largely due to a growing interest for products supporting a
healthy lifestyle. HDS are widely marketed with various health claims. These phytopreparations
are easily available to consumers through several distribution channels: OTC in pharmacies, in
supermarkets, herbalist’s shops or via the Internet.
In this context, problems related on undeclared, unauthorized [1] or toxic botanicals in HDS is of
growing importance, because HDS have generally not been through a rigorous drug testing
process. Furthermore, there is a need to conduct anti-fraud analyses by confirming the presence of
the declared botanicals. Chemical methods already exist for the detection of plants but they are
usually specific for a few plants only.
In this study, a generic method was developed for the multi-target screening of biomarkers in
HDS. This method uses high performance liquid chromatography coupled to hybrid mass
spectrometry (Q-Trap) with an Information Dependent Acquisition (IDA) approach generating
MS/MS spectra which can be compared with an in-house library.
Botanical preparations were unselectively extracted by sonication in methanol. Each plant was
characterized with at least one biomarker, which in turn was identified with its retention time, two
specific transitions and their corresponding ratio as well as three Enhanced Product Ion (EPI)
scans.
With this method, almost hundred biomarkers were targeted by two successive analyses in the
positive and negative ElectroSpray Ionisation (ESI) modes which finally allowed to screen more
than 60 plants.
[1] Swiss guidance on the categorization of botanicals intended for use as therapeutic preparations or food
supplements (2010). List of botanicals, version 1.3, Federal Office of Public Health, Switzerland.
49
SL28
LC-MS/MS determination of marker metabolites in the ethanolic
extract and in mother tinctures of Vitex agnus castus fruits
Angela Mari, Paola Montoro, Sonia Piacente
Dipartimento di Scienze Farmaceutiche e Biomediche, Università di Salerno, via ponte don Melillo, 84084
Fisciano (SA), Italy
[email protected]
Vitex agnus-castus L. (Verbenaceae), chasteberry, is one of the most popular and effective
herbs used for the prevention and treatment of pre-menstrual syndrome (PMS) and
hyperprolactinemia because of its hormone-like effect [1]. Although its mechanism of action has
only partially been determined, dopaminergic activities were found in polar and apolar fractions
of the fruits of V. agnus-castus [2]. The chemical composition of polar extracts is characterized by
the presence of iridoids and flavonoids [3]. Among these latter, casticin, a methoxylated flavonol,
is considered by European Pharmacopoeia (Ph. Eur.) the reference compound for standardization
of dry extracts of the species. A large number of V. agnus-castus products routinely used by
women against PMS are commercially available, most of them being mother tinctures whose
standardization is required.
Many analytical studies are reported about the constituents of the essential oil of chasteberry
fruits but very few reports occur about LC/MS studies of the polar fraction of these fruits [4].
Targeted mass spectrometry approach is considered one of the most powerful way to obtain
metabolite profiling of biological samples so it represents an ambitious tool in metabolomic
quantitative analysis field [5]. While MS itself provides sensitive detection and the ability to
identify metabolites based on MS or MS/MS spectra, LC-MS techniques avoiding signal
suppression phenomena, provide identification of isobaric ions based on differences in their
retention times. According to this, HPLC/ESI-triple quadrupole (QqQ)-MS using a very sensitive
and selective mass tandem experiment such as Multiple Reaction Minitoring is considered one of
the most suitable techniques for quantification of plant metabolites.
Our investigation deals with the characterization of the ethanolic extract obtained from the
fruits of chasteberry. An analytical method for the quantitative determination of seven
compounds, previously reported in literature [3], was developed by using HPLC/ESI-QqQMSMS
in order to obtain a full metabolite profile of the extract. Five commercially available mother
tinctures of V. agnus-castus fruits were analysed under the same analytical conditions and their
chromatographic profiles were compared. In spite of the small contents in casticin, all the extracts
exhibited higher amounts of the iridois aucubin and agnuside. That is the reason why, according
to our results, these two compounds could be considered more suitable than casticin, suggested by
the Ph. Eur, for standardization procedures.
[1] Liu J., Burdette J.E., Xu H., Gu C., van Breemen R.B., Bhat K.P.L., Booth N., Constantinou A.I., Pezzuto J.M., Fong H.H.S.,
Farnsworth N.R., and Bolton J. L. (2001). Evaluation of Estrogenic Activity of Plant Extracts for the Potential Treatment of
Menopausal Symptoms. J. Agric. Food Chem, 49: 2472-2479.
[2] Lucks B.C., Sorensen J., Veal L. (2002). Vitex agnus-castus essential oil and menopausal balance: a self-care survey. Compl.
Ther. in Nursing and Midwifery, 8: 148–154.
[3] Chen S.N., Friesen B.J., Webster D., Nikolic D., van Breemen R.B., Wang Z.J. Fong H.H.S., Farnsworth N.R., Pauli G.F.
(2010). Phytocostituents from Vitex angus-castus fruits. Fitoter, 82(4): 528-33.
[4] Hoberg E.; Meier B.; Sticher O. (2001). Quantitative high performance liquid chromatographic analysis of casticin in the fruits
of Vitex agnus castus. Pharm. Biol, 39(1): 57-61.
[5] Wolfender J.L., Glauser G., Boccard J. and Rudaz S. (2009). MS-based Plant Metabolomic Approaches for Biomarker
Discovery. Nat. Prod. Com, 4 (10): 1417-1430.
50
International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September 2011
PSE-APIVITA Award Lecture
51
Contribution to Phytochemistry in recent years
Sonia Piacente
Dipartimento di Scienze Farmaceutiche e Biomediche, Università di Salerno, via Ponte Don Melillo,
84084 Fisciano (SA), Italy
[email protected]
This presentation will be focused on some of the results of phytochemical investigations
performed in recent years.
-Phenolic constituents of Yucca spp.
Yucca schidigera (Agavaceae) is one of the major commercial source of steroidal saponins. It
possesses the GRAS label which allows its use as foaming agent in soft drink (root beer),
pharmaceutical, cosmetic, food, and feeding-stuffs industries. The main application of yucca
products is in animal nutrition, in particolar as a feed additive to reduce ammonia and fecal odors
in animal excreta. The positive effects of dietary supplementation with yucca products on the
growth rates, feed efficiency, and health of livestock seem to be due not only to the saponin
constituents but also to other constituents. These observations prompted us to investigate the
phenolic constituents of Y. schidigera and of another species of the same genus, Y. gloriosa,
largely cultivated in Eastern Georgia. Yuccaols and gloriosaols, showing very unusual spirostructures made up of one (yuccaols from Y. schidigera) or two (gloriosaols from Y. gloriosa) C15
units linked via a J-lactone ring to a stilbenic moiety related to resveratrol, were isolated [1-2].
Yuccaols showed potent radical scavenging activity, were found to reduce the expression of the
inducible isoform of nitrogen oxide synthase and to inhibit in a dose-dependent manner the
Vascular Endothelial Growth Factor (VEGF)-induced proliferation, migration, and Platelet
Activating Factor (PAF) synthesis in Kaposi’s sarcoma cells.
Antiproliferative pregnane and secopregnane glycosides from Solenostemma argel
-In the frame of a bilateral Italy-Egypt project, Solenostemma argel Hayne (Asclepiadaceae), an
Egyptian wild perennial erect shrub growing in the eastern desert and along the Nile in South
Egypt, used in traditional medicine as a purgative, has been investigated. From the pericarps,
hairy seeds and leaves new 14,15-secopregnane glycosides, namely argelosides A – J, and new
15-keto pregnane glycosides, namely stemmosides C – K, have been isolated [3-4]. On the basis
of the anticancer and cytotoxic activities reported for pregnane derivatives and considering the
very unusual structural features of argelosides and stemmosides, the anti-proliferative activity of
these compounds in Kaposi’s sarcoma (KS) cells has been evaluated. A dose dependence study
was performed to test the effects of argelosides and stemmosides on the VEGF-induced KS cell
proliferation in a concentration range 0.1 – 20 PM. Results indicated that 15-keto-pregnane
glycosides and 14,15-secopregnane glycosides reduced the VEGF-induced KS cell proliferation in
a dose-dependent manner.
-Cardenolides as cytotoxic agents
Over the past few years, there has been a marked increase in the number of reports of
cardiotonic steroids-induced anticancer effects. Cardiac glycosides-mediated effects are linked to
their Na+/K+-ATPase binding properties. In particular calactin-related cardenolides showing trans
fused A/B rings and a single sugar in a unique “dioxanoid” attachment (doubly linked) seem to
markedly bind Na+/K+-ATPase α1 subunit, overexpressed in some cancer cell lines. In this frame
doubly-linked cardiac glycosides isolated from the roots of Pergularia tomentosa were tested in
an vitro growth inhibitory assay including six different human cancer cell lines and for their
ability to inhibit Na+/K+-ATPase activity [5]. The data revealed for these cardenolides a marked
53
cytotoxic activity (IC50 in the range 0.02- 4.00 PM) and a strong correlation between their
citotoxicity and their ability to inhibit sodium pump.
Polyisoprenylated benzophenones from Garcinia cambogia fruits and their effects on cytokine
signalling
Garcinia cambogia L. (Guttiferae) is a small or medium-sized tree whose fruits are used in
traditional Indian medicine as anti-inflammatory remedy. Garcinol, a polyisoprenylated
benzophenone derivative occurring in G. cambogia, has been reported to reduce COX-2 and
iNOS expression as well as NO production, in LPS-stimulated macrophages. In order to provide
deeper insight into the biological effects of garcinol, we carried out its isolation from the fruits of
G. cambogia. This phytochemical investigation afforded garcinol, guttiferones I, J and K, the new
guttiferones M and N along with oxy-guttiferones K, K2, I and M which are the first example of
tetracyclic xanthones derived from the oxidation of polyisoprenilated benzophenones, from
natural source [6]. On the basis of the reported inhibition of STAT-1 activation by garcinol, the
interaction between immobilized STAT-1 and some guttiferones was evaluated by Surface
Plasmon Resonance (SPR). All the tested compounds were able to form complexes with STAT-1,
exhibiting KD in the range 3-20 μM. Concomitantly, we tested the isolated compounds for their
ability to modulate cytokine signalling in different cell lines. The results showed that cytokineinduced STAT-1 and NF-κB activations, as assessed by EMSA, were dose-dependently inhibited
by garcinol, guttiferone M and, to minor extent, by guttiferone K (range 10-20 μM) in most cell
lines.
[1] Balestrieri C., Felice F., Piacente S., Pizza C., Montoro P., Oleszek W.,Visciano V., Balestrieri M. L. (2006)
Relative effects of phenolic constituents from Yucca schidigera Roezl. bark on Kaposi's sarcoma cell
proliferation, migration, and PAF synthesis. Biochemical Pharmacology , 7: 1479-1487.
[2] Bassarello C., Bifulco G., Montoro P., Skhirtladze A., Benidze M., Kemertelidze E., Pizza, C, Piacente, S. (2007)
Yucca gloriosa: A Source of Phenolic Derivatives with Strong Antioxidant Activity. Journal of Agricultural and
Food Chemistry, 55: 6636-6642.
[3] Plaza A., Perrone A., Balestrieri C., Balestrieri M. L., Bifulco G., Carbone V., Hamed A., Pizza C., Piacente S.
(2005) New antiproliferative 14,15-secopregnane glycosides from Solenostemma argel. Tetrahedron , 61: 74707480.
[4] Plaza A., Perrone A., Balestrieri M. L., Felice F., Balestrieri C., Hamed A., Pizza C., Piacente S. (2005) New
unusual pregnane glycosides with antiproliferative activity from Solenostemma argel. Steroids , 70: 594-603.
[5] Piacente S., Masullo M., De Neve N., Dewelle J., Hamed A., Kiss R., Mijatovic T. (2009) Cardenolides from
Pergularia tomentosa display cytotoxic activity resulting from their potent inhibition of Na+/K+-ATPase. Journal
of Natural Products, 72: 1087-1091.
[6] Masullo,M., Bassarello C., Suzuki H., Pizza C, Piacente S. Polyisoprenylated benzophenones and an unusual
polyisoprenylated tetracyclic xanthone from the fruits of Garcinia cambogia. (2008) Journal of Agricultural and
Food Chemistry, 56: 5205-5210.
54
Poster Presentations - International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September 2011
P1
Poster Presentations
Poster Presentations – International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September 2011
56
Poster Presentations - International PSE Symposium on ”Phytochemicals in Nutrition and Health”, 27-30 September 2011
P1
Nutritional and nutraceutical potential of saprotrophic and
mycorrhizal wild edible mushrooms from Northeast Portugal
Lillian Barrosa,b, Cátia Grangeiaa,b, Anabela Martinsb, Isabel C.F.R. Ferreiraa,b
a
CIMO-ESA, Instituto Politécnico de Bragança, Campus de Santa Apolónia, Apartado 1172, 5301-855
Bragança, Portugal.
b
Escola Superior Agrária, Instituto Politécnico de Bragança, Campus de Santa Apolónia, Apartado 1172,
5301-855 Bragança, Portugal.
[email protected]
Mushrooms are appreciated all over the world not only by their texture and flavour, but also by
their chemical, nutritional [1] and functional properties [2,3]. The consumption of wild growing
mushrooms has been preferred to eating of cultivated fungi in many countries of central and
Eastern Europe. Nevertheless, the knowledge of the nutritional value of wild growing mushrooms
is limited. The present study reports the nutritional and nutraceutical potential of five saprotrophic
(Calvatia utriformis, Clitopilus prunulus, Lycoperdon echinatum, Lyophyllum decastes, and
Macrolepiota excoriata) and five mycorrhizal (Boletus erythropus, Boletus fragrans,
Hygrophorus pustulatus, Russula cyanoxantha, and Russula olivacea) wild edible mushrooms.
The nutritional composition was determined on the bases of moisture, proteins, fat, carbohydrate
and ash. For the nutraceutical potential the antioxidant activity was evaluated by different in vitro
chemical and biochemical assays: scavenging effects on DPPH (2,2-diphenyl-1-picrylhydrazyl)
radicals, reducing power, inhibition of E-carotene bleaching and inhibition of lipid peroxidation in
brain cells homogenates by TBARS (thiobarbituric acid reactive substances) assay. The results
were compared to bioactive compounds: phenolics and ascorbic acid (determined by
spectrophotometric techniques); tocopherols (HPLC/fluorescence), sugars (HPLC/RI) and fatty
acids (GC/FID). Mycorrhizal species revealed higher sugars concentration (16–42 g/100 g dw)
than the saprotrophic mushrooms (0.4–15 g/100 g). Furthermore, fructose was found only in
mycorrhizal species (0.2–2 g/100 g). The saprotrophic L. decastes, and the mycorrhizal species B.
erythropus and B. fragrans gave the highest antioxidant potential, mainly due to the contribution
of polar antioxidants such as phenolics and sugars [4]. The bioactive compounds found in wild
mushrooms give scientific evidence to traditional edible and medicinal uses of these species.
FCT (Portugal) and COMPETE/QREN/UE- research project PTDC/AGR-ALI/110062/2009. L. Barros also thanks to
FCT, POPH-QREN and FSE for her grant (SFRH/BPD/4609/2008).
[1] Kalač P. (2009). Chemical composition and nutritional value of European species of wild growing mushrooms: A
review. Food Chemistry, 113: 9–16.
[2] Ferreira ICFR., Vaz JA., Vasconcelos MH., Martins A. (2010). Compounds from Wild Mushrooms with
Antitumor Potential. Anti-cancer Agents in Medicinal Chemistry 10: 424-436.
[3] Ferreira ICFR., Barros L., Abreu RMV. (2009). Antioxidants in Wild 320 Mushrooms. Current Medicinal
Chemistry, 16:1543-1560.
[4] Grangeia C., Heleno SA., Barros L., Martins A., Ferreira ICFR. (2011). Effects of trophism on nutritional and
nutraceutical potential of wild edible mushrooms. Food Research International, 44: 1029–1035.
P2
In vitro inhibition of nitric oxide production and free radical
scavenging activity of non-cultivated Mediterranean vegetables
Filomena Conforti, Mariangela Marrelli, Giancarlo A. Statti, Francesco Menichini
Department of Pharmaceutical Sciences, Faculty of Pharmacy and Nutritional and Health Science,
University of Calabria, I-87036 Rende (CS), Italy
[email protected]
The consumption of non-cultivated botanicals may play a central role in the diet of some
population groups but very few ethnopharmacological and phytopharmacological studies have
exhaustively dealt. Epidemiological studies have indicated that dietary intake of antioxidant
substances from plants is inversely associated with mortality from coronary heart disease and
inflammation [1].
Inflammation is critical in recruiting immune cells and molecules to the site of infection for
defence. Among participating cells, macrophages play a central role in organizing the release of
inflammation mediators, including prostaglandin E2 (PGE2), nitric oxide (NO) and cytokines that
promote host protection, as well as cause pathological consequences such as tissue oedema and
abnormal histological change [2]. The RAW 264.7 mouse macrophage cell line is widely used for
studies of inflammation, due to its reproducible response to lipopolysaccharide (LPS), mediated
by toll-like receptor 4 (TLR4) [3].
Hydroalcoholic extracts from three non-cultivated vegetables traditionally consumed in
Southern Italy, belonging to the Lamiaceae family, were tested for their free radical scavenging
activity (FRSA) in the DPPH (1,1- diphenyl-2-picrylhydrazil radical) screening assay [4] and for
their in vitro inhibition of niric oxide (NO) production [5]. The strongest antiradical activity was
shown for Mentha spicata subsp. glabrata (leaves) with IC50 value 6 Pg/ml. The same extract
showed high in vitro inhibition of nitric oxide production with an activity comparable to a
reference drug, indomethacin. M. spicata subsp. glabrata had relatively high polyphenols content
334 mg per g of extract.
Dietary plants are being recognized as a source of numerous bioactive compounds. Many of
them were shown to possess antioxidant properties and inhibit redox-sensitive intracellular
signaling pathways leading to cell activation. However, the search for new plants exerting antiinflammatory properties continues, especially among the plants included in the diets regarded as
"health beneficial" ones. Clearly, this research has shown that such local food plants are a treasure
trove for developing leads for new nutraceuticals and pharmaceuticals, especially for
inflammatory conditions.
[1] Giugliano D. (2000). Dietary antioxidants for cardiovascular prevention. Nutrition, metabolism, and cardiovascular diseases,
10: 38-44.
[2] Wang L., Tu Y.C., Lian T.W., Hung J.T., Yen J.H., Wu M.J. (2006). Distinctive antioxidant and antiinflammatory effects of
flavonols. Journal of Agricultural and Food Chemistry, 54: 9798– 9804.
[3] Cao X.Y., Dong M., Shen J.Z., Wu B.B., Wu C.M., Du X.D., Wang Z., Qi Y.T., Li B.Y. (2006). Tilmicosin and tylosin have
anti-inflammatory properties via modulation of COX-2 and iNOS gene expression and production of cytokines in LPS-induced
macrophages and monocytes. International journal of antimicrobial agents, 27: 431– 438.
[4] Parejo I., Viladomat F., Bastida J., Rosas-Romero A., Flerlage N., Burillo J., Codina C. (2002). Comparison between the
radical scavenging activity and antioxidant activity of six distilled and nondistilled Mediterranean herbs and aromatic plants.
Journal of Agricultural and Food Chemistry, 50: 6882-6890.
[5] Prakash H., Ali A., Bala M., Goel H.C. (2005). Anti-inflammatory effects of Podophyllum hexandrum (RP-1) against
lipopolysaccharides induced inflammation in mice. Journal of pharmacy & pharmaceutical sciences, 8: 107-114.
58
P3
A new clerodane diterpenoid from the aerial part exudate of Salvia
chamaedryoides Cav.
Angela Bisioa, Emanuela Giacomellia, Gianluca Damonteb, Annalisa Salisb, Daniele
Fraternalec, Giovanni Romussia, Sergio Cafaggia, Donata Riccic, Nunziatina De Tommasid
a
Dipartimento di Chimica e Tecnologie Farmaceutiche e Alimentari, Università di Genova, Via Brigata
Salerno 16147 Genova, Italy
b
Dipartimento di Medicina Sperimentale e Centro di Eccellenza per la Ricerca Biomedica, Università di
Genova, Viale Benedetto XV 7, 16132, Genova, Italy
c
Dipartimento di Scienze dell’Uomo, dell’Ambiente e della Natura, Università di Urbino, Via Bramante
28, Urbino, Italy
d
Dipartimento di Scienze Farmaceutiche,Università di Salerno, Via Ponte Don Melillo, 84084 Salerno,
Italy
[email protected]
Salvia chamaedryoides Cav. [1] is a Mexican species whose aerial part exudate showed
herbicide activity against Papaver rhoeas L. and Avena sativa L. in a preliminary test [2]. The
surface exudate, obtained by rinsing the plant material with CH2Cl2, was subjected to repeated
column chromatography on Sephadex LH-20 , silica gel and HPLC-MS and MS2 experiments
followed by semi-preparative RP-HPLC, yielding a new clerodane diterpenoid (1).
1 was identified by IR and NMR analysis, including TOCSY, COSY, HSQC and HMBC
experiments, and ESI-TRAP-MS and HR-MS analysis.
(1)
[1] Epling C. (1940) A Revision of Salvia, subgenus Calosphace. In: Repertorium Specierum Novarum Regni
Vegetabilis. Vol.110. Fedde F., University of California Press: Berkley, California.
[2] Bisio A., Fraternale D., Giacomini M., Giacomelli E., Pivetti S., Russo E., Caviglioli G., Romussi G., Ricci D.,
De Tommasi N. (2010) Phytotoxicity of Salvia spp. exudates. Crop Protection, 29: 1434-1446.
59
P4
Chemical study on Pavetta corymbosa
Augustine A Ahmadu1and Abdulkarim Agunu2
1
Department of Pharmaceutical and Medicinal Chemistry Faculty of Pharmacy, Niger-Delta
University,Wilberforce island, Bayelsa State-NIGERIA.
2
Department of Pharmacognosy and drug development,Faculty of Pharmaceutical Sciences,
Ahmadu Bello University, Zaria-NIGERIA.
[email protected];
The genus Pavetta have long being used in ethnomedicine as anti-malaria, remedy for tuberculosis
and for relieve of stomach pain[1].The literature does not report any phytochemical studies on
Pavetta corymbosa. The dichloromethane and the ethyl acetate extracts were investigated for
phytochemical constituents. Fractionation of the dichloromethane extract by Flash column
chromatography, sephadex LH-20 and Preparative TLC afforded the trietrpenes: α-amyrin,
Lupenol, ursolic acid and a mixture (1:1) of β-sitosterol and Stigmasterol, while the ethylacetate
extract was fractionated over sephadex LH-20 eluted with methanol to give the known
flavonoids: Quercetin, Quercetin 7-O-rhamnoside and Kaempferol. The structures were elucidated
by NMR spectroscopy and compared with literature [2-5] and are reported here for the first time.
[1] Dalziel J.M. and Hutchinson J. (1955). Useful plants of West Africa. Crown Agents for oversea publication,
London
[2] Bishi D.S., Padalia R.C., Joshi S.C., Tewari A. and Mathela S.C. (2010). Constituents of Nepeta clarkei; Hook F.
Indian Journ. Of Chem. Vol.49B:807-811
[3] Ahmed Z., Zahra D.N. and Malik A. (2006). Phytochemical studies on Abuliton kara. Journ.of Chem. Soc.Pak.
28:3,295-297.
[4] Zhang Tao, YE Qi, FENG Chun and Chen Yaoming (2007). Chemical study on Gladiolus gandavensis. Chin. J.
Appl Environ Biol. 13:5,635-640.
[5] Mabry T. J et al (1970). The Systematic Identification of Favonoids. Spring- Verlag Publication New York.
60
P5
Physicochemical characterization and bioactivity of Portuguese
Propolis towards its standardization
Soraia I. Falcãoa, b, Cristina Freirea, Miguel Vilas-Boasb
a
REQUIMTE/Departamento de Química e Bioquímica, Faculdade de Ciências do Porto, Rua do
Campo Alegre, 4169-007 Porto, Portugal
b
Mountain Research Centre (CIMO), Polytechnic Institute of Bragança, Campus de Sta.
Apolónia, Apartado 1172, 5301-855 Bragança, Portugal
[email protected]
Propolis is a sticky dark colored material collected by honeybees, from leaf buds and other
secretions of numerous tree species (alder, birch, palm, pine, poplar and willow) around the hive,
mix with beeswax and used to reinforce the combs and to keep the hive environment aseptic.[1]
This hive product is widely used in folk medicine since ancient times and recently gained
popularity all over the world as an important ingredient of health foods and cosmetics. Propolis is
thought to improve human health and to prevent diseases such as inflammation, heart disease,
diabetes and even cancer.[2]
Phenolic compounds are the major constituents of propolis representing approximately half of
the resin, while beeswax, volatiles and pollen represent 30%, 10% and 5%, respectively. The
chemical composition of this bee product is highly variable and complex, depending strongly on
the plant sources available to the bees at the site of collection as well as with the geographic and
climatic conditions of the site.
This research outlines the extensive physicochemical characterization of propolis from
different Portuguese regions, a country with botanical diversity, in order to establish quality
criteria parameters for commercial purposes and thus for its valorization. For that, forty samples
from continental Portugal and islands, were characterized and subjected to hydro-ethanol
extraction according to our previous work.[3] Several physicochemical parameters like water, ash
and waxes content, color index, spectrophotometric determination of phenolic compounds, and
antioxidant capacity were determined in either crude propolis or ethanol extracts. The results for
crude propolis show that water content ranged from 2% to 14%, ashes values ranged between 0.516.1% and the wax from 2% to 35%. According to the CIELAB color system, samples showing
an orange to dark orange color like in interior north, coast and Azores archipelago presented
higher values of a* and b* coordinates, comparing to the samples from interior center, south and
Madeira island with darker colors. The phenolic composition of the extracts was achieved
measuring the content in total phenols (10-850 mg eq/100g extract), flavones/flavonols (3-73 mg
eq/100g) and flavanones/ dihydroflavonols (2-73 mg eq/100g). The bioactivity of the extracts was
evaluated by the DPPH free radical scavenging activity with the EC50 ranging from 0.008 mg/mL
to 0.093 mg/mL and by the reducing power, with values in the interval of 110 to 757 mg eq/100g
extract. The analyzed samples presented a wide variability for the different physicochemical
parameters which reflect the phyto-geography differences at the site of collection.
[1] Bankova VS., De Castro SL., Marcucci MC. (2000) Propolis: recent advances in chemistry and plant origin.
Apidol., 31: 3-15.
[2] Banskota A H., Tezuka Y., Kadota SH. (2001) Recent progress in pharmacological research of propolis.
Phytother. Res., 15: 561-571.
[3] Falcão S., Vilas-Boas M., Estevinho LM., Barros C., Domingues MRM., Cardoso SM. (2010) Phenolic
characterization of Northeast Portuguese Propolis: usual and unusual compounds. Anal. Bioanal. Chem., 396:
887-897.
