Isolation Of Hydrogen-producing Bacteria And Investigation Of The Enhanced Biotransforming Activity Of Daidzein By Mixed Culture

Isoflavones are the secondary metabolites during the growth of soybeans, mainly composed of daidzein, genistein and glycitein. Isoflavones, one of the plant-produced phytoestrogens have structural and functional similarity to human estrogen by binding with estrogen receptors. It is well known that isoflavones are of various beneficial effects, such as preventing cancer, lowering blood fat, attenuating bone loss, improving women's menopause syndrome, antioxidant activities, and so on. After being absorbed, isoflavones can be degraded into different metabolities. Studies showed that isoflavone metabolites are of higher and wider bioactivities than that of isoflavones.Eight bacterial strains named strain 4, 7, 9, 21, 26, 28, 208 and G30, which were isolated from Equus quagga, Lama guanicoe, Helarctos malayanus, Cervus albirostris, Grus nigricollis, Crossoptilon mantchuricum, and chicken fecal sample respectively, are facultative bacteria with hydrogen producing activity. Based on morphology feature, physiological and biochemical characteristics as well as analysis of the 16S rDNA sequence, bacterial strains 4, 9, 21, 26 and 208 were identified as Klebsiella pneumoniae 4,Klebsiella pneumoniae 9,Klebsiella pneumoniae 21,Klebsiella pneumoniae 26 and Klebsiella pneumoniae 208. Bacterial strain 7 was identified as Klebsiella variicola 7, strain 28 was identified as Paenibacillus polymyxa 28 and strain G30 was identified as Escherichia fergusonii G30, respectively. All of isolated bacteria can produce hydrogen under aerobic conditions.Gram positive bacterial strain Sharpea azabuensis Niu-O16 isolated from bovine rumen was an obligate anaerobic strain, which is capable of biotransforming daidzein into DHD under anaerobic conditions. After long time domestication resistant to oxygen, original obligate anaerobic bacterial strain Niu-O16 can grow in BHI liquid medium and produce DHD from daidzein under aerobic conditions. However, the biotransformation capacity of daidzein is significantly decreased. In this study, enhanced the biotansformation activity of domesticated Niu-O16 was investigated by coculture with isolated hydrogen-producing bacterial strains. The rusults showed that all the biotransforming capacity of daidzein was significantly increased by mixed culture (P<0.01) compared with pure culture of strain Niu-O16, except the mixed culture of strain 28 and Niu-O16 (P>0.05). Among all the mixed cultures, the biotransformation efficiency of daidzein by the mixed culture of strain G30 and Niu-O16 was the highest. The biotransforming ratio of daidzein was up to 82.8%, which is increased by 25.7% compared with that of pure culture of strain Niu-O16 under aerobic conditions. According to the biotransformation kinetics, the biotransforming spead of daidzein by the mixed culture of bacterial strain G30 and Niu-O16 became faster than that of pure culture of strain Niu-O16. When the concentration of substrate daidzein was 0.2 mmol / L, more than 80% of daidzein can be transformed into DHD by the mixed culture within 12 hrs, and almost completely biotransformed within 24 hrs incubation under the aerobic conditions. In addition, the highest biotransfoming concentration of substrate daidzein was up to 1.2 mmol / L by the mixed culture of bacterial strain G30 and Niu-O16.