Method and system for extracting, separating and purifying giant salamander peptide

Abstract

The invention relates to the technical field of giant salamander peptide separation and purification, in particular to a giant salamander peptide extraction and separation and purification method and system.The method comprises the steps that healthy giant salamanders and giant salamander processing equipment are determined, giant salamander processing by-products are obtained, the giant salamander processing by-products are pretreated, to-be-processed by-products are obtained, the to-be-processed by-products are degreased and deodorized, and the giant salamander peptide is obtained. Preparing giant salamander protein dry powder, preparing giant salamander crude peptide liquid, obtaining a target peptide fragment, finely separating the target peptide fragment to obtain target active peptide distillate, freeze-drying the target active peptide distillate to obtain high-purity giant salamander peptide powder, and storing the molecular weight, amino acid sequence and activity verification parameters of the peptide fragment. And extracting, separating and purifying the giant salamander peptide based on the optimized storage database and the giant salamander grease. The extraction efficiency and the separation and purification efficiency of the giant salamander peptide can be improved.

Description

Technical Field

[0001] This invention relates to the field of giant salamander peptide separation and purification technology, and in particular to a method and system for the extraction, separation and purification of giant salamander peptides. Background Technology

[0002] Giant salamander peptides are a class of bioactive peptides extracted from the giant salamander. Peptides possess various biological activities, such as antibacterial, antioxidant, immunomodulatory, and blood pressure-lowering effects. Separation and purification are key steps in the extraction of giant salamander peptides. Through a series of physical and chemical methods, high-purity giant salamander peptides are extracted from complex biological materials.

[0003] Traditional methods for extracting and purifying giant salamander peptides, such as hot water extraction and enzymatic extraction, while simple to operate, suffer from low extraction efficiency and difficulty in obtaining high-purity peptide fragments. Secondly, existing separation techniques, such as gel filtration and ion exchange chromatography, while capable of separating some peptide fragments, are complex, costly, and difficult to implement on a large scale. Therefore, improving the extraction and purification efficiency of giant salamander peptides is an urgent technical problem to be solved. Summary of the Invention

[0004] This invention provides a method for the extraction, separation, and purification of giant salamander peptides, as well as a computer-readable storage medium. Its main purpose is to improve the extraction efficiency and separation and purification efficiency of giant salamander peptides.

[0005] To achieve the above objectives, the present invention provides a method for the extraction, separation, and purification of giant salamander peptides, comprising: The healthy giant salamander and the giant salamander processing equipment were identified. The giant salamander processing equipment includes: a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. Receive a giant salamander peptide extraction instruction, obtain giant salamander processing by-products according to healthy giant salamanders and the giant salamander peptide extraction instruction, perform pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed, wherein the pretreatment operations include: washing and crushing; The processing by-products were degreased and deodorized to obtain degreased by-products and giant salamander oil. Giant salamander protein powder was prepared based on the degreased by-products, and giant salamander crude peptide liquid was prepared based on the giant salamander protein powder. Receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment, and obtain the target active peptide distillate; The target active peptide was freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptide were determined based on the high-purity giant salamander peptide powder. The activity of high-purity giant salamander peptide powder was verified to obtain activity verification parameters. The peptide molecular weight, amino acid sequence and activity verification parameters were stored in a pre-constructed storage database to obtain an optimized storage database. The extraction, separation, and purification of giant salamander peptides were completed based on an optimized storage database and giant salamander oil.

[0006] Optionally, the process of defatting and deodorizing the by-product to obtain defatted by-product and giant salamander oil includes: The by-products to be processed are spread in a material tray to obtain a loading tray. The loading tray is then placed in the extraction vessel to obtain the by-products to be extracted. Static extraction is performed on the extraction byproduct to obtain the first extraction byproduct. The first extraction byproduct is then dynamically circulated using a preset carbon dioxide gas to obtain a carbon dioxide fluid rich in oil. A separation vessel is used to separate a fluid rich in grease and carbon dioxide to obtain separated carbon dioxide gas and grease. The separated carbon dioxide gas is recovered using a carbon dioxide storage tank to obtain recovered carbon dioxide. The recovered carbon dioxide is then used as carbon dioxide gas and returned to the step of dynamically recycling the first extraction byproduct using the preset carbon dioxide gas until a pre-constructed extraction completion command is received. The defatting byproducts were obtained from the extraction vessel, and the oil was collected to obtain giant salamander oil.

