Sharif Digital Repository

A simple optimization methodology is applied to estimate the intrinsic kinetic parameters for both reversible and irreversible unireactant immobilized enzyme systems that follow the Michaelis-Menten mechanism. The method utilizes a direct-search optimization algorithm along with the numerical solution of the governing differential equations. The usefulness and validity of the method is demonstrated by comparing the predicted values of the intrinsic kinetic constants using the proposed method with a series of experimental values reported in the literature for different immobilized enzyme systems with irreversible and reversible reactions  

Nonporous and mesoporous silica-coated magnetite cluster nanocomposites particles were fabricated with various silica structures in order to develop a desired carrier for the lipase immobilization and subsequent biodiesel production. Lipase from Pseudomonas cepacia was covalently bound to the amino-functionalized particles using glutaraldehyde as a coupling agent. The hybrid systems that were obtained exhibited high stability and easy recovery regardless of the silica structure, following the application of an external magnetic field. The immobilized lipases were then used as the recoverable biocatalyst in a transesterification reaction to convert the soybean oil to biodiesel with methanol.... 

In this study, anti-leukemic enzyme L-asparaginase (E.C.3.5.1.1) from Escherichia coli ATCC 11303 was modified by the microencapsulation technique onto calcium alginate beads. Using response surface methodology (RSM), a three-level full factorial design, the values of concentration of sodium alginate, concentration of calcium chloride, and enzyme loading were investigated to obtain the highest residual L-asparaginase (L-ASNase) activity % (immobilized enzyme activity/free enzyme activity). The effects of the studied factors on immobilization were evaluated The predicted values by the model were close to the experimental values, indicating suitability of the model. The results presented that... 

A magnetic nanocomposite was prepared by entrapment of Fe3O4 nanoparticles into the cross-linked ionic liquid/epoxy type polymer. The resulting support was used for covalent immobilization of cellulase through the reaction with epoxy groups. The ionic surface of the support improved the adsorption of enzyme, and a large amount of enzyme (106.1 mg/g) was loaded onto the support surface. The effect of the presence of ionic monomer and covalent binding of enzyme was also investigated. The structure of support was characterized by various instruments such as FT-IR, TGA, VSM, XRD, TEM, SEM, and DLS. The activity and stability of immobilized cellulase were investigated in the prepared support. The... 

A destabilizing effect at pH 7 of sodium phosphate on several lipases immobilized via interfacial activation is shown in this work. This paper investigates if this destabilizing effect is extended to other inactivation conditions, immobilization protocols or even other immobilized enzymes (ficin, trypsin, β-galactosidase, β-glucosidase, laccase, glucose oxidase and catalase). As lipases, those from Candida antarctica (A and B), Candida rugosa and Rhizomucor miehei have been used. Results confirm the very negative effect of 100 mM sodium phosphate at pH 7.0 for the stability of all studied lipases immobilized on octyl agarose, while using glutaraldehyde-support the effect is smaller (still... 

Over the last decade, nanoparticles used as protein carriers have opened new avenues for a variety of biomedical applications. The main concern for these applications is changes in biological activity of immobilized proteins due to conformational changes on the surface of the carrier. To evaluate this concern, the preparation and biocatalyst activity of α-chymotrypsin-Fe 3O4 @ Au core/shell nanoparticles were investigated. First, Fe3O4 @ Au core/shell nanoparticles were synthesized by coprecipitation method and citrate reduction of HAuCl 4. TEM imaging revealed a core size of 13 ± 3 nm and a shell thickness of 4 ± 1 nm for synthesized nanoparticles. X-ray diffraction (XRD) was used to study... 

Bio-batteries are environmental friendly electricity generation devices that utilize energy from biological sources, like glucose[1] [2]. It consists of an anode with digesting enzymes and a cathode comprising oxygen-reducing enzymes. Immobilizing enzymes is a considerable issue in these kinds of devices. In this article, we propose two new methods for immobilizing and shifting particles likewise enzymes used in bio-batteries towards the anode or cathode end. Our experiments illustrate how tiny particles could be trapped in the nodes of the ultrasonic standing waves and by two methods of displacing the nodes, we can transfer the particles towards a desired location. First method is based on... 

In this paper, the performance of immobilized packed bed glucose isomerase enzyme was mathematically modeled. A modified Michaelis-Menten type relation was used to describe the enzyme kinetics. Mass transfer inside the biocatalyst particle and through the bed column was analyzed simultaneously. Using measured data, physicochemical properties including diffusivity, viscosity and density of sugar solutions were correlated with its concentrations and were used to provide precision in solving the set of model equations. Model equations were solved using the Runge-Kutta and Gauss-Seidel algorithms and finite difference numerical method in MATLAB environment. Model output was used to demonstrate... 

