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Journal September 04 2011.Pmd REVIEW ARTICLE J. Food Sci. Technol. Nepal, Vol. 6 (1-9), 2010 ISSN: 1816-0727 Bacillus Fermentation of Soybean: A Review ASHOK KUMAR SHRESTHA1*, NAWA RAJ DAHAL2 and VEDASTE NDUNGUTSE1 1Centre for Nutrition and Food Sciences, The University of Queensland, St Lucia, Queensland 4072, Brisbane, Australia 2Department of Food Technology and Quality Control, Kathmandu, Nepal Soybeans in its natural form have a little direct use as a food due to its poor digestibility as well as beany taste and flavour. Fermentation; however, can improve the eating and nutritional qualities of soybeans. Fermented soybean foods have been an intricate part of oriental diet for a long time. Bacillus subtilis dominated traditionally fermented soyfoods have typical taste, texture and aroma which is popular in Asian and African countries. B. subtilis fermentation of soaked and cooked soybeans brings many physico-chemicals and sensory changes that make it highly digestible and nutritious. This paper reviews various facets of B. subtilis fermented traditional foods, properties of fermenting organisms, preparation of such fermented foods, changes in chemical composition and nutritional properties and improving the quality of these foods. Keywords: Soyabean, Bacillus fermentation, Health benefits, Acceptability, Commercialization Introduction like fermented soy foods reported in literature are: Nattoin Japan, Thua-Nao in Thailand, Douchi in China, Chung- Soybean (Glycine max L.) is one of the nutritionally richest kook-jong in Korea, Tao-si in Philippines, Dawadawain natural vegetable foods known to human kind, records of its Africa (Sarkar et al., 1994; Shrestha and Noomhorm, 2001; Hu food usage dates back to 2838 BC in China. Although having et al., 2010). With its powerful odour and slimy appearance high protein content, minerals, vitamins and bio-actives, it like a rotten food, large number of population do not like it, has little direct use because of high satiety value caused by whereas other finds eating Kinema (and similar products) a high oil content, poor digestibility, green beany taste, long delightful experience. These foods are eaten in the fresh form cooking time and persistent bitterness. Soybeans as a food is or as a fried curry dish along with boiled rice and sometime used as soymilk, soy flour, soy oil, feed for livestock and with boiled rice, and sometimes as soup, pickle or mixed with poultry, soy concentrate, protein isolates, soy yoghurt, tofu other vegetables (Shrestha, 1989). and fermented foods such as Tempeh, soy sauce, Miso , Natto and Sufu (Beuchat, 1984). Fermentation is a proven method Bacillus spp. is the most dominant naturally fermenting agents to improve flavour, texture and nutritional quality of the in soybeans. These hydrolytic bacteria are associated with soybeans. Besides bringing physico-chemical and sensory utilization and reduction of indigestible oligosaccharides and quality changes, fermentation contributes towards the polysaccharides. The organism has also shown to reduce the preservation of food due to release of metabolites that activity of anti-nutrients that hinders availability of proteins discourage the growth of pathogenic bacteria in foods. and phytochemicals present in soybeans (Sarkar et al., 1997; Fermentation involves a range of microorganisms such as Esteves et al. 2010). B. subtilis fermentation is accompanied lactic acid bacteria, acetic acid bacteria, yeasts, moulds and a by covering intact granules with white-coloured viscous range of bacteria. It also covers wide range of products such substance, slimy appearance, softer texture, and unique as staples, adjuncts to staples, condiments and beverages rotten flavour. It also completely removes the beany odour that use substrates such as cereals, pulses, soybeans, flowers, of raw soybeans and increases sensory quality of the product milk, meat etc. (Tamang and Sarkar, 1988). (Sarkar and Tamang, 1994; Shrestha and Noomhorm, 2001; Hu et al., 2010). The current paper will review various aspects There are many traditional B. subtilis fermented soybeans of Bacillus fermentation of soybeans, particularly chemical foods in various parts of the world. One of the common changes that lead to improvement of nutritional and sensory examples is Kinema which is traditionally consumed by the qualities of soybeans. It will also investigate the potential non-Brahmin Nepali inhabiting Nepal; Darjeeling and Sikkim benefits of Bacillus fermentation on human health, storability of India; and some parts of Bhutan. It is popular among and other aspects as a food. Lepchas and Bhutias who call it Satlyangserand Bari, respectively (Sarkar et al., 1994). Yoshida (1988) reported the Soybean fermentation and fermenting Bacilli origin of Kinema in Southern