[0007] Optionally, the preparation of giant salamander protein powder from defatting byproducts includes: The defatting byproduct was quick-frozen to obtain a pre-frozen defatting byproduct. The pre-frozen defatting byproduct was freeze-dried using a pre-constructed freeze-drying chamber to obtain an initial freeze-dried product. The temperature of the initial freeze-dried product was monitored to obtain the initial cold trap temperature. When the initial cold trap temperature reaches the preset standard cold trap temperature, the pre-built vacuum pump is started, and the freeze-drying chamber is evacuated using the started vacuum pump to obtain a vacuum freeze-drying chamber. The initial freeze-dried product in the vacuum freeze-drying chamber is heated using a preset initial heating temperature to obtain a preliminary dried product. The state data of the preliminary dried product is obtained, wherein the state data is either a porous state or a non-porous state. If the state data is non-porous, the preliminary dried product is used as the initial freeze-dried product, and the process returns to the step of heating the initial freeze-dried product in the vacuum freeze-drying chamber using a preset initial heating temperature until the state data is porous. If the state data is porous, the pre-dried product is continuously heated using a preset secondary drying time to obtain the secondary dried product. Giant salamander protein powder was obtained using secondary drying products and a pre-constructed inert gas.

[0008] Optionally, the step of preparing giant salamander crude peptide solution from giant salamander protein powder includes: To prepare a low cosolvent, the low cosolvent and giant salamander protein powder are mixed according to a preset mixing ratio to obtain an initial mixture. The initial mixture is then ultrasonically treated using a pre-constructed ultrasonic cell disruptor to obtain an ultrasonic mixture. The ultrasonic cell disruptor has preset ultrasonic parameters. The pH value of the ultrasonic mixture is measured to obtain the pH value of the mixture. The pH value of the mixture is adjusted to obtain the pH value of the standard mixture. The standard pH value mixture is then identified based on the pH value of the standard mixture. The standard pH mixture was subjected to temperature adjustment to obtain a standard temperature mixture, and appropriate amounts of alkaline protease and flavor protease were obtained. The standard temperature mixture was subjected to initial enzymatic hydrolysis using a preset first enzymatic hydrolysis time and an appropriate amount of alkaline protease to obtain the first enzymatic hydrolysis mixture. The first enzymatically hydrolyzed mixture was enzymatically hydrolyzed using a preset second enzymatic hydrolysis time and an appropriate amount of flavor protease to obtain an enzymatically hydrolyzed mixture; Enzyme inactivation was performed on the enzymatically hydrolyzed mixture using preset enzyme inactivation parameters to obtain an enzymatically hydrolyzed mixture. The enzymatically hydrolyzed mixture was then transferred to a pre-constructed centrifuge tube to obtain a mixture to be centrifuged. The mixture to be centrifuged was then centrifuged to obtain crude peptide solution of giant salamander.

[0009] Optionally, obtaining the target peptide fragment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instructions includes: The pre-constructed tangential flow ultrafiltration system was set up according to the giant salamander peptide separation and purification instructions to obtain a configured tangential flow ultrafiltration system, which includes: a first-stage ultrafiltration membrane, a second-stage ultrafiltration membrane and a third-stage ultrafiltration membrane; The crude peptide solution of giant salamander was subjected to macromolecular ultrafiltration using a first-stage ultrafiltration membrane to obtain the first giant salamander peptide solution. The first giant salamander peptide solution was subjected to medium-molecular ultrafiltration using a second-stage ultrafiltration membrane to obtain the second giant salamander peptide solution. The second giant salamander peptide solution was ultrafiltered using a third-stage ultrafiltration membrane to obtain a third giant salamander peptide solution. The protein concentration of the third giant salamander peptide solution was then determined, and it was determined whether the protein concentration of the peptide solution was equal to the preset standard protein concentration of the peptide solution. If the protein concentration of the peptide solution is not equal to the protein concentration of the standard peptide solution, the third giant salamander peptide solution is adjusted to obtain an updated giant salamander peptide solution. The updated giant salamander peptide solution is used as the third giant salamander peptide solution. The process of performing protein detection on the third giant salamander peptide solution is repeated until the protein concentration of the peptide solution is equal to the protein concentration of the standard peptide solution. If the protein concentration of the peptide solution is equal to that of the standard peptide solution, then the third giant salamander peptide solution is taken as the target peptide segment.

