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Method for determining alcohol concentration by using enzyme cycling method and alcohol determination kit

A technology of ethanol concentration and cyclic enzymatic method, applied in the field of medical testing, can solve the problems of poor specificity and large interference in measurement, and achieve the effect of high accuracy and good stability

Inactive Publication Date: 2012-07-11
NINGBO MEDICAL SYSTEM BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The enzymatic automatic analysis method is divided into alcohol oxidase method and alcohol dehydrogenase method. The reagent composition of the oxidase method is simple, and the sample does not need pretreatment, but the specificity is poor. Methanol interferes greatly with the determination, and false positives may occur

Method used

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  • Method for determining alcohol concentration by using enzyme cycling method and alcohol determination kit
  • Method for determining alcohol concentration by using enzyme cycling method and alcohol determination kit
  • Method for determining alcohol concentration by using enzyme cycling method and alcohol determination kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] The ethanol assay kit of the present embodiment is a single reagent and consists of the following components:

[0028]

[0029] After the reagents are all dissolved and prepared, they are divided into bottles and freeze-dried to make dry powder reagents; before use, add purified water and reconstitute for use.

[0030] The testing conditions for the reagent of the present invention to measure ethanol in a sample are as follows: temperature: 30-37° C.; light path of cuvette: 1.0 cm. The main detection wavelength is 505nm, and the secondary wavelength is 700nm.

[0031] The method for measuring ethanol in a sample by using the ethanol determination reagent of the present invention is as follows: add the single reagent prepared above to the sample (calibration tube as the sample), read the absorbance A0 immediately after mixing, and read the Absorbance A1, ΔA=A1-A0. Among them, the sample volume is 5-10 μl, and the single reagent volume is 300 μl.

Embodiment 2

[0033] The ethanol assay kit of the present embodiment is a double reagent, consisting of reagent 1 and reagent 2, wherein:

[0034] Reagent 1

[0035] Tris buffer 0.2M

[0036] Trehalose 5g / L

[0037] NAD + 6g / L

[0038] Reagent 2

[0039]

[0040] After the reagents are all dissolved and prepared, they are divided into bottles to make liquid double reagents, which can be used directly.

[0041] Test condition is the same as embodiment 1.

[0042] The method for measuring ethanol in a sample by using the ethanol determination reagent of the present invention is as follows: add reagent 1 to the sample (calibration tube as a sample) and mix well, incubate at 30-37°C for 3-5min and read the absorbance A0, immediately add reagent 2 and mix well , After reacting at 30-37°C for 5 minutes, read the absorbance A1, ΔA=A1-A0. Among them, the sample volume is 30 μl, the reagent 1 volume is 225 μl, and the reagent 2 volume is 75 μl.

Embodiment 3

[0044] The ethanol determination kit of the present embodiment is three reagents, is made up of reagent 1, reagent 2 and reagent 3, wherein:

[0045] Reagent 1

[0046] 3-Trihydroxymethylmethylamine-2-hydroxypropanesulfonic acid buffer 0.5M

[0047] Glycerin 3g / L

[0048] NADP + 5g / L

[0049] Reagent 2

[0050]

[0051] Reagent 3

[0052] Oxidized INT 10g / L

[0053] After all the reagents are dissolved and prepared, they are divided into bottles to make liquid three reagents, which can be used directly.

[0054] Test condition is the same as embodiment 1.

[0055] The method for measuring ethanol in a sample by using the ethanol determination reagent of the present invention is as follows: add reagent 1 and reagent 2 to the sample (calibration tube as a sample) and mix well, read the absorbance A0 after incubating at 30-37°C for 3-5min, and immediately add the reagent 3 Mix well, and after reacting at 30-37°C for 5 minutes, read the a...

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Abstract

The invention provides a method for determining alcohol concentration by using an enzyme cycling method and an alcohol determination kit. The method comprises the following steps of: generating reduced nicotinamide adenine dinucleotide I by using nicotinamide adenine dinucleotide I and alcohol under the action of alcohol dehydrogenase, reacting the reduced nicotinamide adenine dinucleotide I with oxidized iodonitrotetrazolium (INT) under the action of diaphorase to generate red reduced INT, detecting a change in the absorbance of a final reactant, and then calculating the alcohol concentration of a sample. The kit consists of the following components: 0.05 to 0.5M of a buffer solution, 0.1 to 10g / L of a stabilizer, 0.05 to 10g / L of nicotinamide adenine dinucleotide I, 0.5 to 50KU / L of alcohol dehydrogenase, 0.05 to 50KU / L of diaphorase, 0.1 to 10g / L of oxidized INT and 0.1 to 10g / L of a preservative. The method is high in reaction specificity, detection specificity and sensitivity, wide in linear range and low in cost; and the kit is high in stability and accuracy.

Description

technical field [0001] The invention relates to the technical field of medical testing, in particular to a method for measuring ethanol concentration by a circulating enzyme method, and an ethanol measuring kit prepared by using the method. Background technique [0002] Ethanol (alcohol) exists in most alcoholic beverages, and its metabolic rate mainly depends on the content of enzymes in the body, with large individual differences. As we all know, drinking a small amount of alcohol can promote blood circulation, but a large amount of alcohol intake can lead to alcohol dependence, which can cause damage to mental and multiple organ functions, among which the damage to liver function and mental function is the most common and serious. The addiction, abuse and dependence of ethanol is one of the most serious socioeconomic and public health problems in the world today, and alcohol-related liver disease has become an important cause of death in western countries. Therefore, the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/31
Inventor 邹炳德沃燕波
Owner NINGBO MEDICAL SYSTEM BIOTECHNOLOGY CO LTD
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