Optimal Conditions for Continuous Immobilization of Pseudozyma hubeiensis (Strain HB85A) Lipase by Adsorption in a Packed-Bed Reactor by Response Surface Methodology
This study aimed to develop an optimal continuous process for lipase immobilization in a bed reactor in order to investigate the possibility of large-scale production. An extracellular lipase of Pseudozyma hubeiensis (strain HB85A) was immobilized by adsorption onto a polystyrene-divinylbenzene support. Furthermore, response surface methodology (RSM) was employed to optimize enzyme immobilization and evaluate the optimum temperature and pH for free and immobilized enzyme. The optimal immobilization conditions observed were 150？min incubation time, pH 4.76, and an enzyme/support ratio of 1282？U/g support. Optimal activity temperature for free and immobilized enzyme was found to be 68°C and 52°C, respectively. Optimal activity pH for free and immobilized lipase was pH 4.6 and 6.0, respectively. Lipase immobilization resulted in improved enzyme stability in the presence of nonionic detergents, at high temperatures, at acidic and neutral pH, and at high concentrations of organic solvents such as 2-propanol, methanol, and acetone. 1. Introduction Biocatalyst enzymes play an important role in biotechnological applications due to their extreme versatility with respect to substrate specificity and stereoselectivity and exhibit many other features that render their use advantageous when compared to conventional chemical catalysts. As an example, fat and oil hydrolysis using NaOH as a catalyst requires high pressure and temperature to achieve high efficiency (97-98%); in contrast, the same process can be carried out effectively at normal temperature and pressure using lipases, with significant decrease in wastewater production. Lipases (triacylglycerol acylhydrolase; EC 3.1.1.3) catalyze the hydrolysis of triglycerides to glycerol and fatty acids, as well as a variety of reactions in nonaqueous medium (e.g., transesterification, esterification, and interesterification). Such enzymatic properties allow a series of biotransformation reactions that lead to multiple industrial applications in foods, flavors, pharmaceuticals, detergent formulation, oil/fat degradation, cosmetics, and environmental remediation [1–4]. However, soluble enzymes usually exhibit lower stability than chemical catalysts and often cannot be recovered and reused. This severely hinders their application in practice. Nevertheless, this problem can be overcome by enzyme immobilization, which enhances thermal and operational stabilities, ease of handling, and prevention of aggregation and autolysis. Besides, immobilized lipases (IE) on solid support allow recoverability and reuse thus significantly
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