Method for predicting activity of amylase in inactivation process or after inactivation

Abstract

The invention relates to the technical field of food biology, in particular to a method for predicting the activity of amylase subjected to an inactivation process or inactivated amylase. The invention provides a method for predicting an inactivation process based on required amylase activity, which comprises the following steps: providing the required amylase activity, presetting an inactivation temperature, and calculating the inactivation time by adopting an amylase activity-inactivation time prediction model according to the required amylase activity. The invention also provides a method for predicting the activity of the inactivated amylase based on the inactivation process, which comprises the following steps: providing the inactivation temperature and the inactivation time, and calculating the activity of the inactivated amylase by adopting the amylase activity-inactivation time prediction model according to the inactivation temperature and the inactivation time. The method disclosed by the invention has relatively good predictive capacity, and is beneficial to accurately controlling the inactivation process conditions; by optimizing the inactivation process, the production time and cost can be reduced, and the production efficiency is improved; the enzyme activity is strictly controlled or the enzyme is inactivated, so that the food quality safety is ensured.

Description

Technical Field

[0001] The present application relates to the field of food biotechnology, and in particular to a method for predicting an inactivation process or the activity of amylase after inactivation. Background Art

[0002] Amylases are a class of enzymes that catalyze the decomposition of starch and other carbohydrates into simpler sugar molecules. They are widely found in nature, including plants, animals and microorganisms. Amylase activity refers to the ability of these enzymes to catalyze the decomposition of starch, which can be measured in a variety of ways, such as the speed of the enzymatic reaction, the stability of the enzyme, and the affinity of the enzyme for the substrate. Amylases decompose starch molecules by hydrolyzing α-1,4- and α-1,6-glycosidic bonds to produce smaller sugar molecules such as maltose and glucose. Amylase ASa is derived from Aspergillus oryzae and is the main production strain for brewing soy sauce and beer. It can be mass-produced for the production of sugar and baked goods, so it often appears in the processing and production of various starchy foods. Amylase HEa is derived from Bacillus licheniformis, which is widely found in soil and water and is very easy to exist in food processing pipelines. The amylase produced will hydrolyze starch in food, causing product quality deterioration. Amylase is very important in the food industry, textile industry, and papermaking industry. It is used in bread making, beer brewing, starch syrup production, removing starch slurry from fabrics, improving the quality and strength of paper, etc.

[0003] The inactivation process of amylase activity is of great significance to the food processing, biotechnology and pharmaceutical industries. By precisely controlling the inactivation process conditions, it is possible to ensure that the activity of amylase reaches the expected level, thereby ensuring the quality consistency of the product; optimizing the inactivation process can reduce production time and cost and improve production efficiency; in food processing, inactivating amylase can prevent excessive decomposition of starch, avoid excessive sweetness or texture changes in the product, and ensure food safety; by predicting the inactivation process conditions, over-inactivation can be avoided and the biological activity of the enzyme can be maintained. However, methods for accurately predicting the inactivation process are still being identified.

[0004] Whether it is amylase used in production or amylase produced after bacterial contamination, the enzyme activity needs to be strictly controlled or inactivated to ensure food quality and safety. The activity of amylase is controlled by heating temperature and time, thereby controlling product quality. However, there is currently a lack of methods to predict amylase activity. Summary of the invention

[0005] Based on the above-mentioned defects of the prior art, the present application obtains a prediction model and prediction method that can predict the inactivation process or the activity of amylase after inactivation by studying the relationship between a series of inactivation temperature gradients and time-temperature gradients and the activity of amylase after inactivation. It can accurately predict the required inactivation time and inactivation temperature based on the required amylase activity, and can also predict the activity of amylase after inactivation based on the inactivation time and inactivation temperature, thereby ensuring that the activity of amylase reaches the expected level, thereby ensuring the quality consistency of the product, reducing production time and cost, and improving production efficiency.

[0006] The first aspect of the present application provides a method for predicting an inactivation process based on the required amylase activity, comprising:

[0007] 1) Provide the required amylase activity;

[0008] 2) Preset the inactivation temperature and calculate the inactivation time according to the required amylase activity using an amylase activity-inactivation time prediction model.

[0009] The second aspect of the present application provides a device for predicting an inactivation process based on the required amylase activity, comprising:

[0010] 1) A data input module, used to input the required amylase activity;

[0011] 2) A calculation module is used for the following calculations: a preset inactivation temperature, and according to the required amylase activity, an amylase activity-inactivation time prediction model is used to calculate the inactivation time.

[0012] The third aspect of the present application provides a method for predicting the activity of inactivated amylase based on the inactivation process, comprising:

[0013] 1) Provide inactivation temperature and inactivation time;

[0014] 2) According to the inactivation temperature and inactivation time, the amylase activity after inactivation was calculated using the amylase activity-inactivation time prediction model.

