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Method for calculating oxalate decarboxylase activity by determining carbon dioxide concentration

A technology of oxalate decarboxylase and carbon dioxide, applied in the field of determination of oxalate decarboxylase activity, can solve the problems of cumbersome manometer method and high requirements for instruments

Inactive Publication Date: 2008-08-13
DONGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The manometer method is cumbersome and requires high equipment, while the enzymatic method is a development direction

Method used

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  • Method for calculating oxalate decarboxylase activity by determining carbon dioxide concentration
  • Method for calculating oxalate decarboxylase activity by determining carbon dioxide concentration
  • Method for calculating oxalate decarboxylase activity by determining carbon dioxide concentration

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] According to the following table 1, the experiment was carried out.

[0050] Table 1

[0051]

[0052][1], CO 2 The composition of the reagent: 8.0mM phosphoenolpyruvate, 1.6mM NADH, 1000-1200U / L phosphoenolpyruvate carboxylase and 200-300U / L malate dehydrogenase, Tris-HCl buffer, pH 8.0.

[0053] [2], see formula (1).

[0054] calculate:

Embodiment 2

[0056] According to the following table 2, the experiment was carried out.

[0057] Table 2

[0058]

[0059] [1] and [2] see Example 1.

[0060] calculate:

Embodiment 3

[0062] According to Table 3 below, experiments were carried out.

[0063] table 3

[0064]

[0065] [1] and [2] see Example 1.

[0066] calculate:

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Abstract

The present invention relates to a method to calculate oxalate decarboxylase activity by detecting carbon dioxide concentration, which comprises 1) mixing substrate, oxalate decarboxylase, and HAC buffer at a temperature between of 30-37 degree C for 0.5 to 1 hour; 2) confecting Tris-HCl buffer at pH values between 7.3-8.5 containing phosphate enol type pyruvate, phosphoenolpyruvate carboxylase, NADH, malate dehydrogenase at a temperature between of 30-37 degree C, after incubation, detecting A1 at 380 nm location; 3) adding the reaction solution of step 1) into the solution of step 2), reacting at a temperature between of 30-37 degree C and detecting A2 at 380 nm location; 4) calculating NADH decrement according to the formula of delta A380nm=A1-A2, and calculating the generated carbon dioxide consistence according to the reaction principles to calculate oxalate decarboxylase activity. The said method is a good method for dectecting oxalate decarboxylase which is credibility, high sensitivity, and the valve detected thereby is close to the value detected by the classical method for detecting oxalate decarboxylase.

Description

technical field [0001] The invention belongs to a method for measuring the activity of oxalate decarboxylase, in particular to a method for determining the activity of oxalate decarboxylase by measuring the carbon dioxide concentration of an enzyme reaction product. Background technique [0002] At present, the determination methods of oxalate decarboxylase activity mainly include HPLC, spectrophotometry, and pressure measurement. Enzyme activity can be obtained by measuring the amount of formic acid produced by the decomposition of substrate oxalate, the amount of carbon dioxide, or the amount of reduction in oxalate decomposition. Formic acid can be measured by HPLC or formate dehydrogenase method, carbon dioxide can be measured by manometer, and oxalic acid can be measured by HPLC or oxalate oxidase method. The HPLC method mainly has relatively high requirements on instruments and samples, which is not conducive to popularization; the oxalate oxidase in the enzymatic det...

Claims

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Application Information

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IPC IPC(8): G01N21/31G01N21/75
Inventor 洪枫朱翠侠
Owner DONGHUA UNIV
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