part of China. While it spread, Fermenting bacilli encompasses a group of bacteria with this food settled into a niche as seasonings in East Nepal, various similarities, or dissimilarities (Neway, 1989). They are North East India, Burma, Thailand and in Japan. Some Kinema mostly gram positive, aerobic, rodshaped, heat resistant and *Corresponding author, E-mail: [email protected] spore forming bacteria. Bacillus can produce different variety 1 Shrestha et al., : J. Food Sci. & Technol. Nepal, 6 (1-9), 2010 of enzymes including polysaccharides, proteins and nucleic acid hydrolysing enzymes during fermentation (Neway, 1989). B. subtilis is a group of related strains under the family Bacillaceae. It is also called hay Bacillus because it is easily isolated from hay by enrichment culture. This bacteria is largely non pathogenic and grows quickly in inexpensive media. When cultivated in liquid medium, organisms often accumulate on the surface, giving rise to a skin or pellicle. The colonies are irregular and have a curled or hair like structure giving what is sometimes called "Medusa head" appearance. On microscopic examination tangled coils of long chains of bacilli may be found (Burrows, 1973). Priest (1989) reported the amylolytic, proteolytic and lipolytic (de- esterification) activity of this organism. It has ability to secrete high levels of protein into the growth medium. Marshall (1990) reported B. subtilis as one of the few organisms that has been used to remove the bitter taste of soybeans. The lack of pathogenecity among strains of B. subtilis and any of its close relatives has resulted Food and Drug Administration granted the organism as GRAS (Generally Regarded as Safe). The genus B. subtilis is the most dominant bacterium Figure 1. Flow chart of traditional fermentation of B. naturally present in raw soybeans (Sarkar and Tamang, 1995). There are a number of naturally fermented soybeans foods subtilis fermented soybeans (Shrestha, 1989) where this species grow without adding inoculum. Some examples of these soy based foods are: Japanese Hikiwari- Method involves wrapping soaked-boiled soybeans by Natto, Itohiki-Natto;Thai Thua-Nao;Chinese Shui-Toushi; banana (Musa paradisica L.) or Smith leaves (Leucosceptrum Nigerian Daddawa or Dawadawa; and other legume/oilseed canum) or even polythene bags in bamboo lined basket which based fermented products from different regions of Africa is covered with sack clothe and fermented in warm place for 2 (Shrestha, 1997). These traditionally fermented soyfoods are to 3 days (Tamang and Sarkar, 1988). littered with several other microbial species such as Often a little bit of firewood ash is added to boiled and crushed Enterococcus faecium, Candia parapsilosis, Geotrichum soybeans before final fermentation commences. The candidum etc., but Bacillus remains the sole organism fermentation method also varies, for example, in more carrying out the fermentation (Sarkar and Tamang, 1994). sophisticated methods pure culture is used to inoculate the Because soybeans fermentation is still mainly based on boiled soybeans. Shrestha and Noomhorm (2001) isolated B. traditional way of preparation where it is spontaneous, end subtilis from Kinema and hygienically prepared Kinema by products will be different depending on microbial species using the pure culture. In this method soaked beans were that are predominant during fermentation. It will also be autoclaved at 121°C for 35 min, cooled to 30 to 35°C, inoculate different from one region to another and country-to-country with pure culture, ferment at 37°C for 48 hours at 85% RH. Hu depending on fermentation methods. However, where a single et al. (2010) and Chukeatirote et al. (2010) also prepared Thua- species have been used either for experimental or for Nao using pure culture of B. subtilis. Efforts have been made standardisation purposes, production of consistent products to increase the shelf life (2-5 days) by drying fresh product have been obtained and this will be considered in the next into small balls or ground these dry beans into powders sections. (Shrestha and Athapol, 2001; Chukeatirote et al., 2010). Sarkar Methods of soybeans fermentation and colleagues have extensively studied the influence of process variables and inoculum composition on the sensory Most of the Bacillus fermented soy products are still prepared as well as product qualities (Sarkar et al., 1993; Sarkar et al., by traditional methods. So the method of preparation depends 1994; Sarkar et al., 1994; Sarkar and Tamang, 1995). on countries/locality, family convenience, and materials Ikemnebonech, (1986) reported a number of factors such as, available. The basic steps of product preparation such as, water activity, salt concentration, pH and temperature have soaking,
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