[0010] Optionally, the step of performing protein detection on the third giant salamander peptide solution to obtain the protein concentration of the peptide solution includes: The first and second peptide solutions were extracted from the third giant salamander peptide solution. The first peptide solution was subjected to ultrafiltration and centrifugation to obtain the sample residue. The BCA method and Bradford method samples were extracted from the sample residue. The protein concentration of the sample determined by the BCA method was obtained by BCA method determination, and the protein concentration of the sample determined by the Bradford method was obtained by Bradford method determination. The concentration of the peptide solution was obtained based on the Bradford method and the BCA method. The pre-set exogenous standard protein was added to the second peptide solution sample to obtain the internal standard peptide solution sample. The internal standard peptide solution sample was subjected to ultrafiltration and centrifugation to obtain the internal standard sample residue. The internal standard concentration was obtained based on the internal standard sample residue. The exogenous internal standard concentration of the exogenous standard protein was obtained. The recovery rate of the internal standard was calculated based on the internal standard concentration and the exogenous internal standard concentration. If the recovery rate of the internal standard is not within the preset range, the protein concentration of the peptide solution is calculated based on the recovery rate of the internal standard and the concentration of the peptide solution. If the recovery rate of the internal standard is within the range of the internal standard recovery rate, then the concentration of the peptide solution measured shall be taken as the protein concentration of the peptide solution.

[0011] Optionally, obtaining the concentration of the exogenous internal standard for the exogenous standard protein includes: Determine the protein addition volume and concentration of the exogenous standard protein, and calculate the total amount of exogenous standard protein based on the protein addition volume and concentration. The total amount of exogenous standard protein is the product of the protein addition volume and the exogenous standard protein concentration. Obtain the total sample volume of the second peptide solution sample, and calculate the exogenous internal standard concentration based on the total sample volume and the total amount of exogenous standard protein. The exogenous internal standard concentration is the product of the total sample volume and the total amount of exogenous standard protein.

[0012] Optionally, the fine separation of the target peptide to obtain the target active peptide distillate includes: The target column set and SMB system are determined, and each target column in the target column set is connected in series with the SMB system to obtain the integrated SMB system; The target chromatographic column set in the integrated SMB system is divided into functional regions, which include multiple functional regions and correspond one-to-one with the target chromatographic column. Representative chromatographic columns were extracted from the integrated SMB system and connected to a pre-constructed high-performance liquid chromatography system to obtain a pre-configured high-performance liquid chromatography system. The target peptide is introduced into a prepared high-performance liquid chromatography system to obtain the target peptide to be separated. The target peptide to be separated is subjected to a phase B gradient elution operation to obtain the retention time of the target active peptide. Using the retention time of the target active peptide as the switching time, parameters are set for each functional region in the functional region set to obtain a parameterized functional region set, which includes multiple parameterized functional regions. The target peptide to be separated is separated by using a parameterized functional region set and switching time to obtain an initial target peptide. The initial target peptide is then detected in real time to obtain activity signals and ultraviolet signals. If the activity signal and ultraviolet signal meet the preset normal signal parameter conditions, the target active peptide distillate is collected using a pre-constructed fraction collector.

[0013] Optionally, the activity verification of the high-purity giant salamander peptide powder to obtain activity verification parameters includes: Obtain the stop solution, enzyme solution, substrate solution and buffer solution. Prepare the reaction solution based on high-purity giant salamander peptide powder. Mix the enzyme solution, substrate solution, buffer solution and reaction solution to obtain the initial mixed reaction solution. The initial mixed reaction solution is subjected to a water bath reaction to obtain a mixed reaction solution. The termination solution is then added to the mixed reaction solution to obtain a termination reaction solution. The solution used to terminate the reaction was centrifuged to obtain the supernatant. The absorbance of the supernatant was measured to obtain the absorbance of the sample, as well as the absorbance of the substrate control and the blank control. The ACE inhibition rate was calculated based on the absorbance of the sample, the absorbance of the substrate control, and the absorbance of the blank control, and the ACE inhibition rate was used as the activity verification parameter.