[0015] The fourth aspect of the present application provides a device for predicting the activity of inactivated amylase based on an inactivation process, comprising:

[0016] 1) Data input module, used to input inactivation time and inactivation temperature;

[0017] 2) A calculation module, which is used for the following calculations: according to the inactivation time and the inactivation temperature, the inactivated amylase activity is calculated by using an amylase activity-inactivation time prediction model.

[0018] The fifth aspect of the present application provides a computer-readable storage medium having a computer program stored thereon, which, when executed by a processor, implements the method for predicting the inactivation process based on the required amylase activity as in the first aspect above, or the method for predicting the activity of the amylase after inactivation based on the inactivation process as in the third aspect above.

[0019] In a sixth aspect, the present application provides an electronic terminal comprising a memory and a processor, wherein the memory comprises a computer program, and when the computer program is executed by the processor, the method for predicting an inactivation process based on the required amylase activity as in the first aspect above, or the method for predicting the activity of amylase after inactivation based on the inactivation process as in the third aspect above, is implemented.

[0020] The seventh aspect of the present application provides a computer program product, including a computer program, which, when executed by a processor, implements the method for predicting the inactivation process based on the required amylase activity as in the first aspect above, or the method for predicting the activity of amylase after inactivation based on the inactivation process as in the third aspect above.

[0021] In an eighth aspect, the present application provides a method for inactivating amylase, comprising placing the amylase at 55-65°C for 5-30 minutes, or placing the amylase at above 85°C for more than 10 minutes, wherein the amylase is selected from ASa or HEa.

[0022] The beneficial effects of this application are:

[0023] The method provided in the present application can accurately predict the inactivation process conditions required to obtain the desired amylase activity, and can also accurately predict the amylase activity after inactivation under specific inactivation process conditions. The ASa fitting equation provided in the present application has a goodness of fit R-square value >0.97, and the HEa fitting equation has a goodness of fit R-square value >0.98, which has good prediction ability and good prediction accuracy; the method of the present application is conducive to accurately controlling the inactivation process conditions, which can ensure that the activity of the amylase reaches the expected level, thereby ensuring the quality consistency of the product; optimizing the inactivation process can reduce production time and cost, and improve production efficiency; it is also conducive to strictly controlling enzyme activity or inactivating enzymes to ensure food quality and safety. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The standard curve of reducing sugar.

[0025] Figure 2 The effect of temperature on amylase ASa at 70-100℃.

[0026] Figure 3 The effect of temperature on amylase ASa at 55-65℃.

[0027] Figure 4The effect of temperature on amylase HEa at 85-100℃.

[0028] Figure 5 Schematic diagram of the method for predicting the inactivation process based on the required amylase activity of the present application.

[0029] Figure 6 This is a schematic diagram of the method for predicting the activity of inactivated amylase based on the inactivation process of the present application.

[0030] Figure 7 This is a schematic diagram of a device for predicting a deactivation process based on required amylase activity according to the present application.

[0031] Figure 8 This is a schematic diagram of a device for predicting the activity of inactivated amylase based on the inactivation process of the present application.

[0032] Fig. 9 A schematic diagram of an electronic terminal of the present application. DETAILED DESCRIPTION

[0033] The following describes the implementation of the present application through specific examples, and those skilled in the art can easily understand other advantages and effects of the present application from the contents disclosed in this specification. The present application can also be implemented or applied through other different specific implementations, and the details in this specification can also be modified or changed in various ways based on different viewpoints and applications without departing from the spirit of the present application.

[0034] Before further describing the specific implementation methods of the present application, it should be understood that the scope of protection of the present application is not limited to the specific specific implementation methods described below; it should also be understood that the terms used in the examples of the present application are for describing specific specific implementation methods rather than for limiting the scope of protection of the present application; in the specification and claims of the present application, unless otherwise expressly stated herein, the singular forms "a", "an" and "the" include plural forms.

[0035] When the embodiments give numerical ranges, it should be understood that, unless otherwise specified in the present application, the two endpoints of each numerical range and any numerical value between the two endpoints can be selected. Unless otherwise defined, all technical and scientific terms used in the present application have the same meaning as those generally understood by those skilled in the art. In addition to the specific methods, equipment, and materials used in the embodiments, according to the grasp of the prior art by those skilled in the art and the record of the present application, any method, equipment, and material of the prior art similar or equivalent to the methods, equipment, and materials described in the embodiments of the present application can also be used to realize the present application.

[0036] By studying the relationship between a series of inactivation temperature gradients and time-temperature gradients and the activity of amylase after inactivation, this application obtains a prediction model and prediction method that can predict the inactivation process or the activity of amylase after inactivation. It can accurately predict the required inactivation time and inactivation temperature based on the required amylase activity, and can also predict the amylase activity after inactivation based on the inactivation time and inactivation temperature, thereby ensuring that the activity of amylase reaches the expected level, thereby ensuring the quality consistency of the product, reducing production time and cost, and improving production efficiency.

[0037] The present application first provides a method for predicting an inactivation process based on the required amylase activity, comprising:

[0038] 1) Provide the required amylase activity;

[0039] 2) Preset the inactivation temperature and calculate the inactivation time according to the required amylase activity using an amylase activity-inactivation time prediction model.

