Operational Stability Of Enzyme Research Articles

Biocatalytic membranes in anti-fouling and emerging pollutant degradation applications: Current state and perspectives

• Biocatalytic membrane processes produce environmentally friendly and energy-efficient processes. • Enzyme immobilization is the simplest solution to long-term operational stability, activity and reuse. • Modified immobilization methods improve the performance of enzymes over common immobilization methods. • Biocatalytic membranes compared with electrocatalytic and photocatalytic membrane processes. • Biocatalytic membranes have the potential to be used in antifouling and emerging pollutant degradation applications. Biocatalytic membranes, which are fabricated by taking advantage of the synergetic effect of membranes and enzymes, have been one of the attractive treatment methods as sustainable and environmentally friendly. In biocatalytic membrane systems, enzymes can be free or immobilized, but the number of immobilized applications increases due to the advantages of immobilization. Immobilization is one of the critical points to improve the storage and operational stability of enzymes. Additionally, immobilized enzymes have a significant benefit over free enzymes in terms of reusability. This review summarizes recent advances in the preparation of biocatalytic membranes with the different immobilization methods of enzymes in/on membranes, paying particular attention to recent approaches for anti-fouling and emerging pollutant degradation applications. Also, some future outlooks were briefly presented.

Thermal stability of xylose reductase from Debaryomyces nepalensis NCYC 3413: deactivation kinetics and structural studies

Abstract Deactivation kinetics, as well as the operational stability of enzymes, is crucial in economic feasibility of industrial enzymatic processes. In this study, thermal inactivation profiles of Debaryomyces nepalensis NCYC 3413 xylose reductase (DnXR) were obtained at different temperatures. Thermodynamic parameters of the deactivation process were determined by measuring the residual activity of the enzyme. Structural characteristics of the enzyme were determined using circular dichroism, differential scanning calorimetric techniques. Enzyme retained 75–80% activity at 35 °C after incubation for 5 h. At higher temperatures, the residual activity reduced at a faster rate where only 10% of the initial activity was retained in 10 min at 50 °C. Values of ΔH* (-300.47 kJmole−1), ΔG* (−530.66 kJ mole−1) and ΔS* (714.51 × 10−3 kJ K−1 mole−1) indicated that thermal deactivation of DnXR is enthalpy driven. In the presence of cofactor NADPH, melting temperature of the enzyme increased by 2 °C with a decrease in rate of thermal deactivation. Substrate found not to have a positive effect on thermal stability. This is the first report providing information on the thermal deactivation kinetics, thermodynamic parameter estimation and the influence of cofactor on thermal stability of Debaryomyces nepalensis NCYC 3413 xylose reductase.

Enzyme immobilization in a photosensitive conducting polymer bearing azobenzene in the main chain

A new photosensitive and thermosensitive monomer, namely bis(4-(3-thienyl ethylene)-oxycarbonyl)diazobenzene (TDAZO), was synthesized. The photochemical and thermal cis–trans isomerization of the monomer has been investigated. The rate constants of the photoisomerization of TDAZO in ACN and DCM were 0.195 and 0.308 min−1, respectively. For spectroelectrochemical investigation and enzyme immobilization application, TDAZO copolymerized with thiophene and pyrrole. Electrochemical and spectroelectrochemical properties of P(TDAZO-co-Th) were investigated and invertase was immobilized in P(TDAZO-co-Py) copolymer. Immobilization of enzymes was carried out by the entrapment of the enzyme in conducting polymer matrices during electrochemical polymerization of pyrrole through thiophene moieties of the TDAZO. Optimum conditions for this electrode, such as pH, temperature, kinetic parameters (K m and V max) and operational stability were investigated. Kinetic parameters invertase-immobilized in copolymer were smaller than free enzyme. The optimum operational temperature was 10 °C higher for immobilized enzyme than that of the free enzyme. Due to strong interaction between enzyme and diazo group in the polymer main chain, thermal, pH and operational stability of enzyme has been enhanced.

Poly-lysine supported cross-linked enzyme aggregates with efficient enzymatic activity and high operational stability

In this study, the operational stability of enzymes in a cross-linked aggregate (CLEA) formed with poly-lysine was examined. Chymotrypsin, subtilisin, citrate synthase and laccase, which are structurally and mechanistically diverse, were used as model enzymes. The preparation of poly-lysine supported CLEA was completed within 3 h. The immobilized enzymes were more stable than free enzymes at high temperature, in the presence of a chemical denaturant or in an organic solvent and were recycled without appreciable loss of activity. In addition, the immobilized proteases showed higher or similar hydrolytic activity in acidic pH than in neutral pH. This immobilization method was also applicable to the multi-subunit protein. These results suggest that the poly-Lys supported CLEA can be used as catalysts with own enzymatic activity and high operational stability.

