Amylase activity assay

There are many methods for amylase activity detection, which vary according to the application field and detection purpose. The following are some common methods for amylase activity detection:

1. DNS method (3,5-dinitrosalicylic acid method)
The DNS method is one of the most commonly used methods for detecting amylase activity. The principle is that amylase catalyzes starch degradation to produce reducing sugars, which react with DNS reagents to produce reddish-brown compounds. By measuring the absorbance of the compound, the activity of amylase can be quantified.

2. Iodine-starch colorimetric method
This method uses the property that starch produces blue when it encounters iodine. By adding different amounts of amylase preparations to a certain amount of starch, after a certain period of time, the test solution that does not appear blue and has the least amount of enzyme preparation can be considered to have just completely decomposed the starch. The activity of the amylase is calculated based on the amount of enzyme preparation, the amount of starch, and the reaction time.

3. Capillary electrophoresis method
Capillary electrophoresis can efficiently separate the products of amylase-catalyzed degradation. By measuring the peak area of ​​each product in the sample, the activity of amylase can be indirectly inferred. This method has the advantages of high resolution and rapidity.

4. ELISA (Enzyme-Linked Immunosorbent Assay)
ELISA can detect specific amylases and perform quantitative analysis through the signals generated by antigen-antibody reactions. This method has high sensitivity and is suitable for detecting low concentrations of amylase, but requires specific antibodies.

5. HPLC (High Performance Liquid Chromatography)
HPLC can be used to separate and quantify the degradation products after the action of amylase, thereby indirectly measuring the activity of amylase. This method has high accuracy and is suitable for analyzing complex samples.

6. Biochemical Oxidation
In fermentation or wastewater treatment, biochemical oxidation is often used to determine the activity of amylase in samples. By measuring the biochemical oxygen demand (BOD) of the sample, the ability of amylase to degrade starch can be reflected.

7. Conversion Assay
This method expresses the activity of amylase by measuring its conversion rate. The conversion rate of an enzyme refers to the mass of starch that can be converted by a unit mass of amylase within a certain period of time. In actual operation, the activity of amylase can be obtained through a series of experimental steps and calculations.

8. Reducing sugar production method
This method uses a-amylase to randomly act on a-1,4-glycosidic bonds in starch to generate reducing sugars such as glucose and maltose. These reducing sugars can react with 3,5-dinitrosalicylic acid and change its color. By measuring the color change of the solution after the reaction, the activity of amylase can be calculated.

It should be noted that different detection methods have their own advantages and disadvantages. The selection of a suitable detection method depends on the type of sample, the accuracy requirements of the analysis, and the experimental conditions. At the same time, when performing amylase activity detection, the operating procedures should be strictly followed to ensure the accuracy and reliability of the test results.