CN115326638A - Detection method and application of amylase in oyster sauce - Google Patents
Detection method and application of amylase in oyster sauce Download PDFInfo
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- CN115326638A CN115326638A CN202210971167.7A CN202210971167A CN115326638A CN 115326638 A CN115326638 A CN 115326638A CN 202210971167 A CN202210971167 A CN 202210971167A CN 115326638 A CN115326638 A CN 115326638A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N11/00—Investigating flow properties of materials, e.g. viscosity, plasticity; Analysing materials by determining flow properties
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N11/00—Investigating flow properties of materials, e.g. viscosity, plasticity; Analysing materials by determining flow properties
- G01N2011/006—Determining flow properties indirectly by measuring other parameters of the system
- G01N2011/0093—Determining flow properties indirectly by measuring other parameters of the system thermal properties
Abstract
The invention discloses a detection method and application of amylase in oyster sauce, belonging to the technical field of food detection; respectively adding an oyster sauce sample to be detected and blank starch paste into a starch base solution to obtain a test sample and a control sample; preserving the temperature of the sample at 50-60 ℃, respectively recording the viscosity values before and after heat preservation, and calculating the viscosity change rate before and after heat preservation; and then comparing the viscosity change rates before and after the temperature preservation, and determining whether the test sample contains the amylase or not according to the comparison result. The method provided by the invention is simple to operate, quick and efficient, low in requirement on equipment and high in accuracy; the method can be widely used for detecting amylase in substances related to the oyster sauce production process chain, so that the body state stability of the oyster sauce is improved, and the storage stability of the oyster sauce is ensured.
Description
Technical Field
The invention belongs to the technical field of food detection, and particularly relates to a detection method and application of amylase in oyster sauce.
Background
The existence of amylase can lead the body state of the finished oyster sauce to be thinned in the process of storage or shelf sale, and the appearance and the sale of the product are seriously influenced. However, the field with more demand for amylase detection is the medical field and the biological field, and the demand for amylase detection in the food field is relatively less. Although the existing amylase detection methods are various in types, most of the detection methods have the characteristics of high detection cost and great operation difficulty.
For example, in the prior art, CN113834757a, "a method for rapidly detecting high temperature resistant amylase trace residue of oyster sauce raw materials", discloses a method for rapidly detecting high temperature resistant amylase trace residue of oyster sauce raw materials, in which a raw material standard sample with gradient enzyme activity is prepared, a destructive test is carried out to find critical enzyme activity, a broomrad viscometer is used to determine viscosity maps of modified starch solutions of a standard sample and a sample to be detected, and a viscosity change curve is compared to judge whether the sample to be detected contains amylase; the method needs to find a standard sample with the highest enzyme activity as a critical enzyme activity comparison, and needs to use a Brabender viscometer for analysis, so the detection method is complex, the equipment is expensive, and the detection cost is high. In addition, the method comprises the steps of preparing buffer solution, culturing a blank control and a test sample in an oven at 37 ℃ for 72 hours, respectively taking 1mL of the blank control and the test sample in a white porcelain dish after the culture is finished, adding iodine solution, shaking uniformly, and judging whether the amylase is contained according to a color development result; because the method needs to prepare buffer solution, uses various chemical reagents and needs 72 hours of detection time, the detection method provided by the method is complex in operation, low in efficiency and low in result accuracy.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a rapid, simple, low-cost and high-accuracy detection method for amylase in oyster sauce and application thereof.
In order to achieve the purpose, the invention adopts the technical scheme that: a detection method of amylase in oyster sauce comprises the following steps:
(1) Preparation of test samples: adding an oyster sauce sample to be tested into the starch base solution, and uniformly mixing to obtain a test sample;
(2) Preparation of control sample: adding the blank starch paste into the starch base solution, and uniformly mixing to obtain a control sample;
(3) Keeping the temperature of the test sample and the control sample at 50-60 ℃, respectively recording the viscosity value at the beginning of heat preservation and the viscosity value at the end of heat preservation, and calculating the viscosity change rate before and after heat preservation;
(4) Comparing the change rates calculated in the step (3), and determining whether the test sample contains amylase or not according to the comparison result.
