Cytotoxicity and Apoptosis-Inducing Effect of Soybean Broth Cultured with Microorganisms Used in the Production of Fermented Soybean Foods on Human Monoblastic Leukemia U937 Cells

Soybean broth (SB) is a waste product in the manufacturing of boiled soybean-based foods. We cultured the SB with six pure strains of microorganisms (Aspergillus oryzae, Aspergillus sojae, Rhizopus oligosporus, Zygosaccharomyces rouxii, Bacillus subtilis (natto), and Tetragenococcus halophilus) used commercially in the production of fermented soybean foods, including miso, soy sauce, natto and tempeh, and examined their cytotoxicity and apoptosis-inducing effect on U937 cells. Ethyl acetate extracts of all cultures showed cytotoxicity against U937 cells, although that of SB did not in the range of concentrations tested. Further, ethyl acetate extracts of SB after culturing with A. sojae (ESBA) and ethyl acetate extracts of SB after culturing with B. subtilis (natto) (ESBB) elicited DNA fragmentation. The DNA fragmentation elicited by ESBA was prevented by pretreatment with a general caspase inhibitor (Z-Asp-CH2-DCB), implying induction of caspase-dependent apoptosis. Assays for caspase activity showed that apoptosis induction was associated with caspase-3, -8, and 9, which are the principal caspases involved in apoptosis. Thus, SB was conferred cytotoxicity upon culturing with the microorganisms used in this study; in particular, SB cultured with A. sojae induced apoptosis in U937 cells.