61
P6
A new cytotoxic dihydrophenantrene against human cancer cells
from Dioscorea membranacea roots
Arunporn Itharat a, Srisopa Ruangnoob, Pagakrong Thongdeeyingc
a
Department of Applied Thai Traditional Medicine ,
Ph.D program on Nutraceutical, Graduate School,
c
Division of Herbal and Food Research, Faculty of Medicine, Thammasart University, Rungsit,
Klongloung, Pathumthanee, 12121 Thailand
b
[email protected];
The roots of Dioscorea membrancea has been used as anticancer drug in Thai traditional
medicine (1). Bioassay-guided isolation was used to separate the cytotoxic ingredients from the
ethanolic extract of Dioscorea membranacea by testing against five human cancer cell lines, i.e.
large cell lung carcinoma COR-L23, liver cancer cell HepG2, prostate cancer cell PC3, breast
cancer cell line MCF-7, cervical cancer cell line Hela, one normal human lung cell MRC5 using
the SRB assay(2).. Two known dihydrophenantrene compounds [2,4 dimethoxy-5,6-dihydroxy9,10-dihydrophenanthrene (1) and 5-hydroxy-2,4,6-trimethoxy-9,10-dihydrophenanthrene (2)]
and a new dihydrophenantrene compound as 5,6,2 -trihydroxy 3,4-methoxy, 9,10dihydrophenanthrene (3) were isolated. 1 showed the highest cytotoxic activity against lung,
breast and prostate cancer cell lines (IC50= 14.89 , 17.49 and 19.04 µM respectively), and 2
showed selective cytotoxic activity against prostate cancer (IC50= 23.54µM). 3 showed selective
cytotoxic against only breast cancer cells (IC50= 31.41 µM).
[1] Itharat, A.,.Houghton,P.J., E.Eno-Ammguaye, Burke, P.J.,..Sampson, J.H and Raman, A.
(2004) In vitro cytotoxic activity of Thai medicinal plants used traditionally to treat cancer. J
Ethnopharmacology, 90 :33-38.
[2] Skehan, P., Storeng, R., Scudier., D., Monks, A., Mc Mahon, J., Vistica, D., Warren, J.T.,
Bokesch, H., Kenney, S. and Boyd, M.R. (1990). New colorimetric cytotoxicity assay for
anticancer-drug screening. J Natl Cancer Inst 82: 1107-1112.
62
P7
Effects of daily consumption of Rosmarinus officinalis and
Hypericum vesiculosum infusions on rodent cognitive function and
cerebral acetylcholinesterase activity
Ferlemi A.V.1, Lamari F.N.1, Iatrou G.2, Margariti M.2
1
Department of Pharmacy, University of Patras, 2Department of Biology, University of Patras
[email protected]
Cholinergic system is involved in the regulation of several central nervous system functions,
like learning/memory. Acetylcholinesterase (AChE) at the synaptic gap plays a key role in
cholinergic neurotransmission since it hydrolyses the available acetylcholine. Rosmarinus
officinalis (rosemary) is a perennial shrub, native to the Mediterranean region, with many culinary
and other traditional uses. Hypericum vesiculosum is an endemic species of Greek flora, which
has not been yet studied, although other Hypericum have anti-depressant and wound healing
properties. Aim of our project was to study the effects of daily consumption of rosemary and H.
vesiculosum infusions on cognitive function and AChE activity [salt-soluble (SS) and detergentsoluble (DS) fraction] in whole brain and cerebellum of adult mice. Animals were separated in 3
groups: control-group, Ro-treated group (2% w/v of R. officinalis tea for 4 weeks) and Hv-treated
group (2% w/v of H. vesiculosum tea for 4 weeks). Mice body weight was measured at regular
time intervals. Effects on learning/memory were assessed using the step-through latency test and
on AChE activity by the colorimetric method of Ellman. Our results demonstrate that tea drinking
affect neither mouse and tissue weight nor learning/memory indices. There is a tendency of
improvement in the cognitive function of Ro-mice, although not statistically significant.
Moreover, in Ro-mice, AChE activity was lower (p<0,05) [19,29% (SS), 43,76% (DS) and
25,74% (SS), 19,83% (DS) in whole brain and cerebellum] than the control group. H. vesiculosum
did not affect effectively learning and memory but decreased AChE activity [(13,73% (SS),
8,79% (DS) and 18,53% (SS), 15,43% (DS) in whole brain and cerebellum respectively]
compared to the controls. Conclusively, our data suggest that daily consumption of both infusions
for 4 weeks induced significant decrease of cerebral AChE activity, although the complex
function of learning/memory was not affected.
63
P8
Cytotoxic activity against lung and prostate cancer cells of the
ethanolic extract from Antidesma thwaitesianum Müll. Arg. leaves
Bhanuz Dechayonta, Pakakrong Thondeeying b, Arunporn Itharatc
a
Graduated School,
Division of Herbal and Food Research
c
Department of Applied Thai Traditional Medicine, Faculty of Medicine, Thammasat University,
Klongluang , patumtanee, Thailand
[email protected]
b
Antidesma thwaitesianum Müll.Arg. is belongs to the family Euphorbiaceae[1]. It is mostly
found in northeast of Thailand. This plant is traditionally used to treat a various diseases such as
anemia, leucorrhoea, stimulate blood circulation, diuretic and abscess. It was reported that the
fruits of A. thwaitesianum contain high antioxidant activity [2] but leaves not were reported. Thus,
the objectives of this research were studied cytotoxicity of crude extract and fractions from leaves
of A. thwaitesianum against two types of human cancer cell lines ; large cell lung carcinoma
(CORL-23) and prostate cancer cell lines (PC3), by using SRB assay [3].
The ethanolic extract of leaves and the fractions were separated by using vacuum liquid
chromatography (VLC) as hexane (1,000 ml as F1), hexane – chloroform (500:500 ml as F2),
chloroform(500 ml 3 times as F3, F4, F5), chloroform- methanol (750:750 ml as F7) and then
methanol (2000 ml as F8). After running VLC, the different fractions were collected to test
cytotoxic activity.
The results indicate that crude extract showed the specific cytotoxicity against COR-L23
(IC50= 27.91 μg/ml) but it exhibited low cytotoxicity against PC3 (IC50= 75.41 μg/ml). The
activity of fractions shows that the F5 showed the highest cytotoxicity against COR-L23 (IC50=
6.52 µg/ml). We concluded that the principle cytotoxic compound should be isolated from
fraction F5.
[1] Hoffmann P., (1999). The genus Antidesma (Euphorbiaceae) in Madagascar and the Comoro Islands. KewBull,
54:877–85.
[2] Puangpronpitag, D., Areejitranusorn, P., Boonsiri, P., Suttajit, M., Yongvanit, P., (2008) Antioxidant Activities of
Polyphenolic Compounds Isolated from Antidesma thwaitesianum Müll. Arg. Seeds and Marcs. Journal of Food
Science, 73: 648-653.
[3] Skehan, P., Storeng, R., Scudiero, D., Monks, A., McMahon, I., Vistica,D., Waren, J.T., Bokesch, H., Kenney, S.,
Boyd, M.R., (1990). New colorimetric cytotoxicity assay for anticancer drug screening. Journalof National
Cancer Institute, 82: 1107–1112.
64
P9
Comparison of antimicrobial, antioxidant, anti-inflammatory
activities and total phenolic contents of female and male trees from
Carica papaya roots.
Jitpisute Chunthorng-Orna , Arunporn Itharat b
a
Graduate School,
Pathumthani, 12120, Thailand
[email protected]
b
The female (CPF) and male tree (CPM)of Carica papaya roots are used to treat HIV patients
by Thai folk doctors of Thailand. In this research, the water and ethanolic extracts of Carica
papaya roots (female and male trees) were tested in vitro biological effects such as the
antimicrobial activity using disc diffusion assay and minimal inhibitory concentration (MIC) by
using the modified reasazurin assay (1,2) against four species of microorganisms: Bacillus subtilis,
Escherichia coli, Staphylococcus aureus and Candida albicans. The antioxidant activity also
tested by DPPH assay(3). Their anti-inflammatory activity were tested by nitric oxide (NO)
inhibitory activity using RAW264.7 cells (4). Total phenolic content also test by Folin-Ciocalteau
colorimetric method (5). The results found that the ethanolic extract of CPF showed antibacterial
activity against only one of Bacillus subtilis (clear zone= 7 mm , MIC value = 2.5 mg/ml),
contrast with the ethanolic extract of CPM showed antibacterial activity against only one of
Staphylococcus aureus (clear zone= 7.3 mm, MIC value = 2.5 mg/ml). The both water extract of
CPF and CPM had no antibacterial activity. The ethanolic extract and the water extract of CPM
showed antioxidant activity (EC50 of 8.48 and 41.66µg/ml respectively), while the ethanolic
extract and the water extract of CPF had no antioxidant activity (EC50 of > 100µg/ml). The
ethanolic extract of CPM showed highest total phenolic content (83.78 mg/g), as for the water
extract of CPM, CPF and the ethanolic extract of CPF showed low total phenolic content (32.53,
16.71 and 6.86 mg/g respectively). For the water extract of CPF exhibited the highest activity
against the NO inhibitory effect, followed by the ethanolic extract of CPF, the water extract of
CPM and the ethanolic extract of CPM respectively, but the water of CPM showed only one of
toxicity with RAW264.7 cells. These results can supports the traditional used as combination of
female and male trees from Carica papaya roots for treatment of HIV patients because the
ethanolic of CPM showed high antioxidant and total phenolic content , and the both extract of
CPF showed high antibacterial and anti-inflammatory .
[1] Lorian, V. 1996. Antibiotics in Laboratory Medicine. 3rd ed.Williams&Wilkins, Baltimore. McNicholl, B.P.,
McGrath, J.W. and Quinn, J.P. 2006. Water Res, 41 : 127–133.
[2] Sarker, S.D., Nahar, L. and Kumarasamy, Y. 2007. Microtitre plate-based antibacterial assay incorporating
resazurin as an indicator of cell growth, and its application in
the in vitro antibacterial screening of
phytochemicals. Methods,
42:321–324.
[3] Yamazaki, K. Hashimoto, A. Kokusenya, Y. Miyamoto, T. and Sato, T. 1994. Electrochemical method for
estimating the antioxidative effect of methanol extracts of crude drugs. Chem. Pharm. Bull., 42: 1663-1665.
[4] Folin, O. and Ciocalteu, V. 1927. On tyrosine and tryptophan determination in proteins. J. Biol Chem., 27: 627–
650.
[5] Tewtrakul, S. & Subhadhirasakul, S. (2008) Effects of compounds from Kaempferia parviflora on nitric oxide,
prostaglandin E2 and tumor necrosis factor-alpha productions in RAW264.7 macrophage cells. Journal of
Ethnopharmacology, 120, 81–84.
65
P10
Alkaloids of Hippeastrum papilio
Jean Paulo de Andrade1,2, Strahil Berkov1, Kelly B. da Silva2, Liana G. Sachet2, Francesc
Viladomat1, Carles Codina1, José Angelo S. Zuanazzi2, Jaume Bastida1
1
Departament de Products Naturals, Biologia Vegetal i Edafologia, Facultat de Farmàcia, Universitat de
Barcelona, Av. Joan XXIII s/n, E-08028 Barcelona, SPAIN.
2
Faculdade de Farmácia, Universidade Federal do Rio Grande do Sul, Av. Ipiranga 2752, 90610-000,
Porto Alegre, RS, BRAZIL.
[email protected];
The Amaryllidaceae alkaloids constitute a large group of isoquinoline alkaloids with a wide
range of chemical structures and interesting biological properties [1]. The use of galanthamine in
palliative therapy for mild-to-moderate Alzheimer’s Disease (Reminyl®) has prompted a search
for analogous compounds bearing the galanthamine-type skeleton [2] as well as new
Amaryllidaceae species for a sustainable production of this compound [3].
The genus Hippeastrum is endemic to South America, principally Brazil. Earlier studies have
reported the isolation of galanthamine from Hippeastrum species and cultivars, but mainly as a
minor compound.
Hippeastrum papilio, found in the South of Brazil, has been chosen for phytochemical study.,
We have identified six known alkaloids from bulbs and leaves, namely galanthamine (2), which
was found in relatively high quantities, narwedine (3), haemanthamine (4), 11-hydroxyvittatine
(5), 8-O-demethylmaritidine (6) and vittatine (7). We also report the isolation of the new
compound, 11β-hydroxygalanthamine (1).
Alkaloid 1 is an epimer of the alkaloid previously described as habranthine, which was isolated
from Pancratium maritimum with an uncertain stereochemistry of the hydroxyl substituent at
position 11. Using modern 2D NMR, we provide the correct assignment of 1, confirming that the
compound hitherto known as habranthine is in fact 11α-hydroxygalanthamine. Interestingly, 11βhydroxygalathamine (1) has demonstrated a significant ability to inhibit the acetylcholinesterase
enzyme.
[1] Bastida, J.; Lavilla, R.; Viladomat, F. Chemical and biological aspects of Narcissus alkaloids. In: The alkaloids:
Chemistry and Biology. 2006. v.63. p.99-179.
[2]Berkov, S.; Codina, C.; Viladomat, F.; Bastida, J. N-alkylated galanthamine derivatives: potent
acetylcholinesterase inhibitors from Leucojum aestivum. Bioorg Med Chem Lett 2008, 18, 2263-2266.
[3] Berkov, S.; Bastida, J.; Viladomat, F.; Codina, C. Development and validation of GC-MS method for rapid
determination of galanthamine in Leucojum aestivum and Narcissus ssp.: a metabolomic approach. Talanta, 2010,
83, 1455.
66
P11
Activity of compounds extracted from Salvia aurea against
methicillin- resistant Staphylococci.
Schito AM.a, Piatti G.a, Bisio A.b, Giacomelli E.b, Romussi G.b, Pruzzo C.c
a
Dept. of Integr. Surgical and Diagnostic Sciences, University of Genoa, Largo R. Benzi 10, 16132 Genoa,
Italy
b
Dept of Chemistry and Pharmaceutical and Food Technologies, University of Genoa, Via Brigata
Salerno 16147 Genova, Italy
c
Dept of Biology, University of Genoa, Viale Benedetto XV, 5 16132 Genoa, Italy - [email protected]
Methicillin-resistant Staphylococcus aureus (MRSA) and S.epidermidis (MRSE) are Gram positive
pathogens that represent a leading cause of serious human diseases worldwide and are responsible for
substantial morbidity and mortality, both within the healthcare system and in the community [1,2]. We
describe here the activity of three compounds, 12 methoxy-carnosic acid (MCA), 7-methoxy-rosmanol
(MR) and rosmanol (R), derived from S.aurea, on 5 clinical S.aureus strains (2 MRSA and 3 MSSA) and
5 S.epidermidis strains (3 MRSE and 2 MSSE). The three molecules, identified by IR and NMR analysis,
were obtained from the surface exudates by rinsing the fresh aerial part plant material with CH2Cl2 and
repeated column chromatography on Sephadex LH-20 and silica gel. Table 1 shows the minimal inhibitory
concentrations (MICs) of the compounds, determined by standard methods [3]. Killing curve assays, at 4 x
MIC concentrations, were performed on S.aureus and S.epidermidis according to recommended
procedures [3]. A description of the activity expressed on a representative MRSA and MRSE strain is
shown in Fig 1 and Fig 2. MC and R displayed a clear bacteriostatic effect while MR possessed a
bactericidal activity on the same pathogen. On S.epidermidis all tree compounds manifested a
bacteriostatic effect. [31]Our results indicate that MCA, MR and R isolated from S.aurea represent
molecules capable of inhibiting or killing methicillin-resistant Staphylococci. Investigation on their
mechanisms of action are highly warranted.
Tab. 1: MIC values for the indicated compounds
are expressed in μg/ml
Fig 1: S. aureus Oxa-R - time kill curves of a rapresentative MRSA
strain. Compounds were tested at 4 X their respective MIC (log
UFC/ml vs. hours).
Fig 2: S. epidermidis Oxa-R - time kill curves of a rapresentative
MRSE strain. Compounds were tested at 4 X their respective
MIC(log UFC/ml vs. hours).
[1] Mandell, Douglas and Bennett (2005) Principles and Practice of Infectious Diseases, 5th ed.; Mandell, G.,Bennett, J.; Dolin,
R., Eds.; Churchill Livingstone: Philadelphia; pp 2147− 2152 and 2069− 2089
[2] Witte W, Cuny C, Klare I, Nübel U, Strommenger B., Werner G (2008) Emergence and spread of antibiotic-resistant Gram-positive bacterial
pathogens. Int J Med Microbiol. Jul;298(5-6):365-77.
[3] Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; sixteenth informational supplement.
Document M100-S16. Wayne, PA: CLSI; 2006.
67
P12
Influence of benzylaminopurine on antioxidant activity and phenolic
content on in vitro propagated Lamium album L.
Milena Dimitrova, Zhenya Yordanova, Veneta Kapchina-Toteva
Department of Plant Physiology, Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8 Dragan
Tsankov Blvd., 1164 Sofia, Bulgaria
Lamium album L., commonly known as “white dead nettle” is perennial herb from Lamiaceae
family widely used in folk and official medicine. Lamium album L. contains variety of
compounds: phenolics, iridoids, triterpenes, saponines, fatty acids, phytoecdysteroids, essential
oils, tannins, amines. The effect of different concentration (0,1-3,0 mg.l-1) of cytokinin BA (6benzylaminopurine) on in vitro propagation and the amount of secondary metabolites and total
antioxidant activity in Lamium album L. was examined. All concentrations of BA suppressed root
formation and stimulated callusogenesis. Stimulation of shoot number was observed on MS
medium supplemented with 1.6 and 2,6 mg.l-1 BA compared with control plants propagated on
MS medium without plant growth regulators. Shoots cultivated on MS medium supplemented
with 1.4 and 2.0 mg.l-1 BA showed increased content of total phenols. The highest concentrations
of flavonoids were obtained in MS medium supplemented with BA from 1.6 up to 2.0 mg.l-1. The
highest content of antioxidant activity was achieved on MS medium supplemented with BA 1.4
and 1.6 mg.l-1 compared with control plants. In addition, the amount of secondary metabolites and
antioxidant capacity was established in formed callus in underground part of in vitro propagated
Lamium album plants. Significant stimulation of total phenols in callus was observed on MS
medium supplemented with BA in concentrations - 0.5, 0.8 mg.l-1 and 2.2, 2.6 mg.l-1. The highest
content of antioxidant activity was achieved on MS medium supplemented with BA - 0.2, 0.5 and
0.8 mg.l-1. The amount of flavonoids in callus was approximately threefold lower than that
established in shoots. Therefore the higher concentrations of BA enhanced content of phenolic
compounds with high antioxidant activity in shoots, but lower concentrations of BA (0.2, 0.5 and
0.8 mg.l-1) stimulated accumulation of secondary metabolites in formed callus.
This work was financially supported by the grand № DTK-02-29/2009 of Ministry of Education,
Bulgaria.
68
P13
Currant seed extracts showing protective effect on human
lymphocytes DNA
Dejan Gođevaca, Vele Teševićb, Vlatka Vajsa, Slobodan Milosavljević, b Gordana Zdunić c,
Miroslava Stankovićd
a
Institute of Chemistry, Technology and Metallurgy, Njegoševa 12, University of Belgrade, Serbia
Faculty of Chemistry, Studentski trg 12-16, University of Belgrade, Serbia
c
Institute of Nuclear Science ″Vinča″, University of Belgrade, Serbia
d
Institute for Medicinal Plants Research, Tadeuša Košćuška 1, 11000 Belgrade, Serbia
[email protected]
b
On the basis of LC/UV/MS analysis, 35 compounds from the extracts of seeds of six cultivars
of currants (two cv. of black, red and white currant) were identified. Black currants cultivars
contained protocatechuic acid and p-hydroxibenzoic acid, and traces of nitril containing phenolic
acids. Synapoyl glucoside is characteristic for cv. Malling Jewel. Dihydrophaseic acid glycosides
and carboxymethylindol glycosides were present mainly in white and red currant cultivars.
Blackcurrant seeds contained higher amount of flavonoids, especially rutin, isoquercetin and
taxifolin.
The currant seed extracts were tested for in vitro protective effect on chromosome aberrations
in peripheral human lymphocytes using cytochalasin-B blocked micronucleus (CBMN) assay. [1]
The frequency of MN was scored in binucleated cells, and the best antioxidant potential showed
the cultivars of black currants. At a concentration of only 0.17 mg/ml, extract of the seed cv.
Malling Jewel effected a decrease in the frequency of MN for 60% compared with control cell
cultures.
These findings could support the potential benefits of currant seed extracts and their
constituents in a multitude of disease states, including cancer, cardiovascular disease, and
neurodegenerative disorders.
[1] Fenech M., Morley AA. (1993) The cytokinesis-block micronucleus technique: a detailed description of the
method and its application to genotoxicity studies in human populations. Mut. Res. 285: 35-44
69
P14
Inhibitory effect of Kaempferia parviflora on NFкB, Rel A, NOS,
D mRNA expression
COX-2 and TNF-D
Chutha Sae-wong, Supinya Tewtrakul, Pimpimon Tansakul
Faculty of Pharmaceutical Sciences, Prince of Songkla University, Songkhla 90112, Thailand
[email protected]
Nuclear factor-kappa B (NFкB) is dimeric complexes of p50/RelA (p50/p65) which plays an
important role in controlling inflammatory gene activation[1] such as inducible nitric oxide
synthase (iNOS) and cyclooxygenase (COX-2)[2] which are involve in inflammation process. To
clarify the anti-inflammatory mechanism of the active compounds from Kaempferia parviflora,
the effects of 5,7-dimethoxyflavone, trimethylapigenin and tetramethylluteolin on the expression
of mRNA were examined using semi-quantitative analysis of RT-PCR technique. The result
revealed that 5,7-dimethoxyflavone decreased cellular iNOS, COX-2 and Rel A mRNA level and
this compound also affected the β-actin mRNA expression. It was indicated that inhibitory effect
of this compound cause of fault positive effect. Trimethylapigenin dose-dependently suppressed
iNOS, COX-2 and Rel A mRNA level, while inhibitory effect on NFкB was partly affected at the
highest concentration. Tetramethylluteolin selective inhibited Rel A, but slightly inhibited iNOS
mRNA, while the expression of COX-2, NFкB and TNF-D was not affected [Figure 1]. In
conclusion, the mechanism of anti-inflammation of K. parviflara is due to the suppressive effect
of trimethylapigenin mainly through the inhibition of COX-2 enzyme and the selective inhibitory
effect of tetramethylluteolin through Rel A mRNA expression.
Conc
N
C
3
10
30 100
N
C
3
10 30 100
N
C
3
10
30 100 µg/ml
iNOS
COX-2
NFкB
Rel A
TNF-D
β-actin
(A)
5, 7-Dimethoxyflavone
(B)
Trimethylapigenin
(C)
Tetramethylluteolin
Figure 1 Effect of 5, 7-dimethoxyflavone (A), trimethylapigenin (B) and tetramethylluteolin (C) on the expression of
mRNA
[1] Lima T.M., Gorjao R., Hatanaka E., Cury-Boaventula M.F., Silva E.P.P., Procopio J., et al. (2007). Mechanisms
by which fatty acids regulate leucocyte function. Clinical Science 113: 65-77.
[2] Haridas, V., Arntzenm, C.J., Gutterman, J.U. (2001). Avicins, a family of triterpenoid saponins from Acacia
Victoriae (Bentham), inhibit activation of nuclear localization and ability to bind DNA. Proceedings of the
National Academy of Sciences of the United States of America 98: 557-562.
70
P15
Antioxidant activity of Uva di Troia Canosina ad acino piccolo:
comparison of different extraction methods
Filomena Corboa, Angelo Lovecea, Claudio Brunoa, Flavia la Forgiab, Sergio Fontanab,
Mariagrazia Rosellia, Antonio Laghezzaa, Giovanni Lentinia, Carlo Franchinia, Solomon
Habtemariamc
a
Dipartimento Farmaco Chimico, Università degli Studi di Bari, via Orabona 4, 70125 Bari, Italy
Farmalabor S.R.L. Ricerca e Sviluppo via Pozzillo, Z.I. 70053 Canosa di Puglia (Bari), Italy
c
Pharmacognosy Research Laboratories, Medway School of Science, University of Greenwich Central
Avenue Chatham Maritime Kent ME4 4T
b
Over the past few years a big effort has been done for the valorisation of food, agricultural
residues, and wastes by creating added-value using bio-products, bio-fuels, or food ingredients.
Grape is one of the world’s largest fruit crops with a global production of around 55 million
tonnes in 2008 and 26 million tonnes of wine.1 This also generates an enormous amount of waste,
including grape skins and seeds.
Polyphenols, contained in grapes, exhibit a wide range of biological effects as antioxidants,
antimicrobials, and modulators of various enzyme systems. Beneficial effects for human health
have been demonstrated in experimental studies on LDL-cholesterol oxidation, platelet
aggregation, cardiovascular diseases,2–4 and type 2 diabetes.5
Skins and seeds of Uva di Troia Canosina cultivar from red-vinification were used in this
work. The dried marc was extracted by both traditional and microwave-assisted extraction (MAE)
techniques, the extracts obtained were analyzed by capillary zone electrophoresis (CZE) and
evaluated as antioxidant agents by the DPPH assay.
[1] OIV (2009). Organisation internationale de la vigne et du vin. <http://www.oiv.int>
[2] Leifert, W.R., Abeywardena, M.Y., (2008) Cardioprotective actions of grape polyphenols, Nutrition Research, 28,
729–737
[3] Anastasiadi, M., Pratsinis, H., Kletsas, D., Skaltsounis, A.L., Haroutounian, S.A., (2009) Bioactive non coloured
polyphenols content of grapes, wines and vinification by-products. Evaluation of the antioxidant activities of their
extracts. Food Research International, doi:10.1016/j.foodres.2009.11.017
[4] Iriti, M., Faoro, F., (2010) Bioactive chemicals and health benefits of grapevine products. Bioactive Foods in
Promoting Health, 581, 620
[5] A. M. Hodge, D. R. English, K. O’Dea and G. G. Giles, (2006) Diabetic Med., 23, 690–697
71
P16
Nitric oxide inhibitory effect and antioxidant activity of the extracts
from Musa sapientum bracts and flowers
Pathompong Phuakleea, Arunporn Itharatb
a
Graduated School, Faculty of Medicine, Thammasat University.
b
Department of Applied Thai traditional Medicine, Faculty of Medicine, Thammasat University,
Thailand
[email protected]
Musa sapientum Linn. (Musaceae) or banana has been commonly used as health food and
drugs in Thai traditional medicine. For the example, bract and flower parts of Musa sapientum
Linn. were used as drugs and health foods for postpartum care. Thus, the objective of this
research was investigated anti-inflammatory activity by determination on inhibitory effect on
Nitric oxide production on RAW264.7 cell which were stimulated by lipopolysaccharide (LPS)[1]
and tested antioxidant activity by using DPPH radical scavenging assay[2] of bract and flower
parts of banana extracts. The dried bracts and flowers of Musa sapientum were extracted by
macerated with 95% ethanol, 50%ethanol, soaking in water and boiling in water or decoction. The
results were found that the water extract by soaking of flower showed the highest antiinflammatory activity, with an IC50 of 9.75 µg/ml. The decoction extract of flower showed the
highest antioxidant activity, with an EC50 of 12.35 μg/ml, followed by the 95% ethanolic and
soaking extract of flowers, with EC50 values of 15.57 and 22.86 μg/ml, respectively. It was
concluded that the flower of Musa sapientum Linn. may be considered potentially useful as a
source of natural anti-inflammatory and antioxidant ingredients and it was support using in Thai
traditional medicine as food and anti-inflammation for postpartum care.
[1] Tewtrakul, S., and Subhadhirasakul, S. Effects of compounds from Kaempferia parviflora on
nitric oxide, prostaglandin E2 and tumor necrosis factor-alpha productions in RAW 264.7
macrophage cells. Journal of Ethanopharmacology, 2008; 120:81-84.
