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Minimally Processed Baby Leaf Vegetables: Phytonutrient Characterization and Nutritional Stability

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Minimally Processed Baby Leaf Vegetables: Phytonutrient Characterization and Nutritional Stability Minimally processed baby leaf vegetables: Phytonutrient characterization and Nutritional Stability Joana Manuela da Silva Santos Porto, 2014 PhD Thesis Minimally processed baby leaf vegetables: Phytonutrient characterization and Nutritional Stability Joana Manuela da Silva Santos Thesis submitted to Faculdade de Farmácia da Universidade do Porto for Doctor Degree in Pharmaceutical Sciences - Nutrition and Food Science Specialty Supervised by: Professora Doutora Maria Beatriz Prior Pinto Oliveira Porto January, 2014 © Authorised the partial reproduction of this thesis (subject to the approval of the publishers of journals in which the articles were published) only for research purposes through a written declaration of the person concerned that such pledges. iv This work has been supported by Fundação para a Ciência e a Tecnologia (FCT) through grant no. PEst-C/EQB/LA0006/2011 and to the PhD grant (SFRH/BD/ 66476/2009) financed by Programa Operacional Potencial Humano - Quadro de Referência Estratégico Nacional - Tipologia 4.1 - Formação Avançada (POPH-QREN) subsidised by Fundo Social Europeu (FSE) and national funds from Ministério da Ciência, Tecnologia e Ensino Superior (MCTES). v The work presented in this thesis was developed in the following research laboratories: REQUIMTE, Laboratory of Bromatology and Hydrology, Department of Chemical Sciences, Faculty of Pharmacy of University of Porto; FOODOMICS Laboratory at Institute of Food Science Research – CIAL (CSIC-UAM) at Madrid; REQUIMTE-GRAQ, Department of Chemical Engineering of Instituto Superior de Engenharia do Porto, Instituto Politécnico do Porto; and Agrarian School - Instituto Politécnico de Viana do Castelo, Ponte de Lima. vi Acknowledgements I would like to express my special acknowledgement: To my supervisor Professor Doctor Beatriz Oliveira for the opportunity to developed this project. Her guidance, constant encouragement, availability and enthusiasm throughout this PhD were essential to complete this work. To Fundação para a Ciência e Tecnologia (FCT) by the PhD grant (SFRH/BD/66476/2009) financed by Programa Operacional Potencial Humano - Quadro de Referência Estratégico Nacional - Tipologia 4.1 - Formação Avançada (POPH-QREN) subsidised by Fundo Social Europeu (FSE) and national funds from Ministério da Ciência, Tecnologia e Ensino Superior (MCTES), which made possible the accomplishment of the present work. To Professor Doctor Elena Ibáñez for accepting me in her laboratory and for the her sympathy, support and all the help during my traineeship in CIAL. Thank you for sharing your knowledge with me and for the opportunity to work with her fantastic research group. To Doctor Miguel Herrero for sharing his scientific knowledge with me and for the direct laboratory supervision, precise guidelines, scientific accuracy and friendship during my days at FOODOMICS laboratory. To Professor Doctor Maria Teresa Oliva-Teles for the availability, guidance and concern demonstrated during my traineeship in ISEP. To Professor Doctor Cristina Delarue-Matos for receiving me in her research group. Thanks for the warm welcoming and concern during my days in ISEP. The Iberian Salads S.A, in the name of his production director, for providing the vegetable samples used in this study. To Drª. Ana Paula Vale, director of ESA- Instituto Politécnico de Viana do Castelo for the help provided in the dietary fibre determinations and for receiving me in your school. To all collaborators in my PhD work, especially to Doctor José Mendiola and Margarida Almeida. To all the collaborators and especially to my friends at the Laboratory of Bromatology and Hydrology for the support, smiles, advices and presence during this journey. vii To all collaborators at GRAQ- ISEP and at ESA-IPVC for the warm welcoming during my days in your laboratory. To all the friends at CIAL-Madrid for their sympathy and fellowship (especially to Clara, Maria and Alberto). It was a pleasure meeting and to working with all of you. To my parents and to my sisters (Su e Lita) for the constant support, encouraging words and for always believing that I could do this project. To Paulo for your incentive, support and patience during my intense laboratory and writing journeys during this period. viii Abstract The consumption of fresh vegetables is globally recommended by worldwide health organizations as an effective mean to preserve and improve population’s health status. The high concentration in certain bioactive phytochemicals normally present in vegetables is the reason for those recommendations, being the high intake of vegetables correlated to lower incidence of chronic