WO2019165758A1

WO2019165758A1 - Method for rapidly determining activities of enzymes in cereals and predicting optimum temperature of enzymes

Info

Publication number: WO2019165758A1
Application number: PCT/CN2018/099414
Authority: WO
Inventors: 张晖; 朱玲; 齐希光; 吴港城; 王立; 钱海峰; 李言
Original assignee: 江南大学
Priority date: 2018-02-27
Filing date: 2018-08-08
Publication date: 2019-09-06
Also published as: CN108398356B; CN108398356A

Abstract

Disclosed is a method for rapidly determining the activities of enzymes in cereals and predicting the optimum temperature of the enzymes, which relates to the technical field of cereal quality evaluation and determination methods. The method mainly comprises the following steps: (1) performing pre-treatment; (2) determining the moisture content and weighing a sample; (3) setting a temperature program of a rapid viscosity analyser; (4) determining the intrinsic viscosity of a blank sample and a sample with added enzyme inhibitor; (5) determining the enzyme activity according to an obtained comparison feature spectrum; and (6) determining, according to the difference between the enzyme activities at different temperatures, the optimum temperature of enzymes in cereals. Provided is a novel method for determining the activities and optimum temperature of enzymes contained in cereals. The method has the characteristics of simple and convenient operation, rapidness and accuracy, good repeatability, low cost and time saving, facilitates improving the efficiency of cereal quality analysis, facilitates improving the operability of quality determination, avoids manpower and material resource waste and provides theoretical guidance for cereal processing, storage and application.

Description

Method for quickly determining enzyme activity in grain and predicting enzyme temperature

Technical field

The invention relates to a method for quickly determining the enzyme activity in a grain and predicting the temperature of the enzyme, and belongs to the technical field of quality evaluation and measurement methods.

Background technique

China has abundant grain resources and various varieties. Especially rice and wheat are the main food crops in China, and it is also the grain with the largest circulation and consumption in China. Its yield 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.

The intrinsic enzyme activity of cereals is an important indicator of its quality control and application. There are differences in the activities of cereal enzymes in different varieties and regions, which in turn leads to the application properties of cereals of different varieties and origins. The determination of the enzyme activity can be used to understand and judge the degree of germination damage of the grain. For example, in wheat, the wheat with high germination damage has poor eating quality, and vice versa. At the same time, the grain must be stored for a certain period of time during transportation. In the process of storage, the slow metabolism is inevitable due to the storage conditions. The biochemical reaction occurs mainly under the action of various enzymes. If the enzyme activity in the grain is weakened or lost, it will age, resulting in deterioration of 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 an important reference indicator when measuring changes in the degree of 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, mainly including the iodine colorimetric method and the falling number method, in which the iodine colorimetric method has a high degree of variation, a large error, and a long time consuming. In contrast, the drop 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 high temperature amylase. The activity, while the enzymes in the grain are a mixture, different enzymes have an effect on the quality of the grain, and few have been determined for a wide range of total enzyme activity of the grain.

Summary of the invention

In view of the above problems in the prior art, the applicant of the present invention provides an easy method for rapidly determining the enzyme activity in the grain and predicting the temperature of the enzyme. The invention adopts Rapid Visco Analyser (RVA) as an analytical means. The instrument is mainly used for determining enzyme activity to determine wheat bud damage, and is widely used in the study of viscosity characteristics of starch and its derivatives in the later stage. It is in good agreement with the test results of the falling numerical instrument, and it has a good correlation with the application of grain in different systems. The instrument is easy to operate, the measurement process is short, the sample dosage is small, and the most important is the multi-stage precise control of temperature, which lays a good foundation for the enzyme activity measurement and enzyme temperature prediction.

The technical solution of the present invention is as follows:

A first object of the present invention is to provide a method for rapidly determining the enzyme activity and the enzyme temperature in a grain, which is characterized by the degree of viscosity drop during heating gelatinization of the grain before and after the enzyme is inactivated, that is, the relative value of the peak viscosity drop. The parameters of the enzyme activity and the enzyme temperature, the larger the parameter, the higher the enzyme activity; the enzyme activity refers to the general name of the enzyme which reduces the viscosity of each component in the grain; the suitable temperature of the enzyme refers to the grain caused The temperature at which the degree of viscosity reduction is greatest; the components include at least one of starch, protein, fat, polysaccharide, especially starch.