[2] Yamasaki, K., Hashimoto, A. Kokusenya, Y., Miyamoto, T. and Sato, T. Electrochemical
method for estimating the antioxidative effects of methanol extract of crude drugs, Chem.
Pharm. Bull.1994; 42:1663-1665.
72
P17
Correlation between phenolic content and antioxidant activity of
infusions from eleven Mediterranean plant species
Sandra Gonçalves, Diogo Gomes, Patrícia Costa, Anabela Romano
Institute for Biotechnology and Bioengineering, Centre of Genomics and Biotechnology (IBB-CGB),
Faculty of Sciences and Technology, University of Algarve, Campus de Gambelas, Ed. 8, 8005-139 Faro,
Portugal.
[email protected]
Antioxidants scavenge free radicals by initiating and propagating oxidative chain reactions,
thus delaying or preventing intracellular oxidative damage [1]. Natural antioxidants such as
phenolics and flavonoids have been shown to possess multiple pharmacological activities through
their antioxidant properties, including neuroprotective and anti-ageing activities. Highly
heterogeneous soil and climatic conditions of the Mediterranean region have resulted in an
increased diversity of plant species, most of them with medicinal properties that deserve further
investigation. In this study, the total phenolic content of extracts from eleven Mediterranean
plants and their related antioxidant activities were determined. The extracts were prepared as
infusions (cold and hot), as they are traditionally used in popular medicine. The infusions were
investigated for their effect on deoxyribose degradation, reducing properties and Fe2+-chelating
abilities. The free radical scavenging activity and the total phenolic contents were also determined
by using DPPH and Folin-Ciocalteu assays, respectively. The capacity of hot infusions to prevent
Fe2+-induced lipid peroxidation in mouse´s brain (in vitro) was also investigated. The correlation
between phenolic content and the antioxidant activity, obtained with each method, was calculated
according to Pearson’s test. The phenolic content was significantly higher in hot infusions than in
cold infusions of all the species. Among the species studied, the highest phenolic contents were
observed in Arbutus unedo (1693.1 r 118.2 and 2256.8 r 15.6 µmolGAEgps-1 in cold and hot
infusions, respectively), Myrtus communis (1006.1 r 54.4 and 1244.6 r 30.6 µmolGAEgps-1) and
Pistacia lentiscus (759.5 r 74.4 and 1359.1 r 33.2 µmolGAEgps-1). The infusions from these
species also showed the highest Fe2+ chelating ability, reducing power, as well as DPPH freeradical scavenging and inhibitory activity toward lipid peroxidation. A positive correlation was
found between total phenolic content and antioxidant activity obtained with all the methods
excepting hydroxyl radical-scavenging (deoxyribose degradation). The results of this research
represent a good basis for further analysis of the selected plants to discovery new antioxidant
compounds.
S. Gonçalves and P. Costa acknowledge a grant from Portuguese Science and Technology Foundation (FCT,
SFRH/BPD/31534/2006 and SFRH/BD/63505/2009).
[1] Valko M., Leibfritz D., Moncola J., Cronin M.T.D., Mazura M., Telser J. (2007). Free radicals and antioxidants in
normal physiological functions and human disease. Int. J. Biochem. Cell Biol. 39: 44–84.
73
P18
New natural compounds of Dionaea muscipula as anticancerous
agents
François Gaaschta, Monika Jain a, Marie-Hélène Teiten a, Marc Schumacher a, Gilbert
Kirsch b, Denyse Bagrel c, Mario Dicato a, Marc Diederich a
a
Laboratoire de Biologie Moléculaire et Cellulaire du Cancer, Hôpital Kirchberg, 9 rue Edward Steichen,
L-2540 Luxembourg, Grand Duchy of Luxembourg
b
Laboratoire d’Ingénierie Moléculaire et Biochimie Pharmacologique, Université Paul Verlaine de Metz,
Technopôle, 1 Boulevard Arago, F-57070, Metz, France
c
Laboratoire d’Ingénierie Moléculaire et Biochimie Pharmacologique, Université Paul Verlaine de Metz,
Campus Bridoux, Rue du Général Delestraint, F-57070, Metz, France
Since centuries, natural compounds are used in traditional medicines. Recent studies showed
that such substances are able to block the appearance and the development of many diseases such
as cancers. The discovery and characterization of new molecules will permit to fill out the drug
library of new therapeutic molecules that needs to become greater due to the emergence of new
diseases and the appearance of resistance towards conventional treatments.
The Venus Flytrap (Dionaea muscipula Solander Ex Ellis) is the most well known carnivorous
plant. Inherent from the United States, this plant catches and digests small preys with an active
trap. It was already described that this plant contains many therapeutic molecules previously
described in other plants like myricetin, quercetin, plumbagin and ellagic acid but any specific
natural compound has been discovered till today in Dionaea muscipula. The aim of this project is
to discover and to characterize new and/or specific therapeutic molecules issued from Dionaea
muscipula.
We prepared methanolic leaves extracts of Dionaea muscipula. These extracts were shown to
be uptaken by K562 cells and to be cytotoxic toward five different leukaemia cell lines (HL-60,
Jurkat, K562, Raji, U937) and four adherent cancer cell lines: A549 (lung), MDAMB-231
(breast), PC-3 (prostate), SAOS-2 (bone) but have no effect on HT-29 cells (colon). Purification
of the extract by preparative HPLC-MS has shown that the extract contains 7 major compounds
and that among these different substances, at least one is unknown and exerts cytotoxicity towards
K562 cells. Analysis of cell death by fluorescence microscopy after Hoechst/propidium iodide
staining showed that this cytotoxic fraction induces cell death mainly by necrosis. Preliminary
results showed that this fraction was till now unknown as therapeutic substance and will be
identified by NMR.
74
P19
Effects of a rice bran enzymatic extract on cytokines implicated in
the inflammatory state of adipose tissue in a rat model of obesity.
Candiracci Ma., Parrado Ja., Justo M.L. b, Rodríguez-Rodríguez R. b, Castaño A.a and
Herrera M.D.b
a
Departamento de Bioquímica y Biología molecular.
Departamento de Farmacología. Facultad de Farmacia. Universidad de Sevilla. C/ Profesor García
González, nº 2, 41012- Sevilla. Spain.
b
Obesity is characterized by a chronic low-grade inflammation in adipose tissue that is believed
to contribute to the genesis of insulin resistance, leads to type 2 diabetes, a risk factor for
cardiovascular diseases. The inflammation of the adipose tissue is mainly due an influx of
macrophages that secrete proinflammatory cytokines as tumor necrosis factor alpha (TNF-α),
interleukin-6 (IL-6), interleukin 1 beta (IL-1 β) (1, 2).
At the same time, it is well-known that soluble factors released by adipocytes have roles at the
interface between the immune and the metabolic systems. In this sense, adiponectin is an antiinflammatory factor known to be decreased in individuals with visceral obesity and states of
insulin-resistance (3).
Attenuating inflammation in adipose tissue beneficially modifies metabolic disturbances and
reduces disease risk, even in the absence of obesity reduction (4, 5). We have previously
described that obese Zucker rats fed on Rice Bran Enzymatic Extract enriched-standard diet
(RBEE) ameliorated metabolic parameters related to insulin resistance and cardiovascular disease.
Thus, the main goal of present work was to investigate the ability of RBEE to prevent the
inflammatory state in the adipose tissue.
Our results show that RBEE-diets (1% or 5%), significantly decrease the upregulation of
mRNAs of inflammatory cytokines (TNF-D, IL-6) found in obese Zucker rats. However, RBEEdiets did not reverse the downregulation of adiponectin found in obese Zucker rats. In conclusion
we hypothesize that RBEE has a beneficial effect on inflammatory state in the adipose tissue that
is mainly carried out on macrophages-derived cytokines.
[1] Hotamisligil GS, Shargill NS, Spiegelman BM. (1993) Science, 259:87-91.
[2] Bastard JP, Maachi M, Van Nhieu JT, Jardel C, Bruckert E, Grimaldi A, et al. (2002) J Clin Endocrinol Metab.
87: 2084-9.
[3] Tilg H, Moschen A.R. (2006) Nature Reviews. 6: 772-83.
[4] Kanda H, Tateya S, Tamori Y, Kotani K, Hiasa K, Kitazawa R, Kitazawa S, Miyachi H, Maeda S, Egashira K,
Kasuga M. (2006) J Clin Invest. 2006; 116: 1494–1505.
[5] Todoric J, Loffler M, Huber J, Bilban M, Reimers M, Kadl A, Zeyda M, Waldhausl W, Stulnig TM. (2006)
Diabetologia 49: 2109-19.
75
P20
Metabolic engineering of carotenoid biosynthesis in riceendosperm
Chao Bai1, Sol M Rivera2, Vicente Medina1, Gerhard Sandmann3, Ramon Canela2, Changfu
Zhu1, Teresa Capell1 and Paul Christou1, 4
1
Departament de Producció Vegetal i Ciència Forestal, Universitat de Lleida-CRA, Spain
Departament de Química, UdL, Av. Alcalde Rovira Roure, 191, Lleida, 25198, Spain
3
Molecular Biosciences, J.W. Goethe Universitaet, Frankfurt am Main, Germany
4
Institucio Catalana de Recerca i Estudis Avancats, Barcelona, Spain
[email protected]
2
Vitamin A is an essential nutrient in the human diet. Although many crops are rich sources of
pro-vitamin A, millions of people rely on staple crops that contain minimal levels of this nutrient,
for example rice. Several strategies have been used to enhance carotenoid accumulation in plants.
The amounts of carotenoids in plant tissues and organs do not appear to depend solely on
carotenogenic enzyme activities responsible for their synthesis. The upstream precursor (MEP
derived IPP and GGPP) pathway may also positively influence their accumulation [1], while
downstream degradation pathways may deplete the carotenoid pool [2]. The identification and
characterization of a novel gene mutation in cauliflower (or) which appears to mediate the
differentiation of proplastids and/or non-colored plastids into chromoplasts accompanied by a
concomitant accumulation of carotenoids indicated that creating a metabolic sink to sequester
carotenoids is another important mechanism for their accumulation in plants [3]. In order to
investigate the mechanisms of carotenoid accumulation and enhance carotenoid content in rice
endosperm, we genetically engineered rice (Oryza sativa) with the Arabidopsis thaliana orange
gene (Ator)/A. thaliana 1-deoxy-D-xylulose 5-phosphate synthase (Atdxs); Zea mays phytoene
synthase 1 (Zmpsy1), and Pantoea ananatis phytoene desaturase (PacrtI). Here we report
preliminary results from early experiments which demonstrate that co-transformation is a useful
method for engineering such complex traits in rice plants.
[1] Enfissi EMA., Fraser PD., Lois LM., Boronat A., Schuch W., Bramley PM. (2005) Metabolic engineering of the
mevalonate and non-mevalonate isopentenyl diphosphate-forming pathways for the production of healthpromoting isoprenoids in tomato. Plant Biotechnol J, 3: 17–27
[2] Auldridge ME., Block A., Vogel JT., Dabney-Smith C., Mila I., Bouzayen M., Magallanes-Lundback M.,
DellaPenna D., McCarty DR., Klee HJ. (2006) Characterisation of three members of the Arabidopsis
carotenoid cleavage dioxygenase family demonstrates the divergent roles of this
multifunctional enzyme family. Plant J, 45: 982–993
[3] Li L., Van Eck J. (2007) Perspectives: metabolic engineering of carotenoid accumulation by creating a
metabolic sink . Transgenic Research, 16: 581-585
76
P21
Effect of quercetin on hepatic steatosis in ob/ob mice
Jung-In Kim, Jihye Kim, Haneul Choi, Min-Jung Kang, Soomi Jeong, Minju Kim
School of Food and Life Science, Inje University, Gimhae, South Korea
[email protected]
Obesity and hypertriglycerinemia are risk factors associated with non-alcoholic fatty liver
disease (1). Quercetin (QE) exerts anti-hypertriglyecmic effect in mice fed high cholesterol diet
(2). The aim of this study is to investigate effect of QE on obesity, hyperlipidemia, and hepatic
steatosis in ob/ob mice. Four-week-old male ob/ob mice (n=16) and their lean littermates (Leancontrol, n=8) were purchased. After 1 week of adaptation, ob/ob mice were fed either AIN-93G
diet (ob/ob-control) or a diet containing QE at 0.08% for 10 weeks. Lean-control animals were fed
AIN-93G diet. Animals were sacrificed after an overnight fast. Serum levels of triglycerides,
cholesterol and adiponectin and activity of alanine amininotransferase (ALT) were measured.
Liver lipids were extracted and triglyceride levels were measured. The data were analyzed by oneway ANOVA and Tukey’s test (p<0.05). Consumption of QE did not influence body weight of
ob/ob mice significantly. Serum triglyceride, cholesterol, and ALT and hepatic triglyceride levels
of ob/ob-control group were significantly higher than lean-control group. Consumption of QE
significantly decreased serum triglyceride, cholesterol, and ALT and hepatic triglyceride levels
compared with ob/ob control group. Serum adiponectin of ob/ob mice was lower than lean-control
group. Reduced serum ALT of ob/ob mice was significantly increased by consumption of QE.
These results indicate that QE could alleviate hyperlipidmia and hepatic steatosis in ob/ob mice.
Table 1. Serum lipid profile, adiponectin, and ALT and hepatic triglycerides
Group
Lean-control
ob/ob-control
ob/ob-QE
Serum triglycerides (mg/dL)
64.0 ± 10.6 a
134.3 ± 17.4 c
107.0 ± 14.7 b
Serum cholesterol (mg/dL)
90.4 ± 14.2 a
171.7 ± 25.6 c
142.0 ± 19.8 b
Serum adiponectin (Ɇg/mL)
13.8 ± 2.4 b
7.6 ± 1.3 a
11.0 ± 1.6 b
Serum ALT (IU/L)
49.4 ± 10.4 a
197.6 ± 35.0 c
139.5 ± 24.0 b
Hepatic triglycerides (mg/g liver)
13.0 ± 2.9 a
65.5 ± 1.6 c
42.1 ± 5.9 b
mean ± SD. P<0.05
[1] Angulo P. (2002) Nonalcoholic fatty liver disease. N Engl J Med, 16: 1221-1231
[2] Azuma K, Ippoushi K, Terao J. (2010) Evaluation of tolerable levels of dietary quercetin for exerting its
antioxidative effect in high cholesterol-fed rats. Food Chem Toxicol, 48: 1117–1122
77
P22
Olive oil phenols improve the altered human intestinal permeability.
Study in vitro
P. Trisolini1, A. Ignazzi1, F. Pezzolla1, D. Lorusso1, S. Fiume2, L. Debellis2 and M.A. Maselli1
1
Scientific Institute of Gastroenterology “S de Bellis”, Castellana Grotte, Bari; 2 Dept. of General and
Environmental Physiology, University of Bari.
In ex vivo and in vitro models olive oil phenols Hydroxytyrosol (HT) has shown to have role of
"radical scavenger" and to be also involved in the inhibition of oxidation processes related to
alteration cellular aging and degenerative diseases[1, 2]. However nothing is known about HT
participation on human colonic mucosa transport process. Therefore, we tested the effects of HT
on electrophysiological parameters of non stimulated and stimulated human colon.
Segments of sigmoid colon were obtained from 6 patients (78 mean age yrs, range 75-81 yrs),
undergoing left hemicolectomy for non-obstructive sigmoid cancer. Colonic mucosa were taken
from macroscopically normal areas. Colonic mucosa deprived of the muscle layer was mounted in
Ussing chambers and bathed in oxygenated Krebs solution at 37°C. Functional parameters:
transepithelial voltage (Vt), short-circuit current (Icc) and transepithelial resistance (Rt), were
recorded in resting condition and under stimulation with CARB (100PM), forskolin and IBMX in
the absence or in the presence of HT (1PM, 10PM e 100PM). Statistical analysis was performed
by paired t-Student test. P<0.05 was considered to be statistically significant.
The exposure to HT in non stimulated tissues produced not significant variations of the Vt, Icc
and transepithelial resistance.The exposure to HT in CARB, forskolin and IBMX stimulated
tissues produced a progressive dose dependent reduction of the Vt significant at 100PM (p<0.05)
and 10PM (p<0.01); a not significant reduction of the Icc at the same concentrations and a
significant change in transepithelial resistance at 100PM (p<0.05).
HT could help improve the altered intestinal permeability through a reduction of the transport
activity and paracellular permeation in stimulated and non stimulated human colon respectively.
[1] Dowd Y et al. (2004) Bioc. Pharm. 68:2003-8
[2] Fabiani R et al. (2002) Eur. J. Cancer Prev.11:351-8
78
P23
Phenolic composition of Trifolium scabrum L. aerial parts.
Iwona Kowalska1, Dariusz Jędrejek 1 , Łukasz Cieśla 1 , Sonia Piacente2, Milena Mosullo2,
Wieslaw Oleszek1 and Anna Stochmal 1
1
Department of Biochemistry, Institute of Soil Science and Plant Cultivation, State Research Institute, ul.
Czartoryskich 8, 24-100 Pulawy, Poland
2
Department of Pharmaceutical Sciences, University of Salerno, via Ponte Don Melillo-84084, Fisciano,
Salerno, Italy
[email protected]
Flavonoid-derived plant natural products have long been known to function as floral pigments
for the attraction of insect pollinators, as signal molecules for beneficial microorganism in the
rhizosphere, and as antimicrobial defense compounds. Isoflavones are naturally occurring
diphenolic compounds found mainly in the legume family of plants and represent the main class
of phytoestrogens of current interest in clinical nutrition.
Trifolium pratense L. (red clover) is (besides soybean) one of the plant sources of these
compounds. Extracts of Trifolium pratense L. are commercially available as isoflavone enriched
dietary supplements meant for long-term use as a phytoestrogen source and ’’natural’’ form of
hormone replacement therapy. Preliminary analysis of 57 Trifolium species for isoflavone
concentration showed that Trifolium scabrum L. (rough clover) has higher total isoflavone
contents than Trifolium pratense L. 72.76 and 39.51 mg/g dry matter, respectively [1]. Thus, we
suggest that Trifolium scabrum L. can be considered as natural and very good commercial source
of these compounds.
The aim of the present research was to isolate and identify phenolic constituents of T. scabrum
L. aerial parts. It was cultivated in an experimental field of the Institute of Soil Science and Plant
Cultivation in Pulawy, Poland. Plants were harvested at the beginning of flowering. The plant
material was lyophilized, finely powdered, and used for the successive extraction according to
previously developed techniques [2]. Phenolic fractions were separated by C18 preparative
column chromatography into individual isoflavonoids. Purity of isolated compounds was
identified by spectra analysis (UV, ESI-MS/MS, 1H NMR) and by comparison with the literature
data. Among fifteen isolated and characterized substances from the extract of Trifolium scabrum
L., thirteen were isoflavones or isoflavone glucosides and two were flavonol glucosides.
[1] Oleszek W., Stochmal A., Janda B. Concentration of isoflavones and other phenolics in the aerial parts of
Trifolium species. 55, 8095-8100, 2007.
[2] Stochmal, A., Piacente, S., Pizza, C., Riccardis, F., Leitz, R., Oleszek, W. Alfalfa (Medicago sativa L.)
flavonoids. 1. Apigenin and luteolin glycosides from aerial parts. J. Agric. Food Chem., 49, 753-758, 2001.
79
P24
Evaluation of antioxidant and free radical scavenging potential of
Lamium album L. growing in Bulgaria
Miglena Spasimirova Valyovaa, Milena Angelova Dimitrovab, Yordanka Angelova Ganevaa,
Veneta Mihova Kapchina-Totevab, Zhenya Petkova Yordanovab
a
Department of Organic Chemistry, Faculty of Chemistry, Sofia University “St. Kliment
Ohridski”, 1 James Bourchier Blvd., 1164 Sofia, Bulgaria.
b
Department of Plant Physiology, Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8
Dragan Tsankov Blvd., 1164 Sofia, Bulgaria.
In vitro antioxidant capacity and total phenolic and flavonoid contents of different extracts,
obtained from in vivo and in vitro cultivated plants of Lamium album L. were tested. Lamium
album L. is valuable medicinal plant which possesses astringent, antispasmodic, antiinflammatory, antibiotic and bacteriostatic properties. Three different assays (DPPH-free radical
scavenging activity, ABTS-free radical scavenging activity and Phosphomolybdenum redusing
power) for screening of antioxidant activity were applied. The highest phenolic content was
detected in in vivo methanol extracts, obtained by thermostat and Soxhlet extractions (MET and
MES). The highest concentrations of flavonoids were obtained in in vivo MET and MES (62.05 ±
2.25 and 82.11 ± 4.3, respectively). In vivo MET and MES demonstrated the highest DPPH
scavenging potentials than other extracts with IC50 values of 20.62 ± 0.49 and 29.86 ± 0.15
μg.ml-1, respectively. The highest ABTS scavenging potential in in vivo and in vitro L. album L.
extracts were found for MES and MET (0.65 ± 0.009 and 0.60 ± 0.036 TEAC). The results
indicated that the highest reducing potential towards transition metals was found in in vivo ethanol
extract (237.67 ± 1.66 mM.g-1) obtained by thermostat extractions. The total phenolic content
correlated with the radical scavenging activity. The results revealed that L. album L. could be an
interesting source of natural antioxidants with potential use in food supplements. To the best of
our knowledge this is the first report which indicates total phenolic and flavonoid contents,
antioxidant capacity of in vitro propagated plants of L. album L. by comparison with in vivo
grown plants.
This work was financially supported by the grand No. DTK-02-29/2009 of Ministry of Education Youth
and Science, Bulgaria.
80
P25
Evaluation of the saponin content in lines of common beans
(Phaseolus vulgaris L.) selected in Italy
Aldo Tavaa, Barbara Pintusa, Agata Ursinoa, Bruno Campionb
a
CRA-FLC Centro di Ricerca per le Produzioni Foraggere e Lattiero Casearie, v.le Piacenza 29,
26900 Lodi, Italy
b
CRA-ORL Unità di ricerca per l’Orticoltura, via Paullese 28, 26836 Montanaso Lombardo (LO),
Italy
[email protected]
The common bean (Phaseolus vulgaris L.) is one of the most important staple food worldwide,
mainly thanks to its relatively high content of protein and other valuable nutrients such as
complex carbohydrates, minerals, together with some vitamins that are of particular relevance for
human health. Beans also contain other secondary metabolites such as phytates, protease
inhibitors, polyphenols, lectins, raffinosaccharides and saponins that can affect the nutritional
value of this legume [1].
In a series of breeding studies aimed at obtaining bean grains with improved nutritional
properties, several lines of P. vulgaris were selected and characterized by a low content of
antinutritional factors such as lectines and phytates [2,3].
From a nutritional point of view, among all reported metabolites, saponins might represent a
positive trait due to their ability to lower plasma LDL cholesterol concentration by interfering
with cholesterol absorption [4].
The present paper reports an evaluation of saponin/sapogenin content of Italian lines of
common beans selected for their improved nutritional properties.
[1]Norton G., Bliston FA., Bressani R. (1985) Biochemical and nutritional attributes of grain legumes. In:
Summerfield RJ., Roberts EH. ed. Grain legume crops, Collins, London.
[2] Campion B., Perrone D., Galasso I., Bollini R. (2009) Common bean (Phaseolus vulgaris L.) lines devoid of
major lectin proteins. Plant Breeding, 128: 199-204
[3] Campion B., Sparvoli F., Doria E., Tagliabue G., Galasso I., Fileppi M., Bollini R., Nielsen E. (2009) Isolation
and characterization of an lpa (low phytic acid) mutant in common bean (Phaseolus vulgaris L.). Teor Appl
Genet, 118: 1211-1221
[4] Carr TP., Jesch ED. (2006) Food components that reduce cholesterol absorption. In: Taylor SL. ed. Advances in
food nutrition research. Vol 51. Academic Press, London.
81
P26
Glucosinolates from “Mugnolo”, a variety of Brassica oleracea
Argentieri Maria Pia., Pascale Rossana, Avato Pinarosa
Dipartimento Farmaco-Chimico,Università degli Studi “Aldo Moro”di Bari- Via Orabona 4,I- 70125 Bari
– ITALY
[email protected]
Plants of the Brassicaceae family are widespread in the Mediterranean region where they are
widely consumed cooked or as salad vegetables (cabbage, Brussel sprouts, cauliflower, broccoli,
turnips, radishes, cress) or condiments (horseradish, mustard, capers). These plants are a good
source of bioactive compounds of nutraceutical relevance including flavonoids, minerals,
vitamins, and glucosinolates (GLSs). These are responsible for the plant pungent aroma and bitter
taste and are the most extensively studied bioactive compounds in the Brassicaceae family. GLSs
consist of a large group of sulphur-containing secondary metabolites with cancer protection
properties [1]. In recent years, GLSs, and particularly their hydrolysis products isothiocyanates,
have attracted great attention in the scientific world due to their potential anticarcinogenic effects
[2]. Sulforaphane, the isothiocyanate resulting from the hydrolysis of glucoraphanin, the main
GLS in broccoli (B. oleracea var italica) has been shown to reduce tumor formation induced by
chemical carcinogens in several animal models [3].
Within the Brassica genus, B. oleracea includes a number of varieties which, due their
importance as edible crops, have been studied for their content of GLSs. These include B.
oleracea var gemmifera (Brussels sprouts), var capitata (cabbage), var acephala (kale) and
particularly B. oleracea var italica (broccoli). In this study we have analyzed the metabolic profile
of a poorly studied variety of B. oleracea locally called “mugnolo”, traditionally cultivated in
Southern Italy (Puglia region) and largely consumed in the diet. Results obtained by HPLC and
ESI-MS/MS demonstrate that “ mugnolo” inflorescences are characterized by the presence of the
aliphatic GLSs glucoraphanin (1.79 μmol/g), glucoiberin, glucoerucin, and sinigrin, of the
aromatic GLSs glucobarberin (0.56 μmol/g) and gluconasturtin, and of indole GLSs
glucobrassicin (3.51 μmol/g), neoglucobrassicin, 4-methoxyglucobrassicin, and 4hydroxyglucobrassicin.
We also analyzed the GLS profile of “mugnolo” after cooking in water. The results showed
variations in the GLSs content with a 50% fall of their concentration [4].
[1] Moreno DA, Carvajal M, Lopez-Berenguer C, Garcìa-Viguera C. (2006) Chemical and biological characterization
of nutraceutical compounds of broccoli. J Pharm Biomed; 41: 1508–1522.
[2] Cartea ME, Velasco P. (2008) Glucosinolates in Brassica food: bioavailability in food and significance for human
health. Phytochem Rev; 7: 213–229.
[3] Jeffery EH, ArayaM. (2009) Physiological effects of broccoli consumption. Phytochem Rev; 8: 283–298.
[4] Argentieri M.P., Accogli R., Fanizzi F.P., Avato P. (2011) Glucosinolates profile of “mugnolo”, a variety of
Brassica oleracea L. native to Southern Italy (Salento). Planta Med; 77: 287-292.
