Improvement of vitamin B12 production by anaerobic ecosystem

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Although vitamin B12 has been purified from the cells of Propionibacterim spp.after fermentation of sugars, it is difficult to get a high cell concentration due to strong growth inhibition by propionate and acetate prouced. Acetogens produce acetate from methanol/CO2 as substrates and have a high B12 content in the cells. But acetate also inhibits the bacterial growth.Therefore, we attempted to improve these bacterial cultivations to get high concentration of B12 by using microbial ecosystem which can utilize these acids under anaerobic condition. The results were as follows.1.Cultivation of Propionibacterium freudenreichii in coculture with Paracoccus denitrificans or Rhodobacter sphaeroides under denitrification.(1) .In mixed culture with P.denitrificans, B12 concentration could increase ca.2-fold due to the selective utilization of propionate and acetate by P.denitrificans even in the presence of glucose.(2) .In mixed culture with R.sphaeroides, propionate and acetate were also util … More ized and N2 was produced. Therefore, B12 production increased 4-fold. But the cell concentration of P.freudenreichii was still 10 g/l due to imcomplete utilization of these acids by R.sphaeroides.2.Cultivation of Acetobacterium sp.in coculture with a methanogen, Methanothrix sp.(1) .Although B12 content in the cells of Acetobacterium sp.used was very high (5 mg/g cell) , acetate exhibited strong growth inhibition (Kp=270 mM) .(2) Coculture with Methanothrix sp.was carried out in separate bioreactor system, since growth rate of the methanogen was 10 times lower than Acetobacterium. First, a fixed bed reactor of the methanogen was set up, where polypropyrene/ceramic mixture was packed to adhere the methanogen cells. Then, this reactor was conected to the chemostat culture of Acetobacterium with liquid recycling. By using this combined reactor system where Methanothrix cells consumed acetate, acetate level in the chemostat culture could be kept almost zero and B12 productivity reached to 20 mg/l/day. Less