Abstract
Fermentation of rice grains requires diverse metabolic enzymes to be synchronously synthesized by the microbial community. Although many studies have used a metaproteomic approach to investigate the roles of microorganisms in improving the flavor of fermented foods, their roles in producing compounds with biological activity have not yet been reported. In a previous study the ferment obtained from unpolished black rice (UBR) fermented with a defined microbial starter (De-E11), comprised of Rhizopus oryzae, Saccharomycopsis fibuligera, Saccharomyces cerevisiae, and Pediococcus pentosaceus, (fermented UBR; FUBR) showed a strong melanogenesis inhibition activity in B16F10 melanoma cells. Hence, in this study, the roles of these microorganisms in producing the melanogenesis inhibitor(s) in FUBR was investigated using a metaproteomic approach. The melanogenesis inhibition activity of the FUBR liquid (FR-Liq) was found to increase with longer fermentation times. R. oryzae and S. cerevisiae were the major hosts of proteins related to the biosynthesis of melanogenesis inhibitor(s) in the FUBR. During fermentation, the enzymes involved in the degradation of UBR and in the carbohydrate metabolic process were identified. These enzymes were associated with the process of releasing of bioactive compound(s) from UBR and the synthesis of organic acids from the microorganisms, respectively. In addition, enzymes involved in the synthesis of some known melanogenesis inhibitor(s) and in the degradation of the melanogenesis stimulator (arsenate) were detected. Varying the combination of microorganisms in the De-E11 starter to produce the FR-Liq revealed that all four microorganisms were required to produce the most potent melanogenesis inhibition activity. Taken together with the metaproteomics results, this suggested that the microorganisms in De-E11 synchronously synthesize the FR-Liq with melanogenesis inhibition activity. In conclusion, this information on the metaproteome in FUBR will increase our understanding of the microbial metabolic modes and could lead to knowledge-based improvements in the fermented rice process to produce melanogenesis inhibitor(s).