Glutaraldehyde activation of polymer Nylon-6 for lipase immobilization: Enzyme characteristics and stability
ABSTRACT An extracellular alkaline lipase of a thermo tolerant Bacillus coagulans BTS-3 was immobilized onto glutaraldehyde activated Nylon-6 by covalent binding. Under optimum conditions, the immobilization yielded a protein loading of 228 microg/g of Nylon-6. Immobilized enzyme showed maximum activity at a temperature of 55 degrees C and pH 7.5. The enzyme was stable between pH 7.5-9.5. It retained 88% of its original activity at 55 degrees C for 2h and also retained 85% of its original activity after eight cycles of hydrolysis of p-NPP. Kinetic parameters Km and Vmax were found to be 4mM and 10 micromol/min/ml, respectively. The influence of organic solvents on the catalytic activity of immobilized enzyme was also evaluated. The bound lipase showed enhanced activity when exposed to n-heptane. The substrate specificity of immobilized enzyme revealed more efficient hydrolysis of higher carbon length (C-16) ester than other ones.
SourceAvailable from: Deniz Yildirim[Show abstract] [Hide abstract]
ABSTRACT: Pseudomonas sp. lipase was immobilized onto glutaraldehyde-activated Florisil (R) support via Schiff base formation and stabilized by reducing Schiff base with sodium cyanoborohydride. The immobilization performance was evaluated in terms of bound protein per gram of support (%) and recovered activity (%). A 4-factor and 3-level Box-Behnken design was applied for the acylation of (+/-)-2-(propylamino)-1-phenylethanol, a model substrate, with vinyl acetate and the asymmetric acylations of other (+/-)-2-amino-1-phenylethanols with different alkyl substituents onto nitrogen atom such as (+/-)-2-(methylamino)-1-phenylethanol, (+/-)-2-(ethylamino)-1-phenylethanol, (+/-)-2-(butylamino)-1-phenylethanol and (+/-)-2-(hexylamino)-1-phenylethanol were performed under the optimized conditions. The optimal conditions were bulk water content of 1.8%, reaction temperature of 51.5 degrees C, initial molar ratio of vinyl acetate to amino alcohol of 1.92, and immobilized lipase loading of 47 mg mL(-1). (R)-enantiomers of tested amino alcohols were preferentially acylated and the reaction purely took place on the hydroxyl group of 2-amino-1-phenylethanols. The increase of alkyl chain length substituted onto nitrogen atom caused an increase in the acylation yield and ee values of (S)-enantiomers. Enantiomeric ratio values were >200 for all the reactions. Our results demonstrate that the immobilized lipase is a promising biocatalyst for the preparation of (S)-2-amino-1-phenylethanols and their corresponding (R)-esters via O-selective acylation of (+/-)-2-amino-1-phenylethanols with vinyl acetate.PROCESS BIOCHEMISTRY 05/2013; 48(5-6):819-830. DOI:10.1016/j.procbio.2013.04.019 · 2.52 Impact Factor
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ABSTRACT: Pectinase extracted from Aspergillus niger was immobilized on a chitosan-coated chitin support using various methods: immobilization by adsorption (P-QQSA), adsorption on supports activated by 0.5 and 15% glutaraldehyde (w/v) (P-QQSA 0.5 and P-QQSA 15) and covalent attachment to this support using 1% glutaraldehyde (P-QQSA 1). The optimum conditions selected for immobilization were pH 4.5, incubation time of 4 h and protein concentration of 340 μg/mL. Various characteristics of the immobilized pectinase such as optimum pH, heat stability and reusability were evaluated. As a result of immobilization the enzyme's T50 increased, the best being achieved with immobilization using 15% glutaraldehyde and covalent attachment. The optimum pH of the free and immobilized enzymes were 5 (free), 4.5 (P-QQSA), 4.5–5.0 (P-QQSA 0.5) and 4–5 (P-QSA 1 and P-QQSA 15), respectively. The biocatalysts prepared retained 100% of their original catalytic activity after nine cycles of reuse.Practical ApplicationsPectinases hydrolyze pectin and/or pectic acid. These enzymes have widespread applications in the food industry (processing of fruits), wastewater treatment, textile industries, fruit softening and plant infection processes. The stability of these enzymes depends on the aqueous medium and is easily disrupted to the point where the enzymes cannot function appropriately. Immobilization techniques provide a promising approach to retain their stability. Various methods for immobilization of this enzyme have been described: entrapping in alginate, physical adsorption on anion resin, γ-alumina, particles and nanoparticles of silica and covalent attachment to carriers such as porous glass and nylon. However, the development of new methods and supports for immobilizing enzymes is of special importance in enzyme technology. The present article describes several methods for immobilization of pectinase in chitosan-coated chitin for use in the juice and wine industries.Journal of Food Process Engineering 03/2015; DOI:10.1111/jfpe.12203 · 0.63 Impact Factor
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ABSTRACT: The graphite nanosheets decorated with Fe3O4 nanoparticles (about 15–20 nm), i.e., Fe3O4@graphite nanosheets, have been prepared via a facile one-pot method. The resulting composites with large specific surface area, high affinity, and rapid magnetic response were characterized by means of Fourier transform infrared spectra, Raman spectra, powder X-ray diffraction, scanning electron microscopy, transmission electron microscopy, elemental analysis, and thermogravimetric analysis, which indicated that the Fe3O4 nanoparticles were chemically integrated on the graphite nanosheets. The porcine pancreatic lipase (PPL) was immobilized onto the Fe3O4@graphite nanosheets through covalent bonding using glutaraldehyde as a coupling reagent, and the properties of the resulting immobilized PPL such as enzymatic activity, stability, and reusability were researched at the same time. The result proved that the immobilized PPL displayed excellent properties and demonstrated that the prepared Fe3O4@graphite nanosheets would present a potential application as a novel carrier for enzyme immobilization, enrichment, and separation.Journal of Nanoparticle Research 06/2014; 16(6). DOI:10.1007/s11051-014-2459-2 · 2.28 Impact Factor