[0040] In a specific embodiment of the present application, the inactivation process includes inactivation time and inactivation temperature.

[0041] In a specific embodiment of the present application, the inactivation may be a partial loss of activity or a complete loss of activity.

[0042] Amylase activity refers to the ability of amylase to decompose starch and other carbohydrates under specific conditions. Amylase activity can be expressed as unit activity, specific activity, and relative activity. In a specific embodiment of the present application, the amylase activity is expressed as relative activity, and relative activity is a percentage comparing the enzyme activity under experimental conditions with the maximum activity or standard activity. The maximum activity refers to the highest catalytic activity that an enzyme can achieve under specific conditions. This state means that the catalytic efficiency of the enzyme is the highest and that the substrate can be converted into a product most effectively. The maximum activity or standard activity is usually expressed as unit activity.

[0043] In a specific embodiment of the present application, the activity of amylase can be measured by the following method: prepare a series of reducing sugar standard solutions (such as glucose) of known concentrations. Use a spectrophotometer to measure the absorbance of these standard solutions and draw a standard curve (absorbance vs. reducing sugar concentration). In the amylase reaction system, the amylase is mixed with the starch substrate and reacted for a certain time under suitable temperature and pH conditions. The reaction is terminated, usually by heating to inactivate the enzyme. The reaction sample is treated with an appropriate colorimetric reagent (such as DNS reagent, phenol-sulfuric acid reagent, etc.) to produce a color reaction. The absorbance of the sample is measured, and the reducing sugar concentration in the sample is calculated according to the standard curve. By definition, 1IU of enzyme activity refers to the amount of enzyme that catalyzes 1 micromole (μmol) of substrate to product per minute under standard conditions (usually 25°C, pH 6.0). The calculation formula is:

[0044]

[0045] In the specific implementation of this application,

[0046] In a specific embodiment of the present application, the amylase includes but is not limited to HEa and ASa.

[0047] The prediction model refers to a mathematical model that can predict output variables (targets) based on input variables (features). In a specific embodiment of the present application, the prediction model is constructed by fitting equations.

[0048] In a specific embodiment of the present application, the method of step 2) is: based on the preset inactivation temperature, using the corresponding amylase activity-inactivation time prediction model, substituting the required amylase activity into the prediction model, and calculating the inactivation time.

[0049] The required amylase activity is shown as relative enzyme activity.

[0050] In a specific embodiment of the present application, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, measuring the amylase activity after inactivation, making a fitting curve through OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature.

[0051] In a specific embodiment of the present application, the inactivation temperature can be 55-100°C, for example, it can be 55°C, 60°C, 65°C, 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, or 100°C.

[0052] In a specific embodiment of the present application, the inactivation time is 0 to 30 minutes, for example, it can be 0 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes.

[0053] In certain embodiments of the present application, when the amylase is ASa, the inactivation temperature can be 55-100°C, for example, 55°C, 60°C, 65°C, 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, 100°C; the inactivation time can be 0-30 minutes, for example, 0 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes.

[0054] In other embodiments of the present application, when the amylase is HEa, the inactivation temperature can be 80-100°C, for example, 80°C, 85°C, 90°C, 95°C, 100°C; the inactivation time can be 0-15 minutes, for example, 0 minutes, 5 minutes, 10 minutes, 15 minutes.

[0055] In a specific embodiment of the present application, when the amylase is ASa, the amylase activity-inactivation time prediction model is:

[0056] 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572));

[0057] 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619));

[0058] 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668));

[0059] 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695));

[0060] 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334));

[0061] 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634));

[0062] 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153));

[0063] 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955));

[0064] 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594));

[0065] 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364));

[0066] When the amylase is HEa, the amylase activity-inactivation time prediction model is:

[0067] 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758));

[0068] 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318));

[0069] 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177));

[0070] 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344));

[0071] 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567));

[0072] Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

[0073] Where exp is the natural exponential function, which is defined as e raised to the power of x.

[0074] The present application also provides a device for predicting the inactivation process based on the required amylase activity, such as Figure 7 As shown, including:

[0075] 1) A data input module 11, used to input the required amylase activity;

[0076] 2) A calculation module 12 is used for the following calculations: a preset inactivation temperature, and according to the required amylase activity, an amylase activity-inactivation time prediction model is used to calculate the inactivation time.

[0077] In the specific implementation of the present application, the prediction model, construction method, inactivation time, inactivation temperature, etc. in the device are consistent with the relevant description of the above-mentioned method for predicting the inactivation process based on the required amylase activity. The operating principle of each module can refer to the above-mentioned method for predicting the inactivation process based on the required amylase activity, and will not be repeated here.

[0078] Based on the relationship between amylase activity and inactivation temperature and inactivation time provided in the present application, the inactivation process can be predicted by the amylase activity, and the amylase activity can also be predicted by the inactivation process.