[0014] To achieve the above objectives, the present invention also provides a system for the extraction, separation, and purification of giant salamander peptides, comprising: The raw material preparation module is used to determine healthy giant salamanders and giant salamander processing equipment. The giant salamander processing equipment includes a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. It receives giant salamander peptide extraction instructions, obtains giant salamander processing by-products according to the healthy giant salamanders and giant salamander peptide extraction instructions, and performs pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed. The pretreatment operations include washing and crushing. The crude peptide solution preparation module is used to degrease and deodorize the by-products to be processed, to obtain degreased by-products and giant salamander oil, to prepare giant salamander protein powder based on the degreased by-products, and to prepare giant salamander crude peptide solution based on the giant salamander protein powder. The peptide separation and purification module is used to receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment to obtain the target active peptide distillate, freeze-dry the target active peptide distillate to obtain high-purity giant salamander peptide powder, and confirm the peptide molecular weight and amino acid sequence based on the high-purity giant salamander peptide powder. The peptide data management module is used to verify the activity of high-purity giant salamander peptide powder, obtain activity verification parameters, and use a pre-constructed storage database to store peptide molecular weight, amino acid sequence and activity verification parameters to obtain an optimized storage database. Based on the optimized storage database and giant salamander oil, the extraction, separation and purification of giant salamander peptides are completed.

[0015] To address the above problems, the present invention also provides an electronic device, the electronic device comprising: Memory, storing at least one instruction; The processor executes the instructions stored in the memory to implement the above-described method for the extraction, separation, and purification of giant salamander peptides.

[0016] To address the aforementioned problems, the present invention also provides a computer-readable storage medium storing at least one instruction, which is executed by a processor in an electronic device to implement the above-described method for the extraction, separation, and purification of giant salamander peptides.

[0017] To address the problems described in the background art, this invention identifies healthy giant salamanders and their processing equipment. The processing equipment includes a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. This invention selects healthy giant salamanders as raw materials because the proteins and other components in healthy salamanders are in a normal state, reducing abnormalities caused by disease or unhealthy factors, thus laying the foundation for extracting high-quality giant salamander peptides. The invention receives giant salamander peptide extraction instructions, obtains processing by-products based on healthy giant salamanders and the peptide extraction instructions, and performs pretreatment operations on the by-products to obtain the by-products to be processed. The pretreatment operations include washing and crushing. This invention... The processing byproducts of giant salamanders are pulverized into smaller particles, increasing the specific surface area of ​​the material, which is beneficial for subsequent degreasing, deodorization, and enzymatic hydrolysis, thus improving reaction efficiency. The byproducts are then degreased and deodorized to obtain degreased byproducts and giant salamander oil. Giant salamander protein powder is prepared from the degreased byproducts, and then a crude giant salamander peptide solution is prepared from the protein powder. This invention removes oil from the giant salamander processing byproducts, which on the one hand reduces the impact of oil on subsequent enzymatic hydrolysis and separation purification processes, improving the purity of the giant salamander peptides; on the other hand, the obtained giant salamander oil can be used as a raw material for other products, realizing further resource utilization. The invention also receives giant salamander peptide separation and purification instructions, and prepares the crude giant salamander peptide solution and giant salamander peptide separation... This invention obtains target peptides through purification commands, performs fine separation on these peptides to obtain target active peptide fractions. The fine separation further removes impurities and other unwanted peptides, yielding high-purity target active peptide fractions, thus improving the quality and bioactivity of the giant salamander peptides. The target active peptide fractions are then freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptides are confirmed based on the high-purity giant salamander peptide powder. This invention uses freeze-drying of the target active peptide fraction, which removes moisture at low temperatures, maximizing the preservation of the giant salamander peptide's bioactivity and structural integrity. Simultaneously, the dried powder is easy to store and use, extending the product's shelf life. This invention validates the activity of high-purity giant salamander peptide powder, obtaining activity validation parameters. A pre-constructed storage database is used to store the peptide molecular weight, amino acid sequence, and activity validation parameters, resulting in an optimized storage database. This invention enables the evaluation of the actual biological activity of giant salamander peptides, such as antioxidant and immunomodulatory functions, providing a scientific basis for product quality control and application. Based on the optimized storage database and giant salamander oil, the extraction, separation, and purification of giant salamander peptides are completed. This invention establishes a giant salamander peptide information database, facilitating subsequent research, development, and quality traceability, and also providing data support for the standardized production of giant salamander peptides. Therefore, this invention can improve the extraction and purification efficiency of giant salamander peptides. Attached Figure Description

[0018] Figure 1 This is a schematic flowchart of a method for extracting, separating, and purifying giant salamander peptides according to an embodiment of the present invention. Figure 2 A functional block diagram of a system for the extraction, separation, and purification of giant salamander peptides provided in an embodiment of the present invention; Figure 3 This is a schematic diagram of an electronic device for implementing the extraction, separation, and purification method of giant salamander peptides according to an embodiment of the present invention.

