Inhibition of Metalloproteinase from Bitis arietans Venom In Vitro by the Ethyl Acetate Fraction of Moringa oleifera and Its Chemical Profile

Moringa oleifera Lam., Moringaceae, ethyl acetate soluble fraction has been documented to contain bioactive antivenom chemicals against Bitis arietans venom toxicities. Snake venom metalloproteinase is the principal enzyme of B. arietans venom exhibiting severe haemorrhage and haemostatic disturbances in envenomed victims. In this current study, B. arietans snake venom metalloproteinase was isolated using ion-exchange chromatography and gel filtration while SDS-PAGE was used to determine enzyme molecular weight. M. oleifera ethyl acetate soluble fraction was fractionated using column and thin-layer chromatography into subfractions and tested against B. arietans snake venom metalloproteinase. EchiTab polyvalent antivenom was used as the standard drug. The compounds present in the most active subfraction were identified using GCMS. The molecular weight of isolated enzyme was 43.28 kDa with specific activity of 3.06 μmol/min/mg and percentage yield of 18.5%. A total of 8 subfractions were obtained after fractionation and labelled ethyl acetate subfractions F1-8. All subfractions showed significant ( p < 0.05) inhibition against B. arietans snake venom metalloproteinase activity. However, subfraction F5 demonstrated the highest inhibition against B. arietans snake venom metalloproteinase activity with total inhibition at all concentrations compared to other subfractions and antivenom. Subfraction F5 displayed substantial inhibition against the haemorrhagic, haemolytic, and anticoagulant activities of B. arietans venom with a dose-dependent effect. The major bioactive compounds of subfraction F5 were γ-sitosterol (5.92%), kaempferol (5.9%), quercetin (5.28%), n -hexadecanoic acid (5.23%), and n -pentacosane (5.22%) of which some were previously reported as potential snake venom inhibitors. Results showed that subfraction F5 possesses potential antivenom compounds that could be explored for snakebite treatment. Graphical Abstract