In one embodiment of the present invention, the enzyme having a reducing effect on the viscosity of each component includes at least one of an amylase, a pectinase, a cellulase, a protease, and a lipase.

In one embodiment of the present invention, the enzyme activity refers to a general term for the degree of viscosity reduction of the grain during storage; the enzyme suitable temperature refers to a temperature at which the viscosity of the enzyme acts to a maximum extent during storage.

In one embodiment of the present invention, the enzyme activity refers to a general term for an enzyme which causes a decrease in the viscosity of barley when the barley is transformed into a malt state; and the suitable temperature of the enzyme refers to a temperature at which the malt germination of the barley is most pronounced.

In one embodiment of the present invention, the enzyme activity refers to a general term for the enzyme which is immersed in rice to prepare a rice flour state, resulting in a decrease in the viscosity of the rice flour; the suitable temperature of the enzyme means that the enzyme exhibits the greatest degree of degradation during the rice soaking process. temperature.

In one embodiment of the present invention, the enzyme activity refers to a general term for an enzyme that causes a decrease in the viscosity of wheat flour during processing of wheat to wheat products; and the suitable temperature of the enzyme refers to the maximum degree of viscosity of the wheat flour in the wheat flour. temperature.

In one embodiment of the present invention, the method comprises the steps of: (1) determining the moisture content of the grain sample, converting the dry basis of the sample; (2) determining the inactivation of the enzyme based on the moisture content obtained in the step (1). The viscosity increase of the grain samples before and after 30~100 °C; (3) Calculate the activity of the enzyme by looking for the degree of peak viscosity change during the viscosity change, and (4) Calculate the temperature of the enzyme by the degree of peak viscosity change at different temperature ranges.

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

(1) Pretreatment: the grain sample is subjected to milling treatment and passed through a 100 mesh sieve;

(2) determining the moisture content of the sample, on the basis of 14% moisture, weigh 3-5g dry sample;

(3) Setting the enzyme activity measurement program and the optimal temperature measurement program

(4) Weigh 25 g of deionized water and mix it evenly with the sample as a control. At the same time, 25 g of the enzyme inhibitor solution was weighed and mixed with the powder as a measurement sample. The sample is placed in a rapid viscosity analyzer (RVA) to determine a viscosity profile;

(5) According to the RVA characteristic spectrum, the peak viscosity, the disintegration value, the final viscosity, the retrogradation value, the peak time, the paste temperature and the area are calculated according to the method described in the AACC22-08 standard; wherein, the test sample can be compared with The enzyme viscosity was calculated from the peak viscosity increase of the _control , ie (P _test- P _control ) x 100/P _control , where 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 invention, the procedure for determining enzyme activity is as follows:

Procedure 1: 60 ° C for 1 min, 12 ° C / min speed increased to 95 ° C for 2.5 min, then the same rate dropped to 50 ° C for 1.4 min, the stirring speed was maintained at 160r / min;

Procedure 2: 60 ° C for 5 min, 12 ° C / min speed increased to 95 ° C for 5 min, then the same rate dropped to 50 ° C for 3 min, the stirring speed was maintained at 160r / min;

Among them, Procedure 1 is applicable to the determination of enzyme activity in starch extract; Procedure 2 is applicable to the determination of enzyme activity in whole grain flour.

In one embodiment of the present invention, the procedure for determining the optimum temperature is: setting different holding times separately, holding at 30-90 ° C for 5 min, then increasing to 95 ° C at a rate of 12 ° C / min for 2.5 min, and then The rate was lowered to 50 ° C for 1.4 min and the stirring speed was maintained at 160 r / min.

In an embodiment of the present invention, the grain type in the step (1) comprises: starch extract of rice, wheat, corn, buckwheat, oat, buckwheat and rice, wheat, corn, buckwheat, oat, buckwheat .

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

In one embodiment of the present invention, the enzyme inhibitor in the step (4) includes, but is not limited to, a compound containing a carbon-phosphorus bond, a phosphoric acid derivative containing an organic group, silver nitrate, copper sulfate, and chlorination. A mixed formulation of one or more of copper and iron sulfate.

In one embodiment of the invention, the concentration of the enzyme inhibitor in step (4) is controlled to be between 0.01 and 2 mM.

