CN108398356A - A kind of quick judgement cereal inner enzyme vigor and the method for predicting enzyme thermophilic degree - Google Patents

Abstract

The invention discloses a method for quickly determining enzyme activity in grains and predicting enzyme optimum temperature, and relates to the technical field of grain quality evaluation and determination methods. It mainly includes the following steps: (1) pretreatment; (2) moisture content determination and sample weighing; (3) rapid viscosity analyzer temperature program setting; (4) blank sample and enzyme inhibitor added sample to measure intrinsic viscosity; (5) Determine the enzyme activity according to the obtained comparison characteristic spectrum; (6) Determine the optimum temperature of the enzyme in the grain by the difference of the enzyme activity at different temperatures. The invention provides a novel method for measuring enzyme activity and enzyme temperature in grains, which has the characteristics of simple operation, fast and accurate, good repeatability, low cost and time saving, and is beneficial to improve the efficiency of grain quality analysis , which is conducive to improving the operability of quality determination, avoiding the waste of manpower and material resources, and providing theoretical guidance for grain processing, storage and application.

Description

A method for quickly determining enzyme activity in grains and predicting enzyme optimum temperature

technical field

The invention relates to a method for rapidly determining enzyme activity in grains and predicting enzyme optimum temperature, and belongs to the technical field of quality evaluation and determination methods.

Background technique

my country is rich in grain resources and has a wide variety of varieties, especially rice and wheat are the main grain crops in my country, and they are also the grain species with the largest circulation and consumption in my country. Their output and quality are directly related to agricultural production and people's living standards. As people's requirements for food quality are getting higher and higher, the quality of different grains is also generally concerned by people.

The activity of endogenous enzymes in grains is an important indicator for its quality control and application. There are differences in the enzyme activities of different varieties and origins of grains, which will lead to the application properties of different varieties and origins of grains. The degree of germination damage of grains can be understood and judged through the measurement of enzyme activity. Taking wheat as an example, the edible quality of wheat with high germination damage is poor, and vice versa, the quality is good. At the same time, grains must be stored for a certain period of time during transportation, and due to the influence of storage conditions, slow metabolism will inevitably occur during the storage process, and the biochemical reactions that occur are mainly carried out under the action of various enzymes. If the enzyme activity in the grain is weakened or lost, it will age, resulting in poor quality. Studies have shown that the α-amylase activity of rice has an exponential relationship with its non-reducing sugar content, germination rate, viscosity, and fatty acid value. Therefore, α-amylase is also used as an important reference index when measuring the change of its freshness during grain storage.

At present, there are many methods for the determination of enzyme activity, and the principles of the methods are also different. There are mainly iodine colorimetry and falling number method. Among them, the iodine colorimetric method has a high degree of variation, large errors, and takes a long time. , in comparison, the falling number method has high precision and small coefficient of variation, so it is widely used in the control of wheat quality, but most of its methods use wheat as raw material to measure the activity of α-amylase, especially the activity of high-temperature amylase. However, the enzymes in grains are a mixture, and different enzymes have an impact on the quality of grains, but there are few determinations for the total enzyme activity of a wide range of grains.

Contents of the invention

In view of the above-mentioned problems existing in the prior art, the applicant of the present invention provides a simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme. The present invention uses Rapid Visco Analyzer (Rapid Visco Analyzer, RVA) as analysis means, this instrument is mainly used to measure enzyme activity to judge wheat germ damage at first, later stage is widely used in the research of the viscosity characteristic of starch and its derivatives, it not only It has good comparability with the test results of falling number instrument, and also has good correlation with the application of grain in different systems. The operation of the instrument is simple, the measurement process is short, the amount of sample is small, and the most important thing is that it can realize multi-stage precise control of temperature, which has laid a good foundation for it in the measurement of enzyme activity and the prediction of the optimal temperature of the enzyme.

Technical scheme of the present invention is as follows:

The first object of the present invention is to provide a method for quickly measuring enzyme activity and enzyme temperature in grains, said method is characterized by the degree of viscosity drop during heating and gelatinization of grains before and after the enzyme is eliminated, that is, the relative value of peak viscosity drop The parameters of enzyme activity and enzyme optimum temperature, the larger the parameter, the higher the enzyme activity; said enzyme activity refers to the general term of enzymes that reduce the viscosity of each component in the grain; The temperature at which the viscosity decreases the most; the components include at least one of starch, protein, fat, polysaccharide, especially starch.

