ABSTRACT
Diabetes mellitus is treated with oral glucose-lowering drugs or insulin injections. These treatments have adverse effects and are expensive. The antidiabetic, antioxidant, and anti-hyperlipidemic effects of Irvingia gabonensis stem bark extracts are well documented. This study elucidated the mechanisms of action of I. gabonensis stem bark extracts through in vitro and in silico studies; followed by the isolation and characterization of the active components responsible for the plant's antidiabetic, antipancreatic lipase, and antioxidant activities. Aqueous and ethanol extracts were obtained from I. gabonensis and separately subjected to quantitative analysis of phytochemicals, followed by in vitro antioxidants, antidiabetic, and anti-pancreatic lipase assays. The more effective extract (ethanol extract) was then fractionated and the active components were isolated using different chromatography techniques such as vacuum liquid chromatography, thin-layer chromatography, column chromatography, preparative thin-layer chromatography, high-performance liquid chromatography, and liquid chromatography-mass spectrometry. In addition, the molecular interactions of the active compounds with proteins implicated in diabetes pathogenesis, such as α-glucosidase, α-amylase, pancreatic lipase, and receptor for protein glycation, were studied through bioinformatics (in silico) techniques. IBM SPSS Statistics (version 26.0 Software) was used to analyze the data. The results showed that the aqueous and ethanol extracts had 1.78% and 2.89% yield respectively. The ethanol extract had higher total antioxidant power (488.78±47.31 mgGAE/g) compared to the aqueous extract (319.89±8.73 mgGAE/g). It was also observed that the ethanol extract had higher scavenging and inhibitory activities against diverse free radicals compared to the aqueous extract and a far higher phytochemical content. The in vitro antidiabetic and anti-pancreatic assays of the stem bark showed that the ethanol extract was more potent than the aqueous extract. In addition, the GCMS analysis of I. gabonensis stem bark extracts revealed the presence of hydrophobic compounds known for their potent biological activities. The HPLC analysis of the extracts revealed twelve (12) bioactive compounds including kaempferol, quercetin, ellagic acid, and gallic acid, which are known for their potent antioxidant activities. The molecular interactions of the HPLC identified compounds with target proteins showed that quercetin ranked highest in binding energy with αamylase (-6.6 kcal/mol), α-glucosidase (-6.6 kcal/mol), and pancreatic lipase (-5.6 kcal/mol). Bio-guided fractionation of the ethanol extract of I. gabonensis stem bark revealed the presence of 7 major active compounds and an unknown compound with molecular formula C13H10O6, m/z of 263.0547 and 261.0401. Finally, the use of Schrodinger Meastro software, in the in silico analysis, revealed that four (4) of the active compounds namely; ellagic acid tri-methyl ether, tetra-methyl ellagic acid, methyl gallate, and gallic acid, are potential inhibitors of α-amylase, α-glucosidase, pancreatic lipase, and protein glycation. This study has identified and characterized the most active constituents as well as established the mechanisms of action for the antidiabetic, anti-pancreatic lipase, and antioxidant activities of I. gabonensis stem bark. The findings from this study will assist in the production of cheaper, safer, and more effective therapeutic agents for combating diabetes and its long-term complications.
