Development and validation of high-performance thin-layer chromatography method for quantitative estimation of lepidine in Lepidium sativum L. seed extract and its antioxidant activity

Lepidium sativum, a member of the Brassicaceae family, is traditionally used as an edible herb with notable antioxidant, antibacterial, anticancer, and nutritional properties. This study focused on developing and validating a high-performance thin-layer chromatography (HPTLC) method for the quantitative estimation of lepidine in L. sativum seed extract. The HPTLC analysis was performed using pre-coated HPTLC plates 60 F254 as the stationary phase. The developing mobile phase of n-butanol: water: acetic acid (7:2:1.5 v/v/v) showed good separation of lepidine with an Rf of 0.70 ± 0.02. Densitometric determination of lepidine was detected at 314 nm using a CAMAG TLC scanner III. The method was validated under ICH guidelines. It demonstrated good linearity between 0.5 and 4 µg/band, with an R2 of 0.991. The detection limit (LOD) and the quantification limit (LOQ) were 0.162 µg/band and 0.491 µg/band, respectively. The sonication extract of L. sativum showed that the highest lepidine content of 7.8 ± 0.36 mg/g and exhibited strong antioxidant activity in the DPPH assay, with an IC50 of 14.9 µg/ml. The validated HPTLC method proved to be simple, precise, accurate, cost-effective, and suitable for the rapid quantification of lepidine and supporting the antioxidant potential of L. sativum seed extract.