The detection method of amylase in oyster sauce provided by the invention utilizes the fact that amylase has great influence on the body state of amylase paste, under normal conditions, the body state of the starch paste is stable at 60 ℃, and the starch is slightly gelatinized and thickened in body state but cannot be back-thinned after being placed for a certain time; standing the starch paste at 60 ℃ for a certain time, and judging whether the starch paste contains amylase or not according to the state change condition of the starch paste; the physical state of the starch paste can be reflected by the viscosity index, so that the change of the physical state of the starch paste can be judged according to the change of the viscosity index of the starch paste. On the principle, the method is provided with a corresponding test sample and a corresponding control sample, and the viscosity change rates of the test sample and the control sample are calculated and compared to obtain whether the amylase exists in the oyster sauce; the method provided by the invention is simple to operate, is rapid and efficient, and has low requirements on equipment.
In a preferred embodiment of the detection method of the present invention, in the step (4), the method for determining whether the test sample contains amylase according to the comparison result is: when the viscosity change rate of the test sample is more than or equal to 0, the test sample does not contain amylase; when the viscosity change rate of the control sample is less than or equal to that of the test sample and less than 0, the test sample does not contain amylase; when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase.
In a preferred embodiment of the detection method of the present invention, in the step (3), the incubation time is 15 to 16 hours.
Preferably, in the step (3), the incubation time is 15 hours.
As the content of amylase in the oyster sauce is low and the enzymolysis can be carried out in a long time, the inventor finds out through a large number of experiments that the amylase in the oyster sauce can be guaranteed to carry out enzymolysis when the oyster sauce is kept at the temperature of 50-60 ℃ for 15-16 hours, and the detection efficiency can be considered, so that the preferred time for keeping the temperature is 15 hours. In addition, the temperature of 50-60 ℃ is selected because on one hand, alpha-amylase can still maintain high enough activity at 93-95 ℃ and beta-amylase can be inactivated at more than 70 ℃, so that the temperature of 50-60 ℃ is the optimal action temperature of the amylase; on the other hand, starch continues to gelatinize above 60 ℃, so the final choice is to keep the temperature at 50-60 ℃ for 15 hours.
As a preferred embodiment of the detection method of the present invention, in the step (3), the method for calculating the viscosity change rate before and after the heat preservation includes: the rate of change of viscosity = (viscosity value at the end of incubation-viscosity value at the beginning of incubation)/time of incubation.
For example, when the incubation time is 15 hours, the viscosity change rate = (viscosity value at the end of incubation-viscosity value at the start of incubation)/15.
In a preferred embodiment of the detection method of the present invention, in the step (1), the starch base solution is a blank starch paste, and the mass percentage of the blank starch paste is 5.5-6.5%.
Since the amylase in the oyster sauce is measured, the mass percent of the blank starch paste is set to 5.5-6.5%, and more preferably 6% according to the starch content in the oyster sauce.
As a preferred embodiment of the detection method of the present invention, in the step (1), the preparation method of the blank starch paste comprises the following steps: heating the starch solution to boil, preserving heat and cooling to obtain the empty starch paste.
In a preferred embodiment of the detection method of the present invention, the heat-boiling and heat-maintaining time is 25 to 35 minutes.
In order to reduce detection errors and avoid the influence caused by the change of the starch paste, a blank control is set, the blank starch paste is prepared to be used as a control sample, and the blank starch paste is further used as a starch base solution, so that a control experiment can be well carried out, the detection errors are reduced, and the accuracy is improved.
As a preferred embodiment of the detection method, in the step (1), the mass ratio of the oyster sauce sample to be detected to the starch-based liquid is 1 (8-10).
Preferably, in the step (1), the mass ratio of the oyster sauce sample to be tested to the starch-based liquid is 1:9.
In addition, the invention also provides application of the detection method in detection of amylase in substances related to the oyster sauce production process chain.
As a preferred embodiment of the application of the invention, substances related to the oyster sauce production process chain comprise any one of raw materials for preparing the oyster sauce, semi-finished oyster sauce and finished oyster sauce.
Compared with the prior art, the invention has the beneficial effects that:
according to the method for detecting the amylase in the oyster sauce, the viscosity is used as a detection index to reflect the posture change of the starch paste caused by the amylase, and the change rate of the viscosity value of a test sample is compared with the change rate of the viscosity value of a control sample, so that the information whether the oyster sauce contains the amylase or not is obtained; the method can reduce the interference of the color of the oyster sauce on the detection result and improve the detection accuracy; on one hand, the detection material has low cost, the detection tool is simple, the detection operation difficulty is low, the detection time is short, and the detection purpose can be efficiently, quickly, simply and conveniently achieved; on the other hand, the method provided by the invention has good ductility, and can be suitable for substances involved in an oyster sauce production process chain, so that amylase in the substances involved in the oyster sauce production process chain can be monitored and processed in time, the body state stability of the oyster sauce is improved, and the storage stability of the oyster sauce is ensured.