[0019] Explanation of reference numerals in the attached figures: 10. Electronic device; 11. Processor; 12. Memory; 13. Bus.

[0020] The realization of the objective, functional features and advantages of the present invention will be further explained in conjunction with the embodiments and with reference to the accompanying drawings. Detailed Implementation

[0021] It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

[0022] This application provides a method for the extraction, separation, and purification of giant salamander peptides. The execution entity of this method includes, but is not limited to, at least one electronic device configured to execute the method provided in this application, such as a server or a terminal. In other words, the extraction, separation, and purification method can be executed by software or hardware installed on a terminal device or a server device, and the software can be a blockchain platform. The server includes, but is not limited to, a single server, a server cluster, a cloud server, or a cloud server cluster.

[0023] Reference Figure 1 The diagram shown is a schematic flowchart of a method for extracting, separating, and purifying giant salamander peptides according to an embodiment of the present invention. In this embodiment, the method for extracting, separating, and purifying giant salamander peptides includes: S1. Determine the healthy giant salamanders and the giant salamander processing equipment, wherein the giant salamander processing equipment includes: material tray, extraction vessel, separation vessel and carbon dioxide storage tank.

[0024] It should be clarified that "healthy giant salamanders" refers to giant salamanders that have undergone rigorous screening and are free from diseases, injuries, or other health problems, making them suitable for extracting bioactive substances such as peptides. Giant salamander processing equipment refers to specialized equipment used for processing giant salamanders and extracting peptides, including material trays, extraction kettles, separation kettles, and carbon dioxide storage tanks.

[0025] S2. Receive the giant salamander peptide extraction instruction, obtain the giant salamander processing by-product according to the healthy giant salamander and the giant salamander peptide extraction instruction, perform a pretreatment operation on the giant salamander processing by-product to obtain the by-product to be processed, wherein the pretreatment operation includes: washing and crushing.

[0026] It should be explained that the "giant salamander peptide extraction instruction" refers to the instruction issued by the operator to initiate the giant salamander peptide extraction process. Giant salamander processing by-products refer to raw materials extracted from healthy giant salamanders for the production of bioactive substances such as peptides, such as skin, minced meat, and bones. The pretreatment operation of the giant salamander processing by-products refers to washing the by-products with clean water to remove impurities and surface contaminants, and then placing the washed by-products into a pulverizer for further processing. The by-products to be processed refer to the giant salamander processing by-products after washing and pulverizing.

[0027] S3. Degrease and deodorize the by-products to obtain degreased by-products and giant salamander oil. Prepare giant salamander protein powder based on the degreased by-products, and prepare giant salamander crude peptide liquid based on the giant salamander protein powder.

[0028] Specifically, the process of degreasing and deodorizing the by-products to obtain degreased by-products and giant salamander oil includes: The by-products to be processed are spread in a material tray to obtain a loading tray. The loading tray is then placed in the extraction vessel to obtain the by-products to be extracted. Static extraction is performed on the extraction byproduct to obtain the first extraction byproduct. The first extraction byproduct is then dynamically circulated using a preset carbon dioxide gas to obtain a carbon dioxide fluid rich in oil. A separation vessel is used to separate a fluid rich in grease and carbon dioxide to obtain separated carbon dioxide gas and grease. The separated carbon dioxide gas is recovered using a carbon dioxide storage tank to obtain recovered carbon dioxide. The recovered carbon dioxide is then used as carbon dioxide gas and returned to the step of dynamically recycling the first extraction byproduct using the preset carbon dioxide gas until a pre-constructed extraction completion command is received. The defatting byproducts were obtained from the extraction vessel, and the oil was collected to obtain giant salamander oil.

