Application of Invertase Immobilized on Chitosan Using Glutaraldehyde or Tris(Hydroxymethyl)Phosphine as Cross-Linking Agent to Produce Bioethanol

Invertase was immobilized on chitosan using glutaraldehyde or tris(hydroxymethyl)phosphine as cross-linker. The optimum pH for free and immobilized enzyme was found to be 4.5 and 5.5, respectively. The optimum hydrolysis temperature was 55 degrees C for both the free and immobilized forms. Km and Vmax values for free invertase, and invertase immobilized on glutaraldehyde- and THP-activated chitosan were 15, 19, and 20 mM, respectively, and 238, 204, and 212 mM min, respectively. The THP-immobilized enzyme had the highest pH and thermal stability, higher reusability with 70% retention in activity after 9 batches of reuse and higher storage stability with 90% retention in activity after 12 weeks at 4 degrees C, pH 4.5. Fermentation of cane molasses by yeast to form ethanol in the presence of free invertase at 30 degrees C, pH 5.0 led to an increase in ethanol production by 3% and the production increased by 10.7% when immobilized invertase was used as catalyst.