Detailed Description
To better illustrate the objects, aspects and advantages of the present invention, the present invention will be further described with reference to specific examples.
Example 1
The detection method for detecting the oyster sauce sample 1 to be normally produced and sterilized comprises the following steps:
(1) And (3) detecting the disinfection of tools: sterilizing and inactivating enzymes of the detection tools and instruments in a boiling water bath at 100 ℃ for 30 minutes, and drying for later use;
(2) Preparing blank starch paste: preparing 2000g of starch solution with the mass percent of 6%, heating the starch solution to boiling by using an iron pan and an induction cooker, preserving heat for 30 minutes to obtain blank starch paste, and stirring once every 2 minutes in the heat preservation process; then, 270g of the blank starch paste was filled into a 500mL glass bottle as a starch base solution, and 6 bottles of the starch base solution were prepared in total.
(3) Preparation of test samples: adding 30g of oyster sauce samples to be normally produced and sterilized into the 500mL glass bottle containing the starch base solution in the step (2), uniformly stirring to obtain test samples, and preparing 3 test samples in total;
(4) Preparation of control sample: adding 30g of the blank starch paste obtained in the step (2) into a 500mL glass bottle containing the starch base solution obtained in the step (2), uniformly stirring to obtain a control sample, and preparing 3 control samples in total;
(5) Taking 100g of each sample, detecting and recording the initial viscosity value, then placing the samples in a water bath kettle at 50 ℃ for heat preservation in a water bath for 15 hours, ensuring that the starch paste is completely soaked in the water bath kettle in the water bath process, and detecting and recording the finished viscosity value after the water bath is finished;
(6) Calculation of the rate of change of viscosity: viscosity change rate = (viscosity value at the end of heat preservation-viscosity value at the start of heat preservation)/15;
(7) Comparing the viscosity change rates calculated in the step (6), and when the viscosity change rate of the test sample is more than or equal to 0, determining that the test sample does not contain amylase; when the viscosity change rate of the control sample is less than or equal to that of the test sample and less than 0, the test sample does not contain amylase; when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase;
the results of the tests are shown in table 1;
TABLE 1
As can be seen from Table 1, the viscosity change rate of the test sample is not less than 0, which indicates that the oyster sauce sample 1 to be tested does not contain amylase.
Example 2
The detection method for detecting the oyster sauce sample 2 to be detected comprises the following steps:
(1) And (3) detecting the disinfection of tools: sterilizing and inactivating enzymes of the detection tools and instruments in a boiling water bath at 100 ℃ for 30 minutes, and drying for later use;
(2) Preparing blank starch paste: preparing 2000g of starch solution with the mass percent of 6%, heating the starch solution to boiling by using an iron pan and an induction cooker, keeping the temperature for 35 minutes to obtain blank starch paste, and stirring once every 2 minutes in the heat preservation process; then, 270g of the blank starch paste was filled into a 500mL glass bottle as a starch base solution, and 6 bottles of the starch base solution were prepared in total.
(3) Preparation of test samples: adding 30g of oyster sauce samples to be normally produced and sterilized into the 500mL glass bottle containing the starch base solution in the step (2), uniformly stirring to obtain test samples, and preparing 3 test samples in total;
(4) Preparation of control sample: adding 30g of the blank starch paste obtained in the step (2) into a 500mL glass bottle containing the starch base solution obtained in the step (2), uniformly stirring to obtain a control sample, and preparing 3 control samples in total;
(5) Taking 100g of each sample, detecting and recording the initial viscosity value, then placing the samples in a water bath kettle at 60 ℃ for heat preservation in a water bath for 16 hours, ensuring that the starch paste is completely soaked in the water bath kettle in the water bath process, and detecting and recording the finished viscosity value after the water bath is finished;
(6) Calculation of the rate of change of viscosity: viscosity change rate = (viscosity value at the end of heat preservation-viscosity value at the beginning of heat preservation)/16;
(7) Comparing the viscosity change rates calculated in the step (6), and when the viscosity change rate of the test sample is more than or equal to 0, determining that the test sample does not contain amylase; when the viscosity change rate of the control sample is less than or equal to that of the test sample and less than 0, the test sample does not contain amylase; when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase;
the results of the tests are shown in table 2;
TABLE 2
As can be seen from Table 2, the viscosity change rate of the control sample is not greater than the viscosity change rate of the test sample and is less than 0, which indicates that the oyster sauce sample 2 to be tested does not contain amylase.
