Synergistic probiotic co-fermentation drives metabolic remodeling and functional enhancement in Citrus aurantium L. juice

Synergistic co-fermentation enhanced antioxidant capacity of C. aurantium juice. • Co-fermentation increased β-glucosidase activity and flavonoid accumulation. • Probiotic co-fermentation reshaped aroma profiles and volatile composition. • Co-fermentation promoted volatile and non-volatile metabolic remodeling. • Co-fermentation enriched flavone and flavonol biosynthesis pathway. Citrus aurantium L. ( Citrus aurantium ) is rich in flavonoids and bioactive phytochemicals, but its industrial utilization is limited by suboptimal flavor quality and instability during processing. Herein, monoculture fermentation using Lactiplantibacillus plantarum (a lactic acid bacterium) or Bifidobacterium animalis subsp. lactis (a lactic acid bacterium), together with their synergistic co-fermentation, was employed to investigate flavor and functional modifications in C. aurantium juice. Compared with monoculture fermentation, co-fermentation increased total flavonoid and phenolic contents by 23.7-50.4% and 43.6-65.0%, respectively, while β-glucosidase activity, ABTS and DPPH radical-scavenging activities increased by 10.9-27.3%, 12.6-24.4%, and 12.7-26.4%, respectively. Electronic-nose and headspace solid-phase microextraction-gas chromatography-mass spectrometry analyses revealed that co-fermentation generated a distinct volatile profile characterized by enrichment of aroma-related compounds and altered representation of compounds associated with earthy and camphor-like notes. Untargeted metabolomics revealed extensive remodeling of phenolic, amino acid, and lipid metabolism. The relative abundance of limonin showed a marginal, non-significant decrease of 3.9%, while indoline and L-tyrosine increased by 3.9% and 3.8%, respectively ( p  < 0.001). These changes were accompanied by significant enrichment of the flavone and flavonol biosynthesis pathway ( p  = 0.0082). Correlation analysis further revealed strong associations between key differential metabolites and functional indicators. Collectively, these findings demonstrate that synergistic probiotic co-fermentation drives metabolic remodeling and flavonoid biotransformation in C. aurantium juice, providing a promising strategy for developing high-value functional citrus beverages. Future studies should further evaluate sensory acceptability, storage stability and industrial feasibility to facilitate commercial application.