[0029] It should be explained that a material tray is a container used to hold the by-products to be processed. It is typically a flat-bottomed, shallow dish made of stainless steel or other pressure-resistant and corrosion-resistant materials. It is used to evenly distribute the by-products so that they can contact the carbon dioxide fluid more evenly during extraction, improving extraction efficiency. A loading tray refers to a tray already loaded with the by-products to be processed. An extraction vessel is a high-pressure vessel used for supercritical carbon dioxide extraction. It provides a high-pressure, sealed environment, allowing carbon dioxide to contact the by-products to be extracted in a supercritical state, achieving oil extraction. The by-products to be extracted refer to the giant salamander by-products already loaded in the material tray and placed in the extraction vessel. Static extraction of the by-products to be extracted involves injecting carbon dioxide into the extraction vessel while simultaneously starting a circulation pump, allowing the carbon dioxide to circulate between the extraction vessel and the separation vessel. This process allows the supercritical carbon dioxide to fully permeate the by-products to be extracted. The first extraction by-product refers to the by-product after static extraction. The separation of oil-rich carbon dioxide fluid using a pre-constructed separation vessel refers to allowing the oil-rich carbon dioxide fluid to flow from the extraction vessel into the separation vessel. In the separation vessel, pressure is reduced, decreasing the solubility of CO2, causing the extracted oil and odorous substances to completely precipitate and settle at the bottom. Carbon dioxide gas refers to the carbon dioxide used in the extraction process, typically used in a supercritical state. Oil-rich carbon dioxide fluid refers to carbon dioxide fluid carrying a large amount of oil after dynamic circulation. Carbon dioxide fluid refers to carbon dioxide under specific conditions (such as high temperature and high pressure), possessing the density of a liquid and the diffusivity of a gas, effectively dissolving and carrying target substances (such as oil). A separation vessel is a device used to separate oil-rich carbon dioxide fluid. Separated carbon dioxide gas refers to the carbon dioxide gas separated from the oil-rich carbon dioxide fluid; the separated carbon dioxide gas can be recycled and reused, improving resource utilization. Oil refers to the fatty components extracted from the first extraction byproduct. A carbon dioxide storage tank is a high-pressure container used to store recovered carbon dioxide gas for reuse in subsequent extraction processes. Recovered carbon dioxide refers to the carbon dioxide gas separated from the separation vessel. The extraction completion command is a signal from the system indicating that the extraction process is complete based on preset parameters (such as time, pressure, and temperature). Defatted byproducts refer to the byproducts from which oil and fishy odor have been removed after the extraction process. Giant salamander oil refers to all the oil extracted from the giant salamander byproducts.

[0030] Specifically, the preparation of giant salamander protein powder from defatting byproducts includes: The defatting byproduct was quick-frozen to obtain a pre-frozen defatting byproduct. The pre-frozen defatting byproduct was freeze-dried using a pre-constructed freeze-drying chamber to obtain an initial freeze-dried product. The temperature of the initial freeze-dried product was monitored to obtain the initial cold trap temperature. When the initial cold trap temperature reaches the preset standard cold trap temperature, the pre-built vacuum pump is started, and the freeze-drying chamber is evacuated using the started vacuum pump to obtain a vacuum freeze-drying chamber. The initial freeze-dried product in the vacuum freeze-drying chamber is heated using a preset initial heating temperature to obtain a preliminary dried product. The state data of the preliminary dried product is obtained, wherein the state data is either a porous state or a non-porous state. If the state data is non-porous, the preliminary dried product is used as the initial freeze-dried product, and the process returns to the step of heating the initial freeze-dried product in the vacuum freeze-drying chamber using a preset initial heating temperature until the state data is porous. If the state data is porous, the pre-dried product is continuously heated using a preset secondary drying time to obtain the secondary dried product. Giant salamander protein powder was obtained using secondary drying products and a pre-constructed inert gas.

