Improving the properties of chitosan as support for the covalent multipoint immobilization of chymotrypsin.
Changing gel structure and immobilization conditions led to a significant improvement in the covalent multipoint attachment of chymotrypsin on chitosan. The use of sodium alginate, gelatin, or kappa-carrageenan, activation with glutaraldehyde, glycidol, or epichlorohydrin, and addition of microorganisms followed by cellular lysis allowed the modification of the gel structure. Immobilization yields, recovered activities, and stabilization factors at 55 and 65 degrees C were evaluated. Enzyme immobilization for 72 h at pH 10.05, 25 degrees C and reduction with NaBH 4 in chitosan 2.5%-carrageenan 2.5%, with addition of S. cerevisiae 5% and activation with epichlorohydrin led to the best derivative, which was 9900-fold more stable than the soluble enzyme. This support allowed an enzyme load up to 40 mg chymotrypsin x g gel (-1). The number of covalent bonds, formed by active groups in the support and lysine residues of the enzyme, can explain the obtained results. SEM images of the gel structures corroborate these conclusions.
Adriano
Wellington S
WS
Chemical Engineering Department, Federal University of Sao Carlos, UFSCar, Brazil.
Mendonça
Dany B
DB
Rodrigues
Dasciana S
DS
Mammarella
Enrique J
EJ
Giordano
Raquel L C
RL
D000464:Alginates / Q000737:chemistry; D000818:Animals; D002351:Carrageenan / Q000737:chemistry; D002417:Cattle; D048271:Chitosan / Q000737:chemistry*; D002918:Chymotrypsin / Q000737:chemistry*; D004800:Enzymes, Immobilized / Q000737:chemistry; D004811:Epichlorohydrin / Q000737:chemistry; D004852:Epoxy Compounds / Q000737:chemistry; D005780:Gelatin / Q000737:chemistry; D020723:Glucuronic Acid / Q000737:chemistry; D005976:Glutaral / Q000737:chemistry; D006603:Hexuronic Acids / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D008239:Lysine / Q000737:chemistry; D020005:Propanols / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013696:Temperature
D000464:Alginates; D004800:Enzymes, Immobilized; D004852:Epoxy Compounds; D006603:Hexuronic Acids; D020005:Propanols; D004811:Epichlorohydrin; D020723:Glucuronic Acid; D002351:Carrageenan; D005780:Gelatin; D048271:Chitosan; D002918:Chymotrypsin; D008239:Lysine; C004312:glycidol; D005976:Glutaral