[0079] Therefore, the present application also provides a method for predicting the activity of inactivated amylase based on the inactivation process, comprising:

[0080] 1) Provide inactivation temperature and inactivation time;

[0081] 2) According to the inactivation temperature and inactivation time, the amylase activity after inactivation was calculated using the amylase activity-inactivation time prediction model.

[0082] In a specific embodiment of the present application, the method of step 2) is: based on the inactivation temperature, using the corresponding amylase activity-inactivation time prediction model, substituting the inactivation time into the prediction model, and calculating the amylase activity after inactivation.

[0083] In a specific embodiment of the present application, the calculated inactivated amylase activity is displayed as a relative enzyme activity, which can be converted into a unit enzyme activity by multiplying the maximum enzyme activity of the amylase.

[0084] In a specific embodiment of the present application, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, determining the enzyme activity after inactivation, making a fitting curve through OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature.

[0085] In a specific embodiment of the present application, the inactivation temperature can be 55-100°C, for example, it can be 55°C, 60°C, 65°C, 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, or 100°C.

[0086] In a specific embodiment of the present application, the inactivation time is 0 to 30 minutes, for example, it can be 0 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes.

[0087] In certain embodiments of the present application, when the amylase is ASa, the inactivation temperature can be 55-100°C, for example, 55°C, 60°C, 65°C, 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, 100°C; the inactivation time can be 0-30 minutes, for example, 0 minutes, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes.

[0088] In other embodiments of the present application, when the amylase is HEa, the inactivation temperature can be 80-100°C, for example, 80°C, 85°C, 90°C, 95°C, 100°C; the inactivation time can be 0-15 minutes, for example, 0 minutes, 5 minutes, 10 minutes, 15 minutes.

[0089] In a specific embodiment of the present application, when the amylase is ASa, the amylase activity-inactivation time prediction model is:

[0090] 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572));

[0091] 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619));

[0092] 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668));

[0093] 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695));

[0094] 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334));

[0095] 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634));

[0096] 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153));

[0097] 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955));

[0098] 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594));

[0099] 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364));

[0100] When the amylase is HEa, the amylase activity-inactivation time prediction model is:

[0101] 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758));

[0102] 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318));

[0103] 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177));

[0104] 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344));

[0105] 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567));

[0106] Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

[0107] Where exp is the natural exponential function, which is defined as e raised to the power of x.

[0108] The present application also provides a device for predicting the activity of inactivated amylase based on the inactivation process, such as Figure 8 As shown, including:

[0109] 1) Data input module 21, used to input inactivation time and inactivation temperature;

[0110] 2) A calculation module 22 is used for the following calculation: according to the inactivation time and the inactivation temperature, the inactivated amylase activity is calculated using an amylase activity-inactivation time prediction model.

[0111] In the specific embodiments of the present application, the prediction model, construction method, inactivation time, inactivation temperature, etc. in the device are consistent with the relevant description of the method for predicting the activity of amylase after inactivation based on the above-mentioned inactivation process. The operating principle of each module can refer to the above-mentioned method for predicting the activity of amylase after inactivation based on the inactivation process, and will not be repeated here.

[0112] It should be noted that in the specific embodiments of the present application, the division of the various modules of the device is only a division of logical functions. In actual implementation, they can be fully or partially integrated into one physical entity, or they can be physically separated. These modules can all be implemented in the form of software called by a processing element; they can also be all implemented in the form of hardware; some modules can also be implemented in the form of software called by a processing element, and some modules can be implemented in the form of hardware. For example, the detection module can be a separately established processing element, or it can be integrated in a certain chip for implementation. In addition, it can also be stored in a memory in the form of program code, and a certain processing element can call and execute the functions of the above database acquisition module. The implementation of other modules is similar. In addition, these modules can be fully or partially integrated together, or they can be implemented independently. The processing element described here can be an integrated circuit with signal processing capabilities. In the implementation process, each step of the above method or each module above can be completed by an integrated logic circuit of hardware in a processor element or instructions in the form of software.

[0113] For example, these modules in the embodiments of the present application may be one or more integrated circuits configured to implement the above methods, such as: one or more specific integrated circuits (Application Specific Integrated Circuit, referred to as ASIC), or, one or more microprocessors (digital singnal processor, referred to as DSP), or, one or more field programmable gate arrays (Field Programmable Gate Array, referred to as FPGA) or graphics processing units (Graphics Processing Unit, referred to as GPU), etc. For another example, when a certain module above is implemented in the form of a processing element scheduling program code, the processing element may be a general-purpose processor, such as a central processing unit (Central Processing Unit, referred to as CPU) or other processors that can call program codes. For another example, these modules can be integrated together and implemented in the form of a system-on-a-chip (SOC).

[0114] The present application also provides a computer-readable storage medium having a computer program stored thereon, characterized in that when the program is executed by a processor, it implements the method for predicting the inactivation process based on the required amylase activity as described above, or the method for predicting the activity of the amylase after inactivation based on the inactivation process as described above.

