Evaluation of the catalytic activity of lipases immobilized on chrysotile for esterification

Silva, Jane E. S.; Jesus, Paulo César de

doi:10.1590/s0001-37652003000200003

Cited by 24 publications

(12 citation statements)

References 16 publications

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“…After 3 h ϳ10 to 20% of the alcohols was converted to esters by 3 mg whole-cell biocatalyst (Table 2). These figures are comparable to those obtained for other immobilized lipases (20) and suggest that the displayed ROL could be used as a whole-cell lipase.…”

Section: Discussionsupporting

confidence: 84%

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Enhanced Reactivity of Rhizopus oryzae Lipase Displayed on Yeast Cell Surfaces in Organic Solvents: Potential as a Whole-Cell Biocatalyst in Organic Solvents

Shiraga

Kawakami

Ishiguro

et al. 2005

Appl Environ Microbiol

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Immobilization of enzymes on some solid supports has been used to stabilize enzymes in organic solvents. In this study, we evaluated applications of genetically immobilized Rhizopus oryzae lipase displayed on the cell surface of Saccharomyces cerevisiae in organic solvents and measured the catalytic activity of the displayed enzyme as a fusion protein with ␣-agglutinin. Compared to the activity of a commercial preparation of this lipase, the activity of the new preparation was 4.4 ؋ 10 4 -fold higher in a hydrolysis reaction using p-nitrophenyl palmitate and 3.8 ؋ 10 4 -fold higher in an esterification reaction with palmitic acid and n-pentanol (0.2% H 2 O). Increased enzyme activity may occur because the lipase displayed on the yeast cell surface is stabilized by the cell wall. We used a combination of error-prone PCR and cell surface display to increase lipase activity. Of 7,000 colonies in a library of mutated lipases, 13 formed a clear halo on plates containing 0.2% methyl palmitate. In organic solvents, the catalytic activity of 5/13 mutants was three-to sixfold higher than that of the original construct. Thus, yeast cells displaying the lipase can be used in organic solvents, and the lipase activity may be increased by a combination of protein engineering and display techniques. Thus, this immobilized lipase, which is more easily prepared and has higher activity than commercially available free and immobilized lipases, may be a practical alternative for the production of esters derived from fatty acids.Lipases (EC 3.1.1.3) catalyze the hydrolysis of acylglycerides and other fatty acid esters under aqueous conditions and the synthesis of esters in organic solvents (5, 25). Lipase-catalyzed reactions have high substrate specificity or regioselectivity, so lipase-catalyzed ester synthesis in organic solvents is potentially an industrially feasible alternative to conventional chemical methods (1, 4). However, organic solvents may alter the structure and activity of enzymes that usually function in an aqueous environment.One alternative is to immobilize the enzymes on solid supports in order to increase their function and stability in response to organic solvents or increased temperatures. Enzymes may be stabilized by chemical and physical processes (6,15,20,24). With chemical methods enzymes are immobilized by strong covalent bonding, but changes in protein structure often result. In physical stabilization processes, the interactions between enzymes and solids usually are weaker, resulting in fewer changes in the enzyme's structure.Yeast cell surface engineering (12, 13) is an alternative approach that immobilizes an enzyme on the yeast cell surface. Proteins are immobilized by using an outer shell cell wall protein, the C-terminal half of ␣-agglutinin (10, 26). The gene encoding Rhizopus oryzae lipase (ROL), which has been used to produce diesel fuels (8), was fused to a DNA sequence encoding the C-terminal half of yeast ␣-agglutinin and expressed in Saccharomyces cerevisiae. The transformants have hydrolytic...

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Section: Discussionsupporting

confidence: 84%

“…Enzymes may be stabilized by chemical and physical processes (6,15,20,24). With chemical methods enzymes are immobilized by strong covalent bonding, but changes in protein structure often result.…”

mentioning

confidence: 99%

Enhanced Reactivity of Rhizopus oryzae Lipase Displayed on Yeast Cell Surfaces in Organic Solvents: Potential as a Whole-Cell Biocatalyst in Organic Solvents

Shiraga

Kawakami

Ishiguro

et al. 2005

Appl Environ Microbiol

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“…The immobilization of Mucor javanicus lipases onto chrysotile (magnesium silicate) decreased the yield by 25% after its first reuse and by more than 70% after four reuse cycles (Silva & Jesus, 2003). On contrary, the covalent immobilization of Rhizopus oryzae lipases onto silica gel decreased the residual activity by only 20% after twenty reuse cycles (Lee et al, 2006).…”

Section: Reusability Of the Immobilized Lipasementioning

confidence: 99%

Production, Characterization and Immobilization of a Fusarium solani Lipase by Chitosan Magnetic Nanoparticles

Elsayed

Eweda

EI-Tayeb

et al. 2016

Egypt. J. Microbiol.

