Method for measuring content of cozymase I and derivative thereof

A technology of derivatives and coenzymes, which can be used in the determination/inspection of microorganisms, biochemical equipment and methods, and measurement of color/spectral properties, etc., and can solve problems such as troublesome operation.

Active Publication Date: 2006-11-22
阿里生物技术泰州有限公司
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Problems solved by technology

A method for determining the content of coenzyme I using this principle has been reported, but alcohol dehy...

Method used

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Experimental program
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Embodiment 1

[0022] Embodiment 1: Determination of coenzyme I

[0023] Prepare 0.06M, pH8.9 sodium pyrophosphate buffer solution, which contains 0.03% hydrazine hydrate in volume percentage. Add the substrate and the solution to be measured, the substrate is androsterone, the concentration is 0.02mg / ml, the concentration of the measurement solution is 0.01-0.3mg / ml; on the spectrophotometer, after zeroing at 340nm, add 0.3 U / ml of 3α-steroid dehydrogenase, react until the absorbance value no longer increases, and calculate the content of coenzyme I according to the change in absorbance value. Calculate the content of coenzyme I and its derivatives according to the following calculation formula, the calculation formula is: sample content of liquid to be measured (w / v)=(A-A 0 )×V 1 ×663 / (6220×V 2 )×100%, where A is the final absorbance value of the reaction with 3α-steroid dehydrogenase, A 0 is the final absorbance value without 3α-steroid dehydrogenase, V 1 is the reaction volume, 663 ...

Embodiment 2

[0025] Embodiment 2: Determination of thio-coenzyme I

[0026] The determination is carried out under the protection from light, and 0.03M, pH8.0 sodium pyrophosphate buffer solution is configured, which contains 0.01% hydrazine hydrate in volume percentage. Add the substrate and the solution to be measured, the substrate is sodium cholate, the concentration is 0.005mg / ml, the concentration of the measurement solution is 0.01-0.3mg / ml; on the spectrophotometer, after zeroing at 398nm, add according to the reaction volume 0.05U / ml of 3α-steroid dehydrogenase, react until the absorbance value no longer increases, and calculate the content of coenzyme I according to the change in absorbance value. Calculate the content of coenzyme I and its derivatives according to the following calculation formula, the calculation formula is: sample content of liquid to be measured (w / v)=(A-A 0 )×V 1 ×680 / (11900×V 2 )×100%, where A is the final absorbance value of the reaction with 3α-steroid...

Embodiment 3

[0027] Embodiment 3: Determination of 3-acetylpyridine coenzyme I

[0028] A 0.2M, pH 11.0 sodium pyrophosphate buffer solution is prepared, which contains 0.3% hydrazine hydrate in volume percentage. Add the substrate and the solution to be measured, the substrate is sodium deoxycholate, the concentration is 0.08mg / ml, the concentration of the measurement solution is 0.01-0.3mg / ml; Add 1U / ml of 3α-steroid dehydrogenase, react until the absorbance value no longer increases, and calculate the content of coenzyme I according to the change in absorbance value. Calculate the content of 3-acetylpyridine coenzyme I according to the following calculation formula, the calculation formula is: sample content of liquid to be measured (w / v)=(A-A 0 )×V 1 ×662 / (7800×V 2 )×100%, where A is the final absorbance value of the reaction with 3α-steroid dehydrogenase, A 0 is the final absorbance value without 3α-steroid dehydrogenase, V 1 is the reaction volume, 662 is the molecular weight of...

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Abstract

The disclosed content detection method for coenzyme I and its derivative comprises: preparing sodium pyrophosphate buffer with 8.0-11.0 pH value contained 0.01-0.3v% hydrazine hydrate; adding substrate solution and the target solution; with a spectrophotometer, after the maximal absorption peak, adding 3alpha-steroid dehydrogenase till absorbance stopping; calculating the content according to absorbance calculation.

Description

technical field [0001] The invention relates to a method for determining the content of coenzyme I and its derivatives. Background technique [0002] A change in the characteristic absorption spectrum may occur when the oxidized coenzyme I and the reduced coenzyme I are bidirectionally converted. Under certain conditions, it can change linearly with the concentration of the analyte. Many chemical analysis methods are realized based on this principle, but there are many defects in the chemical properties of oxidized / reduced coenzyme I. The chemical properties of oxidized / reduced coenzyme I derivatives have changed greatly, (mainly manifested as changes in maximum absorption peaks, increases in molar absorbance, changes in reaction Km values, and greatly increased solution stability and stable pH range.) The maximum absorption wavelength of the reduced coenzyme I derivatives is between 340-405nm, and the oxidation-reduction reaction participated by the oxidized / reduced coenzy...

Claims

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Application Information

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IPC IPC(8): G01N21/31C12Q1/26
Inventor 闻树群朱永良
Owner 阿里生物技术泰州有限公司
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