Rhizomucor miehei, Humicola sp., Rhizopus niveus, and Candida antarctica B lipases were immobilized by physical adsorption onto a macroporous polypropylene support. In an esterification reaction, the enzyme efficiency, and therefore cost-effectiveness, is greatly affected by enzyme loading, with an apparent suppression of efficiency at low lipase loadings for both R. miehei and Humicola sp. lipases. This results in the appearance of a pronounced maximum in the efficiency-loading relationship at approximately 100,000 lipase units (LU)/g for R. miehei lipase (10% of its saturation loading) and at approximately 200,000 LU/g for Humicola sp. lipase (50% of its saturation loading). The other lipases studied do not show similar trends. At low loadings, only a small portion of the surface area is occupied and gives the lipase the opportunity to spread; it is hypothesized that the reduction in efficiency at low loadings is due to a distortion of the active molecular conformation caused by the lipase maximizing its contact with the support as a result of its high affinity for the support surface. The relationship between efficiency and loading was different for each of the lipases studied, which may reflect both differences in the strength of the affinity of the lipase for the support and in the ease at which the molecular conformation of the lipase can be distorted.
The rates of synthesis of dodecyl decanoate in hexane have been measured as a function of water activity (a,), for various immobilised preparations of the lipases from Rhizomucor miehei and Candida rugosa. Only very large changes in the amount of enzyme adsorbed to the support affect the shape of the ratela, profile; at the highest loadings the profiles tend to become somewhat flatter. A similar levelling can be obtained by pre-adsorbing an inert protein. The effect is probably due to adjacent protein molecules effectively replacing water ; it does not simply reflect mass transfer or interfacial area limitation. The activityla, profile was essentially the same with most supports tested : polypropylene, anion-exchange resin, celite, anion-exchange modified silica. A hydrophobic porous glass support reduced the rate somewhat at intermediate a, values with both enzymes; a polyamide material had this effect only with the lipase from Rh. miehei. The shape of the activityla, profile was not affected by large differences in purity of the lipase preparation, but did differ between forms that probably differ in glycosylation. Overall, relatively few manipulations of the system can significantly affect the shape of the ratela, profiles, which seem to be mainly an intrinsic property of the enzyme molecules used.
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