diseases. However, in our modern society, the busy lifestyle of today’s consumers does not allow to dispend a lot of time to prepare meals, being their decisions greatly influenced by the perceived convenience of a food product. Minimally processed vegetables become a convenient answer to those who want to include more fresh vegetables in the diet. The evolution in this market has been drove by the consumer demand for high quality fresh products that could deliver, in a convenient way, all the flavour and nutritional benefits associated to fresh vegetables. The minimally processed baby leaf products are one of the latest innovations in this sector, and permitted to add more variety, colours, formats and textures to the range of minimally processed vegetables previously available. These products are constituted by more immature leaves, being usually packaged whole, which grant to the product a different and appealing appearance. As leafy vegetables, they are seen as a healthy source of phytochemicals and showed great acceptance within consumers. However, it has been described that the nutritional quality of these products can be significantly influenced by different factors, like maturity stage and processing conditions. The main objective of this work was the evaluation of the nutritional quality of minimally processed baby leaf vegetables during shelf life, namely, vitamin, mineral, phenolic and carotenoid contents, that are the bioactive compounds normally associated to the health benefits provided by fresh vegetables. To achieve that, different extraction methods were used to extract the very different chemical structures from vegetal matrices. Several analytical methodologies were developed and/or applied, namely chromatographic methods coupled to diode array and mass detectors (vitamins, carotenoids and phenolics determination) and an atomic absorption spectrometry based method (mineral analysis), also coupled to a high resolution UV-Vis detector. The results obtained showed that most of the main phytochemicals that contribute to the nutritional quality of baby leaf samples can suffer significant changes during shelf life. Among the micronutrients evaluated, the greatest changes were found among the vitamins, especially the water-soluble ones. However, the behaviour of each vitamin was strongly dependent on the sample species. Baby leaf vegetables were rich in vitamin C, provitamin A and vitamin E, being the new baby leaf, pea shoots, the richest sample on ix vitamin C and provitamin A. Regarding the samples mineral content, the baby leaf studied revealed to be a good source of potassium and calcium, being their mineral content stable during storage. Organic wild rocket baby leaves were the sample with the highest mineral content. Among the different phytonutrients, the majority of the baby leaves studied showed a higher content of glycosylated flavonoid compounds. However, the phenolic profile of each sample was also greatly dependent on the species. Ruby red lettuce was the richest sample in phenolic compounds. Regarding carotenoid content, β-carotene and lutein, typical from green leafy vegetables, were also found in high amounts. Finally, from all the phytochemicals evaluated, it could be referred that the maturity stage of this young leaves did not represent a nutritional disadvantage, when comparing to the contents published for more mature leaves. Keywords: baby leaf; micronutrients; phytonutrients; nutritional quality; HPLC-DAD-MS; HR-CS-AAS. x Resumo O consumo de vegetais frescos é recomendado pelas organizações mundiais de saúde como um meio eficaz para preservar e melhorar a saúde pública. O elevado teor de fitoquímicos bioativos, normalmente presentes em vegetais, é a razão para essa recomendação, uma vez que um consumo elevado de vegetais tem sido associado a uma menor incidência de doenças crónicas. Contudo, o estilo de vida na sociedade atual não se coaduna com uma preparação demorada de refeições saudáveis e que incluam vegetais frescos em abundância. Atualmente, o consumidor baseia a sua decisão na conveniência que determinado produto alimentar pode oferecer. Os vegetais minimamente processados surgiram como uma opção conveniente para quem procura incluir mais vegetais frescos na dieta. A evolução neste mercado tem sido impulsionada pela maior procura de produtos frescos de alta qualidade e que, de uma forma conveniente, consigam preservar o sabor e os benefícios nutricionais, normalmente associados aos vegetais frescos. As ”baby leaf” são uma das mais recentes inovações do setor dos produtos minimamente processados. Estas surgiram como uma forma de acrescentar variedade, cor, formatos e texturas à gama de vegetais minimamente processados já disponíveis no mercado. Estes produtos são constituídos por folhas num estado de maturação precoce, sendo geralmente embaladas inteiras, conferindo aos produtos uma aparência diferente,
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