Universidad Autónoma de Madrid Facultad de Ciencias Departamento de Química-Física Aplicada Biological Activity and Nutritional Properties of Processed Onion Products María Eduvigis Roldán Marín PhD Thesis Spanish National Research Council (CSIC) Instituto del Frío Department of Plant Food Science and Technology This European Doctorate is based on a literature review and five scientific publications A mis padres, A mi hermano. ‘Biological Activity and Nutritional Properties of Processed Onion Products’ PhD Thesis by María Eduvigis Roldán Marín Supervisors M. Pilar Cano Dolado Profesor de Investigación del CSIC Concepción Sánchez-Moreno González Científico Titular del CSIC Department of Plant Food Science and Technology. Instituto del Frío Spanish National Research Council (CSIC). Madrid. SPAIN I 2 TABLE OF CONTENTS Acompanying papers………………………………IV Preface………………………………........................V Summary………………………………………….VII Resumen…………………………………………...IX Abbrevations…………………………………….XIII Chapter 1 General introduction………………………………….1 Chapter 2 Research questions and hypotheses. Objectives…….27 Capítulo 2 Preguntas científicas e hipótesis. Objetivos……...…33 Chapter 3 Overview of the experimental work…………………39 Chapter 4 ‘Characterisation of onion (Allium cepa L.) by-products as food ingredients with antioxidant and antibrowning properties’ (Paper I)......................51 Chapter 5 ‘Onion high-pressure processing: Flavonol content and antioxidant activity’ (Paper II)......……..…....…53 Chapter 6 ‘Effects of an onion by-product on bioactivity and safety markers in healthy rats’ (Paper III)…..………55 Chapter 7 ‘An exploratory NMR nutri-metabonomic investigation reveals dimethyl sulfone as a dietary biomarker for onion intake’ (Paper IV)…..................57 Chapter 8 ‘Effects of an onion by-product on plasma lipids and platelet aggregation in healthy rats’ (Paper V)....59 Chapter 9 General discussion…………………………………..61 Chapter 10 Conclusions. Future prospects………………………85 Capítulo 10 Conclusiones………………………………………...91 References…………………………………………..95 Agradecimientos-Acknowledgements..……….....111 III Accompanying papers Paper I Roldán E, Sánchez-Moreno C, de Ancos B, Cano MP (2008) Characterisation of onion (Allium cepa L.) by-products as food ingredients with antioxidant and antibrowning properties. Food Chemistry, 108:907-916. Paper II Roldán-Marín E, Sánchez-Moreno C, Lloría R, de Ancos B, Cano MP (2009) Onion high-pressure processing: Flavonol content and antioxidant activity. LWT-Food Science and Technolology, 42:835-841. Paper III Roldán-Marín E, Krath BN, Poulsen M, Binderup M-L, Nielsen TH, Hansen M, Barri T, Langkilde S, Cano MP, Sánchez-Moreno C, Dragsted LO (2009) Effects of an onion by-product on bioactivity and safety markers in healthy rats. British Journal of Nutrition, in press (doi:10.1017/S0007114509990870) Paper IV Winning H, Roldán-Marín E, Dragsted LO, Viereck N, Poulsen M, Sánchez-Moreno C, Cano MP, Engelsen SB (2009) An exploratory NMR nutri-metabonomic investigation reveals dimethyl sulfone as a dietary biomarker for onion intake. Analyst, 134: 2344- 2351. Paper V Roldán-Marín E, Jensen RI, Krath BN, Poulsen M, Kristensen M, Cano MP, Sánchez- Moreno C, Dragsted LO (2009) Effects of an onion by-product on plasma lipids and platelet aggregation in healthy rats. Journal of Agricultural and Food Chemistry (under review) IV Preface This PhD Thesis has been conducted at the Department of Plant Food Science and Technology at Instituto del Frío, Spanish National Research Council (Consejo Superior de Investigaciones Científicas-CSIC), Madrid (Spain); the Department of Toxicology and Risk Assessment, National Food Institute, Technical University of Denmark (DTU), Søborg, Denmark; and the Department of Human Nutrition, Faculty of Life Sciences, University of Copenhagen, Frederiskberg, Denmark. During my PhD Thesis period I was granted by the Spanish Ministry of Science and Innovation with a Predoctoral Fellowship. My main workplace was Instituto del Frío- CSIC (Madrid) under the supervision of Dr. M. Pilar Cano and Dr. Concepción Sánchez-Moreno. I also spent one valuable year in Denmark in short ‘winter’ periods during three following years (2006-2008). During those periods I had the opportunity of working at two universities in Denmark with Professor Lars Ove Dragsted and his research team whom I am very grateful too, not only for their warmly welcome but also for their inexhaustible help, friendship and humility. There, I got the chance of learning everyday something new and discussing all the ‘onion’ results in a highly inspiring and nicely environment. These studies were supported by funding from the Spanish Ministry of Science and Innovation AGL2003-09138-C04-01; 200670I08; Program Consolider-Ingenio 2010, FUN-C-FOOD