Unraveling the anti-obesity potential of Haldina cordifolia bioactive fractions in 3T3-L1 adipocytes differentiation: in vitro, high-performance thin-layer chromatography‒multistage mass spectrometry and in silico studies

The anti-obesity activity of the bioactive fractions of Haldina cordifolia (HC) was investigated in pursuit of a novel herbal plant targeting obesity, as its anti-obesity potential has not yet been investigated. This innovative study examined the HC bioactive fraction to select the most significant fraction using an array of studies targeting obesity. Initially, a quantitative phytochemical assessment including total phenolic, flavonoid, and steroidal contents was performed. Additionally, a series of in vitro antioxidant ABTS, DPPH, nitric oxide, and hydrogen peroxide scavenging assays were implemented, whereas, alpha-glucosidase, alpha-amylase, and pancreatic lipase assays were conducted for the inhibition of enzyme capabilities. The findings demonstrated that fractions F2 and F3 have better efficacy, based on IC₅₀ values. Subsequently, the anti-obesity potential of fractions was assessed utilizing cultured 3T3-L1 adipocytes, oil red O staining, and triglyceride content, as markers of accumulation of lipid which showed significant results. Furthermore, high-performance thin-layer chromatography‒multistage mass spectrometry (HPTLC‒MSⁿ) was employed to identify the phytoconstituents within these fractions, particularly the anti-obesity agent, β-sitosterol. HPTLC‒MSⁿ analysis revealed the presence of five major compounds in fraction F2: caffeine, quercetin, β-sitosterol naringin, and ascorbic acid along with retro Diels–Alder fragmentation. Finally, an in silico study was designed which subsequently established the optimal binding activity of the obtained compounds towards the particular receptors targeting obesity. Overall, fraction F2 showed the most potent activity. The study unveiled the promising anti-obesity potential of bioactive fractions from HC and a potential treatment strategy has been established to combat obesity.