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IPASE producing fungus was isolated and identified as a strain of Fusarium solani based on its 18s rDNA sequence. The enzyme it produces was purified by diethyl amino ethyl sephadex (DEAEsephadex) column chromatography. The specific activity of the pure enzyme was 1.98 U/mg protein. The kinetics study showed that K m and V max values were 0.63µM and 29.4µM/min/mg protein, respectively. The MW was 95.27 kDa. Effects of pH, incubation temperature and organic solvents on the lipase activity were studied. The maximum enzyme activity was obtained at pH 8.5 and incubation temperature 35°C. Hexane and butanol inhibited enzyme activity by 51% and 72.6 %, respectively, while DMSO stimulated the activity by 47.8%. The lipase was immobilized by fusion to chitosan-coated iron oxide magnetic nanoparticles and cross-linked by glutaraldehyde. The reusability and storage period of the immobilized enzyme showed that the enzyme retained 80% of its activity after 15 reuse cycles and retained 97% of activity after 30 days of storage at 4°C. The immobilized lipase was tested for synthesis of sugars-oleate esters and the ester products were analyzed by liquid chromatography tandemmass spectrometry (LC/MS/MS). This investigation identified the potential for use of the obtained F. solani lipase in industrial applications that utilize organic solvents or alkaline pH values, such as detergent industry.

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“…3.1.1.3) are widely used enzymes due to their ability to recognize several substrates and to catalyze many different reactions (Barros et al, 2010;Rodrigues and Fernandez-Lafuente, 2010a). In particular, several flavors (short and long chain aliphatic esters) of industrial interest have been obtained by esterification, transesterification or interesterification reactions using lipases (Gandhi, 1997;Alcantara et al, 1998;Abbas and Comeau, 2003;Silva and Jesus, 2003;Hasan et al, 2006), with applications in juices, cheeses, baked goods, candies and beverages, for instance (Mahapatra et al, 2009). The demand for these chemicals has continuously increased at the rate of 4.3% per year (Dhake et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

Stability of immobilized Rhizomucor miehei lipase for the synthesis of pentyl octanoate in a continuous packed bed bioreactor

Skoronski

Padoin

Soares

et al. 2014

Braz. J. Chem. Eng.

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-The enzymatic synthesis of organic compounds in continuous bioreactors is an efficient way to obtain industrially important chemicals. However, few works have focused on the study of the operational conditions and the bioprocess performance. In this work, the aliphatic ester pentyl octanoate was obtained by direct esterification using a continuous packed bed bioreactor containing the immobilized enzyme Lipozyme ® RM IM as catalyst. Enzymatic deactivation was evaluated under different conditions for the operational parameters substrate/enzyme ratio (5.00, 1.67, 0.83 and 0.55 mmol substrate ·min -1 ·g -1 enzyme ) and temperature (30, 40, 50 and 60 °C). The optimal condition was observed at 30 ºC, which gave the minimum enzymatic deactivation rate and the maximum conversion to the desired product, yielding approximately 60 mmols of ester for an enzyme loading of 0.5 g into the bioreactor. A first-order deactivation model showed good agreement with the experimental data.

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Evaluation of the catalytic activity of lipases immobilized on chrysotile for esterification

Cited by 24 publications

References 16 publications

Enhanced Reactivity of Rhizopus oryzae Lipase Displayed on Yeast Cell Surfaces in Organic Solvents: Potential as a Whole-Cell Biocatalyst in Organic Solvents

Enhanced Reactivity of Rhizopus oryzae Lipase Displayed on Yeast Cell Surfaces in Organic Solvents: Potential as a Whole-Cell Biocatalyst in Organic Solvents

Production, Characterization and Immobilization of a Fusarium solani Lipase by Chitosan Magnetic Nanoparticles

Stability of immobilized Rhizomucor miehei lipase for the synthesis of pentyl octanoate in a continuous packed bed bioreactor

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