CSD2007-00063; and AGL2008-04798-C02-01/ALI, and from the Danish Ministry of Food, Agriculture and Fisheries (NuBI, 3304-FVFP-060696-01; and 3304-FVFP-060706-01). María Eduvigis Roldán Marín Madrid, 26 October 2009 V 6 Summary Nowadays, there is a constant and increasing social concern about the food we daily eat and particularly about vegetables. There is a great demand of minimally processed food or food ingredients with functional properties as a consequence of the current changing healthy and environmental sustainable living. Onion (Allium cepa L.) is an important vegetable traditionally used as a food ingredient in the Mediterranean diet that has a high production, domestic, and foreign trade worldwide. It is consumed raw, cooked or processed into different onion products in the daily diet of many subjects. This Allium vegetable has been proven to have interesting technological properties and beneficial health effects including antioxidant, anticarcinogenic, antimicrobial, prebiotic, hypolipidemic, and antithrombotic properties. Onion biological properties have been related with its bioactive compounds such as flavonols (quercetin and its glucosides), dietary fibre (fructans and fructooligosaccharides (FOS)), and organosulfur compounds (OSCs). The main aim of this PhD Thesis was to study the biological activity and nutritional properties of processed onion products in order to evaluate a possible development of novel and innovative functional onion ingredients that could substitute available synthetic ingredients according to the current consumers demand and global concern towards a healthier diet and living. To achieve this objective, in vitro and in vivo studies have been carried out with onion products of three Spanish onion cultivars: Allium cepa L. var. cepa, ‘Recas’, Allium cepa L. var. cepa, ‘Figueres’, and Allium cepa L. var cepa, ‘Grano de Oro’. The first part of the PhD Thesis aimed to evaluate in vitro effects of food processing and preservation technologies on onion nutritional and technological properties. The first in vitro study analyzed ‘Figueres’ and ‘Recas’ onion by-products (juices, pastes and bagasses) stabilized by sterilization, pasteurization, and freezing technologies. Results demonstrated that processing ‘Recas’ onion wastes to obtain onion pastes and the subsequent stabilization with pasteurization trigger to safe onion by-products with a good bioactive compound content and excellent antibrowning properties. In parallel, a second in vitro study showed that processing fresh ‘Grano de Oro’ onion with a nonthermal technology that combines high hydrostatic pressure with low temperature VII enhances flavonol (quercetin and its glucosides), total phenol extractabilities and maintains the antioxidant activity compared with the unprocessed onion. The second key topic of the investigation was the in vivo evaluation of the biological activity and nutritional properties of ‘Recas’ onion by-products. For that purpose, we studied the pasteurized ‘Recas’ onion paste chosen in the previous in vitro study, and its two derived onion fractions, an onion extract (rich in fructans and FOS) and an onion residue. The biological responses shown in healthy rats fed ‘Recas’ onion by-products revealed that these by-products are not genotoxic and that they exert interesting antioxidant and prebiotic properties. Moreover, they showed antithrombotic effects exerting a potential usefulness of these by-products in a cardiovascular disease (CVD) preventive diet. Onion OSCs and FOS seem to be the bioactive compounds responsible of some of the effects shown. Furthermore, the dimethyl sulfone was identified as a dietary biomarker for onion intake in a nutri-metabonomic study. Being able to detect and quantify specific onion intake biomarkers is highly beneficial in control of nutritionally enhanced functional foods and in human intervention studies. To go deep inside the biological activity evaluation of processed onion products, a human intervention study with overweighed subjects was also conducted using a ‘Recas’ onion product which was added to two four precooked dishes as a food ingredient. Currently, the main health effects are being evaluated. In the future, in vitro and in vivo studies would be required including different onion products processed by high hydrostatic pressure or other nonthermal technologies in order to study in depth the potential improvement of onion bioactive compounds bioaccessibility that could promote preventive or protective effects on CVD and obesity, two of the more prevalent diseases nowadays. In conclusion, the current PhD Thesis shows that pasteurizating onion pastes obtained from onion wastes and high-pressure processing fresh onion are two valuable processing food technologies