Abstract
Crotalaria willdenowiana DC., an endemic and underexplored species from the wastelands and forested slopes of Tamil Nadu, was investigated for its phytochemical and biological potential using ultrasound-assisted extraction (UAE) optimized through Response Surface Methodology (RSM). Twenty methanolic root extracts were prepared under varying conditions of sample weight (0.15-1.841 g), extraction time (9.54-110.54 min), and temperature (26.5-68.5 °C). The optimized extract, obtained at 71.69 min, 57.27 °C, and a sample weight of 1.50 g, showed the maximum phytochemical yield, with TPC of 135.32 mg GAE/g, TFC of 122.85 mg QE/g, and an extraction yield of 24.13%. HPLC confirmed the presence of gallic acid and quercetin. Quantitative HPLC showed the presence of gallic acid (22.66 µg/mL) and quercetin (31.84 µg/mL), confirming the presence of some important phenolic and flavonoid components from the extract. In addition, NMR and FTIR studies confirmed the presence of signature functional groups and structural elements for these bioactive compounds. The extract exhibited 65% DPPH radical-scavenging activity and significant α-glucosidase inhibition (IC50 = 286.25 μg/mL). GC-MS analysis further identified 42 volatile constituents in the optimized extract.