Estudo da hidrólise das proteínas de duas fontes vegetais não convencionais: Moringa oleífera Lam e Pereskia aculeata Miller com proteases obtidas de bioprodutos do figo e o abacaxi: análise do efeito citotóxico e antiproliferativo in vitro em células de adenocarcinoma do colon (SW420)
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Data
2022-03-30
Autores
Arismendi Bustamante, Lady Johana
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Universidade Estadual Paulista (Unesp)
Resumo
Objetivo: estudar a hidrólise das proteínas da farinha das folhas de Moringa oleífera Lam (MOF) e Pereskia aculeata Miller (PAF) com enzimas obtidas dos bioprodutos de figo e/ou acabaxi e o efeito antiproliferativo e citotóxico dos produtos de hidrólise nas células do adenocarcinoma de colon. Métodos: tanto a farinha íntegra das folhas quanto as frações solúveis e insolúveis na água foram hidrolisadas com extratos enzimáticos obtidos de bioprodutos do figo e/ou abacaxi. Os produtos de hidrolise foram avaliados por métodos colorimétricos, eletroforéticos e cromatográficos. A capacidade citotóxica e antiproliferativa dos produtos de hidrólise da MOF com o extrato do figo foi avaliada in vitro nas células de adenocarcinoma SW480 pôr o método colorimétrico com sulforodamina B e por citometria de fluxo e detecção de fosfatidilserina, respetivamente. Resultados: a hidrólise da fração insolúvel e solúvel da MOF com os bioprodutos de figo e abacaxi gerou mudanças na solubilidade e nos perfis eletroforéticos e cromatográficos das proteínas. Se ressalta que a fração insolúvel hidrolisada com o extrato de figo apresentou um incremento significativo de proteína solúvel, resultado que foi detectável pelos diferentes métodos aplicados. Por outra parte, a hidrólise da farinha íntegra (sem separação), permitiu verificar que existe um efeito hidrolítico da enzima ficina sob as proteínas de maior massa molecular (> 97 kDa) da MOF e sob diferentes proteínas da PAF. Dos produtos de hidrólise, a fração solúvel da MOF hidrolisada com ficina por 8h permitiu a obtenção de compostos proteicos com atividade citotóxica, antiproliferativa e pró-apoptótica tempo-dependente no modelo in vitro na linhagem de células SW480. Conclusão: o estudo evidencia que os extratos enzimáticos obtidos de bioprodutos do figo e abacaxi hidrolisam as proteínas da MOF e/ou PAF, e que os produtos de hidrólise mudam segundo a fração da farinha hidrolisada (solúvel, insolúvel ou farinha íntegra). O efeito citotóxico e antiproliferativo da fração solúvel de MOF hidrolisada com figo foi comprovado in vitro nas células de adenocarcinoma de cólon SW480, outros estudos devem ser realizados para as outras frações da MOF e PAF.
Objective: to study the hydrolysis of proteins from the flour of Moringa oleífera Lam (MOF) and Pereskia aculeata Miller (PAF) leaves with enzymes obtained from fig and/or pineapple by-products, and the antiproliferative and cytotoxic effect of the hydrolysis products on adenocarcinoma cells of colon. Methods: the whole flour of the leaves and the soluble and insoluble fractions in water were hydrolyzed with enzymatic extracts obtained from fig and/or pineapple byproducts. The hydrolysis products were evaluated by colorimetric, electrophoretic, and chromatographic methods. The cytotoxic and antiproliferative capacity of the hydrolysis products of MOF with fig extract was evaluated in vitro in SW480 adenocarcinoma cells by the colorimetric method with sulphorodamine B and by flow cytometry and detection of phosphatidylserine, respectively. Results: the hydrolysis of the insoluble and soluble fraction of MOF with the fig and pineapple by-products generated changes in the solubility and in the electrophoretic and chromatographic profiles of the proteins. It is noteworthy that the insoluble fraction hydrolyzed with the fig extract showed a significant increase in soluble protein, a result that was detectable by the different methods applied. On the other hand, the hydrolysis of whole flour (without separation) allowed to verify that there is a hydrolytic effect of the ficin enzyme on the higher molecular mass proteins (> 97 KDa) of MOF and on different proteins of the PAF. From the hydrolysis products, the soluble fraction of MOF hydrolyzed with ficin for 8h allowed the obtaining of protein compounds with time-dependent cytotoxic, antiproliferative and pro-apoptotic activity in the in vitro model in the SW480 cell line. Conclusion: the study shows that the enzymatic extracts obtained from fig and pineapple by-products hydrolyze the MOF and/or PAF proteins, and that the hydrolysis products change according to the fraction hydrolyzed (soluble, insoluble, or whole flour). The cytotoxic and antiproliferative effect of the soluble fraction of MOF hydrolyzed with fig was proven in vitro in SW480 colon adenocarcinoma cells, further studies should be performed for the other fractions of MOF and PAF.
Objective: to study the hydrolysis of proteins from the flour of Moringa oleífera Lam (MOF) and Pereskia aculeata Miller (PAF) leaves with enzymes obtained from fig and/or pineapple by-products, and the antiproliferative and cytotoxic effect of the hydrolysis products on adenocarcinoma cells of colon. Methods: the whole flour of the leaves and the soluble and insoluble fractions in water were hydrolyzed with enzymatic extracts obtained from fig and/or pineapple byproducts. The hydrolysis products were evaluated by colorimetric, electrophoretic, and chromatographic methods. The cytotoxic and antiproliferative capacity of the hydrolysis products of MOF with fig extract was evaluated in vitro in SW480 adenocarcinoma cells by the colorimetric method with sulphorodamine B and by flow cytometry and detection of phosphatidylserine, respectively. Results: the hydrolysis of the insoluble and soluble fraction of MOF with the fig and pineapple by-products generated changes in the solubility and in the electrophoretic and chromatographic profiles of the proteins. It is noteworthy that the insoluble fraction hydrolyzed with the fig extract showed a significant increase in soluble protein, a result that was detectable by the different methods applied. On the other hand, the hydrolysis of whole flour (without separation) allowed to verify that there is a hydrolytic effect of the ficin enzyme on the higher molecular mass proteins (> 97 KDa) of MOF and on different proteins of the PAF. From the hydrolysis products, the soluble fraction of MOF hydrolyzed with ficin for 8h allowed the obtaining of protein compounds with time-dependent cytotoxic, antiproliferative and pro-apoptotic activity in the in vitro model in the SW480 cell line. Conclusion: the study shows that the enzymatic extracts obtained from fig and pineapple by-products hydrolyze the MOF and/or PAF proteins, and that the hydrolysis products change according to the fraction hydrolyzed (soluble, insoluble, or whole flour). The cytotoxic and antiproliferative effect of the soluble fraction of MOF hydrolyzed with fig was proven in vitro in SW480 colon adenocarcinoma cells, further studies should be performed for the other fractions of MOF and PAF.
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Moringa oleífera Lam, Pereskia aculeata Miller, bioprodutos, proteases, proteínas, citotoxicidade