[0115] The computer-readable storage medium may be a computer-readable storage medium for predicting an inactivation process based on a desired amylase activity, or may be a computer-readable storage medium for predicting the activity of an inactivated amylase based on an inactivation process.

[0116] The computer-readable storage medium may include a read-only memory, a random access memory, an EEPROM, a CD-ROM or other optical disk storage device, a magnetic disk storage device or other magnetic storage device, a flash memory, a USB flash drive, a mobile hard disk, or any other medium that can be used to store a desired program code in the form of an instruction or data structure and can be accessed by a computer. In addition, any connection can be appropriately referred to as a computer-readable storage medium. For example, if the instruction is sent from a website, a server or other remote source using a coaxial cable, an optical fiber cable, a twisted pair, a digital subscriber line (DSL), or wireless technologies such as infrared, radio, and microwaves, the coaxial cable, optical fiber cable, twisted pair, DSL, or wireless technologies such as infrared, radio, and microwaves are included in the definition of the medium. However, it should be understood that the computer-readable storage medium does not include carrier waves, signals, or other temporary media, but is intended to be directed to non-temporary, tangible storage media.

[0117] Those skilled in the art can understand that all or part of the steps of implementing the above-mentioned method embodiments can be completed by hardware related to the computer program. The computer program can be stored in a computer-readable storage medium. When the program is executed, the steps of the above-mentioned method embodiments are executed; and the aforementioned storage medium includes: ROM, RAM, magnetic disk or optical disk and other media that can store program codes.

[0118] The present application also provides an electronic terminal, comprising a memory and a processor, wherein the memory comprises a computer program, characterized in that when the computer program is executed by the processor, the method for predicting an inactivation process based on the required amylase activity as described above, or the method for predicting the activity of amylase after inactivation based on the inactivation process as described above is implemented.

[0119] The electronic terminal may be an electronic terminal for predicting the inactivation process based on the required amylase activity, or may be an electronic terminal for predicting the activity of the inactivated amylase based on the inactivation process.

[0120] The schematic diagram of the electronic terminal is as follows Fig. 9 As shown, the electronic terminal includes: a processor 31, a memory 32, a communicator 33, a communication interface 34 and a system bus 35; the memory 32 and the communication interface 34 are connected to the processor 31 and the communicator 33 through the system bus 35 and complete communication with each other, the memory 32 is used to store computer programs, the communication interface 34 is used to communicate with other devices, and the processor 31 and the communicator 33 are used to run computer programs so that the terminal executes the above method.

[0121] The system bus 35 may be a Peripheral Component Interconnect (PCI) bus or an Extended Industry Standard Architecture (EISA) bus, etc. The system bus may be divided into an address bus, a data bus, a control bus, etc. For ease of representation, only one thick line is used in the figure, but it does not mean that there is only one bus or one type of bus.

[0122] The communication interface 34 is used to implement communication between the database access device and other devices (such as a client, a read-write library, and a read-only library).

[0123] The memory 32 may include a random access memory (RAM), and may also include a non-volatile memory (non-volatile memory), such as at least one disk memory.

[0124] The processor 31 can be a general-purpose processor, including a central processing unit (CPU), a network processor (NP), etc.; it can also be a digital signal processor (DSP), an application specific integrated circuit (ASIC), a field programmable gate array (FPGA), or other programmable logic devices, discrete gates or transistor logic devices, and discrete hardware components.

[0125] The electronic terminal may be a mobile phone, a computer device, a tablet device, a personal digital processing device, a factory back-end processing device, etc.

[0126] The present application also provides a computer program product, including a computer program, which, when executed by a processor, implements the method for predicting the inactivation process based on the required amylase activity as described above, or the method for predicting the activity of the amylase after inactivation based on the inactivation process as described above.

[0127] The computer program product may be a computer program product for predicting the inactivation process based on the required amylase activity, or may be a computer program product for predicting the amylase activity after inactivation based on the inactivation process.

[0128] In various embodiments, the present application can be implemented as a system or method for performing the above-mentioned technology alone or in any combination. In another embodiment, the present application can be implemented as a computer program product including a computer-readable storage medium and a computer program code encoded on the medium for performing the above-mentioned technology. It should be understood that various programming languages ​​can be used to implement the present application and are not limited to a specific programming language.

[0129] The present application also provides a method for inactivating amylase, comprising placing the amylase at 55-65° C. for 5-30 minutes, or at a temperature above 85° C. for more than 10 minutes, wherein the amylase is selected from ASa or HEa.

[0130] In certain embodiments of the present application, when the amylase is ASa, the method is placing the amylase at 55°C for 30 minutes, or placing the amylase at 60°C for 10 minutes, or placing the amylase at 65°C for 5 minutes.

[0131] In other embodiments of the present application, when the amylase is HEa, the method is to place the amylase at a temperature above 85° C. for more than 10 minutes.

[0132] The method for inactivating amylase provided in the present application can completely inactivate the activity of amylase.