In one embodiment of the present invention, the enzyme that reduces the viscosity of each component includes at least one of amylase, pectinase, cellulase, protease, and lipase.

In one embodiment of the present invention, the enzyme activity refers to the general term for the degree of viscosity decrease of grains during storage; the enzyme optimum temperature refers to the temperature at which the degree of viscosity decrease is the greatest during the storage process.

In one embodiment of the present invention, the enzyme activity refers to the general term for enzymes that cause the barley viscosity to decrease when the barley is transformed into a malt state; the enzyme optimum temperature refers to the temperature at which the barley germinates and the malt viscosity decreases most obviously.

In one embodiment of the present invention, the enzyme activity refers to the general term of the enzymes that soak rice to the state used to prepare rice flour to reduce the viscosity of rice flour; the enzyme optimum temperature refers to the maximum degree of enzyme degradation during rice soaking temperature.

In one embodiment of the present invention, the enzyme activity refers to the general term for enzymes that cause the viscosity of wheat flour to decrease during wheat processing to wheat products; temperature.

In one embodiment of the present invention, the method includes the following steps: (1) measuring the moisture content of the grain sample, and converting the sample dry basis; (2) taking the moisture content obtained in step (1) as a benchmark, measuring The corresponding viscosity growth value of the grain samples before and after at 30-100°C; (3) calculate the activity of the enzyme by looking for the degree of peak viscosity change during the viscosity change process, (4) calculate the optimum temperature of the enzyme through the degree of peak viscosity change in different temperature ranges.

In one embodiment of the present invention, described method comprises the following specific steps:

(1) Pre-treatment: the grain sample is ground and passed through a 100-mesh sieve;

(2) Determining the moisture content of the sample, taking 14% as the moisture basis, weighing 3-5g of the dry sample;

(3) Set enzyme activity determination program and optimal temperature determination program

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The sample is placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum;

(5) According to the RVA characteristic spectrum, refer to the method described in the AACC22-08 standard to calculate the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area; wherein, the test sample can be compared with the The peak viscosity increase value of the control sample is used to calculate the enzyme activity, i.e. (P _test -P _contrast ) × 100/P _contrast , wherein P represents the peak viscosity;

(6) Calculate the optimum temperature of the enzyme according to the area in the RVA characteristic spectrum, that is, the temperature corresponding to the maximum growth of the area formed by the characteristic curve is the optimum temperature.

In one embodiment of the present invention, the program of assaying enzyme activity is as follows any one:

Program 1: Keep warm at 60°C for 1 minute, increase the speed at 12°C/min to 95°C and keep it for 2.5 minutes, then decrease the same rate to 50°C and hold it for 1.4 minutes, and keep the stirring speed at 160r/min;

Program 2: Keep warm at 60°C for 5 minutes, increase the speed at 12°C/min to 95°C and hold for 5 minutes, then decrease the same rate to 50°C and hold for 3 minutes, and keep the stirring speed at 160r/min;

Among them, procedure 1 is suitable for the determination of enzyme activity in starch extract; procedure 2 is suitable for the determination of enzyme activity in whole grain flour.

In one embodiment of the present invention, the procedure for determining the optimum temperature is as follows: set different holding times respectively, keep warm at 30-90°C for 5 minutes, then rise to 95°C at a speed of 12°C/min and keep for 2.5 minutes, then similarly The speed was lowered to 50°C for 1.4 minutes, and the stirring speed was kept at 160r/min.

In one embodiment of the present invention, the types of cereals in the step (1) include: rice, wheat, corn, buckwheat, oats, quinoa and starch extracts of rice, wheat, corn, buckwheat, oats, quinoa .

In one embodiment of the present invention, the sample weight of the starch extract in the step (2) is weighed as 3-4g, while the sample weight of the whole grain flour is controlled at 4-5g, and the same batch of comparative samples needs to weigh the same A mass (dry basis) of the sample was used as a control.

In one embodiment of the present invention, the enzyme inhibitors described in step (4) include but are not limited to: compounds containing carbon-phosphorus bonds, phosphoric acid derivatives containing organic groups, silver nitrate, copper sulfate, chloride One or more mixed compounds of copper and iron sulfate.

In one embodiment of the present invention, the concentration of the enzyme inhibitor in step (4) is controlled at 0.01-2mM.

The invention also provides the application of the method in grain quality control, storage, breeding, fermentation and determination of variety and origin.

