Amylose-like polysaccharide accumulation and hyphal cell-surface structurein relation to citric acid production by Aspergillus niger in shake culture

When 120 mg glucose/ml was used as a carbon source, in shake culture Asperg illus niger Yang no. 2 maximally produced only 15.4 mg citric acid/ml but a ccumulated 3.0 mg extracellular polysaccharide/ml. The polysaccharide secre ted by mycelia of Yang no. 2 in shake culture was confirmed to be an amylos e-like alpha-1,4-glucan by hydrolysis analysis with acid, amylase and gluco amylase. However, in static cultures, such as semisolid and surface culture s free from physical stresses caused by shaking damage, Yang no. 2 produced more citric acid but did not accumulate the polysaccharide. With cultivati on time in shake culture, the amount of extracellular polysaccharide and th e viscosity of the culture broth increased. The increase of shaking speed c aused a remarkable increase in the accumulation of extracellular polysaccha ride, e.g. 11.2 mg extracellular polysaccharide/ml was accumulated in the m edium at a shaking speed of 200 rpm. The addition of 2.0 mg carboxymethylce llulose (CMC)/ml as a viscous additive to the medium reduced drastically th e amount of extracellular polysaccharide accumulated to 1.5 mg/ml, but incr eased the citric acid produced to 52.0 mg/ml. However, intracellular polysa ccharide accumulation kept up a steady rate of 0.26 mu g/mg dried mycelium through the entire period of cultivation. The addition of 3.0 mg polysaccha ride/ml purified from the culture broth to the medium at the start of a cul ture resulted in a decrease of extracellular polysaccharide accumulation bu t an increase of citric acid accumulation. From electron-microscopic observ ation, cell surfaces of hyphae cultivated with CMC were smooth, while hypha e cultivated without CMC had fibrous and granular polysaccharide on the cel l surface. These results suggested that Yang no. 2 secreted the polysacchar ide on the cell surface as a viscous substance and/or a shock absorber to p rotect itself from physical stresses caused by shaking damage in shake cult ure.