Multipoint covalent immobilization of microbial lipase on chitosan and agarose activated by different methods

• Published in: Journal of molecular catalysis. B, Enzymatic Enzymatic, 2008-04, Vol.51 (3), p.100-109
• Main Authors: Rodrigues, Dasciana S., Mendes, Adriano A., Adriano, Wellington S., Gonçalves, Luciana R.B., Giordano, Raquel. L.C.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Candida antarctica type B lipase; Chitosan; Enzyme immobilization; Enzyme stabilization; Fundamental and applied biological sciences. Psychology; Immobilization of enzymes and other molecules; Immobilization techniques; Methods. Procedures. Technologies
• ISSN: 1381-1177; 1873-3158
• DOI: 10.1016/j.molcatb.2007.11.016

In this work Candida antarctica lipase type B (CALB) was immobilized on agarose and chitosan. The influence of activation agents (glycidol, glutaraldehyde and epichlorohydrin) and immobilization time (5, 24 and 72 h) on hydrolytic activity, thermal and alkaline stabilities of the biocatalyst was evaluated. Protein concentration and enzymatic activity in the supernatant were determined during the immobilization process. More active derivatives were attained when the enzymatic extract was first purified through dialysis. The highest activities achieved were: for agarose-glyoxyl (with glycidol), 845 U/g of gel, after 72 h of immobilization; for chitosan-glutaraldehyde and agarose-glutaraldehyde, respectively, 1209 U/g of gel and 2716 U/g of gel, after 5 h of immobilization. Thermal stability was significantly increased, when compared to the soluble enzyme: 20-fold for agarose-glyoxyl (with glycidol)-CALB, 18-fold for chitosan-glutaraldehyde-CALB and 21-fold for agarose-glutaraldehyde. The best derivative, 58-fold more stable than the soluble enzyme, was obtained when CALB was immobilized on chitosan activated in two steps, using glycidol and glutaraldehyde, 72 h immobilization time. The stabilization degree of the derivative increased with the immobilization time, an indication that a multipoint covalent attachment between enzyme and the support had really occurred.