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

The beneficial technical effect of the invention is that the invention provides a novel method for determining the enzyme activity and the enzyme temperature in the cereal, and the enzyme activity and the enzyme temperature in the grain can be determined while studying the gelatinization property of the grain. Compared with the chemical colorimetric method, it has the characteristics of simple operation, fast and accurate, good repeatability, small error, low cost and time saving. Compared with the falling numerical instrument, the method can determine the enzyme activity of the total enzyme in the grain, and can understand the optimum temperature of the enzyme through comparison of different temperatures, and further understand the quality of the grain, which is beneficial to improve the quality analysis of the grain. Efficiency is conducive to improving the operability of quality measurement, avoiding waste of manpower and material resources, and providing theoretical guidance for grain processing, storage and application.

DRAWINGS

Figure 1 is a measurement of rice amylase activity measured in Example 1;

2 is an optimum enzyme activity temperature of the rice flour measured in Example 2;

3 is a comparison of the optimum enzyme activity temperature measured in Example 2 with other temperatures, and the change in rice hardness at different soaking temperatures;

Figure 4 is a measurement of corn flour enzyme activity determined in Example 3;

Figure 5 is a measurement of wheat flour enzyme activity measured in Example 4;

Fig. 6 is a graph showing the measurement of enzyme activity before and after germination of barley measured in Example 5.

Detailed ways

Example 1

A simple method for rapidly determining enzyme activity in a grain and predicting the temperature of the enzyme, characterized in that the method comprises the following specific steps:

(1) Pre-treatment of rice: extracting rice starch, the starch content is 95%, and passing through a 100 mesh sieve.

(2) The moisture content of the sample was measured, and on the basis of 14% moisture, 3 g of the dry sample was weighed.

(3) The specific procedure is as follows: 1 min at 60 ° C, 2 h at 12 ° C / min for 2.5 min, then the same rate is reduced to 50 ° C for 1.4 min, the stirring speed is maintained at 160 r / min.

(4) Weigh 25 g of deionized water and mix it evenly with the sample as a control. At the same time, 25 g of the enzyme inhibitor solution was weighed and mixed with the powder as a measurement sample. The sample was placed in a Rapid Viscosity Analyzer (RVA) to determine the viscosity profile.

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

The results are shown in Fig. 1. The peak viscosity of rice starch was significantly increased after enzyme inactivation, that is, the high enzyme activity was present in rice starch, and the peak viscosity was increased from 6500 cp to 7579 cp, and the calculated enzyme activity was 16.6.

Example 2

(1) Pretreatment: The rice sample is subjected to milling treatment and passed through a 100 mesh sieve.

(2) The moisture content of the sample was measured, and 4 g of the dry sample was weighed on the basis of 14% moisture.

(3) The specific procedure is as follows: set different holding time, heat at 30-90 °C for 5 min, then increase to 95 °C for 2.5 min at 12 °C/min, then drop to 50 °C for 1.4 min at the same rate, stir. The speed is maintained at 160r/min.

(5) Calculate the optimum temperature of the enzyme according to the degree of viscosity drop in the RVA characteristic spectrum, that is, the temperature at which the viscosity decreasing trend is most obvious (the maximum value of the area where the characteristic curve is formed) is the optimum temperature.

The results are shown in Fig. 2. Under different temperature incubations, the difference in peak viscosity before and after enzyme digestion is different. It can be seen from the figure that the optimum temperature of the enzyme in rice flour is 60-70 °C.

The rice hardness test method (three-point measurement method, TA.XT-Plus, P/36 probe) proposed by Japanese Okabe scholars and widely used in the world is used to determine the hardness of rice as a function of immersion temperature and time. At the optimum temperature for enzyme action, the hardness of rice decreased most significantly (from 30499g to 11210g), while at lower temperatures, such as 30°C, the decrease was not significant (32174g to 25524g). The optimum temperature measuring method proposed by the invention can be applied to guide the processing control of rice.

Example 3

(1) Pretreatment: The corn sample was subjected to milling treatment through a 100 mesh sieve.

(2) The moisture content of the sample was measured, and 5 g of the dry sample was weighed based on 14% of water.

(3) The specific procedures are as follows:

The temperature was kept at 60 ° C for 5 min, and the speed was increased to 95 ° C for 5 min at 12 ° C / min. Then the same rate was lowered to 50 ° C for 3 min, and the stirring speed was maintained at 160 r / min.

