Immobilization of urease

FIGURE 6-11 Urea electrode, based on the immobilization of urease onto an ammonium ion-selective electrode. 

Jackbean urease was immobilized on kaolinite and montmorillonite [98]. The amounts of urease required for maximum immobilization were 70 and 90 mg g 1 of kaolinite and montmorillonite, respectively. The Km values of immobilized urease (25.1-60.8 mM) were of the same order of magnitude as that of free urease (29.4 mM) but one order of magnitude higher than those of soil urease (1.77-2.90 mM). Immobilization of urease on clay surfaces leads to increases in the kinetic constants. 

Sahoo, B., Sahu, S.K., Pramanik, P., 2011. A novel method for the immobilization of urease on phosphonate grafted iron oxide nanoparticle. J. Mol. Catal. B Enzym. 69, 95-102. 

I.G. Mourzina, T. Yoshinobu, Y.E. Ermolenko, Y.G. Vlasov, M.J. Schoning and H. Iwasaki, Immobilization of urease and cholinesterase on the surface of semiconductor transducer for the development of light-addressable potentiometric sensors, Microchim. Acta, 144(1-3) (2004) 41-50. 

The gas-sensing electrodes also are used for the potentiometric measurement of biologically important species. An enzyme is immobilized at or near the gas probe. The gas sensor measures the amount of characteristic gas produced by the reaction of the analyzed substance with the enzyme. For example, an enzyme electrode for urea [NH2C(0)NH2] determination is constructed by the immobilization of urease onto the surface of an ammonia-selective electrode. When the electrode is inserted into a solution that contains urea, the enzyme catalyzes its conversion to ammonia ... 

Carlsson J, Axen R, Brocklehurst K, and Crook EM. Immobilization of urease by thiol-disulphide interchange with concomitant purification. Eur. J. Biochem. 1974 44 189-194. 

Suggest how the covalent immobilization of urease onto particles of an ethylene-maleic anhydride copolymer might be expected to change (a) its apparent pH optimum, and (b) its apparent Km for urea. 

Immobilization of urease to a silanized Si3N4 gate by glutaraldehyde... 

Sometimes even membrane transport and mechanical properties are affected by the harsh chemical treatments required in the immobilization procedure. Figure 7.35 shows how immobilization of urease by diazotization on heterogeneous polysulfone flat membranes reduces the membrane hydraulic permeability by 50%.74 It also shows how the higher permeability membranes are more affected by the immobilization procedure. After the enzyme is immobilized, it is wise to check the integrity of the membrane (e.g., the permeability to the species of interest and the strength).71 74... 

Immobilization of Urease on Sweetzyme Pellets For co-immobilization of urease on the Sweetzyme pellets, 500 ml of 1 g/1 urease solution and 2 g of Sweetzyme pellets was added to a 1-1 beaker [35]. The beaker was left on the benchtop at room temperature for 24 h. The pellets were separated from the solution by decanting and gravity filtration and dried on a paper towel at room temperature for 24 h or until dry. Co-immobilized pellets were stored at 4 °C until use. Activity of immobilized urease was measured at pH 7.5 and 25°C using a standard assay procedure that measures the rate of ammonia liberation [43]. The urease activities obtained with our immobilization procedure were in the range of 550-577 U/g pellets, where a unit liberates 1 pmol of ammonia per minute under the assay conditions. 

Simultaneous isomerization and fermentation (SIF) is expected to drive the isomerization forward and lead to higher xylose utilization. Unfortunately, the pH optima for the isomerization ( 7.5) and the fermentation ( 4.5) steps are vastly different. The approach we have taken is to develop a novel technique for SIF that is capable of sustaining two different pH-microenvironments in a single vessel—one optimal for xylose isomerization and the other optimal for fermentation of xylulose. The technique involves co-immobilization of urease with xylose isomerase. We have shown that it is possible to sustain a significant pH gradient between the bulk liquid and the core region of the pellet by adding urea to the fermentation broth. 

Chellapandian, M. and Krishnan, M.R.V., Chitosan-poly (glycidyl methacrylate) copolymer for immobilization of urease. Process Biochem., 33, 595, 1998. 

Akkaya, A., Usian, A.H. Sequential immobilization of urease to gfycidyl methacrylate grafted sodium alginate. J. Mol. Catal. B Enzym. 67, 195-201 (2010)... 

In addition to nucleic acids, enzymes have also been incorporated into conducting a polymer matrix for biosensor applications. As a transducer, a CP can convert the chemical response into an electric current. To enhance the sensitivity and the response time, fabrication of CPs/enzyme nanocomposites with large surface area is a meaningful objective. Syu and Chang demonstrated the immobilization of urease onto PPy nanotubes over carbon paper substrate by a physical entrapment approach [124]. The composite electrodes exhibited a detection sensitivity for the determination of urea of 53.74 mVdecade and a detection limit on the urea concentration of 1.0 pM. Furthermore, the composite electrode shows rapid response, storage stability and reusability. Lipase can also be covalently immobilized... 

Immobilization of Urease. The urease (E.C. 3.5.1.5) was immobilized on three different carriers 4-aminobenzylcellulose, 4-aminobenzoyl-cellulose, poly(vinyl 4-aminobenzoate). 

Maia MD, de Vasconcelos EA, Conceigao de Mascena Diniz Maia PF, da Costa Maciel J, Rizzardi Cajueiro KR, da Paz Carvalho da Silva M, da Silva Jr EF, Fireman Dutra RA, Freire VN, de Filho JL (2007) Immobilization of urease on vapour phase stain etched porous silicon. Process Biochem 42(3) 429 33... 

Immobilization of urease onto an IME device can produce an impedimetric enz3mie biosensor for determination of urea [4]. The enzyme urease can be immobilized by a similar covalent tethering strategy to the interdigit surface via S-aminopropyltrimethoxysilane monolayer and sufib-NHS and EDC as crosslinkers. This creates an enzyme... 

Hashimoto T, Yoko T., Sakka S. Third-order nonlinear optical susceptibihty of aFe203 thin film prepared by the sol-gel method. J. Ceram. Soc. Jpn 1993 101 64-68 Hatayama H., Swabe T, Kurokawa Y. Immobilization of urease on composite fiba- by using a gel formation of celluse acetate and titanium iso-propoxide. J. Sol-Gel Sci. Technol. 1996 7 13-17 Hattori A., Toda H. Highphotocatalytic activity of F-doped TiO film on glass. J. Sol-Gel Sci. Technol. 2001 22 47-52... 

Later, the same idea was applied to determine urea in blood serum [19]. The system was formed by two urea biosensors, obtained by immobilization of urease enzyme (EC 3.5.1.45) onto ammonium-selective membranes, plus ISEs for... 

The determination of urea in blood is even possible when it contains inhibitors such as fluoride ions, which are often used to inhibit glycolysis in blood samples. The appropriate immobilization of urease on the sensor removes interference from fluoride ions [102] and enables direct determination of urea over the whole range of biological concentrations. This biosensor can also be coupled with a glucose oxidase electrode to measure urea and glucose concentrations simultaneously in serum samples [113]. 

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