Catalytic Efficiency Of Lipase Research Articles

Lipase catalyzed esterification of formic acid in solvent and solvent-free systems

Abstract Esterification reaction between formic acid and alcohols (C4, C8) catalyzed by lipases was performed in solvent and solvent-free systems. High Performance Liquid Chromatography (HPLC) was used to study the kinetic parameters (Michaelis-Menten) and the reaction conditions by monitoring the ester synthesis. The optimal conditions for formate ester synthesis were found to be: 0.5 M of formic acid, 1.5 M of alcohol, 2% of Novozym 435 at 40 °C and 400 rpm in acetonitrile solvent without molecular sieves. In solvent-free system, the optimal conditions were found to be: 1 M of formic acid, 10 M of alcohol, 2% of Novozym 435 at 40 °C and 400 rpm without molecular sieves. These optimal conditions resulted in maximum ester yield about 90% for 8 h in solvent system and 5 h in solvent-free system. Among all esterification reactions, the esterification reaction of formic acid with octan-1-ol in solvent-free system was the better regarding the quantity of esterified formic acid, catalytic efficiency of lipase and the required reaction time. Under these optimal conditions, the biocatalyst could be reused for six reaction cycles keeping around 97% of its initial activity after treatment process with n-hexane. Consequently, this enzymatic procedure in solvent-free system could provide formate esters for food and cosmetic applications.

Study of a new spectrophotometric end-point assay for lipase activity determination in aqueous media

A new spectrophotometric end-point method to evaluate lipase activity in aqueous media is described. This procedure is based on blocking the enzyme reaction by adding chloroform:isoamyl alcohol (24:1) as a denaturating agent, and removing the precipitate by centrifugation. Emulsifier screening showed that the chemical component with the least effect on lipase activity from Thermomyces lanuginosus, Mucor javanicus, Aspergillus niger, Rhizomucor miehei, Penicillium camemberti and Burkholderia cepacia, in the proposed method, was 0.1% (w/w) gum arabic. In contrast, Triton X-100, SDS, sodium cholate and Tween 80 reduced lipase activity, depending on the microbial source of the latter. A comparative study with other end-point methods based on the addition of chemical reagents (NaOH, Na2CO3, THAM or acetone–ethanol) or in thermal treatments (chilling or heating) was performed. The proposed procedure was shown to be more accurate in comparison with other methods that were tested. Furthermore, the effectiveness of the method was demonstrated in a study of the specificity of six commercial lipases toward p-nitrophenyl decanoate (C10) and p-nitrophenyl palmitate (C16). The study also analyzed the kinetic behavior and catalytic efficiency of lipase from T. lanuginosus and B. cepacia toward p-nitrophenyl decanoate and p-nitrophenyl palmitate.

The catalytic efficiency of lipase in a novel water-in-[Bmim][PF6] microemulsion stabilized by both AOT and Triton X-100

In the water-in-[Bmim][PF6] microemulsion stabilized by both AOT and Triton X-100, the lipase-catalyzed hydrolysis of 4-nitrophenyl butyrate (p-NPB) was investigated to evaluate the catalytic efficiency of lipase in this novel microemulsion. The structural parameters of the microemulsion and the conditions of the enzymatic reaction affect the catalytic activity of lipase, especially the concentration of Tris–HCl buffer. Under optimum conditions, the catalytic activity of lipase in the present microemulsion is much higher than that in H2O saturated [Bmim][PF6]. When the partitioning of the substrate in the microemulsion is taken into account, the catalytic efficiency of lipase in this novel microemulsion is 14.3 times that in H2O saturated [Bmim][PF6] due to the significant decrease of the Michaelis constant in the microemulsion. Due to the large interface, high water activity, and probably the activating effect of the imidazolium cation in the water pool, the present microemulsion is demonstrated to be a promising medium for the lipase-catalyzed hydrolytic reaction. To demonstrate an important biocatalytic application in the IL-based microemulsion, the lipase-catalyzed synthesis of the flavoring agent benzyl acetate via transesterification of vinyl acetate with benzyl alcohol was also studied in the medium. Due to the high dispersion of lipase, large interface and removal of the byproduct, a maximum yield of 94% was obtained, indicating that the novel microemulsion is really important and useful.

Role of the cosurfactant in water-in-oil microemulsion: interfacial properties tune the enzymatic activity of lipase

The enzymatic activity of Lipase VII from Candida rugosa was studied in the quaternary water-in-oil microemulsion cetyltrimethylammonium bromide (CTAB)/water/pentanol (POH)/hexane (C 6). The enzyme-catalyzed hydrolysis of p-nitrophenylbutirate ( p-NPB) was found essentially independent from the water/surfactant ratio. On the other hand, the enzyme kinetics was strongly affected by the cosurfactant/surfactant ratio. The interfacial composition and the reverse micelles size, as well as the substrate partition between organic bulk and surfactant aggregates, have been determined through pulsed gradient spin-echo NMR measurements. The correlation of the microemulsion structure with enzyme activity has therefore been possible. The lipase affinity for p-nitrophenylbutirate is lower in microemulsion compared to water because the hydrophobic effect is weaker in the former system. The turnover number is unaffected from the reverse micelles size and from the interfacial bromide concentration, while is strongly dependent on the interfacial composition. The data presented indicate a marked influence of the extent of interfacial surface on the catalytic efficiency of lipase.

The temperature-dependent interfacial inactivation of porcine pancreatic lipase Effect of colipase and bile salts

This paper confirms and extends the previous observation that colipase and bile salts stabilize pancreatic lipase against inactivation at its water/substrate interface. It is shown that colipase and bile salts above their critical micellar concentration offer better protection than either of them alone. Colipase has no effect on the catalytic efficiency of lipase against an emulsified substrate in the absence or presence of bile salts. Its reported activation of pancreatic lipolysis at high temperatures in the absence of bile salts is, most likely, fully explained by its protective effect on lipase inactivation. Colipase at high concentrations relative to lipase inhibits the enzyme activity in a competitive fashion. The temperature-dependent surface inactivation of lipase has certain consequences for the methodology of lipase activity determination.