Physical, Volatile, and Flavour Precursor Changes in Faba Bean During Rhizopus oryzae Fermentation

Faba bean (Vicia faba L.) is increasingly recognised as a nutritious and sustainable food ingredient, although its characteristic beany and green odours can limit consumer acceptance. Fermentation offers a potential strategy to modify the sensory and functional properties of legume-based foods. In this study, faba beans of different particle sizes, such as split beans (S), 4000-5000 μm (5K) and 1000-2000 μm (2K), were fermented with Rhizopus oryzae for up to 48 h to produce tempeh-like products. Mycelial development and colour were assessed by qualitative visual assessment and colour measurement, texture was evaluated at 40 and 48 h, volatile compounds were profiled using headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS), and free amino acids and fatty acids were quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Mycelial coverage increased throughout fermentation, with the S and 5K samples showing more extensive and uniform colonisation than 2K samples. Fermentation significantly altered colour, particularly through reduced yellowness, whereas texture showed no consistent differences between 40 and 48 h. Volatile profiles became increasingly diverse after 40 h, with fermentation time as the principal source of variation and particle size contributing to differences at 40 and 48 h. Changes in fatty acid and free amino acids were consistent with lipid transformation and amino acid catabolism. Several volatiles associated with beany and green odours increased rather than decreased during fermentation, providing no evidence for their direct removal. Overall, R. oryzae fermentation substantially altered the volatile profile of faba bean, suggesting that beaniness may be potentially masked by newly formed odourants rather than elimination of individual beany volatiles.