Preparation method of immobilized enzyme nanofiber membrane for detecting alcohol content in saliva

Abstract

The invention discloses a preparation method of an immobilized enzyme nanofiber membrane for detecting the alcohol content in spit. The preparation method comprises the steps of: covalently binding alcohol oxidase through a polymer such as polyethylene glycol or polyethyleneimine as an arm molecular multi-site; further fixing a plurality of subunits of the alcohol oxidase by the polyethyleneimine, polylysine, poly-histidine or chitosan and the like through physical action among charges; then absorbing indigo blue as a chromogenic substrate; generating hydrogen peroxide after ethanol in a sample is oxidized by the alcohol oxidase in detection; leading the indigo blue to fade by the hydrogen peroxide, and conveniently calculating the ethanol concentration in the sample by comparing the fading degree of the indigo blue with a standard color atla. The preparation method is high in sensitivity, simple and convenient to operate, and low in price, can be repeatedly used for a plurality of times, and can be conveniently used for investigating and treating drunk driving by a law-enforcing department; and the reliable evidence is provided for the law enforcement process.

Description

technical field [0001] The invention belongs to the technical field of alcohol concentration detection, in particular to a preparation method of an immobilized enzyme nanofiber membrane for detecting alcohol content in saliva. Background technique [0002] Many industries require qualitative or quantitative detection and analysis of ethanol content. For example, various countries have strict requirements on the blood alcohol content of motor vehicle drivers. In my country, a driver whose blood alcohol concentration reaches 0.2mg / ml is considered to be drunk driving, and when the alcohol concentration reaches 0.8mg / ml, it is considered to be drunk driving. In recent years, tragedies of drunk driving causing tragic accidents are not uncommon. Although the traffic law enforcement department has stepped up law enforcement and punishment, there are always drivers who take risks; in addition, some drivers drive illegally because they cannot accurately judge their own alcohol cont...

AI Technical Summary

Problems solved by technology

This preparation method is simple to operate, but due to the dense filter paper, the enzyme is only physically adsorbed on the surface of the filter paper, resulting in poor thermal stability and acid-base stability, resulting in large detection errors and inaccurate detection results.

At the same time, because the surface of the filter paper is dense, the specific surface area is small, the amount of adsorbed enzyme is small and it is easy to fall off, which makes the detection sensitivity low and the detection error is large.

Moreover, alcohol oxidase is composed of eight subunits, and any subunit of the enzyme falls off during immobilization and use, which will cause the loss of enzyme activity to varying degrees and increase the detection error; while all eight subunits are immobilized On the base carrier, the spatial structure of the enzyme will be destroyed, and the steric hindrance of the enzyme when it acts on the substrate will be increased, thereby affecting the activity of the enzyme and reducing its detection sensitivity and accuracy.

Therefore, the current immobilized alcohol oxidase detection system fails to meet the requirements of accuracy, sensitivity, and repeated use.

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