Thin layer chromatography‐densitometric method for quantification of apigenin, oleanolic acid, and β‐sitosterol in clerodendrum serratum (Bharangi) root extract

Abstract

The present involves the development of an accurate, specific, and precise high‐performance thin‐layer chromatographic method for the identification and quantification of three markers apigenin, oleanolic acid, and β‐sitosterol in Clerodendrum serratum (Bharangi) root extract. The thin layer chromatographic development was carried out using an aluminum plate, pre‐coated with silica gel 60 F254 with toluene–chloroform‐ethyl acetate‐formic acid (5:4:1:0.1 v/v/v/v) as the mobile phase. Densitometric quantification was performed at 334 nm for apigenin and 662 nm for oleanolic acid and β‐sitosterol after derivatization with anisaldehyde‒sulfuric acid reagent. The optimized mobile phase resulted in chromatographic separation of apigenin, oleanolic acid, and β‐sitosterol bands at Rf of 0.19, 0.50, and 0.63, respectively. The method was found to be linear in the concentration range of 200–1200 ng/band for apigenin, and 400–2400 ng/band for oleanolic acid and β‐sitosterol, respectively. The recoveries were found to be 99.34%–99.92% for apigenin, 99.38%–99.81% for oleanolic acid, and 99.49%–99.66% for β‐sitosterol. The optimized method was found to be accurate, reproducible, robust, and specific and it was applied for the quantification of apigenin, oleanolic acid, and β‐sitosterol in hydroalcoholic root extract of Clerodendrum serratum.