Example 3
The detection method for detecting the oyster sauce sample 3 to be detected comprises the following steps:
(1) And (3) detecting the disinfection of tools: sterilizing and inactivating enzymes of the detection tools and instruments in a boiling water bath at 100 ℃ for 30 minutes, and drying for later use;
(2) Preparing blank starch paste: preparing 2000g of starch solution with the mass percent of 5.5%, heating the starch solution to boiling by using an iron pan and an induction cooker, preserving heat for 30 minutes to obtain empty starch paste, and stirring once every 2 minutes in the heat preservation process; then, 300g of the blank starch paste was filled into a 500mL glass bottle as a starch base solution, and 6 bottles of the starch base solution were prepared in total.
(3) Preparation of test samples: adding 30g of oyster sauce samples to be normally produced and sterilized into the 500mL glass bottle containing the starch base solution in the step (2), uniformly stirring to obtain test samples, and preparing 3 test samples in total;
(4) Preparation of control sample: adding 30g of the blank starch paste obtained in the step (2) into a 500mL glass bottle containing the starch base solution obtained in the step (2), uniformly stirring to obtain a control sample, and preparing 3 control samples in total;
(5) Taking 100g of each sample, detecting and recording the initial viscosity value, then placing the samples in a water bath kettle at 55 ℃ for heat preservation in a water bath manner for 15.5 hours, ensuring that the starch paste is completely soaked in the water bath kettle in the water bath process, and detecting and recording the finished viscosity value after the water bath is finished;
(6) Calculation of the rate of change of viscosity: viscosity change rate = (viscosity value at the end of heat preservation-viscosity value at the start of heat preservation)/15.5;
(7) Comparing the viscosity change rates calculated in the step (6), and when the viscosity change rate of the test sample is more than or equal to 0, determining that the test sample does not contain amylase; when the viscosity change rate of the comparison sample is not more than that of the test sample and is less than 0, the test sample does not contain amylase; when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase;
the results of the tests are shown in table 3;
TABLE 3
As can be seen from Table 3, the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, which indicates that the oyster sauce sample 3 to be tested contains amylase.
Comparative example 1
The oyster sauce sample 3 to be detected is detected in the comparative example, and the only difference from the example 3 is that in the step (5), the sample is placed in a water bath kettle at 80 ℃ for heat preservation in a water bath for 15.5 hours;
the results of the tests are shown in table 4;
TABLE 4
As can be seen from Table 4, the viscosity change rate of the test sample is greater than or equal to 0, which indicates that the oyster sauce sample 3 to be tested does not contain amylase.
Comparative example 2
The oyster sauce sample 3 to be detected is detected in the comparative example of the invention, and the only difference of the other examples 3 is that in the step (5), the sample is placed in a water bath kettle at 55 ℃ for 10 hours of water bath heat preservation.
The results of the test are shown in table 5;
TABLE 5
As can be seen from Table 5, the viscosity change rate of the test sample is greater than or equal to 0, which indicates that the oyster sauce sample 3 to be tested does not contain amylase.
Test example 1
The test example of the invention detects the sample containing amylase, and the specific detection method comprises the following steps:
(1) And (3) detecting the disinfection of tools: sterilizing and inactivating enzymes of the detection tools and instruments in a boiling water bath at 100 ℃ for 30 minutes, and drying for later use;
(2) Preparing blank starch paste: preparing 4000g of starch solution with the mass percent of 6%, heating the starch solution to boiling by using an iron pan and an induction cooker, preserving heat for 30 minutes to obtain blank starch paste, and stirring once every 2 minutes in the heat preservation process; then, 270g of the blank starch paste was filled into a 500mL glass bottle as a starch base solution, and 12 bottles of the starch base solution were prepared in total.
