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Vitamin B12 Production in Tempeh by Non-sequential Mixed Fermentation at Laboratory and Bench Scales
Dissertation Abstract:
A study was conducted to produce tempeh with high vitamin B12 by incorporating selected strains of Propionibacterium shermanii. Soybeans of the Phraputthabath variety were soaked in acidified distilled water (1% of 80% lactic acid) for two hours at 70"C, dehulled by hand, and dried at 80°C until the beans were superficially dry. The soybean cotyledons were sterilized and inoculated with Rhizopus oligosporus powdered inoculum at a rate of 0.25g/50 g soybean and cell suspension of P. shermanii 1250 at 1 ml/50 g soybean. The mixture was incubated at 35°C for 24 hours.
Lactobacillus casei did not affect the vitamin B12 production of P. shermanii 1250 in tempeh when added to the soaked beans and also when added during the fermentation.
The optimum conditions for vitamin B12 production by P. shermanii 1250 in tempeh at the laboratory scale consisted of 55 percent bean moisture content (50 g in petri dish), mixed culture of 5 percent P. shermanii 1250 and 0.5 percent R. oligosporus inoculum, and incubation temperature of 35°C for 24-28 hours. The highest yield of vitamin B12 at optimum conditions was 308.4 :g/100g.
In general, compounds such as protein, fat, ash, and amino nitrogen increased during fermentation while carbohydates (by difference), soluble carbohydate, and fiber decreased as fermentation time progressed. In addition, concentration of most of the amino acids, especially lysine and methionine, was higher in vitamin B12-tempeh than in the unfermented soybean.
The optimum conditions used for bench scale production were similar to those for the laboratory scale. Bench scale production of high vitamin B12-tempeh, performed in stainless steel trays (30 cm x 45 cm x 3 cm), yielded 297.47 ng vitamin B12/100 g of mixture at optimum conditions of two em thickness of sterilized soybean mass (1,400 g/tray) covered with non-perforated aluminum foil, and incubation period of 24 hours at 35°C. The chemical composition of high vitamin B12-tempeh produced in trays at optimum conditions was also similar to that produced in laboratory scale.
The mass balance in the production of high vitamin B12-tempeh in the tray at optimum conditions showed that 30.14 percent of total solids were lost during processing. Some intermediate metabolites involved in the pathway of vitamin B12 by P. shermanii 1250 in tempeh (as detected by gas chromatography) included acetic acid, which increased, and propionic acid which decreased as fermentation time progressed. Moreover, succinic acid and glycine decreased during fermentation.
This research was the first to elucidate that mixed fermentation with R. oligosporus and P. shermanii 1250 enhanced high vitamin B12 production in tempeh. Non-sequential mixed fermentation which differed from the ordinary mixed fermentation was demonstrated.
Several advantages in tempeh fermentations were found. namely: 1) sporulation of Rhizopus, which affected the texture and appearance of tempeh. was reduced; 2) the ammonia smell normally found in high amount of traditional tempeh, which makes the product unacceptable especially to Westerners, was also reduced during fermentation; 3) Propionibacterium, which is normally used as starter in cheese-making, provides a favorable flavor acceptable to non-Asians. The product is acceptable to both Orientals and Westerners because of the effect of this mixed fermentation process.