Column chromatography for preparing rosmarinic acid rich extract from Orthosiphon aristatus

Abstract

Chromatographic techniques were used to prepare rosmarinic acid rich extract from Orthosiphon aristatus. Such extract is highly required for herbal product development because of its remarkable bioactivities. Therefore, the plant crude extract was fractionated using solvent mixture, ethyl acetate and ethanol in column chromatography. The solvent system was developed using thin layer chromatography. The results found that 60-99.9% ethyl acetate in ethanol could separate rosmarinic acid with the retention factor (Rf) ranged from 0.2–0.3. Of the solvent systems, 70% ethyl acetate could produce the highest yield of rosmarinic acid, 4.5% w/w with 100% purity. The concentration of rosmarinic acid was quantified by high performance liquid chromatography integrated with tandem mass spectrometry. The subsequent combined fractions could recover 70% of total rosmarinic acid from crude extract. The bed height of the silica gel packed column increased from 16 to 24 and 32 cm did not affect the performance of rosmarinic acid elution in column chromatography. The silica gel absorbent could be repeatedly used up to five times to recover rosmarinic acid from crude extract. Column chromatography can be used to recover rosmarinic acid (>85%) using the binary solvent system (70% ethyl acetate:30% ethanol) effectively.