UHPLC-ESI-QTOF-MS/MS Based Identification, Quantification, and Assessment of in Silico Molecular Interactions of Major Phytochemicals from Bioactive Fractions of Clerodendrum glandulosum Lindl. Leaves

Abstract

Clerodendrum glandulosum Lindl., is an ethnopharmacologically important species of the North-eastern region of India. Leaves of this plant are consumed as food and traditionally used as medicine to cure hypertension, diabetes, and other metabolic syndromes. This study was designed to explore the antioxidant potential in the Clerodendrum leaves guided by in-vitro activity, which is responsible for the therapeutic benefits. Leaves were extracted with 95 % methanol and further fractionated with solvents of varying polarities (e. g., petroleum ether, ethyl acetate, and butanol). Total phenolic and total flavonoid content of the crude extract/ fractions were measured by Folin-Ciocalteu and AlCl₃ methods, respectively. Crude extract/ fractions were screen for in-vitro antioxidant and cytoprotective activities to determine the most bioactive fraction. Simultaneously, the chemical constituents of these fractions were identified and characterized using UHPLC-ESI-QTOF-MS/MS. Subsequently, major phenolic compounds identified were subjected to in-silico molecular docking with pro-oxidant enzymes to elucidate possible biological functions. Both ethyl acetate and butanol fractions showed the presence of a high concentration of phenolic and flavonoid content along with the best antioxidant and cytoprotective properties compared to all other fractions. Chemical profiling of these active fractions revealed the presence of different phenolic compounds, among which verbascoside was the principal compound. These major phytoconstituents also exhibited strong binding interactions with the crucial amino acid residues of the target proteins, which complemented the in-vitro bioactivities. In conclusion, this study offers structured information on antioxidant phytochemicals present in C. glandulosum leaves, which would be worthwhile for future investigations on the therapeutic properties at the molecular level.

Keywords: antioxidant; cytoprotective; molecular docking; oxidase; phenylethanoid glycosides; xanthine.