UPLC-HRMS/MS-BASED CHEMOPROFILING AND BIOACTIVITY EVALUATION OF PHLOGACANTHUS CORNUTUS EXTRACT
Untargeted LC-HRMS/MS metabolomics has become a key approach for characterizing the complex chemical composition of plant extracts; however, integrating multiple annotation platforms into a coherent chemoprofilingframework remains challenging. In this study, an integrated workflow combining MZmine, MS-DIAL, GNPSfeature-based molecular networking, and SIRIUS was applied to systematically annotate metabolites inPhlogacanthus cornutus, a recently described species with limited phytochemical information. Integrated analysis of positive and negative UPLC-MS/MS revealed that the metabolite profile of Phlogacanthus cornutus is dominatedbyiridoid glycosides (including harpagoside and harpagide), hydroxylated diterpenoids, oxylipins, monoacylglycerols, and minor phenolic constituents. Merging polarity-specific ion forms improved annotation coherence and reducedredundancy relative to single-mode analysis. Bioactivity assays demonstrated concentration-dependent nitric oxideinhibition (IC50 = 19.27 ± 1.76 µg/mL) without detectable cytotoxicity, together with measurable antioxidant activity in ABTS .+ (maximum of 230 mg Trolox/g extract) and FRAP (≈11.2-15.3 mg Fe 2+ /g extract at 150 ppm) assays. These findings provide the first LC-MS/MS-based chemoprofile of Phlogacanthus cornutus and demonstratethe utility of integrating complementary annotation platforms for bioactivity-oriented metabolomics studies. Beyondestablishing a phytochemical and bioactivity reference for this understudied species, the proposed workflowmaysupport future natural product discovery, comparative chemotaxonomic studies, and systematic prioritizationof bioactive metabolites in medicinal plant research.
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