Development and Validation of High Performance Thin Layer Chromatography Method for Standardization of Polyherbal Formulation Using Mangiferin and Gallic Acid as Markers

Abstract

Oxitard capsules, a polyherbal formulation, is an effective natural antioxidant that prevents oxidative stress-related tissue damage and is helpful for coronary artery disease. The aim of the present work was systematic development and validation of the high-performance thin-layer chromatographic method for simultaneous quantitative estimation of mangiferin and gallic acid from oxitard capsules. Both markers, mangiferin and gallic acid were separated on a pre-coated silica gel 60 F254 thin-layer chromatographic plate, using toluene:ethyl acetate:formic acid:methanol (4:6:0.8:2 v/v/v/v) as mobile phase. The retention factor of mangiferin and gallic acid was found to be 0.28±0.03 and 0.78±0.03, respectively. The linear regression analysis data for the calibration plots of drugs scanned at isosbestic point 299 nm showed a good linear relationship with r2=0.9931 and r2=0.9901, over the concentration range of 300 to 700 ng/spot and 500 to 900 ng/spot for mangiferin and gallic acid, respectively. The validation parameter followed for simultaneous estimation of these polyherbal drugs were linearity, precision, accuracy, robustness, ruggedness and sensitivity (limit of detection and limit of quantitation). The method was found to be precise, accurate, sensitive and robust meeting all the parameters as per International Council for Harmonisation guideline Q2 (R1). Thus, the developed high-performance thin-layer chromatography method can conveniently be employed for simultaneous detection and quantification of mangiferin and gallic acid in commercial polyherbal formulation.