82
P27
Antioxidant properties of Saffron against H2O2- & selenite-induced
oxidative stress: in vitro & in vivo studies
Fotini N. Lamari1, Magdalini A. Papandreou2, Andriana P. Dimakopoulou1, Evangelos
Drimtzias3, Anastasia-Barbara Ferlemi1, Olga Makri3, Paul Cordopatis1, Costas
Georgakopoulos3, Marigoula Margarity2
1
Department of Pharmacy, 2Department of Biology, 3School of Medicine, University of Patras,
26504 Patras, Greece
[email protected]
Imbalance between oxidative challenge and antioxidant protection is implicated in normal
(ageing) and pathological processes. Crocus sativus has been used for medicinal purposes for
millennia. Its phytochemical composition is unique due to the presence of water-soluble
carotenoids, crocins, which are glycosidic esters of crocetin. The antioxidant properties of saffron
were studied in human neuroblastoma SH-SY5Y cells treated with H2O2, in neonatal rats treated
with sodium selenite -a well known cataractogenic model-, and in aged mice. Results in vitro
showed that both saffron and crocetin provided strong protection in rescuing cell viability,
repressing ROS production and decreasing caspase-3 activation against H2O2-induced toxicity in
SH-SY5Y cells. Experiments in aged mice showed that mice treated intraperitoneously with
saffron (60 mg/kg body weight (BW)) exhibited significant improvement in learning and memory
(passive avoidance test), accompanied by significantly lower brain lipid peroxidation and higher
reduced glutathione. Salt- and detergent-soluble AChE activity remained unchanged. Thus, the
significant cognitive enhancement conferred by saffron administration in aged mice, is more
closely related to the antioxidant reinforcement. Moreover, saffron administration to 9 days old
Wistar rat pups (60 mg/kg BW, 10th and 11th postnatal days) significantly protected them against
selenite-induced (20 μmol/kg BW on 9th day) cataract formation. Saffron treatment protected lens
from selenite-induced lipid peroxidation and protein proteolytic degradation. Region-specific
cerebral protective effects were also afforded to saffron-treated rats against selenite-induced
toxicity; lipid peroxidation levels were significantly lower and catalase levels being significantly
higher in cerebral cortex, while total antioxidant power was increased in all tested brain regions.
Our data suggest that saffron and its constituents could be effective against various oxidative
stressors.
83
P28
Surface sterilization and in vitro shoot multiplication of Dioscorea
membranacea Pierre ex Prain&Burkill
Yaowapha Jirakiattikula and Arunporn Itharatb
a
Department of Agricultural Technology, Faculty of Science and Technology, Thammasat
University, Rangsit Campus, Thailand
b
Applied Thai Traditional Medicine Centre, Faculty of Medicine, Thammasat University, Rangsit
Campus, Thailand
[email protected]
Dioscorea membranacea Pierre ex Prain & Burkill is a Thai medicinal plant which its rhizome
is used for treatments of inflammation, lymphopath, cancers, veneral diseases, leprosy and
dermopathy [1]. The rhizome requirement for medicine ingredient preparations has been
considerably increased therefore mass multiplication through plant tissue culture technique needs
to be applied. In order to establish an efficient in vitro regeneration system for D. membranacea,
explant surface sterilization and culture medium for multiplication were examined. Single nodes
were sonicated for 0, 15, 30, 45 and 60 min followed by surface sterilization with 10% (v/v) and
5% (v/v) sodium hypochlorite for 15 and 10 min, respectively. The results showed that the
highest survival percentages were 28.4% and 22.0% after sonication for 30 and 45 min,
respectively which were not significantly different between the treatments. On the other hand,
contamination totally occurred when sterilized without or with sonication for 15 min. For shoot
multiplication, the in vitro nodes were cultured on MS medium supplemented with 2 or 3 mg/l BA
with or without 0.2 or 0.5 mg/l NAA for 4 weeks. It was found that MS medium supplemented
with 2 or 3 mg/l BA gave the greatest generated shoot number, shoot length, and number of
shoots and nodes which were significantly different among the treatments.
[1] Tewtrakul, S. and A. Itharat. (2007) Nitric oxide inhibitory substances from the rhizome of Dioscorea
membranacea. Journal of Ethnopharmacology, 109: 412-416
84
P29
Activity of Plectranthus barbatus extract against inflammatory
response in human neutrophils
Pedro Luís Faléa, Mariana Alves Filipea, Lia Ascensãob, Maria Luísa Serralheiroa, Lurdes
Miraa
a
Centro de Química e Bioquímica, FCUL, Campo Grande, 1749-016 Lisboa, Portugal
Centro de Biotecnologia Vegetal, (IBB), FCUL, Campo Grande, 1749-016 Lisboa, Portugal
[email protected]
b
Plectranthus barbatus Andrews (Lamiaceae), known as “falso boldo”, is used in South Africa
and South America for a wide range of therapeutical purposes. Previous in vitro studies showed
that a P. barbatus extract, prepared as decoction, has promising antioxidant and anticholinesterase
properties, and the extract composition is almost unaltered during a in vitro simulated digestion,
especially in the content of its main component, rosmarinic acid [1]. In addition, after
administration of the extract to rats its components were found circulating in plasma [2].
Inflammation is a complex biological response to harmful stimuli involving the vascular
system, the immune system and the injured cells. Although it has primarily a protective function,
the destructive effects can largely surpass the gravity of the stimulus. Therefore, excessive
inflammatory responses are often associated with diseases, as atherosclerosis, allergies and
myopathies. The high production of reactive oxygen species, such as hypochlorous acid, by
activated neutrophils, for the purpose of killing invading microorganisms, is responsible for the
tissue damage associated with the inflammation process.
In the present work, neutrophils were isolated from blood of human donors and were activated
by phorbol 12-myristate 13-acetate, to stimulate the production of reactive oxygen species. The
hypochlorous acid was measured by the taurine chlorination method, in the presence and in the
absence of P. barbatus extract and rosmarinic acid.
P. barbatus extract was able to scavenge hypochlorous acid produced by the activated
neutrophils in a dose-dependent manner. The IC50 for the hypochlorous acid scavenging activity
was determined as 10.7±3.1 µg.mL-1 for the P. barbatus extract, and 3.62 ± 0.42 µM for the
isolated rosmarinic acid. The content of rosmarinic acid in the plant extract seems to be the main
responsible for the hypochlorous scavenging activity.
The low IC50 values shown by the plant extract and its main component for the scavenging of
hypochlorous acid, produced by human neutrophils, together with the fact that rosmarinic acid can
be found circulating in plasma after intragastric administration [2], allow to consider that P.
barbatus herbal tea and rosmarinic acid may be useful in the treatment of inflammation related
health problems.
[1]Porfírio S., Falé PL., Madeira PJA., Florêncio MH, Ascensão L., Serralheiro M.L.M. (2010)
Antiacetylcholinesterase and antioxidant activities of Plectranthus barbatus tea, after in vitro gastrointestinal
metabolism. Food Chem., 122: 179-187.
[2]Falé PL., Madeira PJA., Florêncio MH., Ascensão L., Serralheiro MLM. (2011) Function of Plectranthus barbatus
herbal tea as neuronal acetylcholinesterase inhibitor. Food Funct., 2: 130-136.
85
P30
Elucidation of rate limiting steps of ascorbate biosynthesis in maize
Georgina Sanahujaa*,Gemma Farrea, Ludovic Bassiéa, Changfu Zhua,Teresa Capella, Paul
Christoua,b
a
PVCF, Universitat de Lleida, Av. Alcalde Rovira Roure, 191, Lleida, 25198, Spain;
ICREA Institucio Catalana de Recerca I Estudis Avançats, Passeig Lluis Companys, 23, Barcelona 08010, Spain
b
[email protected]
L-Ascorbate (AA), also known as vitamin C, plays an important role as an antioxidant and it
also has various physiological roles. In plants it is implicated in photoprotection of the
photosynthetic apparatus, regulation in cell division and conferring tolerance to environmental
stresses. Plants and many animals can synthesize AA, but humans lack the final enzyme in AA
biosynthesis, L-gulonolactone oxidase. As a result, they depend on dietary sources of AA.
Regulation of AA synthesis varies among plant species and different tissues. As a prelude to a
more refined engineering of the AA pathway in plants we investigated rate-limiting steps in the
maize AA biosynthetic pathway. We carried out a targeted transcriptome profiling of critical
endogenous AA pathway genes in four distinct maize varieties, representing diverse germplasm.
We will present data from quantitative Real Time PCR (qRT-PCR) indicating a broad variation in
the levels of mRNA accumulation in the different varieties of maize we analyzed. We have thus
identified rate limiting steps in the AA in different maize varieties. This information will aid in
the design of more targeted genetic engineering strategies to enhance vitamin C in maize and
other plants.
We acknowledge financial support from Ministry of Science and Innovation-MICINN and European Research
Council Advanced Grant BIOFORCE.
86
P31
Antioxidant activity and antiproliferative properties of Ficus carica
cv. Dottato fruits.
Mariangela Marrelli, Filomena Conforti, Giancarlo A. Statti, Francesco Menichini
Department of Pharmaceutical Sciences, Faculty of Nutrition and Health Sciences, University of Calabria,
I-87036 Rende (CS), Italy
[email protected]
The fig tree is intensively cultivated in the Mediterranean region, from Turkey to Spain and
Portugal; it is also grown commercially in parts of USA and, to a smaller extent, in India and
Japan. Ficus carica Linn. may be important not only as an edible plant: in traditional medicine the
fruits were utilised for their antipyretic, purgative and anti-inflammatory properties [1].
Epidemiological studies have shown that there is a clear significant positive association
between intake of natural products and reduced rate of heart disease mortalities, common cancers
and other degenerative diseases. Free radicals present in human organism cause oxidative damage
to various molecules, such as lipids, proteins and nucleic acids, being involved in the initiation of
those diseases [2]. Antioxidant compounds, such as phenolics, scavenge free radicals, inhibit the
oxidative mechanism that lead to degenerative illnesses. Important findings also support the free
radical hypothesis in skin carcinogenesis [3].
In the present study we describe the evaluation of antioxidant and antiproliferative activities of
various phytocomplexes obtained from fruits of Ficus carica cv. Dottato from Calabria, Cosenza
district (Italy), collected in three different months. The work aimed to assess how the stage of
ripeness influences the biological activity of examined plant, since the phonologic stage is an
important factor that influences the compositional quality of fruits and vegetables.
The antioxidant activity of extracts from Ficus carica cv. Dottato was investigated by the Ecarotene bleaching test [4]. The first harvest (June) of the total extracts exhibited the best
inhibition of peroxidation with an IC50 of 0.004 mg/ml after 30 minutes of incubation. Among
single fractions obtained from total extracts the dichloromethane fraction from II harvest (July)
showed the best activity with an IC50 value of 0.020 mg/ml. The phototoxicity of F. carica
samples was investigated for the first time on the human tumor cell line C32 (amelanotic
melanoma) [5]. Dichloromethane fractions showed the best antiproliferative activity with an IC50
<5 μg/ml independently of the harvest.
The present study suggests that all the fruits extracts have moderate to potent antioxidant and
antiproliferative activity. June is the best month to collect F. carica cv. Dottato fruits in order to
have the best phytocomplex that showed the higher antioxidant activity.
[1] Patil VV., Patil VR. (2011) Ficus carica Linn.-An overview. Res J Med Plant, 5: 246-253
[2] Garcia-Alonso M., de Pascual-Teresa S., Santos-Buelga C., Rivas-Gonzalo JC. (2004) Evaluation of the
antioxidant properties of fruits. Food Chem, 84: 13-18.
[3] Sander CS., Chang H., Hamm F., Elsner P., Thiele JJ. (2004) Role of oxidative stress and the antioxidant network
in cutaneous carcinogenesis. Int J Dermatol, 43: 326-335.
[4] Ismail A., Marjan ZM., Foong CW. (2004) Total antioxidant activity and phenolic content in selected vegetables.
Food Chem, 87:581-586.
[5] Barraja P., Diana P., Montalbano A., Dattolo G., Cirrincione G., Viola G., Vedaldi D., Dall'Acqua F. (2006)
Pyrrolo[2,3-h]quinolinones: A new ring system with potent photoantiproliferative activity. Bioorg Med Chem, 14:
8712-8728
87
P32
Some morphology and genetic relationship
membranacea using molecular technique
of
Dioscorea
Nantana Jaengsuwan
Department of Applied Biology, Thepsatri Rajabhat University, Lop-Buri, Thailand, 15000
[email protected]
Huakhaoyen in Thai name is commomly used in Thai Traditional Medicine as cancer drug [1],
it showed the best seller in Thai Pharmacy drugstore. However, it is often collected difference
types [2]. Thus, the objective of this research is to investigate some morphology and genetic
relationship of Dioscorea membranacea using molecular technique of 22 Huakhaoyen (Dioscorea
membranacea) samples collected from different provinces in Thailand including Nakhon
Ratchasima, Lop Buri, Suphan Buri, Uthai Thani and Chumphon. The morphology in this study
was studied by shape, leaves, fruit, stem color, stipule, epidermal cell and stomata size. Most
characteristics were not distinctively classified. There were two types of stem color, blank and
white. The amplified fragment length polymorphism (AFLP) and the software programs computer
were used to construct the genetic relationship[3,4,5]. The results of DNA fingerprint from AFLP
technique could be classified into a phylogenetic tree. The tree was divided into two groups. The
first group consisted of 1 sample from Huakhaoyen (be identified as Smilax corbularia Kunth)
which was a marker in this study. The second group consisted of 22 samples from all of
Huakhaoyen (Dioscorea membranacea) in the research. The mean Polymorphic Information
Content (PIC) score of AFLP was 0.22. There was genetic diversity of Huakhaoyen (Dioscorea
membranacea) in Thailand
[1] Itharat, A., Houghton, P.J., Ammguaye, E.E., Burke, P.J., Sampson, J.H. and Raman, A. (2004). In vitro cytotoxic
activity of Thai medicinal plants used traditionally to treat cancer. J. Ethnopharmacology, 90 :33-38.
[2] Boonyaratanakornkit, L. and Chantarateptawan, V. (1993). Identification and Specification of Khaao-Yen-Neua,
Khaao-Yen-Tai. Thai J. Pharm. Sci., 17: 79-90.
[3] Rohlf, F.J. 1998. NTSYS-pc: numerical taxonomy and multivariate analysis system. Version 2.0. New York:
Setauket.
[4] Worapa S., Norihiko T., Aree W., Piyada T., Paisan L., Akito K. and Duncan A. V. 2006. Genetic Diversity of the
Vigna Germplasm from Thailand and Neighboring Regions Revealed by AFLP Analysis. Genet. Res. Crop Evol.
53: 1043-1059
[5] Xiaowei N., Yelin H., Lin W., Renchao Z., Shulin D., Darong W., Bosun W., Guohua S., Tian T. and Suhua S.
2006. Genetic diversity of the endangered Chinese endemic herb Primulina tabacum (Gesneriaceae) revealed by
amplified fragment length polymorphism (AFLP). Genetica. 127: 177-183.
88
P33
Metabolic engineering of terpenoid biosynthesis in transgenic
tobacco plants
Bruna Miralpeix1, René Hoefer2, Danièle Werck2, Lemeng Dong3, Harro J. Bouwmeester3,
Teresa Capell1, Paul Christou1,4
1
Departament de Producció Vegetal I Ciència Forestal (PVCF), Universitat de Lleida, Av. Alcalde Rovira
Roure, 177, Lleida, E-25198, Spain.
2
Institut de Biologie Moléculaire des Plantes, CNRS-UPR2357, Université de Strasbourg, 28 Rue Goethe,
67083 Strasbourg Cedex, France.
3
Laboratory of Plant Physiology, Wageningen University, P.O. Box 658, 6700 AR Wageningen, the
Netherlands.
4
Institució Catalana de Recerca I estudis avançats (ICREA Barcelona, Spain)
[email protected]
Catharanthus roseus is a rich source of alkaloids; more than 130 different compounds have so
far been reported, of which vincristine is highly important. Vincristine is mainly used to treat
acute leukaemia, Hodgkin’s disease, rhabdomyosarcomas, neuroblastoma, breast carcinoma,
among other lymphomas. C. roseus leaves accumulate about 5.8 Pg/g (fresh weight) vinblastine
and 0.9 Pg/g vincristine. These amounts represent the lowest level of any medicinally useful
alkaloid produced on a commercial basis. Most of the effort to-date to enhance the accumulation
of these and other molecules have concentrated on non-engineered cell lines under elicitation,
precursor feeding, and culture media optimization. Although these classical methods have resulted
in a thorough characterization of the pathway, metabolic engineering offers a promising approach
for improved product composition and increased yields in transgenic plants and cultured cells.
We engineered into tobacco plants the early part of terpenoid biosynthesis aiming to create
plants accumulating 10-hydroxygeraniol. A number of transgenic plants were recovered after
transformation with a geraniol synthase (GES) from Valeriana officialis and a geraniol 10hydroxylase (CYP76B6) from C. roseus; or GES with G10H from Arabidopsis. Molecular,
expression and metabolite analysis is currently ongoing to identify transgenic plants accumulating
10-hydroxygeraniol and other metabolites. Subsequent experiments will involve introduction of
additional heterologous transgenes into these plants to recreate the biosynthetic pathway to the
important metabolites secologanin and strictosidine.
The research leading to these results has received funding from the European Community's
Seventh Framework Programme FP7/2007-2013 under grant agreement n° 222716
89
P34
Quali-quantitative profiling of the polar extracts of the flowers of
Ammi visnaga plants grown in different conditions
Assunta Napolitanoa, Hela Kallel Sellamib, Milena Masulloa, Samira Smitib, Sonia Piacentea,
Cosimo Pizzaa
a
Dipartimento di Scienze Farmaceutiche, Facoltà di Farmacia, Università degli Studi di Salerno,
via ponte Don Melillo, 84084 Fisciano, SA, Italy
b
University of El-Manar II, Faculty of Sciences, Unity of Research of Vegetal Ecology, Campus
Academia, 2092 Tunis, Tunisia.
Ammi visnaga (L.) Lam. is an annual and biennial herb, belonging to the Umbelliferae family.
It is widely spread in the Mediterranean region and in particular in the north of Tunisia, where it is
known as “guabeba” and used as medicinal plant. Egyptians referred to this plant as “khella”, on
the basis of its major active metabolite, the furanochromone khellin. Khellin products are
commercially available in European market for oral and parenteral administration in the treatment
of bronchial asthma and angina pectoris as vasodilator agent [1]. Along with furanochromones
like khellin, visnagin, and khellol-glucoside, Ammi visnaga is a valuable natural source of
pyranocoumarins such as visnadin, a coronary vasodilator 5–10 times stronger than khellin [2].
Considering that the level of these compounds is greatly influenced by genetic factors and
environmental conditions [3], the study of the metabolite expression of A. visnaga plants
originating from so far unexplored Mediterranean areas, such as Tunisia, prompted us to carry out
the first phytochemical investigation of Tunisian species grown both in spontaneous way and
cultivated in restrained and in abiotic stress conditions, to highlight qualitative and quantitative
differences of metabolite composition in comparison with literature data for non endemic species
and to assess the response of A. visnaga plant to stressed environmental conditions.
To define and to promptly compare the qualitative metabolite profiles obtained for the
methanol extract of the flowers of both spontaneous and cultivated plants, stressed and not
stressed, an analytical approach based on HPLC-ESIMS technique was applied. Subsequently, the
flower methanol extract of spontaneous A. visnaga was purified by different chromatographic
steps. The structures of isolated compounds were elucidated by extensive spectroscopic methods
such as 1D- and 2D-NMR experiments as well as ESIMSn analysis. Twenty-two compounds
belonging to the furanocoumarin, pyranocoumarin, furanochromone, and chromone classes,
including two novel furanochromones, were identified. Finally, a quantitative comparison by LCMS between the metabolite content of the flowers of A. visnaga grown in spontaneous and in
restrained and stressed environmental conditions was carried out.
[1] Gunaydin K., Beyazit N. (2004) The chemical investigations on the ripe fruits of Ammi visnaga (Lam.) Lamarck
growing in Turkey. Nat. Prod. Res., 18: 169-175
[2] Kursinszki L., Troilina J., Szõke E. (1998) Determination of visnagin in Ammi visnaga hairy root cultures using
solid-phase extraction and high-performance liquid chromatography. Microchem. J., 59: 392-398
[3] Gunaydin K., Erimb F.B. (2002) Determination of khellin and visnagin in Ammi visnaga fruits by capillary
electrophoresis. J. Chromatogr. A, 954: 291–294
90
P35
Harnessing valuable botanicals from Areca catechu pericarps
Adinpunya Mitra, Mahesh Kumar
Natural Product Biotechnology Group, Agricultural and Food Engineering Department Indian
Institute of Technology Kharagpur, Kharagpur – 721 302, India
[email protected]
Areca catechu Linn. is a tropical palm belongs to Arecaceae family. Areca fruit is an ovoid
drupe, which becomes orange/reddish when ripened. A. catechu plant is being used as herbal
medicine in many countries because of the presence of different natural products including
phenolics, alkaloids and carotenes. While exploring the possibility of utilizing pericarp wastes of
A. catechu as a promising source for harnessing natural products of economic importance, a wide
range of secondary metabolites were detected upon fruit maturity.
Young pericarp when extracted with methanol and subjected to column chromatography using
Sephadex LH-20 as matrix, yielded 4-hydroxybenzaldehyde as the major soluble phenolics. The
second major soluble phenolics found in the young pericarp extract was 4-hydroxybenzoic acid.
Interestingly, 4-hydroxybenzoic acid was detected earlier from A. catechu as one of major cell
wall-bound phenolic acids, and the content appeared to be the highest (~4 mg/g cell wall dry
mass) among 20 Arecaceae species studied. By contrast, a number of carotenes were detected and
isolated from the soluble pericarp extracts of ripened Areca fruit. Among them, β-carotene was
the major one. The second major carotene in the pericarp tissue was found to be ζ-carotene, which
was identified on the basis of spectral information available in the literature. TLC coupled with
UV-VIS spectroscopy formed the basis for separation, identification and quantification of the
pericarp carotenoids. A reverse phase thin layer chromatography (RP-TLC)-based separation was
standardized for the first time for pericarp carotenoids of A. catechu, and the method was
validated subsequently. Solid phase extraction was also performed for rapid separation of βcarotene, using Sep-Pak® cartridge. The total carotenoid content of pericarp tissue was 0.04 mg/g
dry mass of pericarp tissue using β-carotene as standard. The β-carotene content found in TLCseparated and Sep-Pak® cartridge-separated fractions was 0.11 mg/g and 0.12 mg/g dry mass,
respectively. Further it was found that β-carotene only constitutes around 30% of the total
carotenoid content in ripened A. catechu pericarp as estimated through quantitation of TLC
separated and Sep-Pak® cartridge-separated fractions. Qualitative determination of free-radical
scavenging activity showed scavenging capacity maximum at 30 min, when 1,1-diphenyl-2-picryl
hydrazyl (DPPH) was used as a free-radical. ζ-carotene, also showed antioxidant activity, and
even stronger than that of β-carotene.
91
P36
Exact mass and QToF fragmentation database of bioactive
Amaryllidaceae alkaloids
Laura Torras-Claveriaa, Strahil Berkova, Francesc Viladomata, Carles Codinaa, Jaume
Bastidaa
a
Departament de Productes Naturals, Facultat de Farmàcia, Universitat de Barcelona, Avda.
Joan XXIII s/n, 08028, Barcelona, Catalonia, Spain
A particular feature of the Amaryllidaceae family is the consistent presence of an exclusive
group of alkaloids, which are known as Amaryllidacae alkaloids. These compounds are
responsible for many pharmacological activities, including antiviral, antitumour and
cholinesterase inhibition, and show considerable structural diversity. The nine main skeleton types
of Amaryllidaceae alkaloids are Norbelladine, Lycorine, Homolycorine, Crinine, Haemanthamine,
Narciclasine, Tazettine, Montanine and Galanthamine. Amaryllidaceae plants have been used for
thousands of years as herbal remedies, and in the last 50 years alkaloids from their extracts have
been the object of active chemical investigation [1].
Classical phytochemical structural elucidation is time- and solvent-consuming, and often
results in the isolation and identification of very few compounds. Although gas chromatography
coupled to mass fragmentation technique (GC-MS) has been usually used for identification of
Amaryllidaceae alkaloids, sometimes new compounds are present in concentrations too low for
NMR analysis and identification. In these cases, GC-MS combined with exact mass analysis and
fragmentation obtained from quadrupole time-of-flight with electrospray ionisation (ESI-QToF)
can be an effective method for the structural elucidation of new Amaryllidaceae alkaloids [2].
This work presents the exact mass and QToF fragmentation data base of several
Amaryllidaceae alkaloids. Five groups of alkaloids were analysed (Homolycorine, Lycorine,
Haemanthamine, Galanthamine and Other), with between 3 and 19 compounds per group. The
exact mass of the [M+H]+ ion as well as its Product Ion Scan (PIS) fragmentation were obtained
for each alkaloid. This method could also distinguish between some stereoisomers of
Haemanthamine-type alkaloids, which is not possible by GC-MS.
The valuable structural information of Amaryllidaceae alkaloids obtained by ESI-QToF can
contribute to the phytochemical investigation of Amaryllidaceae plants and allows a faster
identification of unknown bioactive alkaloids without prior isolation, requiring smaller amounts of
plant material and reducing the chemical and time consumption.
[1] Bastida J., Lavilla R., Viladomat F. (2006) The Alkaloids, Elsevier, Chicago.
[2] Torras-Claveria L., Berkov S., Jáuregui O., Caujapé J., Viladomat F., Codina C., Bastida J. (2010) Metabolic
profiling of bioactive Pancratium canariense extracts by GC-MS. Phytochemical Analysis, 21: 80-88
92
P37
Suitability of ancient and modern cereals to produce novel functional
fermented foods with a high content of antioxidant compounds
Maura Ferri1, Andrea Gianotti2 and Annalisa Tassoni1
1
Dept. Experimental Evolutionary Biology, University of Bologna,Via Irnerio 42, 40126, Bologna,
Italy: [email protected]
2
Dept. Food Sciences, University of Bologna, Viale Fanin 44, 40100, Bologna, Italy
Epidemiological studies have strongly suggested that consumption through the diet of fruits,
vegetables and grains, plays a crucial role in the prevention of heart diseases, cancer, diabetes and
Alzheimer pathologies. This effect seems to be related to the high levels of phytochemicals
present in these foods, that combat oxidative stress helping to maintain a balance between
oxidants and antioxidants. Therefore, increased consumption of foods containing high levels of
antioxidants has been recommended.
The traditional sourdough fermentation process of cereals performed by Lactic Acid Bacteria
(LAB), not only improves flavour and texture of the products, but also enables consumption of
whole meal cereals, which are well known for their high nutritional quality and health-promoting
properties including antioxidant protection [1]. In particular Lactobacillus plantarum during
fermentation of vegetal matrices showed an increase of phenolic compounds.
This work is aimed to obtain functional cereal based fermented foods with high content of
antioxidant compounds starting from two different cereals, durum wheat and Kamut khorasan.
The fermentation process was performed by selected L. plantarum strains and their ability to
increase the quali/quantitative profile of antioxidant compounds was considered. Total
polyphenols, total flavonoids, antioxidant activities and detailed polyphenolic profile, were
analysed. Preliminary results indicated, in most of the fermented flour samples, an increase of
total polyphenols levels and antioxidant activities as consequence of the L. plantarum action.
After fermentation a wider range of polyphenols were synthesized by the L. plantarum both in
durum wheat and Kamut matrices. In particular, a large spectrum of catechins (such as
epicatechin, epigallocatechin and catechin), were detected. These findings open the way to future
applications of L. plantarum strains with the aim of formulating new functional foods with
improved healthy properties.
