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Method for determining concentration of oxalic acid by determining concentration of carbon dioxide through enzymatic method

A technology of carbon dioxide and concentration, which is applied in the field of enzyme analysis of oxalic acid concentration, can solve the problems of high cost of oxalate oxidase and restrictions on wide-scale popularization and application, and achieve the effect of low cost, close accuracy and high sensitivity

Inactive Publication Date: 2010-04-07
DONGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the high cost of oxalate oxidase, the wide-scale application of this method has been greatly restricted.

Method used

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  • Method for determining concentration of oxalic acid by determining concentration of carbon dioxide through enzymatic method
  • Method for determining concentration of oxalic acid by determining concentration of carbon dioxide through enzymatic method
  • Method for determining concentration of oxalic acid by determining concentration of carbon dioxide through enzymatic method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033]

[0034] Note: [1] CO 2 The composition of the assay reagent: 8.0mM phosphoenolpyruvate, 1.6mMNADH, 1000-1200U / L phosphoenolpyruvate carboxylase and greater than 200-300U / L malate dehydrogenase, 66mM Tris-HCl buffer solution, pH 8.0; [2] see formula (1).

[0035] calculate:

Embodiment 2

[0037]

[0038] Note: [1] CO 2 Composition of assay reagents: 8.0mM phosphoenolpyruvate, 1.6mM NADH, 1000-1200U / L phosphoenolpyruvate carboxylase and greater than 200-300U / L malate dehydrogenase, 66mM Tris-HCl Buffer, pH 8.0; [2] See Equation (1).

[0039] calculate:

Embodiment 3

[0041]

[0042] Note: [1] CO 2 The composition of the determination reagent: 8.0mM phosphoenolpyruvate, 1.6mMNADH, 1000-1200U / L phosphoenolpyruvate carboxylase and malate dehydrogenase greater than 200-300U / L, 66-100mM Tris- HCl buffer, pH 8.0; [2] See formula (1).

[0043] calculate:

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Abstract

The invention relates to a method for determining the concentration of oxalic acid by determining the concentration of carbon dioxide through an enzymatic method. The method comprises the following steps: (1) carrying out reaction of the mixture of oxalic acid, oxalate decarboxylase and acetate buffer solution for 30-60min at a temperature of 20-37 DEG C; (2) preparing Tris-HCI buffer solution containing phosphoenolpyruvate, phosphoenolpyruvate carboxylase, NADH, malic dehydrogenase, with a pH value of 7.3-8.5, incubating at a temperature of 37 DEG C and determining A1 at the position of 380nm; (3) adding the reaction solution prepared in the step (1) to the solution prepared in the step (2) for reaction for 5min at a temperature of 37 DEG C and determining A2 and A3 at the position of 380nm; and (4) calculating the decrement of NADH according to the formula: delta A 380nm = delta A to be determined - delta A blank, and calculating the concentration of the generated carbon dioxide according to a reaction principle so as to calculate the concentration of the oxalate decarboxylase. The method has simple operation, low cost and high sensitivity; the enzyme reagent adopted in the method is cheaper; and the method has similar accuracy compared with that of the existing kits for determining oxalic acid and has good practicability.

Description

technical field [0001] The invention belongs to the field of enzyme analysis of oxalic acid concentration, in particular to a method for determining the concentration of oxalic acid by measuring carbon dioxide concentration by enzymatic method. Background technique [0002] At present, the determination methods of oxalic acid content mainly include titration, atomic absorption method, colorimetry, enzymatic method and high pressure liquid chromatography (HPLC), in addition to ion chromatography, capillary electrophoresis (CE) and so on. [0003] The titration method was established the earliest, and as a classic method is still widely used in food analysis, but it has not developed much, and its sensitivity is low, and the defects of cumbersome operation and time-consuming have not been improved. Colorimetry is cheap and highly sensitive, but it is difficult to completely extract oxalic acid by pre-precipitation, and the operation is cumbersome and has poor repeatability; ca...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/527C12Q1/32G01N21/33
Inventor 洪枫
Owner DONGHUA UNIV
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