Microbial Intervention in the Soaking Step during Tempeh Preparation to Modulate B Vitamin Contents: A Metabolomics Approach

B vitamins are integral in maintaining human health. Owing to its high B vitamin contents, tempeh serves as an excellent plant-based option to achieve an adequate intake of these nutrients through diet. In order to investigate the metabolomic profile of tempeh and modulate the contents of vitamins B₂ and B₃ in it, we employed microbial interventions using two species of lactic acid bacteria (LAB) and various inoculum sizes in the soaking step during tempeh production. We thoroughly analyzed the tempeh samples using a wide-targeted metabolomics approach via gas chromatography-mass spectrometry. A total of 86 metabolites, including sugars, amino acids, fatty acids, and organic acids, were annotated in all the samples. Principal component analysis revealed that the two LAB species led to alterations in the tempeh metabolome with sugars and amino acids separating it. Additionally, variations in inoculum size influenced the metabolomic profile of tempeh as well. Furthermore, the highest B vitamin concentrations in the tempeh samples were observed when 5% v/v <i>Lactiplantibacillus plantarum</i> was incorporated during the soaking process. Specifically 5.312±0.315 mg of vitamin B₂ per 100 g of food and 10.334±0.138 mg of vitamin B₃ per 100 g of food were detected, which corresponded to a 1.22-fold and 1.08-fold increase compared to the contents observed in tempeh obtained from water-soaked soybean. These findings will assist in advancing tempeh production and will also contribute to innovation in the broader field of fermented food processing.