[1] Gianotti A. et al. (2011) Role of cereal type and processing in whole grain in vivo protection from oxidative
stress. Frontiers in Bioscience, 16: 1609-1618
93
P38
Antioxidant and cytotoxic activities against lung cancer cells of Thai
Medicinal plants for cancer treatment in Kumpramong Temple
Onmanee Prajuabjindaa, Arunporn Itharatb
a
Student of Master Degree of Medical Science Program (Appiled Thai Traditional Medicine) Faculty of
Medicine, Thammasat University.
b
Department of Appiled Thai Traditional Medicine, Faculty of Medicine, Thammasat University, Thailand
[email protected]
Thai medicinal preparation of Kumpramong Temple has been used for cancer treatment more
than ten years ago. It composed with eleven plants. This research were investigated on
antioxidant and cytotoxic activities against lung cancer cell of Thai medicinal plants used to treat
cancer of Kumpramong Temple. The extract procedures was maceration method by 95% ethanol
and dry by evaporator. The antioxidant activity determined by using DPPH radical scavenging
assay [1]. The cytotoxic activity against large cell lung carcinoma (CORL-23) determined by SRB
assay [2]. The results showed that the ethanolic extract of Hydnophytum formicarum showed the
highest antioxidant activity (EC50 = 3.47±0.23 µg/ml) than Thai medicinal preparation of
Kumpramong Temple (EC50 = 4.98+0.06 µg/ml). The ethanolic extract of Tectona grandis stems
showed good cytotoxic activity against CORL-23 by IC50 values 7.78 ± 0.75 µg/ml than Thai
medicinal preparation of Kumpramong Temple.
This results should be supported using of those plants in Thai traditional medicine to treated
cancer patients although the cytotoxic activity of Kumpramong preparation showed less activity
but it showed good evidence as antioxidant activity which is good for treatment cancer patients.
[1].Skehan, P., Storeng, R., Scudiero, D., Monks, A., McMahon, I., Vistica,D., Waren, J.T., Bokesch, H., Kenney, S.,
Boyd, M.R., 1990. New colorimetric cytotoxicity assay for anticancer drug screening. Journal of National Cancer
Institute 82, 1107–1112.
[2].Yamazaki, K. Hashimoto, A. Kokusenya, Y. Miyamoto,T. and Sato, T. 1994. Electrochemical method for
estimating the antioxidative effect of methanol extracts of crude drugs, Chem. Pharm. Bull., 42: 1663-1665
94
P39
Phytochemical investigation of phenolic compounds of elderflowers
growing at different altitudes
Marlene Monschein, Franz Bucar
Institute of Pharmaceutical Sciences, Department of Pharmacognosy, University of Graz, Universitätsplatz
4/1, A-8010 Graz, Austria
[email protected]
Elderflowers, the flowers of Sambucus nigra L. (Common elder, Caprifoliaceae) are used as
herbal tea to treat colds and fever and to prepare elderflower cordial for food purposes. They are
known for their diaphoretic and secretolytic effects. Sambucus nigra is distributed almost all over
Europe, west and central Asia as well as in northern Africa. The aim of this study was to
investigate to which extent the different environmental conditions at various altitudes influence
the formation of phenolic substances in elderflowers. The changes of the levels of flavonoids and
phenolic acids in samples collected at two altitudinal levels (800 m and 1000 m) have been
studied over two growing periods (year 1: 2007 and year 2: 2008, respectively) in order to identify
individual compounds as markers. The dried, powdered plant material (10 to 14 samples per
altitude and year) was extracted with methanol 80 % (v/v) using ASE (Accelerated Solvent
Extraction) and used for further qualitative and quantitative HPLC-PDA-ESI-MS- analysis [1].
Qualitative analysis resulted in the identification of 5 hydroxycinnamic acids including 3- and
5-O-caffeoylquinic acid, 3,5- and 4,5-O-dicaffeoylquinic acid, and 7 flavonol glycosides of
isorhamnetin, kaempferol and quercetin. Quantitative analysis of elderflowers showed statistically
significant to highly significant elevated levels of rutin, narcissin, 3,5-O-dicaffeoylquinic acid and
4,5-O-dicaffeoylquinic acid in samples collected at higher altitude. Increased levels but no
statistical significance of chlorogenic acid and astragalin could be determined. The assumption
that the content of flavonol 3-O-glycosides with adjacent hydroxyl groups in ring B is increased in
plants growing at higher altitude could not be confirmed [1, 2]. Furthermore, the hypothesis that
plants at higher altitude produce more free radical scavengers due to the harsher climatic
conditions and increased ultraviolet radiation could not be verified in general [2]. The radical
scavenging capacity, measured by the DPPH assay, showed IC50 values ranging from 9.38 to
14.37µg/ml. A highly significant rise of radical scavenging capacity linked to increasing altitude
could only be recorded in year 2 (p = <0,001).
Acknowledgements: The Dr.-Heinrich-Jörg foundation and the Office of Research Management and Service,
University of Graz are gratefully acknowledged for financial support.
[1] Rieger, G., Muller, M., Guttenberger, H. und Bucar, F. (2008) Influence of altitudinal variation on the content of
phenolic compounds in wild populations of Calluna vulgaris, Sambucus nigra, and Vaccinium myrtillus. J. Agric.
Food Chem. 56(19): 9080-9086.
[2] Ganzera, M., Guggenberger, M., Stuppner, H. und Zidorn, C. (2008) Altitudinal variation of secondary metabolite
profiles in flowering heads of Matricaria chamomilla cv. BONA. Planta Med. 74(4): 453-457.
95
P40
Growth of Dioscorea membranacea Pierre ex Prain & Burkill and
dioscorealide B accumulation at different harvest times
Panumart Rithichaia, Arunporn Itharatb
a
Department of Agricultural Technology, Faculty of Science and Technology, Thammasat University,
Klongluang, Pathumthani 12121, Thailand
b
Applied Thai Traditional Medicine Center, Faculty of Medicine, Thammasat University, Klongluang,
Pathumthani 12121, Thailand
[email protected]
The rhizome of Dioscorea membranacea Pierre ex Prain & Burkill, popularly knows as KhaoYen-Tai, has been used to treat cancer in Thai traditional medicine. Dioscorealide B (DB), a
naphthofuranoxepin is the bioactive compound isolated from its rhizome which reduces
inflammation and possesses cytotoxic effect against human breast cancer cells [1, 2]. In order to
produce rhizomes for medicinal used, the appropriate harvest time must be investigated.
Therefore, the aim of the present study was to examine growth of D. membranacea and DB
accumulation at different harvest times. The 10-16 cm in length of mother rhizomes (MR) was
planted individually in the pot. Growth of aboveground shoot, rhizome development and DB
accumulated in rhizomes were determined at 3 months interval until 15 months after planting
(MAP). Results revealed that dry weight and the number of aboveground shoots dramatically
increased throughout the experiment. MR accumulated 4.0 %w/w DB at the beginning of
experiment. The degradation of MR started at 9 MAP and completely degraded at 12 MAP. The
daughter rhizomes (DR) were formed at 3 MAP and gradually developed until the final of
experiment. At 15 MAP, DR had 41.8 g dry weight/plant, 14.8 cm in length and 2.7 %w/w DB
accumulation.
[1] Itharat A., Houghton PJ., Ammgusye EE., Burke PJ., Sampson JH., Raman A. (2004) In vitro cytotoxic activity
of Thai medicinal plants used traditionally to treat cancer. Journal of Ethnopharmacology, 90: 33-38.
[2] Hiransai P., Ratanachaiyavong S., Itharat A., Graidist P., Ruengrairatanaroj P., Purintrapiban J. (2010)
Dioscorealide B suppresses LPS-induced nitric oxide production and inflammatory cytokine expression in RAW
264.7 macrophages: The inhibition of NF-ĸB and ERK ½ activation. Journal of Cellular Biochemistry, 109: 10571063.
96
P41
Strong free radical scavenging phenolics from Acacia nilotica pods
Mariateresa Maldinia, Paola Montoroa, Arafa I. Hamedb, Usama A. Mahalelb, Wieslaw
Oleszekc , Anna Stochmalc, and Sonia Piacentea.
a
Dipartimento di Scienze Farmaceutiche, Università degli Studi di Salerno, Via Ponte Don Melillo, 84084
Fisciano, Salerno, Italy,
b
Faculty of Science, South Valley University, Aswan 81528, Egypt
c
Department of Biochemistry, Institute of Soil Science and Plant Cultivation, ul. Czartoryskich 8, 24-100
Pulawy, Poland.
[email protected]
Acacia nilotica Lamarack (Mimosa nilotica, Mimosaceae) is a medium size tree growing in
tropical and subtropical countries and native from Egypt to Mozambique. It is extensively used
for the treatment of various inflammatory diseases and it is reported to be rich in polyphenolic
constituents [1], in particular in catechins which are hypothesized to possess antioxidant
properties and to play a role in the anti-inflammatory activity of several plants [2].
Antinflammatory activity is probably related to their antioxidant activity [3] and to their inhibition
of arachidonic acid metabolism via cyclooxygenase and lypoxygenase pathways [4]. Due to the
complexity of catechin derivatives, the investigation of this class of natural compounds has been
limited by difficulties in their separation. In the present work, rationalization of the phenolics
occurring in the EtOH extract of Acacia nilotica pods, on the basis of ESI-MS and ESI-MS/MS
profiles, has been proposed, together with an LC–ESI-MS qualitative study performed by using a
C18 polar endcapped stationary phase. The fragmentation pattern obtained evidenced the presence
in A. nilotica pods of galloylated- catechins and gallocatechins along with galloylated glucose
derivatives. The structures were confirmed by NMR, after isolation of the pure compounds.
On the basis of the fragmentation patterns, quantitative analyses were performed by LC–ESIMS/MS.
In addition, the radical scavenging activities of the EtOH extract, enriched fractions and pure
compounds were investigated, by using the TEAC assay. The correlation between quantitative
analysis and free radical scavenging activity confirmed the interest of this species as a rich source
of very strong antioxidant principles.
[1] Kaur K., Michael H., Arora S., Härkönen P., Kumar S. (2005) In vitro bioactivity-guided fractionation and
characterization of polyphenolic inhibitory fractions from Acacia nilotica (L.) Willd. ex Del. J Ethnopharmacol.,
99 (3): 353-60.
[2] Benavides A., Montoro P., Bassarello C., Piacente S., Pizza C. (2006) Catechin derivatives in Jatropha macrantha
stems: characterisation and LC/ESI/MS/MS quali-quantitative analysis.Journal of Pharmaceutical and
Biomedical Analysis, 40: 639-647.
[3] Goncalves C., Dinis T., Batist MT. (2005) Antioxidant properties of proanthocyanidins of Uncaria tomentosa bark
decoction: a mechanism for anti-inflammatory activity. Phytochemistry, 66: 89-98.
[4] Zhang W., Liu H., Xie K., Yin L., Li Y., Kwik-Uribe C.L., Zhu X. (2005) Procyanidin dimer B2 [epicatechin-(4β8)-epicatechin] suppresses the expression of cyclooxygenase-2 in endotoxin-treated monocytic cells. Biochemical
and Biophysical Research Communications, 345, 508-515.
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Neuropharmacological study of Eryngium planum L. extract
Thiem B.a, Ożarowski M.a,b, Szulc M.c , Kujawski R.b, Mrozikiewicz P.M.b,d, Mikołajczak
P.L.b,c
a
Department of Pharmaceutical Botany and Plant Biotechnology, University of Medical Sciences, 14 Sw.
Marii Magdaleny , 61-861 Poznań, Poland,
b
Institute of Natural Fibres and Medicinal Plants, 27 Libelta Str., 61-707 Poznań ,
c
Department of Pharmacology, University of Medical Sciences, Rokietnicka 5a, 60-806 Poznań,
d
Laboratory of Experimental Pharmacogenetics, Department of Clinical Pharmacy and Biopharmacy,
University of Medical Sciences, Święcickiego 6, 61-781 Poznań, Poland;
[email protected]
Eryngium planum L.(Apiaceae), used in folk medicine in Europe contains phenolic acids,
triterpene saponins, flavonoids, essential oil, acetylenes and coumarins [1]. The presence of the
active compounds determine their therapeutic activity: diuretic, antidiabetic, expectorant,
spasmolytic, antiinflammatory, antinociceptive, haemolytic, apoptotic and antimycotic. The
activities of Eryngium depend mainly on its high saponin content, but presence of phenolic acids
could play an important role. Triterpene saponins as well as phenolics: chlorogenic, caffeic and
rosmarinic acids and their glucosid derivatives have a wide range of biological and health
promoting properties. Neurobehavioral and neuroprotective effect on CNS have been also
describe for these compounds [2-3]. In traditional herbal medicine numerous plants from
Apiaceae family have been used to treat age related cognitive disorders and neurodegenerative
diseases [4].
The aim of the present studies were to investigate neuroactive properties of methanol extracts
from E.planum roots.
The influence of extract of E. planum radix (ER) on chosen aspects of its pharmacological
profile using rat’s scopolamine-induced amnesia model and spontaneous activity, motor
coordination, anxiety-related reactions and long term memory were assessed [5]. The effect of ER
extract on acetylcholinesterase, butyrylcholinesterase and secretase mRNA expression level in
hippocampus, striatum and frontal cortex of rats were studied on the molecular level. The possible
role of ER extract in the prevention or treatment of neurodegenerative disorders is discussed.
[1] Duke JA et. al (2002) Handbook of medicinal herbs. London CRC Press 277-278
[2] Sparg et al. (2004) Biological activities and distribution of plant saponins. J Ethnopharmacol 94:219-243
[3] Pereira et al. (2005) Neurobehavioral and genotoxic aspects of rosmarinic acid. Pharmacol Research 52:199-203
[4] Adams M et.al. (2007) Plants traditionally used in age related brain disorders – A survey of ethnobotanical
literature. J Ethnopharmacol 113:363-381.
[5] Mikołajczak P et al. (2002) Effects of acamprosate and some polyamine site ligands of NMDA receptor on shortterm memory in rats. Eur. J. Pharmacol. 444:83-96.
98
P43
Using DART-TOFMS for quality assessment of rose hip (Rosa
Canina) fruits
Hana Novotna, Vera Schulzova, Ivana Sosnovcova, Jana Hajslova
Department of Food Chemistry and Analysis, Institute of Chemical Technology Prague, Technicka 3, 166
28 Prague 6, Czech Republic
[email protected]
Rose hip fruits have been traditionally used for the preparation of tea, infusions and jams and
are known as a rich source of many nutritionally valuable substances. The most important of these
is ascorbic acid (vitamin C), with content around 1%. Besides the high content of ascorbic acid,
rose hips have a significant content of other antioxidants, e.g., carotenoids (β-carotene and
lycopene), phenolic compounds, folates, tocopherols, and many others. The high antioxidant
potential of rose hip is used in food supplements, where the addition of rose hips enhances the
effect of synthetic vitamin C. Triterpenic acids also contribute to the antioxidative potential.
Furthermore, these acids show antitumor activity. Recently, a new active compound, a
galactolipid named GOPO®, was discovered in rose hip. This substance shows strong antiinflammatory and antioxidant effects, which can be helpful for the treatment of osteoarthritis.
For metabolomic profiling of rose hips we used a unique ionization source Direct Analysis in
Real Time (DART) coupled with a High Resolution Time of Flight Mass Spectrometer (HRTOFMS). This technique enabled complex quality assessment of rose hip based nutraceuticals and
products. High Resolution TOFMS also enables the identification of many compounds occurring
in rose hip extracts. For quantification of nutritionally important compounds modern analytical
techniques were used.
Acknowledgement: This study was carried out with support from the Ministry of Education, Youth and Sports, Czech
Republic, the project MSM 6046137305 and specific university research (MSMT no. 21/2010)
99
P44
In Vitro cytotoxic activity against human lung cancer cells of the
ethanolic extract from Benjakul preparation and its isolated
compounds
Ruchilak Rattarom 1, Intouch Sakpakdeejaroen 2, Arunporn Itharat 2
1
Student of Doctor of Philosophy, Faculty of Medicine, Thammasat University, Thailand
Department of Applied Thai Traditional Medicine, Faculty of Medicine, Thammasat University, Thailand
[email protected]
2
Benjakul is widely used in Thai traditional medicine as balance health and and have also been
used for cancer treatment by folk doctors(1). It composed with five plants such as Piper chaba
fruit, Piper samentosum root, Piper interuptum stem, Plumbago indica root and Zingiber
officinale rhizome in equal proportions(2). In this study, the ethanolic extract from Benjakul
preparation and its five cytotoxic compounds were isolated by bioassay guide isolation. Cytotoxic
activity was used to test against three types of human lung cancer cell lines (COR-L23, A549 and
NCI-H226) and one normal human lung myofibroblasts cell line (MRC-5) by SRB assay (3). The
results found that the ethanolic extract of Benjakul showed stronger cytotoxic activity against
NCI-H226 than COR-L23 and A549 (IC50 values as 11.62±1.55, 24.26±0.50 and 34.40±0.37
μg/ml, respectively) but less active with normal cell line (MRC-5) (IC50 values 69.92±3.46
μg/ml). The chloroform fraction which was separated by vacumn liquid chromatography, showed
the highest cytotoxic activity against all types of lung cancer cells, so five compounds were
isolated from this fraction (piperine, plumbgin, 6-gingerol, 6-shogaol and myristicin). Plumbagin
exhibited the highest cytotoxic activity against COR-L23, A549 and NCI-H226 (IC50 values
0.42±0.09, 0.53±0.01 and 0.78±0.15 μg/ml, respectively) and 6-shogaol shown the second most
effective cytotoxicity (IC50 values 4.59±0.52, 5.15±0.15 and 4.99±0.03 μg/ml, respectively).
Piperine exhibited specific cytotoxicity against NCI-H226 and COR-L23 more than A549. On the
other hand, 6-gingerol and myristicin exhibited specific cytotoxic activity against A549 higher
than NCI-H226 and COR-L23. These results can support using Benjakul preparation for lung
cancer treatment.
[1] Itharat, A. Singchangchai, P. and Ratanasuwan, P. (1998). Wisdom of Southern Thai traditional doctors.
Research Report of Prince Songkhla University, Songkhla, Thailand.
[2] Sakpakdeejaroen, I and Itharat, A. Cytotoxic compounds against breast adenocarcinoma cells (MCF-7) from
Pikutbenjakul. J Health Res 2009, 23(2), 71-76.
[3] Skehan, P., Storeng, R., Scudier., D., Monks, A., Mc Mahon, J., Vistica, D., Warren, J.T., Bokesch, H.,
Kenney, S. and Boyd, M.R. (1990). New colorimetric cytotoxicity assay for anticancer-drug screening. J Natl
Cancer Inst 82: 1107-1112.
100
P45
Watermelon as a source of bioactive molecules
Marcello Salvatore Lenucci, Stefania Grassi, Anna Eleonora Caprifico, Maria Chiara
Contino, Anna Montefusco, Gabriella Piro, Giuseppe Dalessandro
Di.S.Te.B.A., University of Salento, Via Prov. Lecce-Monteroni, 73100 Lecce, Italy
[email protected]
Watermelon (Cucumis citrullus (L.) Ser.) is a typical summer fruit, known for its refreshing and thirstquenching power. From the nutritional point of view it has attracted considerable interest for the presence
of bioactive molecules with undisputed beneficial properties for human health, such as lycopene, citrulline
and fibers. Many epidemiological studies highlight lycopene involvement in the prevention of degenerative
pathologies such as cardiovascular diseases, diabetes and certain types of cancer due to its high antioxidant
activity, beside other still poorly understood mechanisms of action [1]. Similarly, the water-soluble, nonessential, non-proteic amino acid citrulline is showing a huge potential as therapeutic agent in the
prevention and treatment of various metabolic diseases playing a central role in cell metabolism and organ
functionality [2]. Eventually, the health benefits offered by a high-fiber diet in reducing risk for developing
coronary heart disease, stroke, hypertension, diabetes, obesity, and certain gastrointestinal disorders are
widely recognized [3]. The nutritional importance of these bioactive molecules makes the preparation of
natural extracts and their use in the formulation of high quality supplements, and/or pharmaceuticals, a
valuable alternative to the classical production and distribution chain of watermelon, opening new
opportunities for agri-food processing industry.
The pellet and the supernatant obtained by centrifugation of the homogenized watermelon flesh
(cultivar Dumara) were freeze-dried giving a lycopene rich matrix (~3.3 mg/g f-d.m.), constituted mainly
of clumps of parenchyma cells, and a slightly pink powder containing soluble sugars (principally fructose)
and high amount of citrulline (>10 mg/g f-d.m.), respectively. Both fractions are considerably enriched in
dietary fibers.
Under the right conditions (absence of light and air, and temperature of -20 °C), the matrix can be
stored for a period longer than six months, providing a readily extractable watermelon product available
over time. The citrulline rich powder was stable for several months at room temperature and could be used
as such or after further purification.
Recently, the interest in safe and clean extraction technologies has increased. Supercritical CO2 has
been successfully used to extract lycopene from freeze-dried tomato matrices [4]. The chemical and
physical parameters (pressure, temperature, CO2 flux and particle size) applied on tomato, were used as a
starting point to identify the optimal conditions for lycopene extraction from the freeze-dried watermelon
matrix. The presence of lipids from the watermelon seed endosperm had a positive effect on the amount
and yield of extracted oleoresin. The addition of a lipid co-matrix, consisting of roughly crushed hazelnuts,
did not further increase the extraction yield, whereas the use of ethanol as co-solvent resulted in a
significant increase in lycopene extraction. In the optimal operative conditions, lycopene yield exceeded
75% and an oleoresin containing ~20 mg/g lycopene was obtained.
[1] Kong K.-W., Khoo H.-E., Prasad K.N., Ismail A., Tan C.-P., Rajab N.F. (2010) Revealing the power of the natural red pigment
lycopene. Molecules, 15: 859-987.
[2] Curis E., Nicolis I., Moinard C., Osowska S., Zerrouk N., Bénazeth S., Cynober L. (2005) Almost all about citrulline in
mammals. Amino Acids, 29: 177-205.
[3] Anderson J.W., Baird P., Davis Jr R.H, Ferreri S., Knudtson M., Koraym A., Waters V., Williams C.L. (2009) Health benefits
of dietary fiber. Nutrition Reviews, 67(4): 188-205.
[4] Lenucci M.S., Caccioppola A., Durante M., Serrone L., Leonardo R., Piro G., Dalessandro G. (2010) Optimisation of
biological and physical parameters for lycopene supercritical CO2 extraction from ordinary and high-pigment tomato cultivars.
Journal of the Science of Food and Agriculture, 90: 1709-1718.
101
P46
Adaptation of leaves in Ligularia fischeri (Ledeb.) Turcz to sun,
shade and drought: chlorophylls and phenolic compounds
Sang Min Kim, Je-Seung Jeon, Yu-Jin Jung, Linna Ly, Byung-Hun Um
Functional Food Center, Korean Institute of Science and Technology Gangneung Institute, Gangneung,
Gangwon-do, 210-340, Korea
[email protected]
Ligularia fischeri (Ledeb.) Turcz is a commercial leafy vegetable that has been eaten for a long
time in Korea. The HPLC chromatograms of leaf extracts collected from different areas in Korea
showed a significant variation in caffeoylquinic acid derivatives (CQAs) [1] and two new tri-Ocaffeoylquinic acids were purified and structurally identified by NMR and MS from this plant.
As this plant prefer the low-light and wet environment, in the present study, the effect of sun
and drought was investigated by applying shade condition for 4 weeks to the plants growing under
sun-light. The differences were described on chlorophyll, phenolics levels, containing CQAs,
flavonoids, and anthocyanins.
Sun leaves showed higher levels of leaf thickness, % of dry weight than shade leaves. In
addition, total phenolic and total flavonoid levels in sun leaves were found in higher levels
compared to shade leaves. Especially, four major CQAs (5-CQA, 3,4-, 3,5-, and 4,5-DCQA) were
induced according to the temperature increase, while in shade leaves, these compounds were
continuously decreased in 4 weeks test period. The shade leaves possessed higher chlorophyll
levels on fresh weight basis with bigger leaves in comparison to sun leaves. In a drought
condition, sun leaves turned from green to purple and total anthocyanin content was increased,
while phenolic and chlorophyll levels were decreased slightly.
The results clearly demonstrate that the leaves of L. fischeri show adaptation response to high
irradiance and drought and these characteristics can be useful information in growing this plant
for different commercial application.
[1] Shang YF., Kim SM., Song DG., Pan CH., Lee WJ., Um BH. (2010) Isolation and identification of
antioxidant compounds from Ligularia fischeri. J Food Sci, 75:C530-C535
102
P47
The correlation between the total phenolic and flavonoid contents
and their antioxidant properties of Thai rice cultivars in chemical
and cell-based assays
Pintusorn Hansakula, Umarat Srisawata, Nusiri Lerdvuthisopona, Arunporn Itharatb
a
Department of Preclinical Science, Faculty of Medicine, Thammasat University, Rangsit Campus, Klong
Luang, Pathumthani, 12120 Thailand.
b
Department of Applied Thai Traditional Medicine Center, Faculty of Medicine, Thammasat University,
Rangsit Campus, Klong Luang, Pathumthani, 12120 Thailand.
[email protected]
Free radicals induce oxidative stress in various cell components, leading to some certain
diseases. Due to adverse effects of some synthetic antioxidants, plant-derived antioxidants have
become a profitable alternative to prevent oxidative stress in cells. In this study, the ethanolic
extracts of brown rice and rice bran from Thai rice cultivars: Sangyod red rice and Dawk Mali
105 white rice were evaluated for the correlation between the total phenolic and flavonoid
contents and their antioxidant properties through two chemical assays: DPPH radical-scavenging
and inhibition of lipid peroxidation assays as well as a cell-based assay: scavenging capacity
against intracellular superoxide in cells using dichlo-rofluorescein diacetate (DCF-DA).
All the rice extracts displayed their antioxidant activities in a dose dependent manner through
the DPPH scavenging assay, the anti-lipid peroxidation assay, and the cell-based DCF-DA assay
with EC50 values in the range of 3.54-230.71 μg/ml, 1.09-6.39 mg/ml, and 0.55-1.19 mg/ml,
respectively. The ethanolic extract of Sangyod rice bran and brown rice exhibited the highest and
the second highest inhibitory effect as compared to those of Dawk Mali 105. The total phenolic
and flavonoid contents of all the rice extracts were in the range of 18.97-189.20 mg gallic acid
equivalent per gram rice extract and 18.75-46.13 mg rutin hydrate equivalent per gram rice
extract, respectively. The ethanolic extract of Sangyod rice bran contained the highest content of
both studied compounds. Positive correlations were observed between the phenolic contents and
the three antioxidant assays: the DPPH assay (R = 0.999), the anti-lipid peroxidation assay (R =
0.752), and the cell-based DCF-DA assay (R = 0.990). Furthermore, significant antioxidant effects
of flavonoid contents were also observed in the DPPH assay (R = 0.909) and the cell-based DCFDA assay (R = 0.836), but not in the anti-lipid peroxidation assay (R = 0.435). Importantly, the
ethanolic extracts of Sangyod extracts were shown to be more effective than their water extracts
(1)
.
These findings suggest that ethanolic extracts of brown rice and rice bran of Sangyod red rice
can be promising sources of potential natural antioxidants. This study is also one of a few reports
that perform a correlation analysis between antioxidant activity and total phenolic and flavonoid
contents in Thai rice varieties (2).
