Untargeted NMR metabolomics analysis of Ficus racemosa fruit extracts and UHPLC-MS/MS-molecular networking analysis of the active extract

This study aimed to investigate the effects of drying methods and varying ethanol concentrations on the metabolite composition and biological activity of F. racemosa fruit extracts. Metabolomic analyses were conducted using 1 H-NMR spectroscopy, while the most active extract was characterized using UHPLC-MS/MS and feature-based molecular networking (FBMN). A total of 40 and 45 metabolites were tentatively identified through NMR and UHPLC-MS/MS analyses, respectively. The biological activity results indicated that the freeze-dried (FD) sample extracted with 100% ethanol exhibited the highest activity against 2,2-diphenyl-1-picrylhydrazyl radical (DPPH●), as well as α-amylase and α-glucosidase inhibitory activities, with IC 50 values of 21.5, 7.5, and 5.9 μg/mL, respectively. These activities were compared with reference standards, where quercetin exhibited SC 50 values of 5.6 μg/mL for DPPH scavenging and 7.3 μg/mL for α-glucosidase inhibition, while acarbose showed an IC 50 value of 2.0 μg/mL for α-amylase inhibition. Notably, metabolites such as pheophorbide a, phlorizin, bergenin, caffeic acid, luteoloside, chlorogenic acid, vitexin, naringenin, epicatechin, epilupeol, and betulinic acid were identified as major contributors to the observed antioxidant and enzymatic inhibitory activities. This study lays the groundwork for future research into the development of functional foods from F. racemosa fruit, particularly for managing postprandial hyperglycemia in diabetes. • Freeze-drying better preserved bioactive compounds, enhancing enzyme inhibition. • 100% ethanol extract of F. racemosa exhibited the strongest inhibition of α-amylase and α-glucosidase. • PLS analysis identified 20 key metabolites contributing to the activity. • Osajin was identified in the 100% ethanol extract through molecular networking. • F. racemosa extracts demonstrate potential as natural antidiabetic agents.