Phytochemical profiling and therapeutic potential of Baliospermum montanum: A promising inhibitor of COX-2/15-LOX enzymes and NO production in LPS-stimulated RAW 264.7 macrophages

Abstract

IntroductionBaliospermum montanum (BM) is traditionally used to treat various ailments such as inflammation, rheumatoid arthritis, pain, etc. The root part of BM is well documented for its anti-inflammatory activity. However, the present study evaluates the multitargeted anti-inflammatory activity of the less explored BM leaf part. This is the first study investigating the COX-2, 15-LOX dual inhibiting potential and NO production inhibition of BM leaf extracts, fractions and compounds. MethodsThe BM leaf's water, ethanol, and hydroethanolic extracts were evaluated for anti-inflammatory activity by assessing the inhibition of COX-2, 15-LOX, and NO production. Additionally, the antioxidant potential of the extracts was determined through DPPH, ABTS, and FRAP assays. The most active ethanol extract was fractionated into 12 fractions and tested for anti-inflammatory, antioxidant, phenol and flavonoid content. Compounds were isolated from the most active fractions and identified using NMR techniques. The isolated compounds were checked for their anti-inflammatory activity. In addition, HPLC analysis of ethanol extract, fractions, and isolated compounds was carried out. ResultsThe ethanol extract exhibited higher inhibition of COX-2, 15-LOX enzymes and NO production (IC50 35.75 ± 1.95 μg/mL) in LPS-treated RAW 264.7 cells. It also showed higher antioxidant activity. Following the fractionation of the ethanol extract using different solvents, only the ethyl acetate fractions demonstrated inhibition of COX-2, 15-LOX enzymes, and NO production. The ethyl acetate and acetone fractions are excellent sources of antioxidants, phenolic and flavonoid compounds. An effort to find the active principles from these ethyl acetate fractions resulted in the isolation and identification of four potent anti-inflammatory agents, namely, (-)-epiafzelechin, afzelechin-(4α→8)-afzelechin, afzelechin-(4α→8)-epiafzelechin and β-sitosterol-d-glycoside. The first three compounds were reported for the first time from this plant and afzelechin-(4α→8)-afzelechin was found to be a potent inhibitor of the COX-2/15 LOX enzyme as well as NO production (IC50 = 5.9 ± 1.09 µM). ConclusionBased on these findings, we conclude that ethanol extract, fractions, and isolated compounds exert anti-inflammatory activity through multiple therapeutic mechanisms. The present study scientifically supports the traditional use of plant BM for treating inflammation and other related disorders.