[0031] It should be explained that the quick-freezing of defatting byproducts refers to placing the defatting byproducts in quick-freezing equipment (such as a liquid nitrogen freezer) and rapidly lowering the temperature to -80°C within minutes to reduce ice crystal formation and protect protein structure. Pre-frozen defatting byproducts refer to defatting byproducts that have undergone quick-freezing. A freeze-drying chamber is a sealed container used for freeze-drying, providing a low-temperature, low-pressure environment for freeze-drying pre-frozen defatting byproducts. Initial freeze-dried products refer to defatting byproducts that have undergone preliminary freeze-drying in the freeze-drying chamber; at this point, the defatting byproducts have been partially dehydrated but may still contain some moisture. Initial cold trap temperature refers to the initial temperature of the cold trap during the freeze-drying process. The cold trap is used to capture water vapor sublimating from the freeze-drying chamber. The standard cold trap temperature refers to the target temperature of the cold trap set during the freeze-drying process, which is below -50°C. When the cold trap temperature reaches the standard cold trap temperature, it indicates that the freeze-drying chamber environment has stabilized and the next step can be performed. A vacuum pump is a device used to reduce the pressure inside a freeze-drying chamber. By creating a vacuum, the pressure inside the chamber is reduced, promoting the sublimation of moisture and accelerating the freeze-drying process. A vacuum freeze-drying chamber refers to a freeze-drying chamber where the pressure is significantly reduced under the action of a vacuum pump. The initial heating temperature refers to the initial temperature set when heating the initially freeze-dried product in the vacuum freeze-drying chamber. Heating promotes the sublimation of moisture and accelerates the freeze-drying process. The pre-dried product refers to the freeze-dried product after the initial heating treatment. The secondary drying time refers to the time required for further drying after the pre-dried product reaches a porous state. By continuously increasing the temperature, the moisture in the product is further removed to achieve the purpose of drying. The secondary dried product refers to the product after the secondary drying treatment. At this point, the secondary dried product is completely dry with extremely low moisture content and is suitable for long-term storage. An inert gas is a chemically inert gas, such as nitrogen. In this invention, after freeze-drying, an inert gas is introduced into the vacuum freeze-drying chamber to prevent oxidation of the secondary dried product and protect the activity of the protein. Giant salamander protein powder refers to the dried protein powder obtained after freeze-drying and drying treatment.

[0032] Specifically, the preparation of giant salamander crude peptide solution from giant salamander protein powder includes: To prepare a low cosolvent, the low cosolvent and giant salamander protein powder are mixed according to a preset mixing ratio to obtain an initial mixture. The initial mixture is then ultrasonically treated using a pre-constructed ultrasonic cell disruptor to obtain an ultrasonic mixture. The ultrasonic cell disruptor has preset ultrasonic parameters. The pH value of the ultrasonic mixture is measured to obtain the pH value of the mixture. The pH value of the mixture is adjusted to obtain the pH value of the standard mixture. The standard pH value mixture is then identified based on the pH value of the standard mixture. The standard pH mixture was subjected to temperature adjustment to obtain a standard temperature mixture, and appropriate amounts of alkaline protease and flavor protease were obtained. The standard temperature mixture was subjected to initial enzymatic hydrolysis using a preset first enzymatic hydrolysis time and an appropriate amount of alkaline protease to obtain the first enzymatic hydrolysis mixture. The first enzymatically hydrolyzed mixture was enzymatically hydrolyzed using a preset second enzymatic hydrolysis time and an appropriate amount of flavor protease to obtain an enzymatically hydrolyzed mixture; Enzyme inactivation was performed on the enzymatically hydrolyzed mixture using preset enzyme inactivation parameters to obtain an enzymatically hydrolyzed mixture. The enzymatically hydrolyzed mixture was then transferred to a pre-constructed centrifuge tube to obtain a mixture to be centrifuged. The mixture to be centrifuged was then centrifuged to obtain crude peptide solution of giant salamander.

[0033] It should be explained that a low-cosolvent is a mixed solvent composed of two or more components. The low-cosolvent of this invention consists of choline chloride and glycerol in a molar ratio of 1:2, and a preset solvent volume is added to reduce viscosity. The low-cosolvent can effectively disrupt protein structures, expose enzyme cleavage sites, and improve enzymatic hydrolysis efficiency. Solvent volume refers to a volume preset according to requirements. The mixing ratio refers to a preset ratio of the low-cosolvent to the giant salamander protein powder, ensuring that the low-cosolvent can fully act on the giant salamander protein powder and improve the pretreatment effect. For example, the mixing ratio of the low-cosolvent to the giant salamander protein powder is 15:1 (mL / g). The initial mixture refers to the mixture obtained after mixing the low-cosolvent and the giant salamander protein powder according to the mixing ratio. An ultrasonic cell disruptor is a device that uses the cavitation effect of ultrasound to disrupt cells and disperse substances, used to further disrupt cell structures and protein aggregates, producing a synergistic effect with the low-cosolvent. The ultrasonic mixture refers to the initial mixture after ultrasonic treatment. After ultrasonic treatment, the cell structure and protein aggregates in the initial mixture are further disrupted, which facilitates the subsequent enzymatic hydrolysis process.