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

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

Example 4

(1) Pretreatment: A wheat sample was taken for milling treatment and passed through a 100 mesh sieve.

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

The results are shown in Fig. 5. The peak viscosity of the wheat flour was also significantly increased after the enzyme was inactivated, that is, the higher the enzyme activity in the wheat flour, the viscosity after the enzyme was increased from 2616 cp to 3158 cp, and the calculated enzyme activity was 20.7.

Example 5

(1) Pretreatment: The barley samples before and after germination were subjected to milling treatment and passed through a 100 mesh sieve.

(3) The temperature was kept at 60 ° C for 5 min, the speed of 12 ° C / min was increased to 95 ° C for 5 min, and then the same rate was lowered to 50 ° C for 3 min, and the stirring speed was maintained at 160 r / min.

As a result, as shown in Fig. 6, the viscosity of the barley before and after germination was changed, and the viscosity was lowered from 2140 cp to 1603 cp, and the calculated enzyme activity was 33.4.

Example 6

(1) Pre-treatment: collect 1 kind of wheat flour germinated in the rainy season, 2 kinds of normal mature wheat flour, and wheat flour as high-gluten flour (protein content is above 12%);

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

(6) Measure the drop value of the three types of flour used according to the method described in GB/T 10361-2008, measure the viscosity enzyme activity according to the method of the present invention, and at the same time, apply three kinds of flour to make soft under the same operating environment and operation steps. Bread, the bread volume was determined according to the method described in GB/T20981-2007. The bread formula was as follows: 100% flour, yeast 2%, sugar 10%, salt 1%, oil 6%, water 57%.

The falling value of the wheat flour is detected by a falling numerical instrument, and the detection steps are:

(1) determining the moisture content of the flour, and correcting the sample with a moisture content of 7 g of 15%, and weighing the sample according to the measured moisture content;

(2) Turn on the falling value meter and heat the water bath until it boils;

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

(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 the drop value, viscosity enzyme activity and bread specific volume measured in the examples. The drop value is inversely proportional to the surface or enzyme activity, that is, the higher the enzyme activity, the lower the drop value, and the viscosity enzyme activity and the actual enzyme activity. In direct proportion. It can be seen from the table that the trend of the enzyme activity measured by the present invention is completely consistent with the trend measured by the drop value. It is generally believed that the bread has a specific volume of about 4.8, and the bread quality is better (≤7). The bread is too large and too small to be too small. It can be seen from the table that the enzyme activity is closely related to the specific volume. The specific enzyme activity (falling number 250±9.2, viscosity enzyme activity 20.6±0.2) can be used to predict the specific volume of the bread, and then guide the production and processing of bread flour. It can be seen that the enzyme activity error limit determined by the present invention is reduced by a factor of 10.

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

Example 7

The rice is processed at different temperatures to prepare a wet-milled rice flour product.

The specific process is raw material rice-soaking-wet milling-sieving-pressure filtration-crushing-drying, and the difference is that the soaking temperature is controlled to 65 ° C (or the suitable temperature of the enzyme obtained in the measurement of the enzyme of 60 to 70 ° C) , 50 ° C, 80 ° C.

Comparing the processing efficiencies at different soaking temperatures, it was found that the immersion temperature at 50 °C had a content of 7-10% on the sieve (powder), which was significantly higher than that at 65 °C (1-3%) and 80 °C (4- 5%) of the sieved material, at the same time, the gelatinization properties of the prepared wet-milled rice flour product were measured. The results showed that the gelatinization temperature of the rice flour soaked at 65 ° C was lower than that of the rice flour soaked at 50 ° C and 80 ° C. 1-2 ° C.

Example 8

The enzyme activity/enzymatic temperature of the rice under different storage conditions was measured according to the method of Example 1. The results showed that the enzyme temperature was 60-70 ° C, and the grain was stored at 60 ° C at 30 ° C and 40 ° C ( As a control at the non-enzymatic temperature, the results showed that the enzyme activities under different conditions were 16.6 (storage temperature 30 ° C), 17.8 (storage temperature 40 ° C), and 28.1 (storage temperature 60 ° C). Among them, the rice prepared in the rice at 30-40 ° C is white and bright, and the rice stored at 60 ° C has a high enzyme activity, so that the processed rice is yellowish.