[1] Srisawat U., Panunto W., Kaendee N., Tanuchit S., Itharat A., Lerdvuthisopon N., Hansakul P. (2010)
Determination of phenolic compounds, flavonoids, and antioxidant activities in water extracts of Thai red and
white rice cultivars. J Med Assoc Thai. 93 Suppl 7:S83-91.
[2] Butsat S., Siriamornpun S. (2010) Phenolic acids and antioxidant activities in husk of different thai rice varieties.
Food Sci Technol Int. 16(4):329-36.
103
P48
Acetylcholinesterase inhibition and antioxidant activity of three
Asteraceae species from the Portuguese Aromatic Flora
Ana Margarida Rodriguesa, Pedro Luís Faléb, Lia Ascensãoa, Maria Luísa Serralheirob
a
Centro de Biotecnologia Vegetal (IBB), FCUL, Campo Grande, 1749-016 Lisboa, Portugal
Centro de Química e Bioquímica, FCUL, Campo Grande, 1749-016 Lisboa, Portugal
[email protected]
b
The Portuguese Flora is very rich in aromatic species used for centuries in traditional medicine
in the form of "teas", aqueous extracts obtained by infusion or decoction of different parts of
plants, mainly belonging to Lamiaceae and Asteraceae families [1]. In the present work the
antioxidant and acetylcholinesterase inhibitory activities were evaluated for the decoctions of
stems/leaves and flowers of three Asteraceae species (Helichrysum italicum, Santolina impressa
and Solidago virgaurea) collected from natural populations occurring throughout western
Portugal sandy soils. The in vitro metabolism by the action of gastric and pancreatic artificial
juices, as a simulation of the digestion of the extracts, was also studied.
The antioxidant activity of the aqueous extracts was assessed by the DPPH method. The
decoction of H. italicum flowers showed the lowest EC50 value, representing the lowest
concentration required to consume 50% of DPPH (10.14±0.02 µg/mL). The highest EC50 was
obtained with the decoction of S. impressa flowers (14.70±0.06 µg/mL). As observed for the
antioxidant activity, the lowest IC50 for acetylcholinesterase activity was found in H. italicum
flowers decoction (302.52±1.14 µg/mL). The highest IC50 corresponded to the decoction of S.
impressa leaves (578.98±13.70 µg/mL). All the aqueous extracts from flowers of these three
species show a higher acetylcholinesterase inhibitory activity when compared with those found in
their leaves extracts. The decoctions from flowers of H. italicum showed the highest tested
biological activities.
The results of the in vitro metabolism revealed that decoctions did not suffer appreciable
degradation, showing no significant changes both in its chemical composition and biological
activities.
The high activities found in the three Asteraceae herbal teas could have a useful effect in the
treatment of Alzheimer’s disease (AD) patients, as the inhibition of acetylcholinesterase is the
most effective therapy for AD and the antioxidant activity has been connected with the capacity to
scavenge the free radicals formed during the inflammation processes [2].
[1] Neves, J. M., Matos, C., Moutinho, C., Queiroz, G. & Gomes, L. R. (2009) Ethnopharmacological
notes about ancient uses of medicinal plants in Trás-os-Montes (northern of Portugal). Journal of
Ethnopharmacology, 24: 270-283.
[2] Howes, M. J. R., Perry, N. S. L. and Houghton, P. J. (2003) Plants with traditional uses and activities relevant to
the management of Alzheimer’s Disease and other cognitive disorders. Phytotherapy Research, 17: 1-18.
104
P49
Rosmarinus officinalis extract: a promising clinical management of
Krabbe leukodystrophy
Venera Cardilea, Giovanna Pannuzzoa, Adriana Grazianoa, Juan Garbarinob, Nicolas
Troncosoc and Alessandra Russod
a
Department of Biomedical Sciences, University of Catania, V.le A. Doria 6, 95125 Catania, Italy
Department of Chemistry, University T.F. Santa Maria, Casilla 110-V, Valparaiso, Chile
c
Lo Vicuña & Cia., Av. Santa María 5888, Vitacura, Santiago
d
Department of Sciences of Drug, University of Catania, V.le A. Doria 6, 95125, Catania, Italy
[email protected]
b
Krabbe disease or globoid cell leukodystrophy is an autosomal recessively inherited disorder
caused by the deficiency of galactocerebrosidase, the lysosomal enzyme that catalyzes the
hydrolysis of galactose from galactosylceramide and galactosylsphingosine (psychosine).
Psychosine accumulation results in the loss of myelin and oligodendrocytes in the brain of Krabbe
patients as well as twitcher mice (natural model of human Krabbe disease). Several approaches
for the treatment of Krabbe disease are targets of research, but current treatment opinions for the
disease are very limited. Gene therapy might be effective; however, a stable and strong expression
vector is required [1]. Some studies reported that brains from brains from patients with Krabbe
disease and twitcher show expression of the inflammatory cytokine TNF-D, and IL-6, and
established the relationship of psychosine and cytokine-induced cellular redox alterations by
increase in reactive oxygen species (ROS) and the inhibition of peroxisomal functions [1].
Rosmarinus officinalis L. (Labiatae) is used as a folk medicine around the world. In medicine,
the extract is receiving increasing attention due to its anti-inflammatory and antioxidative
constituents [2]. There are several reports that have established carnosic acid as the major
phenolic diterpenoid present in rosemary leaves with antioxidant activity [2]. The aim of the
present research was to demonstrate suppression of the apoptotic effects of psychosine in cultured
oligodendrocyte progenitor mice cells (OLP-II) with a R. officinalis methanolic extract,
containing 39.7 % of carnosic acid. Protection of the OLP-II cells against psychosine was shown
using the MTT test (the ability of the tetrazolium salt MTT to form a dark blue formazan product
by mitochondrial dehydrogenase in viable cells) and assay of expression of p53 and TNF-related
apoptosis-inducing ligand (TRAIL). The results showed a time-dependent and concentrationdependent decrease of OLP-II viability on exposure to psychosine and dose-dependent protection
with the examined extract. They also demonstrated that psychosine-induced p53 induction of
apoptosis and TNF-related apoptosis-inducing ligand receptors could be decreased by this natural
product. These preliminary data suggest that an integration with phytochemicals may constitute a
promising clinical management of the late-infantile and juvenile forms of Krabbe leukodystrophy.
[1] Pannuzzo G., Cardile V., Costantino-Ceccarini E., Alvares E., Mazzone D., Perciavalle V. (2010). A galactosefree diet enriched in soy isoflavones and antioxidants results in delayed onset of symptoms of Krabbe disease in
twitcher mice. Mol. Genet. Metab. 100: 234-40.
[2] Russo A., Lombardo L., Troncoso N., Garbarino J. and Cardile V. (2009). Rosmarinus officinalis L. extract
inhibits human melanoma cell growth. Natural Prod. Commun. 4: 1707-1710.
105
P50
Antioxidant properties of Asplenium trichomanes, Polypodiaceae
Jelica Srdanova, Veljko Krstonosica, Marija Krickovica, Neda Lakica, Biljana Bozina
a
Department of Pharmacy, Faculty of Medicine, University of Novi Sad, Hajduk Veljkova 3, 21000 Novi
Sad, Serbia
[email protected]
Presence of natural antioxidants is of a great importance, not only because of the impact on the
sustainability of oil, but also for important functions in human organism [1]. Maidenhair
spleenwort (Asplenium trichomanes, Polypodiaceae) has a long tradition of usage in traditional
medicine as a cure for respiratory illnesses. Also, it is used for the treatment of psoriasis. Today,
when oxidative stress plays an important role in pathophysiology of diseases of modern age,
antioxidant potential of maidenhair spleenwort could be of a great interest.
The objective of this study was to assess the oxidative stability of sunflower and corn oil and
effectiveness of Asplenium trichomanes to slow down the peroxidation process of oils.
Peroxidation was evaluated through measuring of peroxide values during 45 days. Also, total
phenolic (gallic acid equivalents-GAE) and flavonoid content (quercetine equivalents-QE) and
scavenging activity of 45% ethanol extract on 2,2-diphenyl-1-pycryl hydrazyil radical (DPPHx)
were determined [2].
Leaves of wild-growing plants of Asplenium trichomanes were collected in July of 2010 at
Fruska gora mountain, Republic of Serbia. Voucher specimen of collected plant material (No. A75/10) was confirmed and deposited at the Herbarium of the Laboratory for Pharmacognosy,
Faculty of Medicine, University of Novi Sad.
The results showed that Asplenium trichomanes exhibited protective acitivity against oxidative
deterioration only in sunflower oil (34.21%), comparing to the pure oil, without plant material.
The amount of dry extract (d.e.) in dried plant material was 12.79%. Total amount of phenolics
was 1.11 mg of GEA/g d.e. Flavonoids were present in the amount of 22.19 QE/g d.e. In DPPHtest, assayed extract exhibited notable antioxidant activity (IC50=13.38μg/ml).
The obtained results provide confirmation of the strong antioxidant and protective activity of
Asplenium trichomanes herb extract, partially explaining some of traditional uses of this fern.
[1] Adnan A. et al. (2009) Analytical investigation of oxidative deterioration of sunflower oil stored under different
conditions. Eletronic Journal of Environmental, Agricultural and Food Chemistry, 8(10), [1043-1051].
[2] Bozin B., Mimica-Dukic N., Samojlik I., Anackov G., Igic R. (2008) Phenolics as antioxidants in garlic (Allium
sativum L., Alliaceae). Food Chemistry, 111(4), [925-929].
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Antioxidant activities, gamma oryzanol, vitamin E and total phenolic
content of hom mali 105 rice (Oryza sativa) in different extraction
methods
Sakuntala Uttama a, Intouch Sakpakdeejaroen b, Rujulak Rattarom c, Arunporn Itharat b
a
Student of Ph.D in Medical Science (Nutraceutical)program, Faculty of Medicine, Thammasat
University. Klongluang, Pathumtanee, Thailand
b
Klongluang, Pathumtanee, Thailand
c
Faculty of Pharmacy, Mahasarakham University, Khamreang, Mahasarakham, Thailand
[email protected]
The objectives of this research are to investigate for the antioxidant activities, gamma
oryzanol , vitamin E and total phenolic content of Hom Mali 105 Rice (Oryza sativa) in
different extraction methods of three parts of rice as follow: rice bran, brown rice and white
rice. The methods of extraction were maceration by ethanol 95% (ME) supercritical fluid
extraction (SFE), boiling in water and dry by freeze dryer (BF),Water from washing rice and
freeze dried (WR), expression method (EX) and soxhlet extraction method by using different
solvent as hexane , chloroform and methanol (SXH, SXC and SXM respectively). DPPH assay (1)
was carried for antioxidant activity and gamma oryzanol and vitamin E content were determined
by HPLC (2,3 ). Total phenolic content was determine by using Folin–Ciocalteu reagent (3) The
result found that BF showed the highest of percentage of yield (21.73%).white rice exhibited the
highest antioxidant activity and it reduced free radical scavenging at concentration 100µg/ml as
48.34%. The SXM method gave the highest total phenolic content follow by SXC and ME are
20.6, 17.01 and 12.37 mg Gallic acid equivalent/ g samples of extracts respectively. ME method
of rice bran gave the highest gamma oryzanol and also vitamin E content are 340, and 205 mg per
100 g of extracts respectively. This study is a guide for extraction product form rice the method of
extraction and part of rice should be chosen for production of health product as essential
bionutrients.
[1] Yamasaki, K. Hashimoto, A. Kokusenya, Y. Miyamoto T. and Sato, T. (1994). Electrochemical method for
estimating the antioxidative effect of methanol extracts of crude drugs, Chem. Pharm. Bull 42 : 1663-1665.
[2] Edison B. (2009). Analysis of Tocopherols by High Performance Liquid Chromatography. E-Journal of Chemistry
6(2), 395-398.
[3] Chotimakorn, C., Benjakul, S. and Silalai, N. (2008). Antioxidant components and properties of five long-grained
rice bran extracts from commercial available cultivars in Thailand. Food Chemistry, 111, 636-641.
107
P52
Anti-inflammatory activities of Smilax corbularia Kunth and its
constituents
Srisopa Ruangnooa, Arunporn Itharata
a
Department of Applied Thai Traditional Medicine Centre, Faculty of Medicine, Thammasat
University, Klongluang, Pathumthani, 12120 Thailand
[email protected]
Smilax corbularia Kunth (Smilacaceae) is a Thai medicinal plant locally known as ‘Hua-KhaoYen Neua’ . Its roots were used to treat inflammations such as arthritis [1,2]. Thus, the objective
of this study is to investigate on anti-inflammatory activity of two extracts (ethanolic and water
extracts) of this plant and its constituentsby determination of inhibitory activities against
lipopolysaccharide (LPS) induced nitric oxide (NO) production in RAW 264.7 cell lines. Nitric
oxide in the culture supernatant was measured by Griess reaction [3]. The ethanolic extract of
Smilax corbularia Kunth exhibited the most inhibitory activity, with an IC 50 value of 83.90+3.82
μg/ml. From this extract, three compounds [ engeletin (1) astilbin (2) and quercetin (3),] were
isolated and further investigated for their inhibitory properties of NO production. It was found that
quercetin (3) possessed the highest activity (IC50 = 9.17 μg/ml or 27.12 μM), whereas engeletin
(1) and astilbin (2) exhibited mild activity. The ethanolic , water extracts of Smilax corbularia
Kunth and compounds 1–3 were also evaluated for the inhibitory effect on LPS-stimulated PGE2
release from RAW 264.7 cells. The results found that engeletin (1) and astilbin (2) possessed
potent activity against PGE2release with IC50 values of 19.67+0.01 μg/ml or 43.81 μM and
19.72+0.02 μg/ml or 45.31 μM, respectively, whereas, quercetin (3) exhibited mild activity. The
present study may support using Smilax corbularia Kunth as inflammatory drug by Thai folk
doctors .
[1] Pongbunrod, S. (1976). Mai Thiet Mueang Thai Kasembanakit. Bangkok, pp.120–122.
[2] Tungtrongjit, K. (1978). Pramuan Supphakun Ya Thai. Bangkok, pp. 107–108.
[3] Tewtrakul, S. and Itharat, A. (2007). Nitric oxide inhibitory substances from the rhizomes of Dioscorea
membranacea. J.Ethnopharmacol. 109:412-416.
108
P53
Bioactive compounds from Capparis spinosa subsp. rupestris
Argentieri Maria Piaa, Macchia Francescob, Papadia Paridec, Fanizzi Francesco Paoloc,
Avato Pinarosaa
a
Dipartimento Farmaco-Chimico, Università, Via Orabona 4, I-70125 Bari, Italy
Orto Botanico, Università, Via Orabona 4, I-70125, Bari, Italy
c
Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università, I-73100 Lecce, Italy
[email protected]
b
The genus Capparis includes about 250 species distributed in the Mediterranean regions which
can be distinguished mainly based on their morphological traits such as leaf shape, flowers and
presence/absence of spines. Some species are used as food and have application in medicine for
skin protection [1].
According to the official European food legislation, commercial capers consumed as
condiment are the flower buds of C. spinosa L., which represents a high polymorphic species with
still an uncertain taxonomic position. It has been separated by some Authors into two different
subspecies, namely C. spinosa L. subsp. spinosa and C. spinosa L. subsp. rupestris (Sibth. & Sm.)
Nyman. The latter is also reported as C. spinosa subsp. rupestris (Veill. in Duhamel) Jafri and
classified as a synonymous species of C. orientalis Veill., from which it might be derived [2].
Caper plants used industrially are generally referred to as C. spinosa, however, the less defined
taxonomy and the fact that several cultivars and some species overlap with others may produce
economic implications due to the quality variability of the commercial products.
The objective of the present study was to determine the spectrum of phytochemical
constituents of C. spinosa subsp. rupestris in order to evaluate the presence of bioactive
metabolites and compare the chemical profile of this species with that of other studied Capparis
sp. and especially with C. spinosa subsp. spinosa. Chemical composition of oil and glucosinolates
from seeds, as well as of glucosinolates and flavonoids from the aerial parts of the plant have been
determined and data presented here.
The overall phytochemical data from the analysis of C. spinosa subsp. rupestris (syn. C.
orientalis) do not differentiate this subspecies from C. spinosa subsp. spinosa, their compositional
profiles resulted in fact largely superimposable. However, obtained results show that C. spinosa
subsp. rupestris represents a very rich source of bioactive compounds of nutraceutical relevance,
especially of the antioxidant rutin.
[1] Rivera, D., Inocencio, C., Obón, M.C., Alcaraz, F. (2003). Review of food and medicinal uses of Capparis L.
subgenus Capparis (Capparidaceae). Econom. Bot. 57: 515-534.
[2] Inocencio, C., Rivera, D., Obón, M.C., Alcaraz, F., Barreña, J-A. (2006). A systematic revision of Capparis
section Capparis (Capparaceae). Ann. Missouri Bot. Gard. 93: 122-149.
109
P54
Isolation of bioactive compounds from Saponaria officinalis L.
Barbara Szajwaj1, Łukasz Pecio1, Małgorzata Szumacher-Strabel2, Adam Cieślak2,
Wiesław Oleszek1 and Anna Stochmal1
1
Department of Biochemistry, Institute of Soil Science and Plant Cultivation, State Research Institute, ul.
Czartoryskich 8, 24-100 Pulawy, Poland
2
Department of Animal Nutrition and Feed Management, RUMEN PULS Poznan, Wolynska 33, 60-637
Poznan, Poland
[email protected]
Saponaria officinalis L. commonly named soapwort or bouncing-bet is a perennial plant from
the carnation family (Caryophyllaceae), it’s native range extends throughout Europe, northern
Africa and western to central Asia, but currently is also cultivated in many countries. Soapwort
has been used in medicine as an expectorant in bronchitis, and in folk medicine it is still used for
skin complaints and rheumatic disorders [1-4]. S. officinalis is a rich source of triterpenoid
saponins but only part of them have been known. Thus, the aim of the present research was to
focus on roots, because they contain about 5% of saponins and about 30% of oligosaccharides,
which have not been investigated and until now only two were known.
The methanolic extract (233 g) from 500 g of dry, powdered roots of S. officinalis was
fractionated on C18 preparative column with H2O (162.99 g), 30% MeOH (8.51 g) and 90%
MeOH (49.90 g). Next, 30% and 90% methanolic fractions were suspended in H2O and extracted
with EtOAc. The EtOAc extraction of 30% MeOH fraction yielded 681 mg of phenolic
compounds, while from 90% MeOH fraction 45.41 g of saponins and 2.98 g of catechins were
obtained. The fraction containing sugars (H2O fraction) was applied to a column chromatography
on DEAE-cellulose, and the column was washed with H2O. This afforded 155.06 g of pure sugars.
All groups of compounds (phenolics, sugars and saponins) were analyzed for their composition
using a Waters ACQUITY UPLCTM system coupled to Waters TQ Detector (Waters Corp.) in full
scan mode (m/z = 200-2000) on a HSS C18 (1.0 x 100 mm, 1.8 µm) column.
Future research on these fractions will be focused on the separation of sugars, phenolics,
catechins and saponins into individual compounds. The activity of the fraction containing
saponins and isolated individual saponins will be tested using ruminants animal models. Saponins
demonstrate beneficial effects in defaunation (selective removal of protozoa) of the rumen and
manipulation of the end products of fermentation [5].
[1] Jia Z., Koike K., Nikaido T. (1998) Major triterpenoid saponins from Saponaria officinalis. J. Nat. Prod., 61:
1368-1373
[2] Jia Z., Koike K., Nikaido T. (1999) Saponarioside C, the first α-D-galactose containing triterpenoid saponin, and
five related compounds from Saponaria officinalis. J. Nat. Prod., 62: 449-453
[3] Jia Z., Koike K., Nikaido T. (1999) New triterpenoid saponins and sapogenins from Saponaria officinalis. J. Nat.
Prod., 62: 1655-1659
[4] http://www.ars-grin.gov/cgi-bin/npgs/html/genform.pl
[5] Wina E., Muetzel S., Becker K. (2005) The impact of saponins or saponin-containing plant materials on ruminant
production – a review. J. Agric. Food Chem., 53: 8093-8105
110
P55
The influence of isoflavonoid genistein incorporated in hydroxypropyl-gamma-cyclodextrin on experimental B164A5 murinic
melanoma model on mice
Tiulea Corinaa, Peev Cameliaa, Feflea Stefanaa, Dehelean Cristinab, Soica Codrutac , Andrei
Motocd
a
University of Medicine and Pharmacy Victor Babes, Timisoara, Romania, Department of Pharmacognosy
University of Medicine and Pharmacy Victor Babes, Timisoara, Romania, Department of Toxicology
c
University of Medicine and Pharmacy Victor Babes, Timisoara, Romania, Department of Pharmaceutical
Chemistry
d
University of Medicine and Pharmacy Victor Babes, Timisoara, Romania, Department of Anatomy
[email protected]
b
Genistein (4,5,7-trihydroxyisoflavone) is the aglicon of genistin and has been identified as the
predominant isoflavone in soybean, the vegetal product from Glycine max Fam. Fabaceae. This
compound is a specific inhibitor of protein tyrosine kinase and was found to exhibit
chemopreventive, cardioprotective and antiosteoporotic activities. One of the major problems of
this active compound is its poor water solubility. Because genistein has a poor water solubility
and the compound was injected in saline solution in order to increase the biodisponibility a
complex with hydroxyl-propyl- gama-cyclodextrin (HPGCD) in a molar ratio 1:2 by kneading
method was prepared. In order to test the complex formation scanning electron microscopy and
differencial scanning calorimetry assay were performed. Results suggest that incorporation took
place. Ten C57BL/6J mice, female of eight weeks were s.c. inoculated with 0.2 ml x 10 6 B164A5
melanoma cell suspension prepared after standard protocols using as solvent saline solution. Mice
were divided in two groups and one group was injected with genistein complex s.c.at daily doses
of genistein of 15 mg/kg b. w. for five days from the first day of experiment. Animals were
inspected daily for the development of tumors, which became visible after 10 days from
inoculation. On day 14 mice were sacrificed. Macroscopic examination was made. After that
tissue samples were fixed in 10% formalin solution and were embedded in paraffin and cut at 4
microns. Finally after deparaffinized the samples were stained with H&E (hematoxylin-eosin) and
microscopically analyzed. One can observe a decease in the number and dimension of tumors in
mice treated with the complex genistein:HPGCD than in case of untreated mice. HE analyze
showed that the diagnosis of malignant melanoma on skin level was present in both groups, but in
case of untreated mice liver metastasis were also present.
The conclusion of this study is that genistein incorporated in hydroxyl-propyl- gamacyclodextrin reduces the number and dimension of subcutaneous tumors and also the distant
metastases on a period of study of two weeks.
111
P56
SPOTLight: sustainable production of thapsigargin using light
Corinna Weitzel, Andreas Harald Klem, Tom Manczak, Trine Bundgaard Andersen,
Joachim Møller Larsen, Henrik Toft Simonsen
Department of Plant Biology and Biotechnology, VKR Research Centre Pro-Active Plants, University of
Copenhagen, Thorvaldsensvej 40, 1871 Frederiksberg, Denmark
[email protected]
The sesquiterpene lactone (SQL) thapsigargin is a promising agent for cancer therapy inducing
apoptosis by inhibiting the sarco/endoplasmatic calcium ATPase (SERCA). Covalently linked to a
short peptide, the molecule can be directed to prostate cancer cells where it is activated in the
presence of prostate specific antigen (PSA) [1]. This modification facilitates targeted treatment of
this slowly growing cancer type.
Nature’s only source of thapsigargin are two species belonging to the genus Thapsia
(Apiaceae) that is widely distributed throughout the Mediterranean area. Of the various Thapsia
species only T. garganica and T. gymnesica produce thapsigargin whereas numerous
sesquiterpenes, also some with related structures, can be found in all Thapsia species [2, 3, 4].
Once thapsigargin is established on the market, future demands cannot be ensured by isolating the
drug from T. garganica without endangering wild populations. Since complexity of the chemical
structure precludes organic synthesis, a biotechnological production platform will be the method
of choice [5]. This platform will be created by our group by transferring thapsigargin’s metabolic
pathway into the moss Physcomitrella patens that can be easily genetically transformed, grown in
liquid culture and maintained in simple mineral medium thereby facilitating stable and
inexpensive supply [5].
Until now, little is known about SQL biosynthesis in Apiaceae plants. In contrary, most
investigations have been focused on artemisin or costunolid, both produced by Asteraceae. Even
though SQL isolated from Apiaceae plants contain stereochemical conformations different from
those found in Asteraceae [3], comparison of the biochemical data with that of related SQL
enabled us to propose an enzymatic pathway by which a thapsigargin precursor could be
generated [3]. Several steps in this pathway are believed to be carried out by cytochrome P450’s.
Systematic investigation of the various steps will be possible by utilization of the unique high
throughput sequencing database of T. garganica and T. villosa available in house. So far, this data
lead to the cloning of two sesquiterpene synthases and eight CYP450 candidate genes, which are
currently under investigation concerning their substrate specificity and their role in thapsigargin
biosynthesis.
[1] Søhoel H, Jensen AM, Moller JV, Nissen P, Denmeade SR, Isaacs JT, Olsen CE, Christensen SB (2006) Natural
products as starting materials for development of second-generation SERCA inhibitors targeted towards prostate
cancer cells. Bioorg Med Chem 14: 2810-2815
[2] Christensen SB, Andersen A, Smitt UW (1997) Sesquiterpenoids from Thapsia species and medicinal chemistry
of the thapsigargins. Fortschr Chem Org Naturst 71: 129-167
[3] Drew DP, Krichau N, Reichwald K, Simonsen HT (2009) Guaianolides in apiaceae: perspectives on
pharmacology and biosynthesis. Phytochem Rev 8: 581-599
[4] Drew DP, Rasmussen SK, Avato P, Simonsen HT (2011) Comparison of volatiles and essential oil components
from Thapsia garganica L. and Thapsia villosa L. and discussion of guaianolide biosynthesis in Apiaceae.
Phytochem. Anal. in press
[5] Drew DP, Krichau N, Reichwald K, Simonsen HT (2009) Guaianolides in apiaceae: perspectives on
pharmacology and biosynthesis. Perspec. Med. Chem. 3:1-6
112
P57
Determination of phytoestrogens in natural products by liquid
chromatography–tandem mass spectrometry
Ivana Sosnovcova, Anna Hurajova, Vera Schulzova, Jana Hajslova
Department of Food Chemistry and Analysis, Institute of Chemical Technology Prague, Technická 5, 166
28 Prague 6 – Dejvice, Czech Republic,
[email protected]
Phytoestrogens are plant-derived compounds, which commonly occur in a wide range of foodand feedstuff. Isoflavones, the well known group of phytoestrogens, are the most abundant
in soybeans but they can be also found in alfalfa sprouts and in other legumes. The main
isoflavones presented in plant matrices are daidzein, genistein, glycitein, formononetin and
biochanin A. As suggested in recently published studies, these plants may comprise substantial
part of dairy cows feed, thus presenting a potential source of isoflavones in bovine milk and milk
products. Phytoestrogens represent biologically active compounds showing estrogenic activity
similar to that of sex hormones – estrogens [1]. Epidemiologic studies have suggested a protective
effect of isoflavones against a number of adult chronic diseases, including coronary heart disease
and breast, endometrial, and prostate cancers. Previously, analysis of phytoestrogens in food was
performed using either HPLC-UV or GC-MS technique [2].