[0133] The present application is further described below by way of examples, but the scope of the present application is not limited thereby. Unless otherwise stated, the experimental methods, detection methods, and preparation methods disclosed in the present application all adopt conventional techniques in molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related fields conventional in the art. These techniques have been fully described in the existing literature. The reagents, instruments, and materials used in the examples can all be obtained by conventional means.

[0134] Example 1 Inactivation of amylase

[0135] Experimental methods:

[0136] 1.Raw materials

[0137] Aspergillus oryzae α-amylase, ASa), Bacillus licheniformis Heat Stable α-amylase, HEa, Lactic acid, Sodium lactate, Hydroxypropyl distarch phosphate, Sodium hydroxide, 3,5-dinitrosalicylic acid, Sodium potassium tartrate

[0138] 2. Inactivation

[0139] Preparation of 0.1U amylase solution A:

[0140] Weigh 50 mg of amylase, dissolve it in 10 mL of water, and store it at 4°C for one week.

[0141] Effect of gradient temperature on amylase activity:

[0142] 20 μL of amylase A solution was placed in a preheated test tube and subjected to gradient treatment at different temperatures to prepare amylase solution B. The temperature gradient was set to 100°C, 95°C, 90°C, 85°C, 80°C, 75°C, 70°C, 65°C, 60°C, and 55°C. At each temperature, the gradient time range was set to 0 to 30 minutes. Subsequently, the treated sample was taken out and placed on ice for cooling. After cooling, the enzyme activity was measured to evaluate the effect of gradient temperature on amylase activity.

[0143] 3. Amylase activity detection

[0144] 150mM lactate buffer configuration:

[0145] Weigh 5.0 g of 90% lactic acid and dissolve it in 1 L of water to prepare a 50 mM lactic acid solution. Weigh 60% sodium lactate and dissolve it in 1 L of water to prepare a 50 mM sodium lactate solution. While measuring the pH, mix the lactic acid and sodium lactate solutions, adjust the pH to 4.2, and store at 4°C.

[0146] Substrate solution C configuration:

[0147] Weigh 0.70 g of hydroxypropyl distarch phosphate and dissolve it in 100 mL of lactic acid buffer (50 mM, pH 4.2). After stirring evenly, place it in a 100°C magnetic stirring water bath and cool it to room temperature after 15 minutes. After cooling, divide it into portions and store it at -20°C.

[0148] DNS reagent configuration:

[0149] Weigh 0.48 g of sodium hydroxide, 0.15 g of 3,5-dinitrosalicylic acid, and 9.0 g of potassium sodium tartrate, dissolve in 30 mL of water, and store in a brown reagent bottle at 4°C.

[0150] Detection method:

[0151] Take a certain amount of substrate solution C and preheat it at 45℃ for 10 minutes. Take 180μL substrate C solution and 20μL amylase B solution to react at 45℃ for 2 minutes. After the reaction is completed, add 200μL DNS color developer, keep it at 100℃ for 15 minutes, place it on ice for 5 minutes, centrifuge it at 7000g at room temperature for 10 seconds, and then measure the absorbance value A540.

[0152] Reducing sugar standard curve determination:

[0153] Prepare 0.5-8mM maltose in lactate buffer (50mM, pH4.2). Take 200μL maltose solution, add 200μL DNS colorimetric reagent, keep at 100℃ for 15min, place on ice for 5min, centrifuge at 7000g for 10s at room temperature, and measure the absorbance value A540. Use the built-in function of Microsoft Excel to draw a trend line, and find the linear regression equation based on the trend line.

[0154] Experimental results:

[0155] The relationship between reducing sugar (maltose) concentration and absorbance is shown in Table 1. The reducing sugar standard curve is shown in Figure 1 shown.

[0156] Table 1 Relationship between reducing sugar (maltose) concentration and absorbance

[0157]

[0158] The linear regression equation of maltose (x) and absorbance (y): y=0.2091x-0.0406.

[0159] Goodness of fit R square value: 0.998.

[0160] Example 2 Construction of a model for predicting inactivation process and amylase activity

[0161] Experimental method: At each inactivation temperature, according to the relationship between inactivation time and residual enzyme activity, OriginLab was used to make a fitting curve, and the Boltzmann sigmoid function equation was used for fitting to obtain the fitting equation of the enzyme activity after inactivation with respect to the inactivation time at each inactivation temperature.

[0162] Experimental results:

[0163] like Figure 2 , 3 As shown, under the same temperature conditions, the activity of amylase gradually decreases with time. As the temperature decreases, it takes longer to completely inactivate amylase. At temperatures above 55°C, the activity of ASa amylase will gradually decrease after heating for a period of time. At temperatures above 65°C, the enzyme activity is completely lost within 5 minutes. At 60°C, it takes about 10 minutes for the amylase activity to be completely lost. At 55°C, it takes about 30 minutes for the amylase activity to be completely lost. It is recommended to inactivate amylase ASa in a short time at above 65°C.

[0164] The fitting equation results of amylase ASa are shown in Table 2, and the goodness of fit R square value is >0.97.

