enhanced aglycone production of fermented soybean products by Bacillus species

This study evaluated the effect of starter culture and fermentation period on the isoflavone content of protein-rich soybeans variety TG145. Initially, soybeans were washed, soaked in water for 16 h and autoclaved at 121°C for 40min. Three different bacterial starter cultures (~104 CFU/g) namely Bacillus subtilis BEST195, B. subtilis Asaichiban and B. subtilis TN51 were then added and the fermentation was allowed to proceed at 42°C for 24 h (natto-style) and 72 h (thua nao-style). The quantities of six major isoflavones (daidzin, genistin, glycitin, daidzein, genistein, and glycitein) were then determined in these fermented soybean products using reverse phase HPLC technique. Generally, our results clearly showed that the content of total isoflavones in the fermented products prepared by Bacillus starter cultures greatly increased ranging from 43 to 99% compared to that of the unfermented autoclaved soybeans. In addition, a dramatic increase of aglycones was also observed (> 400%) in the soybean products fermented by Bacillus subtilis strain TN51. This present study suggests a promising use of Bacillus starter cultures in improving isoflavone compounds especially the aglycones which would benefit for novel functional food development. Acta Biol Szeged 53(2):93-98 (2009) Key WordS Aglycones Bacillus fermented soybean isoflavones Accepted Nov 24, 2009 *Corresponding author. E-mail: [email protected] 93 Soybeans (Glycine max (L.) Merr.) are highly regarded as a healthy food in several Asian countries and are widely consumed as soymilk, tofu and fermented products. In these soybeans and soy-products, several phytochemicals are found and they appear to be the active compounds causing many beneficial health effects (Dixon 2004; Wiseman 2006). Soy isoflavones in particular are a group of natural heterocyclic phenols comprising of aglycones, β-glucosides, acetylglucosides, and malonylglucosides. These isoflavone compounds are of great importance due to their pharmacological and antioxidant properties. For example, several researchers have showed the beneficial use of isoflavones; these include prevention of mammary cancer (Gotoh et al. 1998), reduced risk of cardiovascular diseases (Teede et al. 2001), improvement of bone health and menopause symptoms (Potter et al. 1998; Ishimi et al. 2002), antimutagenic effects (Peterson et al. 1998; Park et al. 2003), and antidiabetic effects (Liu et al. 2006). Of these compounds, it appears that the glucoside forms are predominant in soybeans. However, it should be noted that the content and composition of isoflavones are variable depending on many parameters such as soybean variety (Wang and Murphy 1994; Lee et al. 2003, 2007), geographical plantation (Wang and Murphy 1994; Hoeck et al. 2000;, Lee et al. 2003), storage time (Lee et al. 2003), crop year (Wang and Murphy 1994), and food processing techniques (Jackson et al. 2002; Kao et al. 2004, Lee et al. 2007). In Asia, there are several traditional fermented soybean products such as Japanese natto, Korean chungkookjang, Indian kinema and Thai thua nao. Interestingly, numerous studies have reported that the amount of aglycones is much higher in fermented soybeans compared to that of unfermented soybeans; such products include natto (Wei et al. 2008), miso (Yamabe et al. 2007), sufu (Yin et al. 2004, 2005), douche (Wang et al. 2007), and chungkukjang (Kwak et al. 2007). At present, the presence of aglycone isoflavones has attracted special interest due to their bioavailability, high rate of absorption in animals and humans (Izumi et al. 2000; Kano et al. 2006) and high antiproliferative activity on human cancer cells (Peterson et al. 1998). Especially for fermented soybeans, several studies have also revealed the role of microbes in soybean fermentation as the active agents to enrich isoflavone aglycones. This conversion of glucoside isoflavones to aglycones during the fermentation is achieved by the activity of microbial β-glucosidase enzymes (Chien et al. 2006;, Kuo et al. 2006;, Chun et al. 2007, 2008). As part of the programme to improve nutritional value of thua nao, we previously screened and identified a strong proteolytic bacterium Bacillus subtilis sp. TN51 from commercial thua