In the presented study, suitable analytical method for determination of phytoestrogens in
different matrices was developed. The performance characteristics achieved using an ultra
performance liquid chromatography (UPLC) coupled to a tandem mass spectrometer (MS/MS)
were evaluated, low limits of quantification (LOQs) were the key criteria that enable control of
compounds and their metabolites even at very low levels. The main aim of this study was to
monitor the content of phytoestrogens in different plants, soy-based foodstuff and nutraceuticals.
Transfer via feedstuffs into the bovine milk and milk products, as well as the levels of equol
(daidzein metabolite) were investigated.
Acknowledgement: This study was carried out within the projects NPVII 2B08073 supported by the
Ministry of Education, Youth and Sports of the Czech Republic, partly from the projects MSM
6046137305, and Specific University Research (MSMT No. 21/2011).
[1] Krajcova A., Schulzova V., Lojza J., Krizova L., Hajslova J. (2010) Phytoestrogens in bovine plasma and milk –
LC-MS/MS analysis. Czech J. Food Sci. 28, 264-274
[2] Dabrowski W. M., Sikorski Z. E. (2005) Toxins in Food. CRC Press, 85-104
113
P58
Anti-inflammatory effects of Thai Traditional medicine called
Ridsiduangmahakan
Suchada Kittisrisopita, Arunporn Itharata
a
Department of Applied Thai Traditional Medicine, Faculty of Medicine, Thammasat University, 12120,
Pathumthani, Thailand
[email protected]
Ridsiduangmahakan (RSD) preparation was used to treat hemorrhoid patients by Thai
traditional doctors long time ago. Thus, this study was aimed to investigate anti-inflammation of
this preparation by three anti-inflammation pathway: inhibitory activity on LPS induced NO
production, TNF-D release and LPS-stimulated PGE2 in macrophage cells (RAW264.7) [1]. The
preparation was macerated with 95%ethanol and its crude extract characteristic was divided as
two part (solid and oil). The percentage of yields of the ethanolic extract were 16.30% (oil 4.68%
and solid 11.62%). The results of RSD extract (oil, solid and oil+solid) on inhibitory activity on
LPS induced NO production showed IC50 values of 48.1±1.8, 56.9±5.1 and 78.9±0.6 µg/ml,
respectively and no cytotoxicity observed. Indomethacin was used as positive control with IC50
value of 20.32 Pg/ml or 56.78 PM. The inhibitory effects on LPS-induced TNF-D release in RAW
264.7 cells were found that RSD (oil) gave IC50 value better than indomethacin (IC50 value of
31.86r1.86 Pg/ml and 51.41r3.58 Pg/ml or 143.69r10.01 PM, respectively) whereas RSD (solid)
and RSD (oil+solid) did not exhibit inhibitory effect on TNF-D (IC50 values > 100 Pg/ml). In
addition, the inhibitory effects of RSD (oil) on LPS-stimulated PGE2 showed the best IC50 value (
8.85r1.60 µg/ml) followed by RSD (solid+oil) 16.80r4.04 µg/ml and RSD (solid) 21.22r3.29
µg/ml (IC50 value of indomethacin = 1.00±0.43 µg/ml or 2.80r1.20 PM). These results lead to
conclude that RSD preparation is anti-inflammatory drug through COX-2 inhibition pathway
rather than through inhibition on TNF-D and NO production. It was suggested that oil of RSD
preparation should be selected to prepare as antiinflammation drug and also supported using by
folk doctors for treatment hemorrhoid patients.
[1] Tewtrakul, S. & Subhadhirasakul, S. (2008) Effects of compounds from Kaempferia parviflora on nitric oxide,
prostaglandin E2 and tumor necrosis factor-alpha productions in RAW264.7 macrophage cells. Journal of
Ethnopharmacology, 120, 81–84.
114
P59
Varietal and environmental effects on some nutraceuticals in selected
Spanish common beans (Phaseolus vulgaris L.)
Mercedes M. Pedrosa, Mara Sacristán, Carmen Burbano, Alejandro Varela, Blanca
Cabellos, Carmen Cuadrado, Mercedes Muzquiz
Instituto Nacional de Investigación y Tecnología y Alimentaria (INIA), Dpto Tecnología de Alimentos,
Ctra A Coruña KM 7.5, 28850 Madrid, Spain
mmartí[email protected]
Beans are the most important grain legumes for direct human consumption in the world. Their
production is almost twice that of chickpea, which is the second most important grain legume.
They are consumed as mature grain, as immature seed, as well as a vegetable (both leaves and
pods) [1].
Nowadays, the nutritional value of legumes is gaining considerable interest in developed
countries because of the demand for healthy foods [2,3]. Several reports claim that inclusion of
legumes in the daily diet has many beneficial physiological effects in controlling and preventing
various metabolic diseases such as diabetes mellitus, coronary heart disease and colon cancer [4].
Related to nutraceutics foods, there are some preview researches about the antinutrients’s
behavior in legumes, most particularly in beans. These substances are plant secondary
metabolites, now increasingly recognized for their potential benefits for human health, amongst
them are trypsin inhibitors, phytates, phenolic substances and α-galactosides [5]. Recent research
has revealed the possible therapeutic properties of phytate in the prevention breast and colon
cancers, probably due to its anti-oxidant properties. Phytate has also been implicated in the
reduction of cholesterol and other lipids due to its presence in high fiber diets [6]. Moreover,
trypsin inhibitors are effective in preventing, or suppressing, carcinogen-induced transformation
in vitro and carcinogenesis in animals. Prebiotic effect has been recognized in D-galactosides like
as fiber functions and could be used to functional ingredient once it have been extracted and
purified. In addition, phenolic compounds are attributed antioxidant properties.
The aim of this work is to determine the composition on some nutraceutical compounds
(galactosides, inositol phosphates, trypsin inhibitors and phenolic compounds) for some selected
Spanish varieties of common beans and to verify the effect of the environment on the content of
these compounds.
Most of the research on Spanish dry beans has been related to varietal selection. The criteria
for selection have always been resistance to disease or high yields but never phytochemical
composition. So, the results obtained will show a starting point in the hypothesis of selection of
bean varieties with high nutritional value and rich in bioactive components, as well as the
development of new functional foods derived from them.
Acknowledgements: INIA, Project RTA2008-00007-C02.
[1] Sathe S.K. , (2002) Crit. Rev. Biotechnol., 22, 175–223
[2] Shehata N.A., Darwish N., El-Nahry F., Andel-Razek F.A. (1988) Nahrung, 32, 3–8
[3] Simpson H.C., Lousley R.S., Greekie M., Hockaday T.D.R., Carter R.D., Mann J.I. (1981).Lancet, 1, 1-4
[4] Tharanathan R.N., Mahadevamma S. (2003) Trends Food Sci. Technol., 14, 507–518,
[5] Kigel J. (19999 Biotech. Agron. Soc. Environ., 3, 205–209
[6] Midorikawa K., Murata M., Oikawa S., Hiraku Y., Kawanishi S. (2001) Biochem. Biophys. Res. Commun., 288,
552–557
115
P60
Antibacterial activity against biofilm formation of usnic acid and
hyperforin
A. Rosatoa, B.P.I. Schiavoneb, G. Caggianod, M.Piarullib, L. Verottac, F. Corboa, C.
Franchinia
a
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Bari Aldo Moro,
Bari, Italy
b
Doctorate School of Biomedical Sciences, Hygiene Section, University of Bari Aldo Moro, Bari,
Italy
c
Department of Organic and Industrial Chemistry, University of Milano, Milano
d
Department of Biomedical Science, Human Oncology Section,University of Bari Aldo Moro,
Bari, Italy
[email protected]
In the human body, bacteria are present in biofilms in essentially every niche that they colonize
such as skin flora, oropharyngeal and nose flora, intestinal flora, in addition bacteria adhere to
endovascular structures. Thus these microorganisms can compromise a wide number of medical
devices, exposing patients at risk for local and systemic infectious complications.
Many natural products show antimicrobial activity against a number of bacteria, but only a
small number of these are also modulators (inhibitors or disrupters) of the biofilm formation, one
of the limits of many antibiotics to be efficacious [1].
In the light of these evidences we decided to test two natural phloroglucinols: usnic acid, a
secondary lichen metabolite, and hyperforin [2], a phytochemical compound, and some of their
derivatives, against biofilms. The assays were performed on biofilms from Gram+ and Grambacterial strains using in vitro diffusion cell bioassay system [3].
Scanning electronic microscopy (S.E.M.) revealed that the presence of these substances
modified and disrupted the structure of the tested biofilms. In addition, to test cell viability of
bacterial strains, viable counts of the biofilms of Staphylococcus aureus ATCC 29213,
Staphylococcus aureus ATCC 43300 and Enterococcus faecalis ATCC 29212 were performed
[4]. Actually we are supporting these antimicrobial activity on pathogenic oropharyngeal flora
using Streptococcus mutans ATCC 35668 and Streptococcus sanguinis ATCC 10556 forming
biofilms as bacterial strains.
The in vitro model used could represent an useful system to know the interactions that appear
to operate on biofilm. The experimental details and results of the study are reported in this
communication.
[1] Richards, Justin J.; Melander, Christian. (2009) Small molecule approaches toward the non-microbicidal
modulation of bacterial biofilm growth and maintenance. Anti-Infective Agents in Medicinal Chemistry, 8(4),
295-314.
[2] P. Avato , F. Raffo , G. Guglielmi , C. Vitali and A. Rosato. Extracts from St. John’s Worth and their
antimicrobial activity. Phytotherapy Research. 18, 230-232. 2004.
[3] Y. H. Samaranayake, J. Ye, J. Y. Y. Yau, B. P. K. Cheung, and L. P. Samaranayake (2005) In Vitro Method To
Study Antifungal Perfusion in Candida Biofilms J. Clin. Microbiol. 43: 818 - 825.
[4] Mohammed A. S. Alem and L. Julia Douglas (2004) Effects of Aspirin and Other Nonsteroidal AntiInflammatory Drugs on Biofilms and Planktonic Cells of Candida albicans Antimicrob. Agents Chemother. 48:
41 – 47.
116
P61
Antidiabetic effect of extract of Terminalia arjuna in streptozotocininduced diabetic Wistar rats
Tan Benny KH, Yong Edwin WW, Hsu A
Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, MD
11, #05-09, 10 Medical Drive, Singapore 117597
[email protected]
Terminalia arjuna is an important medicinal plant widely used against various ailments.
Numerous studies have shown antioxidant, cardiotonic and hypoglycaemic, properties of the
herb1, 2. However few studies have been conducted to probe its anti-diabetic potential. Oxidative
stress has been implicated in the development of diabetic complications such as hypertension. In
view of the close link between diabetes, oxidative stress and cardiovascular disease, we
investigated the anti-diabetic capability of a 70% methanolic extract of T. arjuna administered
orally for 28 days to streptozotocin (STZ)-induced diabetic rats.
A dose-response study of T. arjuna extract on glucose tolerance in STZ-diabetic rats (results in
Figure 1) was followed up by a 28-day treatment with the extract. The extract significantly
reduced fasting serum glucose level in STZ-diabetic rats at the end of the 28-day period.
Decreased hepatic glucose-6-phosphatase activity and increased muscle glycogen content were
found in the extract-treated rats. GLUT4 expression in skeletal muscle was also enhanced in these
rats. Serum and pancreatic insulin concentrations were not significantly increased while serum
and hepatic triglyceride and total cholesterol levels were significantly decreased. Interestingly, T.
arjuna-treated rats had elevated GSH levels and augmented activities of antioxidant enzymes.
Results from this study provide support for the use of T. arjuna in the treatment and possible
prevention of complications of diabetes mellitus. Future purification and bioassay-guided studies
are planned to identify the bioactive component(s) in the herb plant. Further detailed studies can
then be conducted to elucidate the exact mechanism(s) underlying the potential effectiveness of T.
arjuna as an anti-diabetic herb.
Figure 1. Dose-response effect of T. arjuna extract on glucose tolerance in STZ-diabetic rats
[1] Sunyana Jain, P. P. (2009). Terminalia arjuna: a sacred medicinal plant: phytochemical and pharmacological
profile. Phytochemistry Reviews, 8 (2): 491-502
[2] B. Ragavan, S. K. (2006). Antidiabetic effect of T arjuna Bark Extract in Alloxan-Induced Diabetic Rats. Indian
Journal of Clinical Biochemistry, 21 (2): 123-128.
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P62
Simultaneous determination of salicylates and benzoic acid in food
by SPME-LC-UV-DAD
A. Aresta, G. Lioce, C.G. Zambonin
Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, via Orabona 4, 70126 Bari
[email protected]
Salicylate, acetylsalicylic acid (aspirin) and their derivatives (salicylates) are used as fungicidal
and antimicrobial agents in pharmaceuticals preparations (external use) as well as in the treatment
of inflammatory processes as antipyretic and analgesic drugs (internal use).
Salicylates have been also used in beverages and foods for preservation, but it has been
forbidden since the sixties in several countries due to its toxicity [1].
These chemicals occur also naturally in many plants, including many fruits vegetables, and
herbs. Salicylates in plants act as a natural immune hormone and preservative, protecting the
plants against diseases, insects, fungi, and harmful bacteria.
Salicylates are generally regarded as safe for adults, even if high enough doses are harmful to
everyone. However, most people can handle average amounts of salicylate in food, products and
medications without any adverse affects on their health. Unfortunately, there is a small percent of
the population for which even a small dose of salicylates can be a problem. Some adults and
children may develop symptoms and health problems from salicylates which are dose-related.
This is called ‘Salicylate Sensitivity’ or ‘Salicylate Intolerance’.
Individuals that are sensitive to salicylates may suffer with urticaria, angiooedema, rhinitis,
bronchial asthma and recurrent nasal polyps [2]. The chronic nature of some of these clinical
presentations, without other obvious cause, may suggest an underlying etiology related to dietary
salicylates. A low salicylate diet may be of clinical benefit to such affected individuals. This
cannot be established however, until the salicylate content of different food and drinks is known.
Data on the salicylate content of foods are scarce and contradictory. Our aim was to develop an
accurate analytical method to measure the salicylates content of selected food commodities.
Existing papers on this topic have been essentially based on chromatographic techniques, after
purification of the analytes by means of complicate isolation procedures. A good alternative could
be represented by the use of solid-phase microextraction (SPME), a solventless technique initially
coupled to GC and later interfaced also to LC [3].
Thus, in the present work, a solid-phase microextraction (SPME)–LC–UV-DAD method for
the simultaneous determination of salicylates and benzoic acid in food samples was developed for
the first time using a polydimethylsiloxane/divinylbenzene (PDMS/DVB) coated fiber. The
procedure required very simple sample pretreatments, isocratic elution, and provided highly
selective extractions. The applicability of the method was demonstrated on different food
products, i.e. kiwis, blueberries, lemons, mandarins, oranges, broad beans, and commercial fruit
juices.
[1] T.M. Brown, A.T. Dronsfield, P.M. Ellis and J.S. Parker, Educ. Chem., 35, 1998, 47-51.
[2] S. Jantti-Alanko, E. Holopainen, and H. Malmberg, Rhinology, 8, 1989, 59–64.
[3] C.G. Zambonin, Anal Bioanal. Chem., 375, 2003, 73–80.
118
P63
Formulation and characterization of new packaging material
incorporating chitosan nanoparticles-vitamin E and C for food shelf
life improvement
Calvano C.D.a, Trapani G.b, Barnaba M.a, De Giglio E.a, Cafagna D.a, Aresta A.a, Zambonin
C.G. a, c
a
Dipartimento di Chimica, Università degli Studi di Bari Aldo Moro, via Orabona 4, 70126 Bari
Dipartimento Farmaco Chimico, Università degli Studi di Bari Aldo Moro, via Orabona 4, 70126 Bari
c
Centro Interdipartimentale SMART, Università degli Studi di Bari Aldo Moro, via Orabona 4, 70126 Bari
[email protected]
b
One of the principal causes of food quality deterioration is the oxidation of unsaturated lipids
initiated by free radicals. When lipids are exposed to environmental factors such as air, light and
temperature, oxidation reactions start to produce undesirable flavours, rancid odours, discoloration
and other forms of spoilage. During oxidation, a series of volatile and non-volatile compounds are
originated, altering sensory proprieties, causing rancid flavors and decreasing the nutritional quality
and safety, which may play a role in the development of some diseases [1]; in addition lipid
oxidation could cause a consumer rejection of the food product.
The rate of autoxidation depends on temperature, pH, the degree of unsaturation of the
fatty acids and the number of unsaturated fatty acids in the triacylglycerol or phospholipid
molecule, as well as on the availability of oxygen and transition metal ions. The other major
cause of food spoilage during storage is the bacterial contamination arising from several sources,
such as the animal itself, the external environment and its handling.
Packaging is important to preserve the quality and safety of fruits, vegetables and processed
foods for assured shelf life extension. The new generation of edible coatings is especially designed
to allow the incorporation and/or controlled release of antioxidants, vitamins, nutraceuticals, and
natural antimicrobials [2-4].
Among packaging material, chitosan has been well-known for its excellent film-forming
property, antimicrobial activity, and unique coagulating ability with metal and other lipid and
protein complexes. It seems to be due to the presence of high density of amino groups and hydroxyl
groups in the chitosan polymer structure. Its high binding ability and antimicrobial properties are
both beneficial in developing new applications of this natural polymer in food preservation. Herein,
we aimed at the formulation of chitosan-cyclodextrin nanoparticles (NPs) obtained by the ionic
gelation process in order to load vitamin E and C selected as powerful antioxidant agents. In fact,
vitamin E is important in the prevention of lipid peroxidation, whereas vitamin C reacts effectively
with superoxide and hydroxyl radicals. Vitamins C and E and vitamin precursors (e.g. carotenoids)
should reduce the rate of initiation or prevent the propagation of free radicals as notable nonenzymatic antioxidants.
The NPs will be characterized in terms of light scattering, zeta-potential, vitamins content and by
X-Ray Photoelectron Spectroscopy (XPS) measurements. Afterwards, the NPs loading vitamin C
and E will be dispersed on polymeric films that are usually found as food packaging materials.
[1] R. M. Adibhatla, J.F. Hatcher Antioxidants & redox signaling, 12, 2010, 125-169
[2] M. Vargas, C.Pastor, A.Chiralt, D. J. McClements, C. Gonz lez-Mart nez Crit Rev Food Sci Nut, 48, 2008, 496–511.
[3] P. K Dutta, S. Tripathi, G. K. Mehrotra, J. Dutta, Food Chem 114, 2009,1173-1182.
[4] P.E. Chiu, L.S. Lai, Int J Food Microbiol 139, 2010, 23-30.
119
Complete List of Presentations
PLENARY LECTURES
PL 1
PL 2
PL 3
PL 4
PL 5
PL 6
PL 7
PL 8
PL 9
PL10
PL11
G. Appendino - Food-based drug discovery: overcoming the nutrition-medicine divide.
M. Muzquiz - Bioactive compounds in legumes: pronutritive and antinutritive actions. Implications for
nutrition and health.
nUG~GTGlmzh˅GyGGuGmGGjU
V. Cheynier - Phenolic compounds : from plants to foods.
J.L.Wolfender - New trends in the analysis of natural products for the detailed metabolite profiling of crude
extracts.
Y. Tikunov, S. van Heusden, J. Molthoff , H. van de Geest , A. Ballester, I. Romero, A. Granell, R. de Vos, A.
Bovy - Breeding for improved flavour and health.
S.B. Engelsen, F. Savorani - NMR metabolomics: advantages and disadvantages for studying bioactive
components in food.
A.M. Aura - Phenolic nutritional phytochemicals as biomarkers of plant food intake.
B. A. Halkier,U.H. Mortensen, L. Albertsen, B. Salomonsen, M. Mikkelsen, J. K.Vester, F. Geu-Flores Bioengineering of glucosinolates production in yeast.
V. Lanzotti - Bioactive Natural Compounds from Food Plants.
P. J. Russell - Naturals for the masses – Chemistry approaches for safe commercialization.
SHORT LECTURES
SL 1
SL 2
SL 3
SL 4
SL 5
SL 6
SL 7
SL 8
SL 9
SL10
SL11
SL12
SL13
SL14
SL15
SL16
SL17
SL18
P. Reddell, V. Gordon, P. Parsons, G. Muench, C. Williams- Discovery of New Nutraceuticals from Australia’s
Tropical Rainforests.
J.M.I.Alqudah, K..Reilly, M.T.Gaffney, A.B. Martin-Diana & C.Barry-Ryan - Screening of Irish Fruit and
Vegetable Germplasm for Novel Anti-Tumor and Pesticidal Compounds.
M. Durante, M. Lenucci, L. Rescio, G. Mita, S. Caretto.- Durum Wheat By-Products as Natural Sources of
Valuable Nutrients
L. Brown, H. Poudyal- Anthocyanin-enriched purple carrot juice with low carotenoids attenuates
cardiovascular and hepatic symptoms induced by high carbohydrate, high fat feeding in rats.
M. Lenucci, L. Rescio, G. Piro, G. Dalessandro - Tomato organic lycopene: the way to obtain a unique healthy
product.
F. Bucar, E. Knauder - Isoflavone Composition of Soy and Red Clover Dietary Supplements.
Candiracci, A. Castaño, C. SantaMaría, E. Revilla, M. Dominguez-Barragán, B. Rodríguez-Morgado, D.
García-Antras, M. Carballo, J. Bautista, M.D. Herrera, J. Parrado - Antioxidant and anti-inflammatory
properties of a new soluble extract from grape pomace. M.
S. Nunes, A.T. Serra, M. R. Bronze, Catarina M.M. Duarte, A.A. Matias - Polyphenols-rich extracts obtained
from Prunus avium and opuntia ficus indica as Natural anti-inflammatory modulators for Inflammatory Bowel
Diseases.
T. Magrone, S. Fontana, E. Jirillo.Polyphenols from red wine exert potential beneficial effects to human health.
M.A. Papandreou, M. Tsachaki, S. Efthimiopoulos, D. Klimis-Zacas, M. Margarity, F. N. Lamari Differential antioxidant and anti-amyloid effect of a wild blueberry polyphenol-rich extract: in vitro and in
vivo studies.
V. Schulzova, H. Novotna, L. Vaclavik, A. Hurajova, J. Hajslova - Metabolomic Profiling and Fingerprinting:
New Effective Approach for Plant Characterisation and Authentication.
M. Zotti, M. Colaianna, M.G. Morgese, L.Ciuffreda, P. Tucci, P. Avato, L. - Estrous cycle in female rats
influences the neurochemical and neurobehavioural effects of carvacrol
J. Gruz, L. Spichal - Application of purine derivative LGR-1814 improves functional properties of field-grown
lettuce (Lactuca sativa).
A. Szakiel, C. Pączkowski, H. Koivuniemi, S.Huttunen - Comparison of triterpenoid content of Vaccinium
vitis-idaea berries and leaves collected from natural forest habitats in Finland and Poland.
M. Monschein, K. Ardjomand-Woelkart, I. Wolf, J. Rieder, R. Bauer - Formation of astragaloside IV from
acylated astragalosides during extraction of Astragali Radix
K. Michalska, W.Kisiel - Sesquiterpenoids from aerial parts of Lactuca aculeate
The volatile fraction of herbal teas.Christine Tschiggerl and Franz Bucar
Tassoni A., Ferri M. - Elicitation, production and release of polyphenols in fed-batch fermentation of grape
SL19
SL20
SL21
SL22
SL23
SL24
SL25
SL26
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SL28
cells.
S. M. Cardoso, S.I. Falcão, A.M. Peres, O.R. Pereira, M.R.M. Domingues Insights into olive mill wastewaters
phenolics.
C. Longo, R. Zefferino, A. Leone - Molecular Mechanisms Underlies the Biological Activities of
Phytochemicals: searching the gap
G. Farre, S. Rivera, C. Bai, R. Canela, G. Sandmann, C. Zhu, T. Capell, P. Christou - The Arabidopsis Orange
(Or) gene generates a metabolic sink and enhances accumulation of carotenoids in cereal cells and plants.
S. Bhattacharya, S. Sinha, P. Dey, N. Das, M. K. Maiti - Production of nutritionally desirable fatty acids in
seed oil of Indian mustard (Brassica juncea L.) by metabolic engineering.
H. Poudyal, L. Brown - α-Linolenic acid-rich oil from chia seed (Salvia hispanica L.) induces lipid
redistribution, heart and liver protection in diet-induced obese rats.
M-H. Teiten, A. Gaigneaux, S. Chateauvieux, A.M Billing, J. Renaut, M. Dicato, M. Diederich - Proteomic
identification of differentially expressed proteins in curcumin-treated prostate cancer cells.
J. Hadsbjerg, S. Willkomm, K. Nöske, T.D. Rasmussen, E. Brunstein, K.. Niefind, O.G. Issinger - Natural
compounds as selective inhibitors for protein kinases and their potential application in cancer research.
M. Miele, A.M. Mumot, A. Zappa, P. Romano, L. Ottaggio - Hazel and Other Angiosperm Plants as New
Sources of Compounds with Taxane-Like Activity.
C. Mathon, M.Duret, P. Edder, S. Bieri, P. Christen - Targeted screening of botanicals in herbal dietary
supplements by LC-MS/MS with an Information Dependent Acquisition approach.
A. Mari, P. Montoro, S. Piacente - LC-MS/MS determination of marker metabolites in the ethanolic extract and
in mother tinctures of Vitex agnus castus fruits.
POSTERS
P1
P2
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P5
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P7
P8
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L. Barros, C. Grangeia, A. Martins, I. C.F.R. Ferreira.- Nutritional and nutraceutical potential of saprotrophic
and mycorrhizal wild edible mushrooms from Northeast Portugal.
F. Conforti, M. Marrelli, G. A. Statti, F. Menichini -In vitro inhibition of nitric oxide production and free
radical scavenging activity of non-cultivated Mediterranean vegetables.
A. Bisio, E. Giacomelli, G. Damonte, A. Salis, D. Fraternale, G. Romussi, S. Cafaggi, D. Ricci, N. - De
Tommasi - A new clerodane diterpenoid from the aerial part exudate of Salvia chamaedryoides Cav.
A. A Ahmadu, A. Agunu - Chemical study on Pavetta corymbosa.
S. I. Falcão, C. Freire, M. Vilas-Boas - Physicochemical characterization and bioactivity of Portuguese
Propolis towards its standardization.
A. Itharat, S. Ruangnoo, P. Thongdeeying - A new cytotoxic dihydrophenantrene against human cancercCells
from Dioscorea membranacea roots.
A.V. Ferlemi, F.N. Lamari, G. Iatrou, M. Margariti - Effects of daily consumption of Rosmarinus officinalis
and Hypericum vesiculosum infusions on rodent cognitive function and cerebral acetylcholinesterase activity.
B. Dechayont, P. Thondeeying, A. Itharat - Cytotoxic activity against lung and prostate cancer cells of the
ethanolic extract from Antidesma thwaitesianum Müll. Arg. Leaves.
J. Chunthorng-Orn, A. Itharat - Comparison of antimicrobial, antioxidant, anti inflammatory activities and
total phenolic contents of female and male trees from Carica papaya Roots.
J. P. de Andrade, S. Berkov, K. B. da Silva, L. G. Sachet, F. Viladomat, C. Codina, J. A. S. Zuanazzi, J.
Bastida - Alkaloids of Hippeastrum papilio.
A.M. Schito, G. Piatti, A. Bisio, E. Giacomelli, G. Romussi, C. Pruzzo - Activity of compounds extracted from
Salvia aurea against methicillin- resistant Staphylococci.