The enzyme activity of corn was determined according to the method of Example 3. The result showed that the enzyme activity was 15.8, the enzyme temperature was 65-75 ° C, the corn flour and water were prepared into 10% corn milk, and the temperature was kept at 70 ° C for 2 h. Thereafter, the boiling water was heated for 20 min, and then gel was formed at 4 ° C to be incubated at 30 ° C, 90 ° C for 2 h (non-enzymatic temperature) as a control, and the results showed that the gel strength was 67.5 g by texture analyzer, respectively. /cm ² (70 ° C), 57.8 g / cm ² (30 ° C), 60.4 g / cm ² (90 ° C), it can be seen that at the enzyme temperature, due to the action of the enzyme, the starch is degraded to a certain extent, Thus, the degree of rejuvenation is high, and thus the gel strength is high, while the lower or higher temperature enzymes are not significantly effective, and thus the gel strength is lower than the optimum temperature. At the same time, the gel strength was slightly higher than 30 ° C at 90 ° C due to the high degree of gelatinization of the starch.

The enzyme activity/enzyme temperature of wheat was measured according to the method of Example 4. The result showed that the enzyme activity was 20.7, the enzyme temperature was 50-60 ° C, and the grain was subjected to dip treatment at 55 ° C, followed by drying. , grinding, at 20 ° C, 80 ° C (non-enzymatic temperature) as a control, the results show: the amount of reducing sugar in wheat whole powder is 0.60%, 0.38%, 0.43%, which shows that at the enzyme temperature Next, more starch and polysaccharides are degraded by enzymes into reducing sugars.

The enzyme activities of different germination degree barley were measured according to the method of Example 5. The results showed that the barley enzyme activity of germination for 3 days and 6 days was 29.6, 37.2, respectively, and the α- in barley was determined by chemical method (reducing sugar method). The enzyme activity of β-amylase increased from 300 U/g to 450 U/g. From this, it is understood that the enzyme activity measured by the present invention is consistent with the tendency of the enzyme activity of barley having different germination degrees as determined by the chemical method.

Although the present invention has been disclosed in the above preferred embodiments, the present invention is not limited thereto, and various modifications and changes can be made thereto without departing from the spirit and scope of the invention. The scope of the invention should be determined by the scope of the claims.

Claims

A method for grain quality control, storage, breeding, fermentation, and determination of variety and origin, characterized in that the method is to control grain quality, storage, breeding, fermentation and variety according to enzyme activity and temperature of the enzyme in the grain, The determination of the origin; the determination of the enzyme activity and the enzyme temperature in the grain are as follows:

The degree of viscosity drop when the cereal is heated and gelatinized before and after the enzyme is destroyed, that is, the relative value of the peak viscosity decrease is used as a parameter for characterizing the enzyme activity and the enzyme temperature. The larger the parameter, the higher the enzyme activity; the enzyme activity refers to the grain A general term for an enzyme having a reduced viscosity of each component; the enzyme suitable temperature refers to a temperature at which the degree of viscosity reduction is maximized in the cereal; and each component includes at least one of starch, protein, fat, and polysaccharide.
The method according to claim 1, wherein the determining, storing, breeding, fermenting, cultivating, and producing the grain according to the enzyme activity and the enzyme temperature in the grain comprises:

The rice is stored at a non-enzymatic temperature to ensure that the rice is white and translucent, and the rice is stored at a suitable temperature to prepare a beige yellow color; or

Choose to use corn flour at the enzyme temperature to prepare the gel to increase the gel strength; or,

