A validated high-performance thin-layer chromatography method for the quantification of chlorogenic acid in the hydroalcoholic extract of Gynura cusimbua leaves

A simple, reliable, and reproducible high-performance thin-layer chromatography (HPTLC) method was developed for the quantitative determination of chlorogenic acid in the hydroalcoholic extract of Gynura cusimbua (D. Don) S. Moore leaves (HAGC). HPTLC plates (10 cm x 10 cm) pre-coated with silica gel 60F(254) as the stationary phase and a mobile phase consisting of ethyl acetate-formic acid-acetic acid-water (100:11:11:2.6, V/V) were used for achieving good separation. Densitometric evaluation for fingerprinting and quantification was performed at a scanning speed of 100 mm/s and 6 x 0.45-nm micro-slit dimension at 366 nm in absorption mode with a deuterium lamp. The method was validated in terms of linearity, limits of detection and quantification, precision, accuracy, and robustness parameters. The R-F of chlorogenic acid was 0.43 & PLUSMN; 0.01. The linear regression analysis data for the calibration plots of chlorogenic acid showed a good linearity relationship with r(2) = 0.9996, in the concentration range of 50-250 ng/spot. The limits of quantification and detection were 43.12 and 14.36 ng/band, respectively. The amount of chlorogenic acid in HAGC was found to be 3.37% w/w. The quantification and the method validation of chlorogenic acid have not yet been reported for G. cusimbua which can be employed for the appropriate standardization of the plant extract/formulation.