Integrated High‐Performance Liquid Chromatography, Gas Chromatography‐Mass Spectrometry and Liquid Chromatography‐Tandem Mass Spectrometry‐Based Metabolomics of Bacopa monnieri : Chemometric Insights Int

ABSTRACT

Bacopa monnieri (Brahmi) is a widely used Ayurvedic medicinal herb valued for its cognitive‐enhancing and neuroprotective properties, which are attributed to a complex mixture of triterpenoid saponins, flavonoids and other phytochemicals. Conventional quality‐control approaches based on a limited number of marker compounds may not adequately capture its phytochemical complexity. In this study, an integrated multimodal analytical platform was developed to comprehensively evaluate phytochemical variability among cultivated, natural habitat and commercially available Bacopa monnieri samples. Hydroalcoholic extracts were analysed using high‐performance liquid chromatography (HPLC) for total bacoside quantification, LC‐tandem MS (LC‐MS/MS) for targeted determination of apigenin and luteolin and gas chromatography‐MS (GC‐MS) for untargeted metabolomic profiling. Multivariate chemometric tools, including principal component analysis, hierarchical cluster analysis, correlation analysis and Venn diagram analysis were employed for data interpretation. Extractive yields ranged from 11.83% to 12.85%, whereas total bacoside content varied markedly from 3.15% to 25.25%, indicating substantial source‐dependent differences. The validated LC‐MS/MS method demonstrated excellent linearity ( R ² = 0.997–0.998) and high sensitivity, revealing higher concentrations of apigenin and luteolin in in‐house extracts, while several commercial samples contained these compounds below the limit of quantification. GC‐MS profiling identified diverse metabolites, including triterpenoids, phytosterols, fatty acids and phenolic compounds. Chemometric analyses effectively discriminated samples according to geographical origin and processing history. This integrated analytical strategy provides a robust framework for authenticity assessment, quality evaluation, and standardization of Bacopa monnieri and its commercial preparations.