Production and characterization of a novel high viscosity polysaccharide(Methylan) from methanol by methanol by methylobacterium organophilum

Abstract

A new anionic high molecular weight exopolysaccharide, Methylan, was produced by Methylobacterium organophilum from methanol as a sole source of carbon and energy under the specific culture conditions. For production of the polysaccharide, effects of medium components and other substrates such as sugars and TCA-intermediates with methanol were investigated in batch cultures. The polysacchride production was significantly influenced by the concentration and source of nitrogen. Higher cell growth rate and polysaccharide productivity were achieved with inorganic nitrogen source such as ammonium sulfate. The optimum C/N ratio for polysaccharide production was obtained around 30 with ammonium sulfate as a nitrogen source. When the requirement of metal ions was examined, $Mn^{+2}$ was found to be essential for polysaccharide production. With a deficiency of $Mn^{+2}$, polysaccharide was not accumulated at all. Compared to the case where methanol was the only carbon and energy source, the polysaccharide production was enhanced upto two folds, when glucose malate and succinate, TCA-intermediates were added to the medium. To prevent and overcome the methanol inhibition, and to improve the polysaccharide production, fed-batch cultures with different feeding modes (constant, intermittent and exponential feeding) were investigated. In terms of productivity, there was no significant difference among three different feeding modes of operation. The polysaccharide production was enhanced upto in a range from 0.7 to 1.1 g/l. Supplement of glucose as a mixed substrate, addition of $Fe^{++}$ and $Mo^{++}$ for nitrogen fixation, and condition for overcoming the nitrogen regulation were carried out for mass production of the polysaccharide with intermittent feeding fed-batch fermentation using DO-stat technique. Under the above conditions, the polysaccharide was produced in a range from 3 to 4 g/l with the cell concentration between 6 to 7.5 g/l. Nitrogen regulation in th...