α-Amylase (α-AMY) and α-glucosidase (α-GLU) inhibitors are important for controlling postprandial hyperglycemia (PHG). Bixa orellana (annatto) reported inhibitory activity against these enzymes because of its bioactive compound content. However, an understanding of its inhibitory mechanisms and metabolic profile is necessary to establish its therapeutic potential. The present study aimed to elucidate the inhibitory mechanisms of B. orellana extract (BOE) on α-AMY and α-GLU, identify and quantify its bioactive compounds using metabolomics (untargeted and targeted) analyses, and evaluate their interactions through in silico approaches. BOE exhibited IC50 values of 37.75 and 47.06 mg mL-1 for α-AMY and α-GLU, respectively, indicating mixed and competitive inhibition types. Thirty-six putative compounds were identified by untargeted metabolomics, mainly fatty acids (dethiobiotin, occidentalol, palmitic acid, norbixin, among others). The most significant biosynthetic pathways included secondary metabolites (unclassified), unsaturated fatty acids, phenylpropanoids and flavonoid metabolism. Eighteen compounds were identified and quantified by the targeted analysis, such as l-phenylalanine, gallic acid, protocatechuic acid and naringenin. In silico studies highlighted xanthoangelol, norbixin, myricetin and 26-hydroxybrassinolide as key compounds with the highest binding affinities to enzyme active sites. BOE effectively inhibited α-AMY and α-GLU, with gallic acid, naringenin, xanthoangelol, norbixin and 26-hydroxybrassinolide identified as key bioactive contributors. These findings provide molecular evidence of the inhibitory mechanisms of BOE and support its potential for PHG management and diabetes control. © 2024 Society of Chemical Industry.
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