(3) Preparation of test samples: respectively taking 1mL, 3mL and 5mL of 100g/L alpha-amylase solution, respectively fixing the volume to 100mL, then respectively taking 1mL of the diluted alpha-amylase solution, respectively fixing the volume to 100mL; finally, 30mL of each of the two diluted alpha-amylase solutions (respectively, test samples 1,2 and 3) is added into the 500mL glass bottle containing the starch base solution in the step (2), the mixture is uniformly stirred to obtain test samples, and 3 test samples are prepared at each concentration;
(4) Preparation of a blank sample: adding 30g of tap water which is boiled for 30 minutes into the 500mL glass bottle containing the starch base solution in the step (2), uniformly stirring to obtain a control sample, and preparing 3 blank samples in total;
(5) Taking 100g of each sample, detecting and recording the initial viscosity value, then placing the samples in a water bath kettle at 50 ℃ for heat preservation in a water bath for 15 hours, ensuring that the starch paste is completely soaked in the water bath kettle in the water bath process, and detecting and recording the finished viscosity value after the water bath is finished;
(6) Calculation of the rate of change of viscosity: viscosity change rate = (viscosity value at the end of heat preservation-viscosity value at the start of heat preservation)/15;
(7) Comparing the viscosity change rates calculated in the step (6), and when the viscosity change rate of the test sample is more than or equal to 0, determining that the test sample does not contain amylase; when the viscosity change rate of the control sample is less than or equal to that of the test sample and less than 0, the test sample does not contain amylase; when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase;
the results of the tests are shown in table 6;
TABLE 6
As can be seen from Table 6, the rate of change of viscosity for the test samples < 0. Ltoreq. That for the control samples, indicating that the samples contain amylase, and vice versa, indicating that the method of the invention is feasible and allows the determination of minute amounts of amylase in the samples.
Test example 2
The test example of the invention verifies whether the oyster sauce sample to be tested in examples 1-3 and comparative examples 1-2 contains amylase, the oyster sauce sample 1-3 to be tested is placed at 25-30 ℃ for 30 days, the posture of the oyster sauce is observed, the oyster sauce sample 3 is obviously thinned, the oyster sauce sample 1 and the oyster sauce sample 2 are free from abnormality, the oyster sauce sample 1 and the oyster sauce sample 2 do not contain amylase, the oyster sauce sample 3 contains amylase, and the conclusion obtained by the test example 1-3 is consistent, and the test example 2 and the test example 1 jointly show that the method provided by the invention can effectively detect whether the oyster sauce contains amylase; the results obtained in comparative examples 1-2 are different from those obtained in example 3, which indicates that the incubation temperature and incubation time in the detection method affect the detection accuracy.
Finally, it should be noted that the above embodiments are intended to illustrate the technical solutions of the present invention and not to limit the scope of the present invention, and although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention.
Claims (10)
1. A detection method of amylase in oyster sauce is characterized by comprising the following steps:
(1) Preparation of test samples: adding an oyster sauce sample to be tested into the starch base solution, and uniformly mixing to obtain a test sample;
(2) Preparation of control sample: adding the blank starch paste into the starch base solution, and uniformly mixing to obtain a control sample;
(3) Keeping the temperature of the test sample and the control sample at 50-60 ℃, respectively recording the viscosity value at the beginning of heat preservation and the viscosity value at the end of heat preservation, and calculating the viscosity change rate before and after heat preservation;
(4) Comparing the change rates calculated in the step (3), and determining whether the test sample contains amylase or not according to the comparison result.
2. The method for detecting according to claim 1, wherein in the step (4), the method for determining whether the test sample contains amylase or not based on the comparison result is:
when the viscosity change rate of the test sample is more than or equal to 0, the test sample does not contain amylase;
when the viscosity change rate of the comparison sample is not more than that of the test sample and is less than 0, the test sample does not contain amylase;
when the viscosity change rate of the test sample is less than 0 and less than or equal to that of the control sample, the test sample contains amylase.
3. The detection method according to claim 1, wherein in the step (3), the incubation time is 15 to 16 hours.
4. The detection method according to claim 3, wherein in the step (3), the viscosity change rate before and after the heat preservation is calculated by: the rate of change of viscosity = (viscosity value at the end of incubation-viscosity value at the beginning of incubation)/time of incubation.
5. The detection method according to claim 1, wherein in the step (1), the starch-based liquid is blank starch paste, and the mass percentage of the blank starch paste is 5.5-6.5%.
6. The detection method according to claim 5, wherein in the step (1), the preparation method of the blank starch paste comprises the following steps: heating the starch solution to boil, preserving heat and cooling to obtain the empty starch paste.
7. The detection method according to claim 6, wherein the incubation time for boiling and holding is 25 to 35 minutes.
8. The detection method according to claim 1, wherein in the step (1), the mass ratio of the oyster sauce sample to be detected to the starch-based liquid is 1 (8-10).
9. The use of the detection method according to any one of claims 1 to 8 for detecting amylase in substances involved in oyster production process chain.
10. The use according to claim 9, wherein the substances involved in the oyster cocktail production process chain include any one of raw materials for preparing oyster cocktail, semi-finished oyster cocktail, and finished oyster cocktail.
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