Abstract:
Objective
To study the venom neutralization ability and prophylactic effect of Calotropis gigantea
methanolic root extract (CGMR) against Daboia russelii envenomation using in vitro, in
silico, and in vivo methods.
Background
Snake venom is composite, that contains neurotoxins, disintegrins, hemotoxins, and proteases
in complex with phospholipase A2 [PLA2] enzymes. Hence toxicity of venom is mainly
attributed to PLA2 enzymes or their protein complexes per se. Roots of the plant, Calotropis
gigantea are used extensively as a phyto-antidote by tribal communities to treat snake bite
victims. However the antivenom property of root has not been comprehensively studied so
far. This study scientifically reasserted the use of Calotropis Gigantea R.Br (root extract) in
neutralization of D.russelii venom, alongside investigated the protection rendered by the
extract.
Materials and Methods
Phytochemicals of Calotropis gigantea methanolic root (CGMR) extract were fractionated
into hexane (non-polar fraction) and methanol (polar fraction) based on polarity. The in vitro
PLA2 inhibitory action of crude extract and both fractions was determined using biochemical
assay. Since significant PLA2 inhibition was observed in non-polar fraction, it was subjected
to GC/MS analysis. The compounds obtained through GC/MS analysis were virtually docked
to PLA2 macromolecule by protein-ligand binding simulation programs. The acute and sub acute toxicity of the extract was determined using guidelines 423 and 407 respectively in
mice. The LD50 dose of Daboia russelii snake venom (DRSV) was determined using standard
protocol. The in vivo neutralization ability and the in vitro neutralization (pre-incubated) LD50 of venom was determined to be 11µg/dose and the % survival of group receiving only
venom was 50%. High dose of CGMR effectively neutralized LD50 dose of venom in both in
vivo (%survival = 67.67%) and in vitro neutralization experiment (% survival =100%).
Neutralization of venom was better in in vitro neutralization (pre-incubation). The
histopathological analysis of organs necropsied from CGMR pre-treated animals
demonstrated significant tolerance against venom.
Conclusion
Together these results indicate that CGMR has significant venom neutralizing potential as
traditionally claimed. Further the extract showed significant venom neutralization ability and
protection effects against venom insult. The high venom neutralization ability may be due to
PLA2 inhibitors present in the extract.
