Pattern analysis of chromatograms of Euphorbiae genus

Abstract

A TLC method has been developed for the identification of two bio-markers, myricitrin and quercitrin, in Euphorbia hirta L., the raw material of Euphorbiae Hirtae Herba (飛揚草). These two bio-markers showed good separation with developing solvent composed of dichloromethane: ethyl acetate: methanol: formic acid in a ratio of 5:3:1.5:1, v/v, spraying with 10% sulphuric acid in ethanol, and detected in UV light at 366 nm. Three bio-markers, isoquercitrin, ellagic acid and afzelin, were also added for the fingerprint pattern analysis. HPLC method has been developed for the analysis of the five selected bio-markers in Euphorbia hirta L. samples. The bio-markers were detected with good separation when using a Dikma Platisil ODS column at 40oC, with a binary gradient mobile phase composed of 0.2% phosphoric acid and methanol, and detected at wavelength 360 nm. The HPLC method has been validated to test the reproducibility of fingerprints. TLC analysis indicated some differences between the Euphorbiae genus studied (Euphorbia hirta L., Euphorbia thymifolia L., Euphorbia humifusa W., Euphorbia maculata L., Euphorbia prostrata A. and Euphorbia hypericifolia L.) in a relatively rapid way. Differentiation between the species of same genus and detection of bio-markers in the Euphorbiae genus samples were performed in a more precise and accurate way with the assist of HPLC analysis. Comparison on patterns between Euphorbiae genus, same species and different species, have been briefly investigated. The difference between the fingerprint patterns of the same species samples from different collection regions was also investigated.