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Master's Dissertation
DOI
10.11606/D.6.2019.tde-10012019-171854
Document
Author
Full name
Ana Clara da Costa Pinaffi
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2018
Supervisor
Committee
Torres, Elizabeth Aparecida Ferraz da Silva (President)
Camargo, Adriano Costa de
Hassimotto, Neuza Mariko Aymoto
Purgatto, Eduardo
Title in Portuguese
Polifenóis não-extraíveis provenientes do guaraná (Paullinia cupana): caracterização por MALDi-TOF/TOF e avaliação do potencial e cinética de inibição da alfa-glicosidase
Keywords in Portuguese
α-glicosidase
MALDi
Paullinia cupana
Polifenóis Não-Extraíveis
Abstract in Portuguese
Introdução: Polifenóis não-extraíveis (NEPPs) são uma fração de polifenóis que não são extraídos da forma convencional por estarem associados à parede celular de produtos de origem vegetal. Um corpo crescente de estudos tem evidenciado seus potenciais efeitos benéficos, especialmente associados à saúde intestinal e interações com a microbiota. O guaraná (Paullinia cupana), fruto típico da biota amazônica, é conhecidamente rico em polifenóis da família dos flavanóis, mas ainda existe uma lacuna a respeito da fração de polifenóis não-extraíveis em sua composição. Objetivo: Caracterizar a fração de polifenóis não-extraíveis quanto a sua composição química, e avaliar sua potencial capacidade de inibição enzimática. Métodos: O guaraná em pó foi submetido a extração aquo-orgânica para obtenção da fração extraível, e o resíduo proveniente dessa extração foi submetido a hidrólise ácida e hidrólise básica para obtenção dos NEPPs. A capacidade redutora total (CRT) foi quantificada pelo método de Folin-Ciocalteu. A quantificação de taninos condensados foi realizada pelo método de Porter. A determinação do perfil de fenólicos foi realizada por HPLC-ECD e LC-MS para as frações extraíveis e hidrolisáveis, e MALDi-TOF/TOF para a fração condensada. Os testes enzimáticos foram realizados com base na cinética de estado estacionário. Os testes estatísticos foram realizados utilizando softwares Excel e SPSS. Resultados: O perfil de fenólicos para a fração extraível consiste na presença de catequina e epicatequina como componentes majoritários, com 5,45 ± 0,15 e 5,95 ± 0,22 mg/g de guaraná em pó (base seca), respectivamente, além de proantocianidinas B1 e B2 e trímero de tipo A. Já o perfil fenólico da fração não-extraível contém uma mistura complexa de monômeros como catequina, leucoantocianidina, cianidina e delfinidina. A fração NEPP também contém dímeros, trímeros, tetrâmeros e pentâmeros de flavanóis, tanto de tipo A quanto de tipo B, com alta variabilidade de grau de hidroxilação. O ensaio enzimático com α-glicosidase resultou em valores de IC50 de 9,504 e 1,624 µg EAG/mL para a fração extraível e a não-extraível, respectivamente. O modo de inibição para ambas as frações foi classificado como misto, com valores de Ki e K'i de 0,403 e 1,735 µg/mL para a fração extraível e 0,287 e 0,847 µg/mL para a fração não-extraível. Conclusões: A fração de polifenóis não-extraíveis possui composição variada e complexa quando comparada a fração extraível, e possui potencial de inibição de α-glicosidase que deve ser explorado de maneira mais aprofundada, uma vez que tal potencial é de interesse para o controle de doenças crônicas como o diabetes tipo 2.
Title in English
Non-extractable polyphenols from guarana (Paullinia cupana): MALDi-TOF/TOF characterization and evaluation of potential and kinetics of alpha-glucosidase inhibition
Keywords in English
α-glucosidase
MALDi
Non-extractable Polyphenols
Paullinia cupana
Abstract in English
Introduction: Non-extractable polyphenols (NEPPs) are a portion of polyphenols that cannot be extracted in the conventional way due to being associated with the cell wall of products of plant origin. A growing number of studies have been showing its potential beneficial effects, especially in relation to gut health and microbiota interactions. The guarana (Paullinia cupana), a fruit native of the Amazon rainforest, is known to be rich in polyphenols from the flavanol family, but there is still a gap about non-extractable polyphenols in its composition. Objective: Characterize the non-extractable polyphenol portion in relation to its chemical composition and evaluate its enzymatic inhibition capacity. Methods: The extractable fraction was obtained by aqueous-organic extraction, and the residue from this extraction was treated with acid and alkaline hydrolysis to obtain the NEPPs. The total reducing capacity (TRC) was quantified by the Folin-Ciocalteu method. The quantification of condensed tannins was performed with the Porter method. The phenolic profile was determined by HPLC-ECD and LC-MS for the extractable and hydrolysable fractions, and MALDi-TOF/TOF for the condensed fraction. The enzymatic assay was carried out using steady-state kinetics. The statistical tests were performed using Excel and SPSS. Results: The phenolic profile of the extractable fraction consists of catechin and epicatechin as major components with 5,45 ± 0,15 and 5,95 ± 0,22 mg/g guarana powder (dry weight), respectively, besides B1 and B2 proanthocyanidins and type A trimer. The phenolic profile of the non-extractable fraction contains a complex mixture of monomers like catechin, leucoanthocyanidin, cyanidin, and delphinidin. The NEPP fraction also contains type A and type B dimers, trimers, tetramers, and pentamers of flavanols, with high variability of the degree of hydroxylation. The α-glucosidase enzymatic assay had IC50 values of 9,504 and 1,624 Introduction: Non-extractable polyphenols (NEPPs) are a portion of polyphenols that cannot be extracted in the conventional way due to