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Master's Dissertation
DOI
10.11606/D.60.2016.tde-03102016-143050
Document
Author
Full name
Camila Capel Godinho
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2016
Supervisor
Committee
Gobbo Neto, Leonardo (President)
Cavalheiro, Alberto Jose
Souza, Gloria Emilia Petto de
Title in Portuguese
Análise metabolômica da bioatividade em vias COX e LOX-dependentes de plantas da subtribo Lychnophorinae
Keywords in Portuguese
Asteraceae
Cicloxigenase
espectrometria de massas
LC-MS
Lipoxigenase
Lychnophorinae
Metabolômica
Abstract in Portuguese
Muitas substâncias das espécies de Lychnophorinae (Asteraceae) são relatadas como inibidoras da síntese de mediadores da cascata do processo inflamatório. Nesse processo, duas enzimas são essenciais e atuam no metabolismo do ácido araquidônico, formado em processos inflamatórios: ciclooxigenase (COX) e lipoxigenase (LOX). A análise de impressões digitais metabólicas é um método capaz de fornecer informações sobre o objeto de estudo através da utilização de ferramentas estatísticas, além de possibilitar a correlação desses dados com outros utilizando métodos in silico. O objetivo deste estudo foi analisar 26 espécies da subtribo Lychnophorinae quanto à atividade inibitória in vitro das vias COX- e LOX-dependentes e identificar as substâncias responsáveis por essa atividade através de análises in silico de correlação entre bioatividade e impressão digital metabólica. As impressões digitais metabólicas dos extratos hidrometanólicos das espécies de Lychnophorinae foram obtidas utilizando UHPLC-UV-(DAD)-MS (OrbitrapTM). Os ensaios de triagem de inibição das vias COX-1 e 5-LOX-dependentes revelaram que 20 espécies possuem atividade inibitória dupla para ambas as vias com valores de IC50 menores que 100 ?g.mL-1. Dentre essas, 11 espécies apresentaram valores de IC50 menores que 40 ?g.mL-1, e cinco apresentaram valores de IC50 menores que 10 ?g.mL-1. Utilizando-se as impressões digitais metabólicas e os resultados de inibição enzimática foram realizadas análises estatísticas multivariadas supervisionadas e não supervisionadas (PCA, PLS e OPLS) visando à identificação de substâncias discriminantes (ativas). Através das análises de correlação, obtiveram-se as prováveis substâncias que detém o potencial inibitório. As cinco substâncias com maior potencial discriminante foram identificadas por técnicas de desreplicação e são: a lactona sesquiterpênica (4,5-diidro-15-desoxigoyazensolido), dois flavonóides glicosilados (3-O-(acetil-hexosídeo)-quercetina e 7-O-(cumaroil-hexosídeo)-apigenina), e um hidroxinerolidol. Assim, este estudo revelou o ótimo potencial inibidor das enzimas COX-1 e 5-LOX dos extratos hidrometanólicos das espécies de Lychnophorinae. Além disso, indicou as substâncias mais discriminantes responsáveis por essa atividade, sendo as substâncias acima mencionadas as propostas como as principais responsáveis pela atividade inibidora das enzimas COX e LOX
Title in English
Metabolomic analysis of the bioactivity in COX and LOX pathways-dependent of plants from subtribe Lychnophorinae
Keywords in English
Asteraceae
Cyclooxygenase
LC-MS
Lipoxygenase
Lychnophorinae
mass spectrometry
Metabolomics
Abstract in English
Several compounds from Lychnophorinae species (Asteraceae) are reported as inhibitors of cascade mediators that elicits the inflammatory process. During this process, two enzymes are essential in the metabolism of the arachidonic acid: cyclooxygenase (COX) and lipoxygenase (LOX). The analysis of metabolic fingerprinting is a method that provides information about the object of study by using statistical tools, and enables the correlation of these data with others using in silico methods. This work therefore aimed to analyze 26 species of the Lychnophorinae subtribe for in vitro inhibition of COX-1 and 5-LOX and (to) identify the bioactive compounds responsible for this activity by in silico correlation analysis between bioactivity and metabolic fingerprint. The metabolomic analysis was carried out using UHPLC-DAD-ESI-Orbitrap and a metabolic fingerprint for each extract was obtained. The in vitro inhibition screening assays of COX-1 and 5-LOX revealed that 20 extracts presented dual inhibitory activity on both enzymes with IC50 values lower than 100 ?g.mL-1. Among them, 11 species showed IC50 values lower than 40 ?g.mL-1 and five lower than 10 ?g.mL-1. In order to identify discriminant substances (active), supervised and non-supervised multivariate statistical analysis (PCA, PLS e OPLS) were performed using the metabolic fingerprints and the results of enzyme inhibition. Through correlation analysis, it was possible to locate the substances most likely to be responsible for the pharmacological activity in both enzymes simultaneously; among them, five were chosen as the most likely. The substances were identified by dereplication as: a sesquiterpene lactone (4,5-dihydro-15-desoxygoyazensolide), two flavonoids (3-O-(acetil-hexoside)-quercetin and 7-O-(cumaroil-hexoside)-apigenine), and a hidroxynerolidol. In summary, in this work it was possible to reveal crude extracts with outstanding inhibitory potential of both, COX-1 and 5-LOX, enzymes as well as to propose the most probable compounds responsible for this action, and the compounds mentioned above were proposed as the main responsible for the inhibitory activity.
 
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Release Date
2018-10-03
Publishing Date
2016-12-02
 
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