[0165] Table 2 Fitting equation of amylase ASa

[0166]

[0167]

[0168] like Figure 4 As shown in the figure, under the same temperature conditions, the activity of amylase gradually decreases with time. As the temperature decreases, the time to completely inactivate amylase increases. At temperatures above 85°C, the activity of ASa amylase will gradually decrease after heating for a period of time. At temperatures above 85°C, the enzyme activity is completely lost in about 10 minutes. At 80°C for 40 minutes, the activity of amylase is not completely lost. It is recommended to inactivate amylase HEa at above 85°C.

[0169] At each temperature, the inactivation time of amylase HEa was longer than that of ASa.

[0170] The fitting equation results of amylase HEa are shown in Table 3, and the goodness of fit R square value is: >0.98.

[0171] Table 3 Fitting equations for amylase HEa

[0172]

[0173] Example 3 Amylase Half-Life and Inactivation Time

[0174] In this example, the half-life and inactivation time of amylase were calculated by using the fitting equation obtained in Example 2, and compared with the actually measured half-life and inactivation time to verify the fitting equation obtained in Example 2.

[0175] Experimental methods:

[0176] By using the single variable solution function of Microsoft Excel, the corresponding x value for a given y value can be effectively calculated. First, enter a known y value, such as 7, in cell A1 of the Excel worksheet. Then, enter the calculation formula in cell B1, (the following is an example of using cell calculation, not the formula in the embodiment, for example = 2*C1+3, where C1 is the cell for the x value to be solved. Enter the initial guess value (such as 0) in C1. Then, select <Single Variable Solution> in the <Data> tab of Excel, set the target cell to B1, the target value to A1, and the change cell to C1. After clicking OK, Excel will automatically adjust the value of C1 so that the calculation result of B1 is equal to A1. Finally, check the value in C1, which is the x value to be sought. Record the input y value and the calculated x value for subsequent analysis and verification.

[0177] Solution for half-life x: When y=50, the calculation formulas are the fitting equations in Table 2 and Table 3 respectively. The half-life x is obtained according to the above single variable solution function.

[0178] Solution for the inactivation time x: When y=0, the calculation formulas are the fitting equations in Table 2 and Table 3, respectively. According to the above-mentioned single variable solution function, the inactivation time x is obtained.

[0179] Experimental results: The half-life and inactivation time of amylase are shown in Table 4.

[0180] Table 4 Amylase half-life and inactivation time

[0181]

[0182] The half-life calculated according to the fitting curve is consistent with the half-life shown by the experimental results, and the half-life of the inactivated amylase can be obtained by fitting the curve, indicating that the prediction model of the present application has good accuracy.

[0183] Under the same temperature conditions, the inactivation time of HEa is about 5-9 times that of ASa, and it takes longer to inactivate. The same inactivation time is about 95s, but HEa needs 95℃ and ASa needs 65℃, which is 30 degrees higher than ASa.

[0184] The above embodiments are merely illustrative of the principles and effects of the present application and are not intended to limit the present application. Anyone familiar with the technology may modify or change the above embodiments without violating the spirit and scope of the present application. Therefore, all equivalent modifications or changes made by a person of ordinary skill in the art without departing from the spirit and technical ideas disclosed in the present application shall still be covered by the claims of the present application.

Claims

1. A method for predicting a deactivation process based on a desired amylase activity, comprising: 1) Provide the required amylase activity; 2) Preset the inactivation temperature and calculate the inactivation time according to the required amylase activity using an amylase activity-inactivation time prediction model.

2. The method according to claim 1, characterized in that The method of step 2) is: based on the preset inactivation temperature, using the corresponding amylase activity-inactivation time prediction model, substituting the required amylase activity into the prediction model, and calculating the inactivation time; Preferably, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, measuring the amylase activity after inactivation, making a fitting curve by OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature; Preferably, the inactivation temperature is 55-100° C.; the inactivation time is 0-30 minutes; and the amylase is selected from ASa or HEa.

3. The method according to claim 2, characterized in that When the amylase is ASa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572)); 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619)); 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668)); 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695)); 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334)); 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634)); 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153)); 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955)); 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594)); 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364)); When the amylase is HEa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758)); 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318)); 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177)); 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344)); 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567)); Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

4. A device for predicting the inactivation process based on the required amylase activity, comprising: 1) A data input module, used to input the required amylase activity; 2) a calculation module, which is used to calculate the following: preset inactivation temperature, and calculate the inactivation time according to the required amylase activity using an amylase activity-inactivation time prediction model; Preferably, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, measuring the amylase activity after inactivation, making a fitting curve by OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature; Preferably, the inactivation temperature is 55-100°C; the inactivation time is 0-30 minutes; the amylase is selected from ASa or HEa; Preferably, when the amylase is ASa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572)); 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619)); 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668)); 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695)); 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334)); 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634)); 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153)); 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955)); 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594)); 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364)); When the amylase is HEa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758)); 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318)); 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177)); 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344)); 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567)); Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

5. A method for predicting the activity of inactivated amylase based on the inactivation process, comprising: 1) Provide inactivation temperature and inactivation time; 2) According to the inactivation temperature and inactivation time, the amylase activity after inactivation was calculated using the amylase activity-inactivation time prediction model.

