Optimization of Oligosaccharide Production from Using a Response Surface Methodology and the Immunostimulating Effects of These Oligosaccharides on Macrophage Cells.

Production of oligosaccharides from CCK940 was optimized using a response surface methodology with a central composite design. Culture temperature and the concentrations of sucrose and maltose were used as the main factors. The predicted optimum conditions for the production of oligosaccharides were a culture temperature of 30 °C, a sucrose concentration of 9.6% (/), and a maltose concentration of 7.4% (/). Using these optimal conditions, CCK940 was cultured using a fermenter to produce oligosaccharides, and the resulting oligosaccharides with a degree of polymerization greater than 4 were purified by Bio-gel P2 gel permeation column chromatography and then lyophilized. When macrophages were treated with the purified oligosaccharides at concentrations of 0.1⁻10 mg/mL, no cytotoxicity towards the macrophages was observed. However, nitric oxide production levels were similar to those following treatment with 1 μg/mL lipopolysaccharide. The mRNA expression levels of tumor necrosis factor-α, interleukin-1β, interleukin-6, and inducible nitric oxide synthase were all also increased in a dose-dependent manner following treatment with the oligosaccharides. These data suggest that oligosaccharides produced by CCK940 could be used as an immune enhancer of macrophages.