Long-term activity of covalent grafted biocatalysts during intermittent use of a glucose/O 2 biofuel cell

The operational stability of enzymes in a concentric glucose/O 2 biofuel cell has been significantly improved with the synthesis of grafted enzyme electrodes compared to entrapped enzyme electrodes. The concentric device combined glucose electro-oxidation by glucose oxidase at the anode and oxygen electro-reduction by bilirubin oxidase at the cathode. The entrapped enzyme electrodes were prepared from physical immobilization of the enzymes by a polypyrrole polymer onto the electrode surface. The grafted enzyme electrodes were synthesized by grafting the enzymes via alkyl spacer arms to a poly(aminopropylpyrrole) film onto the electrode surface. From spectrophotometric and electrochemical analyses, it was demonstrated that the spacer arms increased the operational stability and enzyme mobility that favoured electron transfer from their active sites to the electrode. The maximum power output of the assembled biofuel cell was 20 μW cm −2, at 0.20 V with 10 mM glucose in phosphate buffer pH 7.4. The grafted enzyme electrodes presented an unprecedented operational stability as the maximum of power density of the BFC remains constant after intermittent use over a 45-day period. This was a remarkable improvement compared to electrodes with entrapped enzymes, which lost 74% of their initial power density after intermittent use over a 17-day period.

Acid activated montmorillonite: an efficient immobilization support for improving reusability, storage stability and operational stability of enzymes

Three enzymes, a-amylase, glucoamylase and invertase, were immobilized on acid activated montmorillonite K 10 via two independent techniques, adsorption and covalent binding. The immobilized enzymes were characterized by XRD, N2 adsorption measurements and 27 Al MAS-NMR spectroscopy. The XRD patterns showed that all enzymes were interca- lated into the clay inter-layer space. The entire protein backbone was situated at the periphery of the clay matrix. Intercalation occurred through the side chains of the amino acid residues. A decrease in surface area and pore volume upon immobilization supported this observation. The extent of intercalation was greater for the covalently bound systems. NMR data showed that tetrahedral Al species were involved during enzyme adsorption whereas octahedral Al was involved during covalent binding. The immobilized enzymes demon- strated enhanced storage stability. While the free en- zymes lost all activity within a period of 10 days, the immobilized forms retained appreciable activity even after 30 days of storage. Reusability also improved upon immobilization. Here again, covalently bound enzymes exhibited better characteristics than their adsorbed counterparts. The immobilized enzymes could be successfully used continuously in the packed bed reactor for about 96 hours without much loss in activity. Immobilized glucoamylase demonstrated the best results.

High performance enzymatic synthesis of oleyl oleate using immobilised lipase from Candida antartica

High performance enzymatic synthesis of oleyl oleate, a liquid wax ester was carried out by lipase-catalysed esterification of oleic acid and oleyl alcohol. Various reaction parameters were optimised to obtain high yield of oleyl oleate. The optimum condition to produce oleyl oleate was reaction time; 5 min, organic solvents of log P ≥ 3.5, temperature; 40-50oC, amount of enzyme; 0.2-0.4 g and molar ratio of oleyl alcohol to oleic acid; 2:1. The operational stability of enzyme was maintained at >90% yield up to 9 cycles. Analysis of the yield of the product showed that at optimum conditions, >95% liquid wax esters were produced.

Operational Stability of Enzymes.

Operational Stability of Enzymes.

Operational stability of enzymes. Acylase-catalyzed resolution of N-acetyl amino acids to enantiomerically pure L-amino acids.

The method of measuring enzyme deactivation by monitoring necessary addition of fresh enzyme to keep a constant degree of conversion in a CSTR at constant [E] x tau, the product of concentration of active enzyme [E] and residence time tau, was successfully applied to acylase I from porcine kidney and Aspergillus oryzae fungus. Fungal enzyme was found to be more stable than kidney enzyme. Activation by both Co2+ and Zn2+ ions also yielded increased operational enzyme stability: Co2+ and Zn2+ are better stabilizers than activators. Mg2+ and Ca2+ are found to be neither activators nor stabilizers. Fungal acylase partially deactivated by exposition to a metal-free medium in the CSTR was reactivated by addition of Zn2+, demonstrating that loss of Zn2+ from the enzyme molecule is mainly responsible for deactivation in a continuous reactor.