Response surface methodology: Synthesis of inulooligosaccharides with an endoinulinase from Aspergillus niger

Abstract

Oligosaccharides of chain length up to degree of polymerisation (DP) 5 have been synthesised using a purified endoinulinase and response surface methodology with central composite experimental design for the optimisation of the product yield. The endoinulinase was purified through a three-step procedure in a yield of 1.11% and 3.5-fold and its molecular weight was 68.1 kDa by SDS-PAGE. It had optimum pH and temperature values of 6 and 60 °C, respectively with only 10% loss in activity over 60 min under these conditions. From the kinetic studies K m and V max were estimated at 3.53 mM and 666.7 μmol min −1 ml −1, respectively. Using the purified endoinulinase preparation isolated from Aspergillus niger, at a temperature (60 °C), inulin concentration (150 mg ml −1), incubation time (48 h), pH (6.0) and enzyme dosage (60 U ml −1) produced high yields of inulotrioses (70.3 mM), inulotetraoses (38.8 mM), and inulopentaoses (3.5 mM). Response surface regression predicted similar product levels under the same conditions. A comparative analysis of the yields of individual oligosaccharides produced from both the crude and purified enzyme is also presented.