Preparation and Evaluation of Conessine Enriched Extract from Holarrhena antidysenterica Bark for Enhanced Antibacterial Properties

The current study was designed with a view to develop an efficient method to recover extract of Holarrhena antidysenterica bark with high conessine content decipher enhanced antimicrobial action. H. antidysenterica bark powder was extracted by reflux using methanol as a solvent containing varying percentage of di-ethylamine (2.5%, 5%, and 10% respectively). Extract was further subjected to chromatographic separation to isolate pure conessine. Isolated conessine was characterized by 1H and 13C NMR spectroscopy. Estimation of conessine content in the extracts was performed by high performance thin layer chromatography (HPTLC). The comparative antimicrobial properties of all the extracts were assessed in both gram +ve and gram -ve bacteria by performing broth microdilution assay and disc diffusion assay method. A complementary in-silico study was also employed to understand the molecular interaction between conessine and New Delhi metallo-βlactamase 1 by molecular docking. It was observed that, the extract obtained with 5% di-ethylamine containing methanol contains maximum amount of conessine (Approx. 3%). In-vitro antibacterial assay showed that, the same extract exhibited maximum anti-bacterial properties when compared with the other extracts (2.5% and 10% di-ethylamine) which directly corroborates with conessine content. Molecular docking experiment also revealed strong antibacterial action (docking score: -8.1) compared to tetracycline (docking score: -7.5) by blocking with the crucial amino acid residues of New Delhi metallo-β-lactamase 1 protein which may be responsible for antibacterial action. From the current findings it can be concluded that, the extract obtained from methanol containing 5% di-ethylamine is ideal solvent for better recovery of conessine from H. antidysenterica bark with better antimicrobial action