Synergistic effect of bioactive compounds of polyherbal fractions of Clinacanthus Nutans and Elephantopus Scaber in the treatment of wound

Modern healthcare system recognizes herbal medicine as a form of alternative medicine. Everyone in their life span experiences a different kind of wound. Clinacanthus nutans and Elephantopus scaber are well known traditional wound healing herbs. The present research work develops a new polyherbal formulation from the leaves of Clinacanthus nutans and Elephantopus scaber in the treatment of wound. Furthermore, bioactive compounds from different fractions were identified by using chromatography and spectroscopic method. The water soluble was further fractioned to isolate bioactive compounds. The herbal combination of Elephantopus scaber and Clinacanthus nutans were evaluated as an in-vitro antioxidant activity containing equal amount of both medicinal herb with their individual herbal activity followed by fractionation of polyherbal formulation for in-vivo wound healing activities. Antioxidant activity was performed in vitro by DPPH radical scavenging antioxidant activity followed by in-vivo wound healing activities using different wound model. The leaves of Clinacanthus nutans and Elephantopus scaber were extracted with 50% ethanol (EtOH) and separated using silica gel, Sephadex LH-20 to isolate the pure compounds. 2-(aminomethylsulfinyl)-N-hydroxyacetamide was the proposed compound isolated from aqueous fraction of the polyherbal formulation. The characteristics of the isolated compounds were achieved using Infrared spectroscopy (IR) and LCMS scan on aqueous fraction. The result showed that Elephantopus scaber crude extract on the basis of ECso performs a much faster action but with less % inhibition as compared to the combination of the new polyherbal formulation of crude extract. The polyherbal formulation has the highest % inhibition (89.49%) at the same dose as compared to individual Elephantopus scaber (87.66%). Comparison among all crude and fractions of new polyherbal formulation, it was found that the ethyl acetate fraction of polyherbal formulation has the fastest activity, ECso 14.83 J.tg/ml, with inhibition of 89.28%. Furthermore, during evaluation of wound contraction on excision and incision wound model, ethyl acetate fraction possess highest activity with (P < 0.001) and (P < 0.0001) respectively. During burn wound model, aqueous fraction (P < 0.001) possesses highest activity followed by an ethyl acetate fraction (P < 0.0001). Liquid chromatography-Mass spectrometry (LCMS) analysis identifies the presence of several flavonoid-based compounds that work synergistically with other bioactive compounds. These bioactive compounds were further confirmed by liquid chromatography-electrospray ionizationtandem mass spectrometry (LC-ESI-MS/MS). The prescence of bioactive compounds such as apigenin, luteolin, b-sitosterol, 3'-hydroxy-5,6,7,4'-tetra methoxy flavone, quercetin, vanillin, gallic acid, benzoic acid and rutin were confirmed by using High performance liquid chromatography. In conclusion, flavonoid increases the antioxidant activity that surge the rate of wound contraction and work synergistically with other bioactive compounds.