Development and Validation of a High-Performance Thin-Layer Chromatography Method for Simultaneous Qualitative and Quantitative Analysis of Scutellarin and Scutellarein in S. barbata D. Don

Abstract

Background: S. barbata D. Don, a perennial herb from the Lamiaceae family, is renowned for its medicinal properties, with scutellarin and scutellarein being key bioactive constituents. Aim: In this study, we present the development and validation of a High-Performance Thin- Layer Chromatography (HPTLC) method for the simultaneous quantitative and qualitative analysis of scutellarin and scutellarein in the hydroethanol extract of S. barbata D. Don. Methods: The chromatographic conditions were optimized using different solvent systems, and validation was performed as per ICH guidelines. Results: The mobile phase comprising Ethyl acetate: methanol: formic acid: water (20: 2.7: 0.5: 2) and scanning wavelength set at 254 nm was optimized for study, resulting in distinct and well-separated spots for scutellarin and scutellarein with Rf values of 0.31 and 0.96, respectively. The method was rigorously validated to ensure reliability and reproducibility. Calibration curves exhibited excellent linearity for both scutellarin (r² > 0.98) and scutellarein (r² > 0.99) over a concentration range of 10-30 μg/ml. The developed method underwent thorough validation following ICH guidelines. Validation parameters included accuracy, limit of quantification, limit of detection, linearity, precision, and recovery. The determined concentration of scutellarin and scutellarein in the hydroethanolic extract of S. barbata D. Don was found to be 27.00 μg/mg and 20.16 μg/mg of extract, respectively. Conclusion: This proposed method demonstrates its utility for both qualitative and quantitative analyses of scutellarin and scutellarein in the hydroethanolic extract of S. barbata D. Don, offering a reliable and validated approach for quality control and standardization in the herbal product industry.