Figure 6.

Optimization of enzymatic hydrolysis conditions and analysis of the prepared AOS. (A) Effects of the E/S ratio and enzymolysis time on the AOS production. The effects were analyzed by measuring the content of reducing sugars released in the mixture using the DNS method. The graph shows data from triplicate experiments (mean ± standard deviation [S.D.]). (B) HPLC analysis of the degradation products of sodium alginate by AlyC7. The degradation reaction was performed at 30 °C for 90 min in a 200 µL mixture containing 0.5 M NaCl, 50 mM Tris-HCL (pH 9.0), 2 mg/mL sodium alginate, and AlyC7 with an E/S ratio of 50 U/mg. DP1 to DP6 represent mono-, di-, tri-, tetra-, penta-, and hexa-saccharide, respectively. Saturated mannuronate oligosaccharides were taken as the standards. (C) Analysis of the yield of trisaccharides in the degradation products. The inset is a standard curve correlating trisaccharide concentration to peak area, which was generated using commercial saturated mannuronate trisaccharide (DP3) at concentrations of 0.5–6 mg/mL as standards. Figures in (B,C) are representatives of triplicate experiments.