6. The method according to claim 5, characterized in that The method of step 2) is: based on the inactivation temperature, using the corresponding amylase activity-inactivation time prediction model, substituting the inactivation time into the prediction model, and calculating the amylase activity after inactivation; Preferably, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, measuring the amylase activity after inactivation, making a fitting curve by OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature; Preferably, the inactivation temperature is 55-100° C.; the inactivation time is 0-30 minutes; and the amylase is selected from ASa or HEa.

7. The method according to claim 6, characterized in that When the amylase is ASa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572)); 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619)); 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668)); 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695)); 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334)); 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634)); 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153)); 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955)); 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594)); 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364)); When the amylase is HEa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758)); 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318)); 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177)); 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344)); 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567)); Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

8. A device for predicting the activity of inactivated amylase based on an inactivation process, comprising: 1) Data input module, used to input inactivation time and inactivation temperature; 2) A calculation module, used for the following calculation: according to the inactivation time and the inactivation temperature, using the amylase activity-inactivation time prediction model, calculate the inactivated amylase activity; Preferably, the amylase activity-inactivation time prediction model is constructed by the following method: setting a gradient inactivation temperature, setting a gradient inactivation time at each inactivation temperature, inactivating the amylase, measuring the amylase activity after inactivation, making a fitting curve by OriginLab, and fitting using the Boltzmann sigmoid function equation to obtain the amylase activity-inactivation time prediction model at each inactivation temperature; Preferably, the inactivation temperature is 55-100°C; the inactivation time is 0-30 minutes; the amylase is selected from ASa or HEa; Preferably, when the amylase is ASa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=-0.4553+(101.66134+0.4553) / (1+exp((x-8.74957) / 2.58572)); 2)t=95℃, y=0.9623+(100.12221-0.9623) / (1+exp((x-12.15868) / 2.47619)); 3)t=90℃, y=2.19587+(98.96031-2.19587) / (1+exp((x-19.45594) / 3.78668)); 4)t=85℃, y=11.10007+(133.75724-11.10007) / (1+exp((x-14.77117) / 12.85695)); 5)t=80℃, y=4.26256+(110.94108-4.26256) / (1+exp((x-29.13015) / 13.68334)); 6)t=75℃, y=4.98968+(105.0241-4.98968) / (1+exp((x-37.49811) / 11.55634)); 7)t=70℃, y=4.28202+(103.93286-4.28202) / (1+exp((x-58.30212) / 14.08153)); 8)t=65℃, y=-4.48052+(232.58576-4.48052) / (1+exp((x-165.74586) / 524.98955)); 9)t=60℃, y=0.57028+(116.65515-0.57028) / (1+exp((x-131.27608) / 70.14594)); 10)t=55℃, y=2.59329+(104.03222-2.59329) / (1+exp((x-93.86879) / 27.46364)); When the amylase is HEa, the amylase activity-inactivation time prediction model is: 1)t=100℃, y=0.25348+(194.93622-0.25348) / (1+exp((x-3.49558) / 37.46758)); 2)t=95℃, y=1.19324+(128.75097-1.19324) / (1+exp((x-71.02372) / 51.6318)); 3)t=90℃, y=-2.46989+(127.17693+2.46989) / (1+exp((x-130.60228) / 81.4177)); 4)t=85℃, y=1.29439+(120.11128-1.29439) / (1+exp((x-180.22015) / 103.49344)); 5)t=80℃, y=-356.83821+(1299.33271+356.83821) / (1+exp((x+11406.89157) / 11716.65567)); Among them, x is the inactivation time, y is the amylase activity, and t is the inactivation temperature.

9. A computer-readable storage medium having a computer program stored thereon, characterized in that: When the program is executed by a processor, the method for predicting the inactivation process based on the required amylase activity as described in any one of claims 1 to 3 or the method for predicting the activity of the inactivated amylase based on the inactivation process as described in any one of claims 5 to 7 is implemented.

10. An electronic terminal comprising a memory and a processor, wherein the memory comprises a computer program, characterized in that: When the computer program is executed by a processor, the method for predicting the inactivation process based on the required amylase activity as described in any one of claims 1 to 3 or the method for predicting the activity of the inactivated amylase based on the inactivation process as described in any one of claims 5 to 7 is implemented.

11. A computer program product, comprising a computer program, wherein when the computer program is executed by a processor, the method for predicting an inactivation process based on the required amylase activity as described in any one of claims 1 to 3, or the method for predicting the activity of an amylase after inactivation based on the inactivation process as described in any one of claims 5 to 7 is implemented.

12. A method for inactivating amylase, comprising placing the amylase at 55-65°C for 5-30 minutes, or placing the amylase at above 85°C for more than 10 minutes, wherein the amylase is selected from ASa or HEa.

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