JPS634661B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a method for quantifying α-amylase in a sample, and particularly to a method for quantifying α-amylase activity in a sample containing α-amylase using an enzyme electrode. Quantitative measurement of α-amylase activity provides useful knowledge in the field of clinical medicine. Therefore, this type of quantitative method is generally quick and simple;
High selectivity is desired. Conventionally, two types of α-amylase measurement methods are known: a non-enzymatic method and an enzymatic method. for example,
As non-enzymatic methods, a satscarogenic method that measures the increase in reducing sugars by α-amylase, an amyloclastic method that measures the amount of residual starch by an iodine reaction, and a chromogenic method that uses dye-bound starch as a substrate are used. ing. but,
These non-enzymatic methods are characterized by (a) long reaction times; (b)
There are disadvantages such as unstable color development, (c) insufficient reproducibility and selectivity, and (d) need for centrifugation and excessive work. On the other hand, the enzymatic method determines the α-amylase content in a sample by a combination of enzymatic reactions, and compared to the non-enzymatic method described above, it has (a) higher selectivity and (b) a lower reaction rate. It has characteristics such as shortening of time. However, since conventional enzymatic methods use colorimetry as a quantitative method, samples with high α-amylase activity must be diluted before measurement, and pipetting during dilution can contaminate the sample. There are some difficulties. Furthermore, when measuring the activity of α-amylase in blood by a colorimetric method, it is essential to prepare a serum sample by centrifugation, and there is a drawback that, for example, it is impossible to measure the whole blood sample as it is. Therefore, as a result of various studies in order to overcome the problems of the conventional method for quantifying α-amylase content mentioned above, the present inventors have determined that α-amylase reaction is possible by selecting a predetermined reagent and a predetermined enzymatic reaction. Glucose is generated based on
It has been found that α-amylase can be continuously quantified together with glucose in a sample. That is, maltopentaose and α-
Glucosidase is added and the enzymatic reaction is carried out using a phosphate buffer, preferably at pH 7, while a hydrogen peroxide electrode with an immobilized glucose oxidase membrane is used before and after the addition of α-glucosidase. By detecting the reaction current and comparing these current values, the α-amylase content can be quantified. The enzymatic reaction in this case is as follows.

[Table] Glucose oxidase
(3) Glucose

Claims (1)

[Claims] 1. A predetermined amount of sample is reacted with maltopentaose and α-glucosidase, and the resulting glucose is detected as a change in current with an enzyme electrode consisting of a hydrogen peroxide electrode having an immobilized glucose oxidase membrane. α by the enzyme electrode method, which is characterized by
-Method for quantifying amylase. 2. A method for quantifying α-amylase according to claim 1, using an enzyme electrode method using a phosphate buffer with a pH of 7.