scholarly journals The Relative Merits and Applicability of Kinetic and Fixed-Incubation Methods of Enzyme Assay in Clinical Enzymology

1972 ◽  
Vol 18 (12) ◽  
pp. 1449-1454 ◽  
Author(s):  
D W Moss
Keyword(s):  
Enzyme Activity ◽  
Continuous Monitoring ◽  
Enzyme Assay ◽  
Kinetic Method ◽  
Reaction Progress ◽  
Kinetic Principle ◽  
Clinical Enzymology

Abstract The considerations are discussed that make continuous monitoring of reaction-progress curves superior to fixed-incubation methods in the determination of enzyme activity. Provided that they are used with caution and their limitations are appreciated, fixed-incubation methods continue to fulfill a useful, though diminishing, role in clinical enzymology because of their adaptability to existing patterns of automation. The introduction of suitable mechanized equipment will favor the eventual complete adoption of the kinetic method. However, the use of such equipment should not be at the expense of th important characteristics of the kinetic principle.

A Kinetic Method for Determination of the Biosynthetic Activity of Glutamine Synthetase

1992 ◽  
Vol 57 (10) ◽  
pp. 2174-2180
Author(s):  
Vladimír Mikeš
Keyword(s):  
Enzyme Activity ◽  
Glutamine Synthetase ◽  
Kinetic Method ◽  
Kinetic Studies ◽  
Biosynthetic Activity ◽  
Ph Indicator ◽  
Permeabilized Cells ◽  
Cell Extracts

In the present paper a rapid and simple kinetic method for determination of the biosynthetic activity of glutamine synthetase is described based upon the decomposition of ATP, accompanied with acidification of the slightly buffered medium which can be measured by means of a pH indicator. The method can be used for determination of the enzyme activity in whole permeabilized cells, crude cell extracts, as well as for kinetic studies and studies of the effects of inhibitors on the purified glutamine synthetase.

A Rapid Electrophoretic Method for the Determination of the Isoenzymes of Serum Lactate Dehydrogenase

1966 ◽  
Vol 12 (5) ◽  
pp. 308-313 ◽  
Author(s):  
Albert W Opher ◽  
Charles S Collier ◽  
Joseph M Miller
Keyword(s):  
Enzyme Activity ◽  
Lactate Dehydrogenase ◽  
Quantitative Determination ◽  
Serum Lactate ◽  
Serum Lactate Dehydrogenase ◽  
Electrophoretic Method ◽  
The Individual ◽  
Electrophoretic Procedure ◽  
Purple Formazan

Abstract A convenient electrophoretic procedure for the separation and quantitation of lactate dehydrogenase (LDH) isoenzymes is described. The system uses polyacetate Sepraphore III strips.* The areas of activity are shown by incubation with an LDH substrate combined with tetra-nitro-blue-tetrazolium. The reduction of the latter to the purple formazan is quantitatively related to the enzyme activity. Quantitative determination of the individual colored areas is performed by densitometry.

Automatic kinetic method for the determination of reduced glutathione in blood

1992 ◽  
Vol 269 (2) ◽  
pp. 273-279 ◽  
Author(s):  
Rafael Jiménez-Prieto ◽  
Antonio Velasco ◽  
Manuel Silva ◽  
Dolores Pérez-Bendito
Keyword(s):  
Reduced Glutathione ◽  
Kinetic Method

scholarly journals Extracellular α-Galactosidase from Trichoderma sp. (WF-3): Optimization of Enzyme Production and Biochemical Characterization

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Aishwarya Singh Chauhan ◽  
Arunesh Kumar ◽  
Nikhat J. Siddiqi ◽  
B. Sharma
Keyword(s):  
Enzyme Activity ◽  
Enzyme Assay ◽  
Biochemical Characterization ◽  
Catalytic Efficiency ◽  
Culture Conditions ◽  
Significant Loss ◽  
Trichoderma Sp ◽  
Rate Of Reaction ◽  
Higher Temperature ◽  
Substrate Concentrations

Trichoderma spp. have been reported earlier for their excellent capacity of secreting extracellular α-galactosidase. This communication focuses on the optimization of culture conditions for optimal production of enzyme and its characterization. The evaluation of the effects of different enzyme assay parameters such as stability, pH, temperature, substrate concentrations, and incubation time on enzyme activity has been made. The most suitable buffer for enzyme assay was found to be citrate phosphate buffer (50 mM, pH 6.0) for optimal enzyme activity. This enzyme was fairly stable at higher temperature as it exhibited 72% activity at 60°C. The enzyme when incubated at room temperature up to two hours did not show any significant loss in activity. It followed Michaelis-Menten curve and showed direct relationship with varying substrate concentrations. Higher substrate concentration was not inhibitory to enzyme activity. The apparent Michaelis-Menten constant (Km), maximum rate of reaction (Vmax), Kcat, and catalytic efficiency values for this enzyme were calculated from the Lineweaver-Burk double reciprocal plot and were found to be 0.5 mM, 10 mM/s, 1.30 U mg−1, and 2.33 U mg−1 mM−1, respectively. This information would be helpful in understanding the biophysical and biochemical characteristics of extracellular α-galactosidase from other microbial sources.

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