Beneficial technical effect of the present invention is: the present invention provides a kind of assay method of enzyme activity and enzyme optimum temperature contained in the novel grain, can measure the activity and enzyme optimum temperature of the enzyme in the grain while studying the gelatinization characteristic of grain, Compared with chemical colorimetry, it has the characteristics of simple operation, fast and accurate, good repeatability, small error, low cost and time saving. Compared with the falling number instrument, this method can measure the enzyme activity of the total enzyme in the grain, and can understand the optimal temperature of the enzyme through the comparison of different temperatures, and has a better understanding of the quality of the grain, which is conducive to improving the analysis of grain quality. Efficiency is conducive to improving the operability of quality determination, avoiding waste of manpower and material resources, and providing theoretical guidance for grain processing, storage and application.

Description of drawings

Fig. 1 is the determination of the rice amylase activity measured in embodiment 1;

Fig. 2 is the rice flour optimum enzyme activity temperature measured in embodiment 2;

Fig. 3 is that the optimum enzyme activity temperature measured in embodiment 2 is compared with other temperatures, the change of rice hardness under different soaking temperatures;

Fig. 4 is the measured corn flour enzyme activity assay of embodiment 3;

Fig. 5 is the measured wheat flour enzyme activity determination of embodiment 4;

Fig. 6 is the determination of enzyme activity before and after germination of barley measured in Example 5.

Detailed ways

Example 1

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Rice pretreatment: extract rice starch, the starch content is 95%, and pass through a 100-mesh sieve.

(2) Determining the water content of the sample, taking 14% as the water content, weighing 3 g of the sample on a dry basis.

(3) The specific procedure is as follows: keep warm at 60°C for 1min, increase the speed at 12°C/min to 95°C and hold for 2.5min, then decrease the same speed to 50°C and hold for 1.4min, and keep the stirring speed at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) Calculate the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area according to the RVA characteristic spectrum and refer to the method described in the AACC22-08 standard. Wherein, the enzyme activity can be calculated by the increase value of the peak viscosity of the test sample relative to the control sample, that is, (P _test -P _control )×100/P _control , wherein P represents the peak viscosity.

The results are shown in Figure 1. The peak viscosity of rice starch increased significantly after the enzyme was eliminated, that is, there was high enzyme activity in rice starch, and the peak viscosity increased from 6500cp to 7579cp. The calculated enzyme activity was 16,6.

Example 2

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pre-treatment: the rice sample is pulverized and passed through a 100-mesh sieve.

(2) Determining the water content of the sample, taking 14% as the water content, weighing 4 g of the sample on a dry basis.

(3) The specific procedure is as follows: set different holding times respectively, keep warm at 30-90°C for 5 minutes, then rise to 95°C at a speed of 12°C/min and hold for 2.5 minutes, then drop to 50°C at the same rate for 1.4 minutes, stir The speed is kept at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) Calculate the optimum temperature of the enzyme according to the degree of viscosity decrease in the RVA characteristic spectrum, that is, the temperature corresponding to the most obvious viscosity decrease trend (the maximum increase in the area formed by the characteristic curve) is the optimum temperature.

The results are shown in Figure 2. At different temperatures, the difference in peak viscosity before and after enzyme inactivation is different. It can be seen from the figure that the optimum temperature for enzyme in rice flour is 60-70°C.

The rice hardness test method (three-point test method, TA.XT-Plus, P/36 probe) proposed by Japanese Okabe scholars and widely used in the world was used to measure the change of rice hardness with soaking temperature and time. It can be seen from Figure 3 that, At the optimum temperature for enzyme action, the hardness of rice decreased most obviously (from 30499g to 11210g), but at a lower temperature, such as 30°C, the decrease was not significant (32174g to 25524g). The optimal temperature determination method proposed by the invention can be applied to guide the processing control of rice.

Example 3

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pre-treatment: the corn sample is ground and passed through a 100-mesh sieve.

(2) Determining the water content of the sample, taking 14% as the water content, weighing 5 g of the sample on a dry basis.

(3) The specific procedures are as follows:

Incubate at 60°C for 5 minutes, increase the speed at 12°C/min to 95°C and maintain for 5 minutes, then decrease the same rate to 50°C and maintain for 3 minutes, and keep the stirring speed at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) According to the RVA characteristic spectrum, calculate the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area by referring to the method described in the AACC22-08 standard. Wherein, the enzyme activity can be calculated by the increase value of the peak viscosity of the test sample relative to the control sample, that is, (P _test -P _control )×100/P _control , wherein P represents the peak viscosity.

