Isolation and characterization of phenolic compounds from Brazilian green propolis with antiparasitic (Trypanosoma cruzi) activity

Submitted by Leticia Schettini (leticia@ufrrj.br) on 2021-10-06T20:45:05Z No. of bitstreams: 1 2019 - Lucas de Oliveira Pires.pdf: 3208222 bytes, checksum: aa33d01b8f364150cad13dc9a968be0d (MD5) Made available in DSpace on 2021-10-06T20:45:05Z (GMT). No. of bitstreams: 1 2019 - Lucas de Oliveira Pires.pdf: 3208222 bytes, checksum: aa33d01b8f364150cad13dc9a968be0d (MD5) Previous issue date: 2019-03-27 CAPES - Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior CNPq - Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico FAPERJ - Funda??o Carlos Chagas Filho de Amparo ? Pesquisa do Estado do Rio de Janeiro Propolis is a resinous natural product elaborated by honeybees from different parts of plants, such as shoots and flowers. Its chemical composition is very complex and depends on factors such as climate, seasonality, soil, vegetation and also the bee species. Propolis has currently been identified as na importante source of bioactive compounds due to its high biological potential, with anti-inflammatory, antibacterial, antitumoral, antiparasitic and antioxidant activities. In this work, ethanolic and n-hexanic extracts were prepared from three samples of green propolis obtained in different regions of Rio de Janeiro. The quality of these extracts was evaluated using colorimetric methods for the determination of the total phenolic content ? total phenolic and flavonoid contents ? and also the antioxidant capacity was evaluated by capturing organic free radicals (DPPH and ABTS), as well as the ferric reducing antioxidant power (FRAP). The qualitative chemical profile of the extracts was evalutated through HPLCPDA analisys, and the influence of the solvente polarity on the extraction profile was studied. The bioactive compounds 3,5-diprenyl-4-hydroxycinamic acid (Artepillin C), 3-prenyl-4-[(3- phenylpropanoil)-oxy]-cinnamic acid (Baccharin) and 2,2-dimethyl-8-prenylcroman-5- propenoic acid were isolated from the n-hexane exrtract by two chromatographic steps: fracionation of crude extract using the mixture of solventes chloroform/acetone in increasing gradiente concentration, followed by purification of fractions of interest by HPLC-UV in semipreparative scale, using the mixture methanol/water in increasing gradiente concentration. All the isolated constituents were characterized by NMR (1H and 13C), UV spectra and CG-MS techniques, and the contente of these componentes in the crude extract was quantified through HPLC-PDA in reverse phase. The antiparasitic activity of green propolis and its active principles was evaluated against tripomastigoe forms of Trypanosoma cruzi, the parasite responsible for causing Chagas? disease. A pr?polis ? um produto natural resinoso elaborado pelas abelhas mel?feras a partir de diferentes partes das plantas, como brotos e flores. Sua composi??o qu?mica ? muito complexa e depende de fatores como clima, sazonalidade, solo, vegeta??o e tamb?m da esp?cie da abelha produtora. A pr?polis atualmente tem sido apontada como uma importante fonte de subst?ncias bioativas por conta do seu elevado potencial biol?gico, com atividade anti-inflamat?ria, antibacteriana, antiviral, antitumoral, antiparasit?ria e antioxidante. Neste trabalho foram preparados extratos etan?licos e hex?nicos a partir de tr?s amostras de pr?polis verde, obtidas em diferentes regi?es do Estado do Rio de Janeiro. A qualidade desses extratos foi avaliada a partir de m?todos colorim?tricos para a determina??o do teor de subst?ncias fen?licas ? teor de fen?licos totais e flavonoides totais ? e da capacidade antioxidante atrav?s da captura dos radicais livres 2,2-difenil-1-picrilhidrazila (DPPH) e ?cido 2,2?-azino-bis(3- etilbenzotiazolina-6-sulf?nico) (ABTS+.), al?m da capacidade redutora do ?on f?rrico (FRAP ? ferric reducing antioxidant power). O perfil qu?mico dos extratos foi avaliado atrav?s de cromatografia l?quida de alta efici?ncia com detector de arranjo de diodo (CLAE-DAD) com fase reversa (C-18). Os componentes bioativos ?cido 3,5-diprenil-4-hidroxicin?mico (Artepillin C), ?cido 3-prenil-4-[(3-fenilpropanoil)oxi]-cin?mico (bacarina) e ?cido 2,2- dimetil-8-prenilcromano-5-propenoico foram isolados a partir do extrato hex?nico de pr?polis verde atrav?s de duas etapas cromatogr?ficas: fracionamento em coluna flash do extrato bruto utilizando clorof?rmio: acetona, com gradiente crescente de polaridade, seguido da purifica??o das fra??es de interesse por CLAE-UV semipreparativa, tendo como solvente a mistura bin?ria metanol: ?gua em gradiente crescente de concentra??o. Todos os constituintes isolados foram caracterizados por t?cnicas de resson?ncia magn?tica nuclear (RMN 1H e 13C), espectrometria na regi?o do ultravioleta (UV) e cromatografia gasosa acoplada a espectrometria de massas (CG-EM), e o teor desses componentes dos extratos brutos foi quantificado por CLAE-DAD. A atividade antiparasit?ria da pr?polis verde e de seus princ?pios ativos foi avaliada frente ?s formas infectivas (tripomastigotas) de Trypanasoma cruzi, parasito respons?vel por causar a doen?a de Chagas.