being associated with the cell wall of products of plant origin. A growing number of studies have been showing its potential beneficial effects, especially in relation to gut health and microbiota interactions. The guarana (Paullinia cupana), a fruit native of the Amazon rainforest, is known to be rich in polyphenols from the flavanol family, but there is still a gap about non-extractable polyphenols in its composition. Objective: Characterize the non-extractable polyphenol portion in relation to its chemical composition and evaluate its enzymatic inhibition capacity. Methods: The extractable fraction was obtained by aqueous-organic extraction, and the residue from this extraction was treated with acid and alkaline hydrolysis to obtain the NEPPs. The total reducing capacity (TRC) was quantified by the Folin-Ciocalteu method. The quantification of condensed tannins was performed with the Porter method. The phenolic profile was determined by HPLC-ECD and LC-MS for the extractable and hydrolysable fractions, and MALDi-TOF/TOF for the condensed fraction. The enzymatic assay was carried out using steady-state kinetics. The statistical tests were performed using Excel and SPSS. Results: The phenolic profile of the extractable fraction consists of catechin and epicatechin as major components with 5,45 ± 0,15 and 5,95 ± 0,22 mg/g guarana powder (dry weight), respectively, besides B1 and B2 proanthocyanidins and type A trimer. The phenolic profile of the non-extractable fraction contains a complex mixture of monomers like catechin, leucoanthocyanidin, cyanidin, and delphinidin. The NEPP fraction also contains type A and type B dimers, trimers, tetramers, and pentamers of flavanols, with high variability of the degree of hydroxylation. The α-glucosidase enzymatic assay had IC50 values of 9,504 and 1,624 Introduction: Non-extractable polyphenols (NEPPs) are a portion of polyphenols that cannot be extracted in the conventional way due to being associated with the cell wall of products of plant origin. A growing number of studies have been showing its potential beneficial effects, especially in relation to gut health and microbiota interactions. The guarana (Paullinia cupana), a fruit native of the Amazon rainforest, is known to be rich in polyphenols from the flavanol family, but there is still a gap about non-extractable polyphenols in its composition. Objective: Characterize the non-extractable polyphenol portion in relation to its chemical composition and evaluate its enzymatic inhibition capacity. Methods: The extractable fraction was obtained by aqueous-organic extraction, and the residue from this extraction was treated with acid and alkaline hydrolysis to obtain the NEPPs. The total reducing capacity (TRC) was quantified by the Folin-Ciocalteu method. The quantification of condensed tannins was performed with the Porter method. The phenolic profile was determined by HPLC-ECD and LC-MS for the extractable and hydrolysable fractions, and MALDi-TOF/TOF for the condensed fraction. The enzymatic assay was carried out using steady-state kinetics. The statistical tests were performed using Excel and SPSS. Results: The phenolic profile of the extractable fraction consists of catechin and epicatechin as major components with 5,45 ± 0,15 and 5,95 ± 0,22 mg/g guarana powder (dry weight), respectively, besides B1 and B2 proanthocyanidins and type A trimer. The phenolic profile of the non-extractable fraction contains a complex mixture of monomers like catechin, leucoanthocyanidin, cyanidin, and delphinidin. The NEPP fraction also contains type A and type B dimers, trimers, tetramers, and pentamers of flavanols, with high variability of the degree of hydroxylation. The α-glucosidase enzymatic assay had IC50 values of 9,504 and 1,624 µg GAE/mL for the extractable and non-extractable fraction, respectively. The mode of inhibition was classified as mixed for both fractions, with Ki and K'i values of 0,403 and 1,735 µg/mL for the extractable fraction and 0,287 and 0,847 µg/mL for the non-extractable fraction. Conclusions: The non-extractable polyphenols fraction has a varied and complex composition when compared to the extractable fraction, and it has a α-glucosidase inhibition potential that must be explored in a more detailed fashion since said potential is of interest for the control of chronic diseases such as type 2 diabetes. g GAE/mL for the extractable and non-extractable fraction, respectively. The mode of inhibition was classified as mixed for both fractions, with Ki and K'i values of 0,403 and 1,735 µg/mL for the extractable fraction and 0,287 and 0,847 µg/mL for the non-extractable fraction. Conclusions: The non-extractable polyphenols fraction has a varied and complex composition when compared to the extractable fraction, and it has a α-glucosidase inhibition potential that must be explored in a more detailed fashion since said potential is of interest for the control of chronic diseases such as type 2 diabetes. g GAE/mL for the extractable and non-extractable fraction, respectively. The mode of inhibition was classified as mixed for both fractions, with Ki and K'i values of 0,403 and 1,735 µg/mL for the extractable fraction and 0,287 and 0,847 µg/mL for the non-extractable fraction. Conclusions: The non-extractable polyphenols fraction has a varied and complex composition when compared to the extractable fraction, and it has a α-glucosidase inhibition potential that must be explored in a more detailed fashion since said potential is of interest for the control of chronic diseases such as type 2 diabetes.
 
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Publishing Date
2019-01-11
 
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