HPTLC method development and validation for the determination of andrographolide in raw material and tablet containing ethyl acetate fraction of Andrographis paniculata

Abstract

Purpose: To develop and validate a rapid, accurate, and selective High Performance Thin Layer Chromatography (HPTLC), for quantification of andrographolide in raw materials and tablets ethyl acetate fraction of Andrographis paniculata as antimalarial agent. Methods: HPLC was conducted to determine the andrographolide concentration of ethanol extract and Camag linomat 5 in the stationary phase. HPLC measurements were conducted at a wavelength of 228 nm with a ratio of chloroform: methanol (90:10, v/v) in the mobile phase. Results: The validated method was separated andrographolide from other component with good resolution, and obtained retention factor was 0.38 ± 0.03. The data for calibration plot showed good linear relationship, with R2 = 0.998 in the concentration range of 138.0 - 460.0 ng/spot. The limit of detection and quantification were 9.6 ng/spot and 28.8 ng/spot, respectively. The percentage recovery was between 98.0 and 100.5 %. Additionally, the relative standard deviation method was between 1.4 and 1.0 % Conclusion: This method fulfills the validation requirements of selectivity, linearity, accuracy, and precision. Further, it can separate andrographolide from degradants. Thus, HPTLC method can be used to analyze the ethyl acetate fraction of ethanol extract of A. paniculata and its tablet products.