This study reports the importance of physicochemical characteristics of the core-shell structured polymer-coated Fe3O4 cluster@SiO2 nanocomposite particles for designing the biocatalysts used for transesterification of soybean oil. To demonstrate this issue, two different types of polymer molecules such as polyethylenimine (PEI) and polyacrylic acid (PAA) each with two different molecular weights are employed. The highest fatty acid methyl esters (FAMEs) synthesis yield values are achieved by the lipases immobilized onto the low- and high-molecular-weight PEI-coated particles as compared to those of the low- and high-molecular-weight PAA-coated particles (i.e., ∼64.8% and ∼73.1% versus... 

This paper presents a new method for spectrophotometirc detection of sulfide applying fungal peroxidase immobilized on sodium alginate. The sensing scheme was based on decrease of the absorbance of the orange compound, purpurogallin produced from pyrogallol and H2O2 as substrates, due to the inhibition of peroxidase by sulfide. Absorbance of purpurogallin was detected at 420nm by using a spectrophotometer. The proposed method could successfully detect the sulfide in the concentration range of 0.6-7.0μM with a detection limit of 0.4μM. The kinetic parameters of Michaelis-Menten with and without sulfide were also calculated. Possible inhibition mechanism of peroxidase by sulfide was deduced... 

A facile approach for the preparation of core-shell structured poly(acrylic acid) (PAA)-coated Fe3O4 cluster@SiO2 nanocomposite particles as the support materials for the lipase immobilization is reported. Low- or high-molecular-weight (1800 and 100 000, respectively) PAA molecules were covalently attached onto the surface of amine-functionalized magnetic silica nanoacomposite particles. The successful preparation of particles were verified by scanning transmission electron microscopy (STEM), X-ray diffraction (XRD), vibrating sample magnetometer (VSM), thermogravimetric analysis (TGA), zeta potential measurement, and Fourier-transform infrared (FTIR) techniques. Once lipase is covalently... 

Application of immobilized horseradish peroxidase (HRP) in porous calcium alginate (ca-alginate) for the purpose of phenol removal is reported. The optimal conditions for immobilization of HRP in ca-alginate were identified. Gelation (encapsulation) was optimized at 1.0% (w/v) sodium alginate in the presence of 5.5% (w/v) of calcium chloride. Upon immobilization, pH profile of enzyme activity changes as it shows higher value at basic and acidic solution. Increasing initial phenol concentration results in a decrease in % conversion. The highest conversion belongs to phenol concentration of 2 mM. Investigation into time course of phenol removal for both encapsulated and free enzymes showed... 

In the present work, an amperometric inhibition biosensor for the determination of sulfide has been fabricated by immobilizing Coprinus cinereus peroxidase (CIP) on the surface of screen printed electrode (SPE). Chitosan/acrylamide was applied for immobilization of peroxidase on the working electrode. The amperometric measurement was performed at an applied potential of -150. mV versus Ag/AgCl with a scan rate of 100. mV in the presence of hydroquinone as electron mediator and 0.1. M phosphate buffer solution of pH 6.5. The variables influencing the performance of sensor including the amount of substrate, mediator concentration and electrolyte pH were optimized. The determination of sulfide... 

Chondroitinase ABCI (cABCI) is a drug enzyme that can be used to treat spinal cord injuries. Due to low thermal stability of cABCI, this enzyme was immobilized on Fe3O4 nanoparticle to increase its thermal stability. The size and morphology, structure and magnetic property of the Fe3O4 nanoparticles were characterized by the analyses of SEM, XRD and VSM, respectively, and FTIR spectroscopy was employed to confirm the immobilization of cABCI on the surface of Fe3O4 nanoparticles. The results indicated that the optimum conditions for pH, temperature, cABCI-to-Fe3O4 mass ratio and incubation time in immobilization process were 6.5, 15 °C, 0.75 and 4.5 h, respectively, and about 0.037 mg cABCI... 

The presence of pharmaceutical micropollutants in water and wastewater is considered a serious environmental issue. To eliminate these pollutants, biodegradation of pharmaceuticals using enzymes such as laccase, is proposed as a green method. In this study, immobilized laccase was used for the removal of two model pharmaceutical compounds, carbamazepine and diclofenac. Polyvinylidene fluoride (PVDF) membrane modified with multi-walled carbon nanotubes (MWCNTs) were synthesized as a tailor-made support for enzyme immobilization. Covalently immobilized laccase from Trametes hirsuta exhibited remarkable activity and activity recovery of 4.47 U/cm2 and 38.31%, respectively. The results also... 

Chondroitinase ABC I (cABC I) has received notable attention in treatment of spinal cord injuries and its application as therapeutics has been limited due to low thermal stability at physiological temperature. In this study, cABC I enzyme was immobilized on the dextran-coated Fe3O4 nanoparticles through physical adsorption to improve the thermal stability. The nanoparticles were characterized using XRD, SEM, VSM, and FTIR analyses. Response surface methodology and central composite design were employed to assess factors affecting the activity of immobilized cABC I. Experimental results showed that pH 6.3, temperature 24 °C, enzyme/support mass ratio 1.27, and incubation time 5.7 h were the...