Gorse (Ulex europeaus) wastes with 5,6-dimethyl benzimidazole supplementation can support growth of vitamin B₁₂ producing commensal gut microbes

Many commensal gut microbes are recognized for their potential to synthesize vitamin B-12, offering a promising avenue to address deficiencies through probiotic supplementation. While bioinformatics tools aid in predicting B-12 biosynthetic potential, empirical validation remains crucial to confirm production, identify cobalamin vitamers, and establish biosynthetic yields. This study investigates vitamin B-12 production in three human colonic bacterial species: Anaerobutyricum hallii DSM 3353, Roseburia faecis DSM 16840, and Anaerostipes caccae DSM 14662, along with Propionibacterium freudenreichii DSM 4902 as a positive control. These strains were selected for their potential use as probiotics, based on speculated B-12 production from prior bioinformatic analyses. Cultures were grown in M2GSC, chemically defined media (CDM), and Gorse extract medium (GEM). The composition of GEM was similar to CDM, except that the carbon and nitrogen sources were replaced with the protein-depleted liquid waste obtained after subjecting Gorse to a leaf protein extraction process. B-12 yields were quantified using liquid chromatography with tandem mass spectrometry. The results suggested that the three butyrate-producing strains could indeed produce B-12, although the yields were notably low and were detected only in the cell lysates. Furthermore, B-12 production was higher in GEM compared to M2GSC medium. The positive control, P. freudenreichii DSM 4902 produced B-12 at concentrations ranging from 7 ng mL(-1) to 12 ng mL(-1). Univariate-scaled Principal Component Analysis (PCA) of data from previous publications investigating B-12 production in P. freudenreichii revealed that B-12 yields diminished when the carbon source concentration was <= 30 g L-1. In conclusion, the protein-depleted wastes from the leaf protein extraction process from Gorse can be valorised as a viable substrate for culturing B-12-producing colonic gut microbes. Furthermore, this is the first report attesting to the ability of A. hallii, R. faecis, and A. caccae to produce B-12. However, these microbes seem unsuitable for industrial applications owing to low B-12 yields.