Abstract
Sojae Semen Praeparatum (SSP), a traditional Chinese fermented soybean product, was optimized through novel single/double enhancement fermentation using Aspergillus oryzae and Bacillus subtilis. Enhanced fermentation significantly increases the production of flavor amino acids. Using headspace solid-phase extraction microextraction gas chromatography-mass spectrometry (HS-SPEM-GC-MS) technology, 51 critical flavor substances were identified, confirming that enhanced fermentation improves the flavor profile of SSP. Isoflavone quantification revealed that enhancement strategies promoted isoflavone conversion. Enzyme inhibition and antioxidant activities were superior in the double enhancement fermentation group. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis demonstrated a significant increase in SSP's enrichment in the flavone and flavonol biosynthesis pathway. Screening identified 19 key flavonoid components strongly correlated with bioactivity, and enhancement fermentation notably enhancing their accumulation. Metagenomic sequencing revealed 14 key differential microorganisms, essential to flavor development and activity enhancement in SSP. This study offers valuable insights for optimizing fermentation processes to enhance product quality.