Development and validation of RP-HPLC method for simultaneous analysis of andrographolide, phyllanthin, and hypophyllanthin from herbal hepatoprotective formulation

Abstract

A simple, sensitive, and accurate liquid chromatographic method with photodiode array detector was developed for the determination of andrographolide, phyllanthin, and hypophyllanthin. The separation was carried out on a reverse-phase 250 mm × 4.6 mm, 5μ symmetry C8 column (Waters). The gradient was prepared from 0.1% orthophosphoric acid (solvent A) and (1:1) acetonitrile:methanol (solvent B) as mobile phase delivered at a flow rate of 1 mL min-1. A linear behavior was observed between observed peak area response, and concentration of analytes was investigated, with good correlation coefficient. The method established was successfully applied to quantify andrographolide, phyllanthin, and hypophyllanthin from the herbal hepatoprotective formulation.