M. Dimitrova, Z. Yordanova, V. Kapchina-Toteva - Influence of benzylaminopurine on antioxidant activity
and phenolic content on in vitro propagated Lamium album L.
D. Gođevac, V. Tešević, V. Vajs, S. Milosavljević, G. Zdunić , M. Stanković - Currant seed extracts showing
protective effect on human lymphocytes DNA.
C. Sae-wong, S. Tewtrakul, P. Tansakul - Inhibitory effect of Kaempferia parviflora on NFкB, Rel A, NOS,
COX-2 and TNF-D mRNA expression.
F. Corbo, A. Lovece, C. Bruno, F. La Forgia, S. Fontana, M. Roselli, A. Laghezza, G. Lentini, C. Franchini, S.
Habtemariam - Antioxidant activity of Uva di Troia Canosina ad acino piccolo: comparison of different
extraction methods.
P. Phuaklee, A. Itharat - Nitric oxide inhibitory effect and antioxidant activity of the extracts from Musa
sapientum bracts and flowers.
S. Gonçalves, D. Gomes, P. Costa, A. Romano - Correlation between phenolic content and antioxidant
activity of infusions from eleven Mediterranean plant species.
F. Gaascht, M. Jain, M-H. Teiten, M. Schumacher, G. Kirsch, D. Bagrel, M. Dicato, M. Diederich -New
P19
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natural compounds of Dionaea muscipula as anticancerous agents.
M. Candiracci, J. Parrado,M.L. Justo, R.Rodríguez-Rodríguez, A. Castaño, M.D. Herrera - Effects of a rice
bran enzymatic extract on cytokines implicated in the inflammatory state of adipose tissue in a rat model of
obesity.
C. Bai, S. M. Rivera, V. Medina, G. Sandmann, R. Canela, C. Zhu, T. Capell, P. Christou - Metabolic
engineering of carotenoid biosynthesis in riceendosperm.
J-I. Kim, J. Kim, H. Choi, M-J. Kang, S. Jeong, M. Kim- Effect of quercetin on hepatic steatosis in ob/ob mice.
P. Trisolini, A. Ignazzi, F. Pezzolla, D. Lorusso, S. Fiume, L. Debellis, M.A. Maselli - Olive oil phenols
improve the altered human intestinal permeability. Study in vitro.
I. Kowalska, D. Jędrejek, Ł. Cieśla, S. Piacente, M. Mosullo, W. Oleszek, A. Stochmal - Phenolic composition
of Trifolium scabrum L. aerial parts.
M. S. Valyova, M. A. Dimitrova, Y. Ganeva, V. M. Kapchina-Toteva, Z. P. Yordanova - Evaluation of
antioxidant and free radical scavenging potential of Lamium album L. growing in bulgaria.
A. Tava, B. Pintus, A. Ursino, B. Campion - Evaluation of the saponin content in lines of common beans
(Phaseolus vulgaris L.) selected in Italy.
M.P. Argentieri, R. Pascale, P. Avato - Glucosinolates from “Mugnolo”, a variety of Brassica oleracea.
F. N. Lamari, M. A. Papandreou, A. P. Dimakopoulou, E. Drimtzias, A-B. Ferlemi, O. Makri, P. Cordopatis,
C. Georgakopoulos, M. Margarity - Antioxidant properties of Saffron against H2O2- & Selenite-induced
oxidative stress: in Vitro & in Vivo Studies.
Y. Jirakiattikul, A. Itharat - Surface sterilization and in vitro shoot multiplication of Dioscorea membranacea
Pierre ex Prain & Burkill.
P. L. Falé, M. A. Filipe, L. Ascensão, M. L. Serralheiro, L. Mira - Activity of Plectranthus barbatus extract
against inflammatory response in human neutrophils.
G. Sanahuja, G. Farre, L. Bassié, C. Zhu,T. Capell, P. Christou - Elucidation of rate limiting steps of ascorbate
biosynthesis in maize.
M. Marrelli, F. Conforti, G. A. Statti, F. Menichini - Antioxidant activity and antiproliferative properties of
Ficus carica cv. Dottato fruits.
J. Nantana - Some Morphology and Genetic Relationship of Dioscorea membranacea using Molecular
Technique.
B. Miralpeix, R. Hoefer, D. Werck, L. Dong, H. J. Bouwmeester, T. Capell, P. Christou - Metabolic
engineering of terpenoid biosynthesis in transgenic tobacco plants.
A. Napolitano, H. K. Sellami, M. Masullo, S. Smiti, S. Piacente, C. Pizza - Quali-quantitative profiling of the
polar extracts of the flowers of Ammi visnaga plants grown in different conditions.
A. Mitra, M. Kumar - Harnessing valuable botanicals from Areca catechu pericarps.
L. Torras-Claveria, S. Berkov, F. Viladomat, C. Codina, J. Bastida - Exact mass and QToF fragmentation
data base of bioactive Amaryllidaceae alkaloids.
M. Ferri, A. Gianotti, A. Tassoni - Suitability of ancient and modern cereals to produce novel functional
fermented foods with a high content of antioxidant compounds.
O. Prajuabjinda, A. Itharat - Antioxidant and cytotoxic activities against lung cancer cells of Thai Medicinal
Plants for Cancer treatment in Kumpramong Temple.
M. Monschein, F. Bucar - Phytochemical investigation of phenolic compounds of elderflowers growing at
different altitudes.
P. Rithichai, A. Itharat - Growth of Dioscorea membranacea Pierre ex Prain & Burkill and dioscorealide B
accumulation at different harvest times.
M. Maldini, P. Montoro, A. I. Hamed, U. A. Mahalel, W. Oleszek, A. Stochmal, S. Piacente - Strong free
radical scavenging phenolics from Acacia nilotica pods.
B. Thiem, M. Ożarowski, M. Szulc, R. Kujawski, P. M. Mrozikiewicz, P.L. Mikołajczak
Neuropharmacological study of Eryngium planum L. extract.
H. Novotna, V. Schulzova, I. Sosnovcova, J. Hajslova - Using DART-TOFMS for quality assessment of rose
hip (Rosa Canina) fruits.
R. Rattarom, I. Sakpakdeejaroen, A. Itharat - In Vitro cytotoxic activity against human lung cancer cells of the
ethanolic extract from Benjakul preparation and its isolated compounds.
Watermelon as a source of bioactive molecules. Marcello Salvatore Lenucci, Stefania Grassi, Anna Eleonora
Caprifico, Maria Chiara Contino, Anna Montefusco, Gabriella Piro, Giuseppe Dalessandro
S. M. Kim, J-S. Jeon, Y-J. Jung, L. Ly, B-H. Um - Adaptation of leaves in Ligularia fischeri (Ledeb.) Turcz to
sun, shade and drought: chlorophylls and phenolic compounds.
P. Hansakul, U. Srisawat, N. Lerdvuthisopon, A. Itharat - The correlation between the total phenolic and
flavonoid contents and their antioxidant properties of Thai rice cultivars in chemical and cell-based assays.
A. M. Rodrigues, P. L. Falé, L. Ascensão, M. L. Serralheiro - Acetylcholinesterase inhibition and antioxidant
activity of three Asteraceae species from the Portuguese Aromatic Flora.
P49
P50
P51
P52
P53
P54
P55
P56
P57
P58
P59
P60
P61
P62
P63
V. Cardile, G. Pannuzzo, A. Graziano, J. Garbarino, N. Troncoso, A. Russo - Rosmarinus officinalis extract: a
promising clinical management of Krabbe leukodystrophy.
J. Srdanov, V. Krstonosic, M. Krickovic, N. Lakic, B. Bozin - Antioxidant properties of Asplenium
trichomanes, Polypodiaceae.
S. Uttama, I. Sakpakdeejaroen, R. Rattarom, A. Itharat - Antioxidant activities, gamma oryzanol ,vitamin E
and total phenolic content of hom Mali 105 Rice (Oryza sativa) in different extraction methods.
S. Ruangnoo, A. Itharat - Anti-inflammatory activities of Smilax corbularia Kunth and its constituents.
M.P. Argentieri, F. Macchia, P. Papadia, F.P. Fanizzi, P. Avato - Bioactive compounds from Capparis spinosa
subsp. Rupestris.
B. Szajwaj, Ł. Pecio, M. Szumacher-Strabel, A. Cieślak, W. Oleszek, A. Stochmal - Isolation of bioactive
compounds from Saponaria officinalis L.
C. Tiulea, C. Peev, S. Feflea, C. Dehelean, C. Soica, A. Motoc - The influence of isoflavonoid genistein
incorporated in hydroxy-propyl-gamma-cyclodextrin on experimental B164A5 murinic melanoma model on
mice.
C. Weitzel, A. H. Klem, T. Manczak, T. A. Bundgaard, J. M. Larsen, H. S. Toft - SPOTLight: sustainable
production of thapsigargin using light.
I. Sosnovcova, A. Hurajova, V. Schulzova, J. Hajslova - Determination of phytoestrogens in natural products
by liquid chromatography–tandem mass spectrometry.
S. Kittisrisopit, A. Itharat - Anti-inflammatory Effects of Thai Traditional Medicine called
Ridsiduangmahakan.
M. M. Pedrosa, M. Sacristán, C. Burbano, A. Varela, B. Cabellos, C. Cuadrado, M. Muzquiz - Varietal and
environmental effects on some nutraceuticals in selected Spanish common beans (Phaseolus vulgaris L.).
A. Rosato, B.P.I. Schiavone, G. Caggiano, M.Piarulli, L. Verotta, F. Corbo, C. Franchini - Antibacterial
activity against biofilm formation of usnic acid and hyperforin.
KH. Tan Benny, W.W. Yong Edwin, A. Hsu - Antidiabetic effect of extract of Terminalia arjuna in
streptozotocin-induced diabetic Wistar rats.
A. Aresta, G. Lioce, C.G. Zambonin - Simultaneous determination of salicylates and benzoic acid in food by
SPME-LC-UV-DAD
C.D. Calvano, G. Trapani, M. Barnaba, E. De Giglio, D. Cafagna, A. Aresta, C.G. Zambonin - Formulation
and characterization of new packaging material incorporating chitosan nanoparticles-vitamin E and C for
food shelf life improvement
Authors´ Index
A
page
Agunu A.
Ahmadu A.A.
Albertsen L.
Alqudah J.M.I.
Andersen T.B.
Appendino G.
Ardjomand-Woelkart K.
Aresta A.
Argentieri M.P.
Ascensão L.
Aura A-M.
Avato P.
60
50
40
13
112
11
33
118, 119
82, 109
85, 104
36
28, 82, 109
B
page
Bagrel D.
Bai C.
Ballester A-R.
Barnaba M.
Barros L.
Barry-Ryan C.
Bassié L.
Bastida J.
Bauer R.
Bautista J.
Berkov S.
Bhattacharya S.
Bieri S.
Billing A.M.
Bisio A.
Bouwmeester H.J.
Bovy A.
Bozin B.
Bronze M.R.
Brown L.
Bruno C.
Brunstein E.
Bucar F.
Burbano C.
74
41, 76
29
120
57
13
86
66, 92
33
21
66, 92
42
49
44
59, 67
89
29
106
23
16, 43
71
46
20, 35, 95
115
C
page
Cabellos B.
Cafaggi S.
Cafagna D.
Calvano C.D.
Caggiano G.
115
59
119
119
116
Campion B.
Candiracci M.
Canela R.
Capell T.
Caprifico A.E.
Carballo M.
Cardile V.
Cardoso S.M.
Caretto S.
Castaño A.
Chateauvieux S.
Cheynier V.
Choi H.
Christen P.
Christou P.
Chunthorng-Orn J.
Cieśla Ł.
Cieślak A.
Ciuffreda L.
Codina C.
Codruta S.
Colaianna M.
Conforti F.
Contino M.C.
Corbo F.
Cordopatis P.
Costa P.
Cuadrado C.
81
21, 75
41, 76
41,76,86,89
19, 101
21
105
38
15
21, 75
44
22
77
49
41,76,86,89
64
79
110
28
66, 92
111
28
58, 87
19, 101
71, 116
83
68
115
D
page
da Silva K.B.
Dalessandro G.
Damonte G.
Das N.
de Andrade J.P.
De Giglio E.
De Tommasi N.
Debellis L.
Dechayont B.
Dehelean C.
Dey P.
Dicato M.
Diederich M.
Dimakopoulou A.P.
Dimitrova A.M.
Domingues M.R.M.
Dominguez-Barragán M.
Dong L.
66
19, 101
59
42
66
120
59
78
64
111
42
44, 74
44, 74
81
68, 80
38
21
89
Drimtzias E.
Duarte C.M.M.
Durante M.
Duret M.
81
23
15
49
Graziano A.
Gruz J.
105
30
H
page
E
page
Edder P.
Efthimiopoulos S.
Engelsen S-B.
49
25
32
F
page
Falcão S.I.
Falé P.L.
Fanizzi F.P.
Farre G.
Feflea S.
Ferlemi A.V.
Ferlemi A-B.
Ferreira C.F.R.I.
Ferri M.
Filipe M.A.
Fiume S.
Fontana S.
Franchini C.
Fraternale D.
Freire C.
38, 61
85, 104
109
41, 86
111
63
83
57
37, 93
85
78
24, 71
71, 116
59
61
Habtemariam S.
Hadsbjerg J.
Hajslova.J.
Halkier B.A.
Hamed A.I.
Hansakul P.
Herrera M.D.
Hoefer R.
Hurajova A.
Hsu A.
Huttunen S.
71
46
27, 99, 113
40
97
103
21, 75
89
27, 113
117
31
I
page
Iatrou G.
Ignazzi A.
Issinger O-G.
Itharat A.
58
78
46
62,64,65,72
84,94,96,100,
103,107,108,
114
J
page
G
page
Gaascht F.
Gaffney M.T.
Gaigneaux A.
Ganeva A.Y.
Garbarino J.
García-Antras D.
Gelbmann W.
Georgakopoulos C.
Geu-Flores F.
Giacomelli E.
Gianotti A.
Gođevac D.
Gomes D.
Gonçalves S.
Gordon V.
Granell A.
Grangeia C.
Grassi S.
74
13
44
80
105
21
17
83
40
59, 67
93
69
73
73
12
29
57
19, 101
Jaengsuwan N.
Jain M.
Jędrejek D.
Jeon J-S
Jeong S.
Jirakiattikul Y.
Jirillo E.
Jung Y-J.
Justo M.L.
88
74
79
102
77
84
24
102
75
K
page
Kang M-J.
Kim J.
Kim J-I.
Kim M.
Kim S.M.
Kirsch G.
Kisiel W.
Kittisrisopit S.
77
77
77
77
102
74
34
114
Klem A.H.
Klimis-Zacas D.
Knauder E.
Koivuniemi H.
Kowalska I.
Krickovic M.
Krstonosic V.
Kujawski R.
Kumar M.
112
25
20
31
79
106
106
98
91
L
page
La Forgia F.
Laghezza A.
Lakic N.
Lamari F.N.
Lanzotti V.
Larsen J.M.
Lentini G.
Lenucci M.S.
Leone A.
Lerdvuthisopon N.
Lioce G.
Longo C.
Lorusso D.
Lovece A.
Ly L.
71
71
106
63, 83
45
112
71
14, 19, 101
39
103
119
39
78
71
102
M
page
Macchia F.
Magrone T.
Mahalel U.A.
Maiti M.K.
Makri O.
Maldini M.
Manczak T.
Margarity M.
Mari A.
Marrelli M.
Martin-Diana A.B.
Martins A.
Maselli M.A.
Masullo M.
Mathon C.
Matias A.A.
Medina V.
Menichini F.
Michalska K.
109
24
97
42
83
97
112
25, 63, 83
50
58, 87
13
57
78
90
49
23
76
58, 87
34
Miele M.
Mikkelsen M.
Mikołajczak P.L.
Milosavljević S.
Mira L.
Miralpeix B.
Mita G.
Mitra A.
Molthoff J.
Monschein M.
Montefusco A.
Montoro P.
Morgese M.G.
Mortensen U.H.
Mosullo M.
Motoc A.
Mrozikiewicz P.M.
Muench G.
Mumot A.M.
Muzquiz M.
47
40
98
69
85
89
15
91
29
33, 95
19, 101
50, 97
28
40
79
111
98
12
97
14, 115
N
page
Napolitano A.
Niefind K.
Nöske K.
Novotna H.
Nunes S.
90
46
46
27, 99
23
O
page
Oleszek W.
Ottaggio L.
Ożarowski M.
79, 97, 110
47
98
P
page
Pączkowski C.
Pakakrong Thondeeying P
Papadia P.
Pannuzzo G.
Papandreou M.A.
Parrado J.
Parsons P.
Pascale R.
Pecio Ł.
Pedrosa M.M.
Peev C.
Pereira O.R.
31
62, 64
109
105
25, 83
21, 75
12
82
110
115
111
38
Peres A.M.
Pezzolla F.
Phuaklee P.
Piacente S.
Piarulli M.
Piatti G.
Pintus B.
Piro G.
Pizza C.
Poudyal H.
Prajuabjinda O.
Pruzzo C.
38
78
72
50, 79, 90, 97
116
62
81
19, 101
90
16, 43
94
67
R
page
Rasmussen T.D.
Rattarom R.
Reddell P.
Reilly K.
Rescio L.
Revilla E.
Ric de Vos R.
Ricci D.
Rieder J.
Rithichai P.
Rivera S.
Rodrigues A.M.
Rodríguez-Morgado B.
Rodríguez-Rodríguez R.
Romano A.
Romero I.
Romussi G.
Rosato A.
Roselli M.
Ruangnoo S.
Russell P.J.
Russo A.
46
100,107
12
13
15
21
29
59
33
96
41, 76
104
21
75
47, 73
29
59, 67
116
71
61, 108
48
105
S
page
Sachet L.G.
Sacristán M.
Sae-wong C.
Sakpakdeejaroen I.
Salis A.
Salomonsen B.
Samira Smiti S.
Sanahuja G.
Sandmann G.
64
115
70
100,107
59
40
90
86
41, 76
SantaMaría C.
Savorani F.
Schiavone B.P.I.
Schito AM.
Schulzova V.
Schumacher M.
Sellami H.K.
Serra A.T.
Serralheiro M.L.
Simonsen H.T.
Sinha S.
Sosnovcova I.
Spichal L.
Srdanov J.
Srisawat U.
Stanković M.
Statti G.A.
Stochmal A.
Szajwaj B.
Szakiel A.
Szulc M.
Szumacher-Strabel M.
21
32
116
67
27,99,113
74
90
23
85, 104
112
42
99, 113
30
106
103
69
58, 87
79, 97, 110
110
31
98
110
T
page
Tan Benny K.H.
Tansakul P.
Tassoni A.
Tava A.
Teiten M-H.
Tešević V.
Tewtrakul S.
Thiem B.
Tikunov Y.
Tiulea C.
Torras-Claveria L.
Toteva V.K.
Trabace L.
Trapani G.
Trisolini P.
Troncoso N.
Tsachaki M.
Tschiggerl C.
Tucci P.
117
70
37, 93
81
44, 74
69
70
98
29
111
92
68, 80
28
120
78
105
25
35
28
U
page
Um B-H.
Ursino A.
102
81
Uttama S.
107
V
page
Vaclavik L.
Vajs V.
Valyova S.M.
van de Geest H.
van Heusden S.
Varela A.
Verotta L.
Vester J.K.
Viladomat F.
Vilas-Boas M.
27
69
80
29
29
115
116
40
92
61
W
page
Weitzel C.
Werck D.
112
89
Williams C.
Willkomm S.
Wolf I.
Wolfender J-L.
12
46
33
26
Y
page
Yong Edwin W.W.
Yordanova P.Z.
117
68, 80
Z
page
Zambonin C.G.
Zappa A.
Zdunić G.
Zefferino R.
Zhu C.
Zotti M.
Zuanazzi J.A.S.
118, 119
47
69
39
41, 76, 86
28
66
List of participants
Country
Last-name
First-name
Email
AUSTRALIA
BROWN
GORDON
POUDYAL
REDDELL
Lindsay
Victoria
Hemant
Paul
[email protected]
[email protected]
[email protected]
[email protected]
AUSTRIA
BUCAR
MONCHEIN
TSCHIGGERL
Franz
Marlene
Christine
[email protected]
[email protected]
[email protected];
BULGARIA
DIMITROVA
GANEVA
Milena
Yordanka A.
[email protected]
[email protected]
CZECH REPUBLIC
GRUZ
NOVOTNA
SCHULZOVA
SOSNOVCOVA
SPICHAL
Jiri
Hana
Vera
Ivana
Lukas
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
DENMARK
BOLDYRETT
ENGELSEN
HALKIER
ISSINGER
WEITZEL
Brigitte
SØren B.
Barbara A.
Olaf-Georg
Corinna
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
ENGLAND
RUSSELL
Paul J.
[email protected]
FINLAND
AURA
Anna-Marja
[email protected]
FRANCE
CHEYNIER
Veronique
[email protected]
GERMANY
BAUER
Anne Katrin
[email protected]; [email protected]
GREECE
FERLEMI
LAMARI
PAPANDREOU
Anastasia
Fotini N.
Magdalini A.
[email protected]
[email protected]
[email protected]
INDIA
MAITI
MITRA
Mrinal K.
Adinpunya
[email protected]
[email protected]
IRELAND
ALQUADH
Jafar
[email protected]
ITALY
APPENDINO
ARESTA
ARGENTIERI
AVATO
BISIO
CALVANO
CONFORTI
CORBO
DALESSANDRO
DURANTE
FERRI
Giovanni
Antonella
Maria Pia
Pinarosa
Angela
Cosima D.
Filomena
Filomena
Giuseppe
Miriana
Maura
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
GELBMANN
GIACOMELLI
JIRILLO
LANZOTTI
LENUCCI
LEONE
LUCCHETTI
MARELLI
MARI
MIELE
MONTORO
NAPOLITANO
PIACENTE
PISTELLI
ROSELLI
RUSSO
SCHIAVONE
SOSSAI
TASSONI
TAVA
TOMÈ
TRISOLINI
ZOTTI
Wolfgang
Emanuela
Emilio
Virginia
Marcello S.
Antonella
Vito
Mariangela
Angela
Mariangela
Paola
Assunta
Sonia
Luisa
Mariangela
Alessandra
Brigida
Dimitri
Annalisa
Aldo
Franca
Pietro
Margherita
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
LUXEMBOURG
GAASCHT
TEITEN
Francois
Marie-Hèlène
[email protected]
[email protected]
NIGERIA
AHMADU
Augustine
[email protected]
NORWAY
BRUSTAD
Irena
[email protected]
POLAND
KISIEL
KOWALSKA
OLESZEK
STOCHMAL
SZAJWAJ
SZAKIEL
THIEM
Wanda
Iwona
Wieslaw
Anna
Barbara
Anna
Barbara
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
PORTUGAL
BARROS
CARDOSO
DUARTE
FALCAO
FALÈ
GONCALVES
MATIAS
Lilian
Susana
Catarina
Soraia I.
Pedro Luis
Sandra
Ana
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
ROMANIA
TIULEA
Corina
[email protected]
SERBIA
GODEVAC
SRDANOV
Dejan
Jelica
[email protected]
[email protected]
SINGAPORE
TAN BENNY
Kwong Huat
[email protected]
SOUTH KOREA
KIM
KIM
Jung-In
Sang Min
[email protected]
[email protected]
SPAIN
BAI
CANDIRACCI
CASTANO
NAVARRO
de ANDRADE
FARRÈ
HERRERA
MIRALPEIX
MUZQUIZ
PARRADO
PEDROSA
RODRIGUEZ
MORGADO
SANAHUJA
SERRALHEIRO
TORRASCLAVEIRA
Chao
Manila
Angelica
[email protected]
[email protected]
[email protected]
Jean Paulo
Gemma
Gonzalez
M.D.
Bruna
Mercedes
Rubio Juan
Mercedes M.
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
mmartí[email protected]
Bruno
[email protected]
Georgina
Maria Luisa
Laura
[email protected]
[email protected]
[email protected]
SWITZERLAND
CHRISTEN
MATHON
WOLFENDER
Philippe
Caroline
Jean-Luc
[email protected]
[email protected]
[email protected]
THAILAND
ITHARAT
CHUNTHORNGORN
DECHAYONT
HANSAKUL
JAENGSUWAN
JIRAKIATTIKUL
KITTISRISOPIT
PHUAKLEE
PRAJUABJINDA
RATTAROM
RITHICAI
SAE-WONG
SRISOPA
UTTAMA
Arunporn
[email protected]
Jitpisute
[email protected]
Bhanuz
Pinturson
Nantana
Yaowapha
Suchada
Pathompong
Onmanee
Ruchilak
Panumart
Chutha
Ruangnoo
Sakuntala
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
Arnaud
[email protected]
THE NETHERLANDS
BOVY
General Information
Information and registration desk
The information and registration desk will be available at the Conference Venue.
Participants can pick up their personal conference materials at the registration desk, which will be
open as follows:
Tuesday-27 September 2011, 12.00-18.00
Wednesday-28 September 2011, 8.45-18.00
Thursday-29 September 2011, 8.45-18.00
Friday-30 September 2011, 8.45-12.00
Language
The conference language is English. There will be no simultaneous interpretation.
Name badges
Badges will be issued to all participants and accompanying persons and allow access to all
scientific sessions.
Projection
Please contact the local organizers to deliver your Powerpoint projections.
Posters
There will be two poster sessions. Posters with odd numbers (Poster Session 1) will be on
display from Tuesday-27 September to Wednesday-28 September. Posters with even numbers
(Poster session 2) will be on display from Thursday-29 September to Friday-30 September.
Presenting Authors with odd poster numbers are asked to be present to their posters during the
Poster session 1 on Wednesday, 18.30-20.00. Presenting Authors with even poster numbers are
asked to be present to their posters during the Poster session 2 on Thursday, 18.30-20.00.
Refreshments and meals
Coffee, tea and other refreshments are included in the registration fee for all registered
participants. Lunches and dinners are included in the hotel accomodation fee for participants
staying at Grand Hotel Riva del Sole. Registered participants who are not staying at Grand Hotel
Riva del Sole and wish to have lunch/dinner at the Congress may buy tickets at the Registration
desk.
Social Programme
Symposium Dinner will take place at Grand Hotel Riva del Sole on Thursday-29 September,
20:30 pm. Gala dinner is included in the registration fee for all registered participants. Please
confirm your participation at the Registration Desk.
Tuesday-27 September, 9:00 p.m.: “Italian Dragees”: After dinner by the “Confetteria & Museo
Mario Mucci”.
Wednesday-28 September, 9.00 p.m.: Visit to the medieval town of Bitonto
A post Symposium guided sightseeing tour to the UNESCO World Heritage town of Alberobello
will take place on Friday-30 September, 14:00 pm. A buffet with wine tasting will be offered free
of charge to all registered participants.
Certificate of attendance/Payment receipt
All registered participants will receive the Certificate of Attendance and the receipt for the
payment of the registration fee together with the conference materials.
Liability
The Congress organizers cannot be held responsible for any loss, theft, damage or injury to any
person or property during the Congress, whatever the cause may be. The liability of persons and
enterprises providing means of transportation or other services, remains unaffected. Each
Congress participant and accompanying person takes part in all the congress events at his/her own
risk.
Under the High Patronage and with the financial support of:
Regione Puglia
Federazione Ordini
Farmacisti Italiani
Ordine dei Farmacisti
della Provincia di Bari
Ordine dei Farmacisti
della Provincia di Brindisi
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