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Abstract

METHOD DEVELOPMENT AND VALIDATION FOR SIMULTANEOUS ESTIMATION OF GALLIC ACID AND PIPERINE IN HERBAL EXTRACT AND POLYHERBAL FORMULATION BY HPTLC

Smita G. Patil, Varsha M. Jadhav*, Vilasrao J. Kadam

ABSTRACT

This article gives a simple, rapid and accurate High performance thin layer chromatographic method for determination of Gallic acid and Piperine in developed herbal formulation. Merck TLC aluminium sheets of silica gel G60 F254 with the thickness of 200μm was used to carry out the separation. Toluene: ethyl acetate: formic acid(5:3.5:0.5 v/v/v), was used as mobile phase. Analysis of the compounds was carried out by densitometry in the absorbance mode at 290 nm. The selected mobile phase gave the well defined peak at the Rf value of 0.20±0.03 and 0.50 ±0.03 for gallic acid and piperine respectively. Method was then validated for accuracy, linearity, precision, specificity, robustness, limit of detection (LOD) and limit of quantification (LOQ) as per ICH guidelines. The linear regression analysis data for the calibration plots showed good linear relationship with the regression coefficient (r2) of 0.994 and 0.993 for the gallicacid and piperine respectively, in the concentration range of 200-900ng/spot. The limit of detection and limit of quantification were 0.72 and 2.18 ng/spot, respectively for gallic acid and 36.88 and 111.78 ng/ spot, respectively for piperine. In conclusion, statistical analysis of data showed that the method is precise, selective and reproducible for estimation of both compounds. Hence the developed method can be applied in routine quality control analysis for identification and quantification of gallic acid and piperine in their extracts and developed herbal formulation.

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