The results are shown in Figure 4. The peak viscosity of corn flour also increased significantly after deactivating the enzyme, that is, there was high enzyme activity in the corn flour. After deactivating the enzyme, the viscosity increased from 1450cp to 1679cp, and the calculated enzyme activity was 15.8.

Example 4

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pretreatment: take wheat samples for milling treatment, and pass through a 100-mesh sieve.

(2) Determining the water content of the sample, taking 14% as the water content, weighing 5 g of the sample on a dry basis.

(3) The specific procedure is as follows: keep warm at 60°C for 1min, increase the speed at 12°C/min to 95°C and hold for 2.5min, then decrease the same speed to 50°C and hold for 1.4min, and keep the stirring speed at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) According to the RVA characteristic spectrum, the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area can be obtained. Wherein, the enzyme activity can be calculated by the increase value of the peak viscosity of the test sample relative to the control sample, that is, (P _test -P _control )×100/P _control , wherein P represents the peak viscosity.

The results are shown in Figure 5. The peak viscosity of wheat flour also increased significantly after deactivating the enzyme, that is, there was high enzyme activity in the wheat flour. After deactivating the enzyme, the viscosity increased from 2616cp to 3158cp, and the calculated enzyme activity was 20.7.

Example 5

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pretreatment: mill the barley samples before and after germination, and pass through a 100-mesh sieve.

(2) Determining the water content of the sample, taking 14% as the water content, weighing 4 g of the sample on a dry basis.

(3) Keep warm at 60°C for 5 minutes, increase the speed at 12°C/min to 95°C and keep it for 5 minutes, then decrease the same speed to 50°C and keep warm for 3 minutes, and keep the stirring speed at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) According to the RVA characteristic spectrum, calculate the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area by referring to the method described in the AACC22-08 standard. Wherein, the enzyme activity can be calculated by the increase value of the peak viscosity of the test sample relative to the control sample, that is, (P _test -P _control )×100/P _control , wherein P represents the peak viscosity.

The results are shown in Figure 6, the viscosity of the barley before and after germination changed, the viscosity decreased from 2140cp to 1603cp, and the calculated enzyme activity was 33.4.

Example 6

A simple method for quickly determining the enzyme activity in grains and predicting the optimum temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pretreatment: collect 1 kind of wheat flour that germinates in the rainy season, 2 kinds of normally mature wheat flour, and the wheat flour is high-gluten flour (protein content is more than 12%);

(2) Determining the water content of the sample, taking 14% as the water content, weighing 5 g of the sample on a dry basis.

(3) The specific procedure is as follows: keep warm at 60°C for 1min, increase the speed at 12°C/min to 95°C and hold for 2.5min, then decrease the same speed to 50°C and hold for 1.4min, and keep the stirring speed at 160r/min.

(4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The samples were placed in a rapid viscosity analyzer (RVA) to measure the viscosity characteristic spectrum.

(5) According to the RVA characteristic spectrum, the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area are calculated by referring to the method described in the AACC22-08 standard. Wherein, the enzyme activity can be calculated by the increase value of the peak viscosity of the test sample relative to the control sample, that is, (P _test -P _control )×100/P _control , wherein P represents the peak viscosity.

(6) Measure the falling number of the three kinds of flour used according to the method described in GB/T 10361-2008, and measure the viscosity enzyme activity according to the method of the present invention. At the same time, use three kinds of flour to make soft flour under the same operating environment and operating steps Type bread, measure the volume of bread according to the method described in GB/T20981-2007, the bread formula is as follows: 100% flour, 2% yeast, 10% sugar, 1% salt, 6% oil, 57% water.

Use a falling number meter to detect the falling number of wheat flour, and the detection steps are:

(1) measure the moisture content of flour, and take 15% moisture content 7g sample size as correction, take sample according to measured moisture content;

(2) Turn on the falling number measuring instrument and heat the water bath until boiling;

(3) Move the weighed flour sample into a dry and clean viscosity tube, add 25ml of water with an automatic liquid adder, and vibrate evenly;

(4) Put the viscosity tube into the boiling water bath, turn on the stirring, the instrument will automatically complete the test, and record the time displayed on the electronic timer as the falling value.

Table 1 is a comparison table of falling number, viscosity enzyme activity and bread specific volume value measured in the examples. The falling number is inversely proportional to the dough or enzyme activity, that is, the higher the enzyme activity, the lower the falling number, and the viscosity enzyme activity is related to the actual enzyme activity. Proportional. As can be seen from the table, the measured trend of the enzyme activity of the present invention is completely consistent with the measured trend of the falling number. It is generally believed that when the specific volume of bread is around 4.8, the quality of bread is better (≤7), and bread that is too large or too small is not good. It can be seen from the table that the enzyme activity is closely related to the specific volume, and the specific volume of bread can be predicted by measuring the appropriate enzyme activity (falling number 250±9.2, viscosity enzyme activity 20.6±0.2), and then guide the production and processing of bread flour. It can be seen that the error limit of the enzyme activity determined by the present invention has decreased by 10 times.

