Species-specific accumulation of flavonoids and phenolic acids coordinates bioactivities in Gleditsia L.

The spines of Gleditsia species are a traditional medicinal resource in China, rich in flavonoids and phenolic acids as key bioactive compounds. However, the interspecific differences in their metabolomic profiles and the underlying links to bioactivities remain unclear, which limits the precise utilization of these resources. Here, metabolic profiling via UPLC-MS/MS with the MWDB4.0 database and multivariate analyses (PCA, OPLS-DA, HCA) systematically compared spines of G. sinensis, G. japonica var. delavayi, and G. fera. Total flavonoid (TFC) and phenolic (TPC) contents were quantified, and in vitro antioxidant (DPPH, ABTS⁺, FRAP) and enzyme inhibitory (α-amylase, α-glucosidase) activities were assessed. Pearson correlation analysis linked composition to bioactivity. A total of 1314 metabolites were identified, with flavonoids (511) and phenolic acids (246) being most abundant. Among 566 differentially accumulated metabolites (DAMs), 304 common DAMs (153 flavonoids, 53 phenolic acids) showed distinct species-specific accumulation. G. sinensis had the highest TFC (11.50 mg RE/g DW) and TPC, while G. japonica var. delavayi showed the lowest levels of these compounds. Its extract exhibited the strongest antioxidant (ABTS⁺ IC₅₀ = 8.91 μg/mL; FRAP = 17.2076 μmol Fe²⁺/g DW) and enzyme inhibitory (α-amylase IC₅₀ = 28.5 μg/mL; α-glucosidase IC₅₀ = 14.4 μg/mL) activities, significantly surpassing positive controls. TFC strongly correlated with FRAP capacity (r = 0.903, P < 0.001); notably, 4-O-Galloylquinic Acid showed an exceptionally high correlation (r = 0.991). This integrative study reveals species-specific flavonoid and phenolic acid accumulation in Gleditsia spines and their bioactivity mechanisms, providing a foundation for high-value utilization and development of natural hypoglycemic and antioxidant products.