Immobilization of yeast inulinase on chitosan beads for the hydrolysis of inulin in a batch system

- Extracellular inulinase was partially purified from Kluyveromyces marxianus YS-1.
- Partially purified enzyme was immobilized on glutaraldehyde activated chitosan beads.
- Free and immobilized inulinase was used for the hydrolysis of inulin in a batch system.
- To check the operational stability of the immobilized biocatalyst, recycling of the developed immobilized biocatalyst was carried out for inulin hydrolysis.
- The developed immobilized enzyme was successfully used for hydrolysis of inulin for 14 batches.

An extracellular inulinase was partially purified by ethanol precipitation and gel exclusion chromatography from a cell free extract of Kluyveromyces marxianus. Partially purified inulinase exhibited 420 IU/mg specific activity and it was immobilized on chitosan beads. Activity yield of immobilized inulinase was optimized with glutaraldehyde concentration (1-5%), glutaraldehyde treatment time (30-240 min), enzyme coupling-time (2-16 h) and enzyme loading (5-30 IU) as functions. Under the optimized conditions maximum yield 65.5% of immobilized inulinase was obtained. Maximum hydrolysis of inulin 84.5% and 78.2% was observed at 125 rpm after 4 h by immobilized and free enzyme, respectively. A retention-time of 4 h and 5 h was found optimal for the hydrolysis of inulin under agitation (125 rpm) by free and immobilized enzyme, respectively. The recycling of the developed immobilized biocatalyst was carried out after 5 h of inulin hydrolysis in a batch system. The developed immobilized biocatalyst was successfully used for the hydrolysis of inulin for 14 batches. This is the first report on the immobilization of yeast inulinase on chitosan beads for the hydrolysis of inulin in a batch system.