The whole wheat flour is prepared by using the grain at the enzyme temperature to increase the amount of reducing sugar in the whole wheat flour.
A method for rapidly determining enzyme activity and temperature of an enzyme in a cereal, characterized in that the method uses a relative degree of viscosity drop during heating gelatinization of the grain before and after the enzyme is inactivated, that is, a relative value of a decrease in peak viscosity as a characteristic enzyme activity and an enzyme The parameter of temperature, the larger the parameter, the higher the enzyme activity; the enzyme activity refers to the general name of the enzyme which has a lowering effect on the viscosity of each component in the grain; the suitable temperature of the enzyme refers to the maximum degree of viscosity reduction in the grain. Temperature; the components include at least one of starch, protein, fat, and polysaccharide.
The method according to claim 1, wherein said enzyme activity is a general term for an enzyme having a reducing effect on the viscosity of starch in the grain.
The method according to claim 1, wherein the enzyme which has a reducing effect on the viscosity of each component comprises at least one of amylase, pectinase, cellulase, protease, and lipase.
The method according to claim 1, wherein said enzyme activity refers to a general term for the degree of viscosity drop of the grain during storage; and said enzyme suitable temperature refers to a temperature at which a function of the viscosity during storage causes the degree of viscosity drop to the greatest extent.
The method according to claim 1, wherein the enzyme activity refers to a general term for an enzyme which causes a decrease in barley viscosity when the barley is transformed into a malt state; and the suitable temperature of the enzyme means that the barley germination into a malt viscosity is most markedly decreased. temperature.
The method according to claim 1, wherein the enzyme activity refers to a general term for the enzyme which is immersed in rice to prepare a rice flour state, resulting in a decrease in the viscosity of the rice flour; and the suitable temperature of the enzyme means that the enzyme degrades during the rice soaking process. The temperature at which the degree of action is greatest.
The method according to claim 1, wherein the enzyme activity refers to a general term for an enzyme which causes a decrease in viscosity of wheat flour during processing of wheat to wheat products; and the suitable temperature of said enzyme refers to a maximum degree of viscosity of wheat flour by enzyme degradation in wheat flour. The required temperature.
The method according to any one of claims 2 to 9, characterized in that the method comprises the steps of: (1) determining the moisture content of the grain sample, converting the dry basis of the sample; (2) determining the grain sample before and after the inactivation of the enzyme at 30 The corresponding viscosity increase value at ~100 °C; (3) calculating the activity of the enzyme by looking for the degree of peak viscosity change during the viscosity change, and (4) calculating the optimum temperature of the enzyme by varying the peak viscosity at different temperature ranges.
The method according to any one of claims 2 to 10, characterized in that it comprises the following specific steps:

(1) Pretreatment: the grain sample is subjected to milling treatment and passed through a 60-150 mesh sieve;

(2) determining the moisture content of the sample, on the basis of 14% moisture, weigh 3-5g dry sample;

(3) setting the enzyme activity measurement procedure and the optimum temperature measurement procedure;

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

(5) According to the RVA characteristic spectrum, peak viscosity, disintegration value, final viscosity, retrogradation value, peak time, paste temperature and area can be obtained; wherein the enzyme can be calculated by the peak viscosity increase value of the test sample relative to the control sample Vigor, ie (P test- P control ) × 100/P control , where 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.
The method of claim 11 wherein the enzyme activity assay procedure is as follows:

Procedure 1: 60 ° C for 1 min, 12 ° C / min speed increased to 95 ° C for 2.5 min, then the same rate dropped to 50 ° C for 1.4 min, the stirring speed was maintained at 160r / min;

Procedure 2: Incubate at 60 ° C for 5 min, increase the speed at 12 ° C / min to 95 ° C for 5 min, then reduce the temperature to 50 ° C for 3 min at the same rate, and maintain the stirring speed at 160 r / min.
The method according to claim 11, wherein the optimum temperature measuring procedure is: holding at 30-90 ° C for 5 min, followed by raising at a rate of 12 ° C / min to 95 ° C for 2.5 min, and then dropping to the same rate at 50 ° C. The temperature was kept for 1.4 min and the stirring speed was maintained at 160 r/min.
The method according to any one of claims 3 to 4, 10 to 11, wherein the cereal comprises: rice, wheat, corn, buckwheat, oat or buckwheat, or rice, wheat, corn, buckwheat, oats or Starch extract of buckwheat.
The method according to claim 11, wherein the sample weight of the starch extract in the step (2) is 3-4 g, and the weight of the whole grain powder sample is 4-5 g, and the same batch of the comparative sample needs to be weighed the same quality. Or a dry sample as a control.
The method according to claim 11, wherein said enzyme inhibitor in step (4) comprises: a compound containing a carbon-phosphorus bond, a phosphoric acid derivative containing an organic group, silver nitrate, copper sulfate, and chlorination. A mixed formulation of one or more of copper and iron sulfate.
The method according to claim 11 or 16, wherein the concentration of the enzyme inhibitor is from 0.01 to 2 mM.
The method according to any one of claims 3 to 17 for use in grain quality control, storage, breeding, fermentation, and determination of variety and origin.