Fig. 3

Determination of optimal pH a and temperature b for sucrose conversion using crude InuGB-V3. Sucrose transfructosylation and hydrolysis of sucrose by crude inulosucrase were investigated after 24 h of incubation under varied conditions. Activity assays were performed in biological triplicates at a 1 mL scale. The reactions were started by adding 4000 U L⁻¹ crude inulosucrase to prewarmed solutions of 570 g L⁻¹ sucrose supplemented with 1 mM CaCl₂. a The influence of pH on crude InuGB-V3 was investigated by using 25 mM sodium citrate buffer (pH 2.5–3.5), 25 mM sodium acetate buffer (pH 3.5–5.5), or 25 mM potassium phosphate buffer (pH 5.5–8.0) at an incubation temperature of 37 °C. To determine the influence of the buffer substances on the activity, one test was carried out with sodium citrate buffer and one with sodium acetate buffer at pH 3.5. The same applied to the determination of activity at pH 5.5, where a sodium acetate buffer and a potassium phosphate buffer were used. b Reactions to investigate the influence of temperature were performed at 25–65 °C incubation temperature using 25 mM sodium acetate buffer (pH 5.5). Sucrose and fructose concentrations were determined via isocratic HPLC using 65% [v/v] acetonitrile at a flow rate of 1 mL min⁻¹