Activity of different Candida antarctica lipase B formulations in organic solvents.
The activity of different formulations of Candida antarctica lipase B (CALB), such as crude CALB, purified CALB, purified CALB lyophilized with PEG (CALB + PEG) or oleic acid (CALB + OA), and the commercial formulation Novozym 435, was determined in toluene, carbon tetrachloride, and 1,4-dioxane at various water activities (a(w)). The reaction between vinylacetate and 1-octanol was used as the model reaction and both transesterification (formation of 1-octylacetate) and hydrolytic (formation of acetic acid from vinylacetate) activities were determined. For equal amounts of lipase protein, CALB + PEG (and to a lesser extent CALB + OA) displayed higher activity than that of the other formulations; for instance, in toluene (a(w) < 0.1), it was 260-, 13-, and 1.8-fold more active than crude CALB, purified CALB, and Novozym 435, respectively. Moreover, the transesterification activity of CALB + PEG was of the same order of magnitude (51%) of the activity shown by the enzyme in the hydrolysis of vinylacetate in aqueous buffer. These results suggest that PEG and oleic acid could act as lyoprotectants, preventing the formation of intermolecular interactions during the lyophilization process that might be responsible for protein denaturation. No diffusional limitation was observed for CALB + PEG-catalyzed reactions. Purified CALB, in contrast to the other formulations, showed a marked activity increase (2.1 to 7.8-fold) as a function of a(w) and, in 1,4-dioxane, it was 3.5-fold more active when it was added to the solvent after previous dissolution of the lyophilized powder in water.
Secundo
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Istituto di Biocatalisi e Riconoscimento Molecolare, via Mario Bianco 9, 20131 Milano, Italy.
Carrea
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G
Soregaroli
C
C
Varinelli
D
D
Morrone
R
R
D002175:Candida / Q000201:enzymology*; D002251:Carbon Tetrachloride / Q000737:chemistry; D004146:Dioxanes / Q000737:chemistry; D004789:Enzyme Activation / Q000502:physiology; D004800:Enzymes, Immobilized; D004951:Esterification; D005612:Freeze Drying; D005656:Fungal Proteins / Q000737:chemistry; D006868:Hydrolysis; D008049:Lipase / Q000737:chemistry* / Q000378:metabolism*; D019301:Oleic Acid / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D014050:Toluene / Q000737:chemistry
D004146:Dioxanes; D004800:Enzymes, Immobilized; D005656:Fungal Proteins; D019301:Oleic Acid; D014050:Toluene; D011092:Polyethylene Glycols; D002251:Carbon Tetrachloride; C093667:Novozyme 435; D008049:Lipase