Harnessing HPTLC and HPLC Techniques for Identification and Quantitative Estimation of Bioactive constituents in Ayush D Formulation and its three main ingredients viz; Syzygium cumini, Withania coagulans and Caesalpinia bonducella

Ayurveda, an ancient science originating from India, serves as a holistic approach to maintaining human well-being and offers remedies derived from indigenous plants, animal sources, and mineral-based products to address various ailments. The standardization of polyherbal drugs is a crucial aspect in ensuring their quality. High-Performance Thin-Layer Chromatography (HPTLC) fingerprinting profiling and High-Performance Liquid Chromatography (HPLC) plays a pivotal role in identifying key compounds. In this manuscript HPLTC has been used to identify some major compounds such as Ellagic acid and Gallic acid in the Ayush D formulation and Syzygium cumini extract, as well as Withaferin A in Ayush D and Withania coagulans, and Lupeol in Ayush D and Caesalpinia bonducella. The determination of Retention Factor (Rf) values of reference standards was observed in test solutions of Ayush D and its constituent ingredients under ultraviolet (UV) light at 254 nm and 366 nm, as well as white light at 540 nm. High-Performance Liquid Chromatography (HPLC) chromatograms of Ayush D and its ingredients extracts are analyzed in correlation with respective reference standards. The distinctive bioactive constituents inherent to these ingredients facilitate their facile identification in polyherbal formulations. The identification and verification of bioactive constituents are accomplished through HPTLC fingerprint profiling, while the quantification of biomarkers via HPLC serves as an optimal method for evaluating the quality of Ayurvedic formulations during the developmental stages of quality control protocols. This comprehensive approach aids in establishing robust quality assurance measures for Ayurvedic formulations, ensuring their efficacy and adherence to standardized criteria.