Immobilization of cholinesterase and urease

Guilbault, George G.; Das, J.

doi:10.1016/0003-2697(70)90305-2

Cited by 47 publications

(9 citation statements)

References 12 publications

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“…A similar change was also observed in the case of pigeonpea urease immobilized on chitosan beads [8]. A slightly lower apparent K m value was found for jack bean urease immobilized in a polyacrylamide gel (5 mM) [12]; however, urease immobilized in hydrocarbon‐based liquid surfactant membrane exhibited a K m approximately 50 times greater than the soluble enzyme [28].…”

Section: Resultssupporting

confidence: 60%

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Characterization of gelatin‐immobilized pigeonpea urease and preparation of a new urea biosensor

Srivastava

Kayastha

Srinivasan

2001

Biotech and App Biochem

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Urease purified from pigeonpea seeds was immobilized on gelatin beads via cross-linking with glutaraldehyde. The maximum immobilization (75%) was observed at 30 mg/ml gelatin, 0.414 mg of enzyme/bead, 1% (v/v) glutaraldehyde and 4 degrees C. Beads stored in 50 mM Tris/acetate buffer (pH 7.3) at 4 degrees C showed a half-life of 240 days and there was practically no leaching of enzyme (less than 2%) over a period of 30 days. These beads can be reused more than 30 times (with 24 h intervals) without much loss of enzyme activity (i.e. less than 11%). The immobilized urease showed a shift in its optimum pH from 7.3 to 6.5 in Tris/acetate buffer. Optimum temperature also shifted from 47 to 65 degrees C compared with the soluble enzyme. Gelatin-immobilized pigeonpea urease had a higher K(m) (8.3 mM) than that of the soluble enzyme (3.0 mM). The time-dependent temperature inactivation pattern was also found to change from biphasic to monophasic kinetics. The immobilized beads were used for the preparation of a new urea biosensor with a response time of less than 2 min. At least 14 samples of urea can be measured with this biosensor within an hour. The beads, as well as the biosensor, were used to analyse the urea content in clinical samples from the local clinical pathology laboratories. The results obtained with the biosensor were strikingly similar to those obtained with the various commonly employed biochemical/autoanalyzer(R) methods used. These immobilization studies also have a potential role in haemodialysis machines that maintain the urea level in kidney patients and in the construction of a portable/wearable kidney. The easy availability of the pigeonpea urease, the ease of its immobilization on gelatin and a significantly lower cost of the urease described in the present study makes it a suitable product for future applications in therapeutics and diagnostics.

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Section: Resultssupporting

confidence: 60%

“…Enzyme electrodes that measure the produced ammonium ions have been extensively described [12–15]. Immobilization of relevant enzymes at the surface of the sensing electrode is one of the crucial steps in the construction of a biosensor.…”

Section: Introductionmentioning

confidence: 99%

Characterization of gelatin‐immobilized pigeonpea urease and preparation of a new urea biosensor

Srivastava

Kayastha

Srinivasan

2001

Biotech and App Biochem

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“…A similar change was also observed in case of pigeonpea urease immobilized on chitosan beads [6]. A slightly lower apparent Michaelis constant was found for jack‐bean urease immobilized in polyacrylamide gel (5 mM) [19]; however, urease immobilized in hydrocarbon‐based liquid surfactant membrane exhibited an approx. 50 times greater K m than the soluble enzyme [20].…”

Section: Resultssupporting

confidence: 66%

Immobilization of pigeonpea (Cajanus cajan) urease on DEAE‐cellulose paper strips for urea estimation

Reddy

Srivastava

Dey

et al. 2004

Biotech and App Biochem

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Pigeonpea ( Cajanus cajan ) urease was immobilized on 1 cmx1 cm DEAE-cellulose paper strips. The optimum immobilization (51% activity) was observed at 4 degrees C, with a protein concentration of 1.0 mg/strip. The apparent optimum pH shifted from 7.3 to 6.8. Immobilized urease showed an optimal stability temperature of 67 degrees C, compared with 47 degrees C for the soluble urease. Time-dependent kinetics of the thermal inactivation of the immobilized urease were examined and found to be monophasic as compared with the soluble enzyme, which was biphasic. The Michaelis constant ( K (m)) for the DEAE-cellulose-immobilized urease was found to be 4.75 mM, 1.5 times higher than the soluble enzyme. Immobilized strips stored at 4 degrees C showed an increased half-life ( t (1/2)=150 days). There was practically no leaching of the enzyme from the immobilized strips over a period of 2 weeks. These strips were used for estimating the urea content of blood samples; the results obtained matched well with those obtained in a clinical laboratory through an Autoanalyzer(R) (Zydus Co., Rome, Italy). The easy availability of pigeonpea urease, the ease of its immobilization on DEAE-cellulose strips and the significantly lower cost of urease described in the present study makes it a suitable product for future applications in diagnostics.

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“…The latter conjugate is prepared by suspending silastic resin in water containing enzyme and adding catalytic amounts of stannous octoate to induce gel formation. Localized heating during gel formation has been reported to cause up to 80% loss in enzyme activity (Guilbault and Das, 1970). Strandberg and Smiley (1971) reported on entrapment of glucose isomerase in polyacrylamide.…”

Section: Entrapmentmentioning

confidence: 99%

SYMPOSIUM: Immobilized Enzymes in Food Systems

Stanley

Olson

1974

Journal of Food Science

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S The advantages and disadvantages of three general systems for immobilizing enzymes (adsorption, entrapment and covalent bonding) are discussed with emphasis on practicality for food processing applications. Only covalent bonding results in complete fixation. For economy and simplicity, adsorption has many advantages but composition of feeds may cause excessive enzyme loss. The combination of adsorption on a phenol‐formaldehyde resin with covalent crosslinking with glutaraldehyde is described for immobilizing lactase and advantages are discussed.

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Immobilization of cholinesterase and urease

Cited by 47 publications

References 12 publications

Characterization of gelatin‐immobilized pigeonpea urease and preparation of a new urea biosensor

Characterization of gelatin‐immobilized pigeonpea urease and preparation of a new urea biosensor

Immobilization of pigeonpea (Cajanus cajan) urease on DEAE‐cellulose paper strips for urea estimation

SYMPOSIUM: Immobilized Enzymes in Food Systems

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