Table 1 Comparison of the influence of the falling number instrument and the method used in the present invention on the specific volume of bread.

Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Any person familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore The scope of protection of the present invention should be defined by the claims.

Claims (10)

1. A method for quickly measuring enzyme activity and enzyme optimum temperature in cereals, characterized in that, the method is characterized in that the viscosity drop degree is used as the relative value of peak viscosity drop when the heating and gelatinization of cereals before and after the enzyme is eliminated as a characterizing enzyme activity and The parameter of the enzyme suitable temperature, the larger the parameter, the higher the enzyme activity; the enzyme activity refers to the general term of enzymes that reduce the viscosity of each component in the grain; the enzyme suitable temperature refers to the degree of viscosity reduction caused in the grain Maximum temperature; said components include at least one of starch, protein, fat, polysaccharide, especially starch. 2. The method according to claim 1, characterized in that the method comprises the following steps: (1) measuring the moisture content of the grain sample, and converting the dry basis of the sample; (3) Calculate the activity of the enzyme by looking for the degree of peak viscosity change during the viscosity change process, (4) Calculate the enzyme optimum temperature through the degree of peak viscosity change in different temperature ranges. 3. The method according to claim 1 or 2, characterized in that, comprising the following specific steps: (1) Pre-treatment: the grain samples are ground and passed through a 60-150 mesh sieve; (2) Determining the moisture content of the sample, taking 14% as the moisture basis, weighing 3-5g of the dry sample; (3) Set up the enzyme activity determination program and the optimum temperature determination program (4) Weigh 25g of deionized water and mix with the sample evenly, as a control sample. At the same time, weigh 25g of the enzyme inhibitor solution and mix it with the powder as a test sample. The sample is placed in a fast viscosity analyzer to measure the viscosity characteristic spectrum; (5) According to the RVA characteristic spectrum, the peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, pasting temperature and area can be obtained; among them, the enzyme can be calculated by the peak viscosity increase value of the test sample relative to the control sample Vitality, i.e. (P _test - P _control ) × 100/P _control , where P represents peak viscosity; (6) Calculate the optimum temperature of the enzyme according to the area in the RVA characteristic spectrum, that is, the temperature corresponding to the maximum growth of the area formed by the characteristic curve is the optimum temperature. 4. The method according to claim 3, characterized in that, the enzyme activity assay program is as follows: Program 1: Keep warm at 60°C for 1 minute, increase the speed at 12°C/min to 95°C and keep it for 2.5 minutes, then decrease the same rate to 50°C and hold it for 1.4 minutes, and keep the stirring speed at 160r/min; Program 2: Keep warm at 60°C for 5 minutes, increase the speed at 12°C/min to 95°C and hold for 5 minutes, then decrease the same speed to 50°C and hold for 3 minutes, and keep the stirring speed at 160r/min. 5. The method according to claim 3, characterized in that the optimum temperature determination procedure is: 30-90°C for 5 minutes, then rise to 95°C at a rate of 12°C/min and keep for 2.5 minutes, then drop to 95°C at the same rate. Keep warm at 50°C for 1.4min, and keep the stirring speed at 160r/min. 6. The method according to any one of claims 1-3, wherein the grains include: rice, wheat, corn, buckwheat, oats or quinoa, or rice, wheat, corn, buckwheat, oats or quinoa of starch extract. 7. The method according to claim 3, characterized in that, the sample weight of the starch extract in the step (2) is 3-4g, the whole grain powder sample weight is 4-5g, and the same batch of contrast samples needs to be weighed Samples of the same mass or dry basis were used as controls. 8. The method according to claim 3, characterized in that, the enzyme inhibitors described in step (4) include: compounds containing carbon-phosphorus bonds, phosphoric acid derivatives containing organic groups, silver nitrate, copper sulfate, One or more mixtures of copper chloride and iron sulfate. 9. The method according to claim 3 or 8, characterized in that the concentration of the enzyme inhibitor is 0.01-2 mM. 10. The application of the method according to any one of claims 1 to 9 in grain quality control, storage, breeding, fermentation, and determination of variety and origin.