Liquid Chromatography Coupled With High‐Resolution Mass Spectrometry Profiling of Metabolites in Cassia alata Flower Extracts and Their DPPH Free Radical Scavenging Activity

ABSTRACT Cassia alata flowers remain phytochemically under‐characterized relative to the leaves. However, floral tissues can harbour distinct and, in some species, more abundant phenolic and flavonoid profiles, representing an unexploited source of antioxidant phytochemicals with potential functional‐food and nutraceutical relevance. Also, Cassia alata L. (Fabaceae) is a major source of numerous anthraquinones and flavonoids, yet the secondary metabolome of its floral tissues has not yet been thoroughly studied in comparison to its vegetative parts. This research therefore aimed to examine the phytoconstituents of C. alata flower extracts and evaluate their antioxidant potential. This represents, to our knowledge, the first liquid chromatography coupled with high‐resolution tandem mass spectrometry (LC‐HRMS/MS) and 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH)‐based comparative profiling of C. alata flower extracts prepared with three solvents of differing polarity. Aqueous, 70% methanol, and 70% ethanol extracts of C. alata flowers were prepared and characterized using LC‐HRMS in negative ionization mode. The antioxidant potential was determined using the DPPH radical scavenging assay. LC‐HRMS enabled the tentative identification of 42 metabolites (in aqueous, methanolic, and ethanolic extracts), which were mainly flavonoids, phenolic acids, and glycosides. Some notable metabolites determined were Quercetin, Luteolin, Isomyricitin, Kaempferol‐dihexoside, Kaempferol‐ O ‐rutinoside, and Sophoricoside. The DPPH assay showed that all the extracts exhibited free radical scavenging activity in a concentration‐dependent manner. The ethanolic extract exhibited the strongest activity, with the lowest IC 50 value of 12.10 ± 2.22 µg/mL, followed by the aqueous extract, while the methanolic extract showed the weakest activity among the plant extracts. The observed antioxidant potential of the ethanolic extract may be associated with the wide diversity of tentatively identified flavonoid‐related metabolites. These results suggest that C. alata flower extracts may serve as potential sources of antioxidant phytochemicals for food and health‐related applications.