[0034] Importantly, the ultrasonic parameters refer to the pre-set parameters of the ultrasonic cell disruptor when processing the mixture, including power and time. The pH measurement of the ultrasonic mixture is performed using a pH meter. The pH value of the mixture refers to its acidity or alkalinity; pH ​​affects the efficiency and effectiveness of subsequent enzymatic hydrolysis and needs to be adjusted to a suitable range. Adjusting the pH value of the mixture involves adding dilute sodium hydroxide solution dropwise if the pH value is lower than the preset target pH value, and adding dilute hydrochloric acid dropwise if the pH value is higher than the preset target pH value. The target pH value is the pre-set pH value. The standard mixture pH value refers to the pH value achieved after adjustment, used to ensure that the enzymatic hydrolysis reaction proceeds under the standard mixture pH conditions. The standard temperature mixture refers to the mixture that has reached the enzymatic hydrolysis reaction temperature after temperature adjustment. The steps for obtaining appropriate amounts of alkaline protease and flavor protease are as follows: The weight of the standard temperature mixture is obtained; the addition ratios of alkaline protease and flavor protease are determined; the appropriate amount of alkaline protease is obtained by multiplying the weight of the standard temperature mixture by the addition ratio of alkaline protease; the appropriate amount of flavor protease is obtained by multiplying the weight of the standard temperature mixture by the addition ratio of flavor protease. Appropriate amount of alkaline protease refers to the amount of enzyme added calculated based on the weight of the standard temperature mixture. Appropriate amount of flavor protease refers to the amount of enzyme added calculated based on the weight of the standard temperature mixture.

[0035] Understandably, the first enzymatic hydrolysis time refers to the time during which the alkaline protease acts on the mixture. For example, the first enzymatic hydrolysis time is 2 hours. The first enzymatic hydrolysate refers to the mixture after alkaline protease hydrolysis. The second enzymatic hydrolysis time refers to the time during which the flavor protease acts on the first enzymatic hydrolysate. The enzymatically hydrolyzed mixture refers to the mixture after flavor protease hydrolysis. Enzyme inactivation parameters refer to the conditions used to inactivate enzyme activity, including enzyme inactivation temperature and enzyme inactivation time. Enzyme inactivation temperature and enzyme inactivation time refer to the temperature and time corresponding to the enzyme inactivation operation performed on the enzymatically hydrolyzed mixture, respectively. The enzymatically hydrolyzed mixture refers to the mixture after the enzyme inactivation operation, in which the enzyme activity has been inactivated and the enzymatic hydrolysis reaction has stopped. A centrifuge tube is a container used for centrifugation operations to separate enzymatically hydrolyzed mixtures. The mixture to be centrifuged refers to the enzymatically hydrolyzed mixture transferred to a centrifuge tube, ready for centrifugation. Giant salamander crude peptide liquid refers to the liquid containing peptide fragments obtained from the supernatant after centrifugation.

[0036] S4. Receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment, and obtain the target active peptide distillate.

[0037] It should be explained that the giant salamander peptide separation and purification command is a command issued by the system to start the giant salamander peptide separation and purification process.

[0038] Specifically, the step of obtaining the target peptide fragment according to the crude peptide solution of the giant salamander and the giant salamander peptide separation and purification instructions includes: The pre-constructed tangential flow ultrafiltration system was set up according to the giant salamander peptide separation and purification instructions to obtain a configured tangential flow ultrafiltration system, which includes: a first-stage ultrafiltration membrane, a second-stage ultrafiltration membrane and a third-stage ultrafiltration membrane; The crude peptide solution of giant salamander was subjected to macromolecular ultrafiltration using a first-stage ultrafiltration membrane to obtain the first giant salamander peptide solution. The first giant salamander peptide solution was subjected to medium-molecular ultrafiltration using a second-stage ultrafiltration membrane to obtain the second giant salamander peptide solution. The second giant salamander peptide solution was ultrafiltered using a third-stage ultrafiltration membrane to obtain a third giant salamander peptide solution. The protein concentration of the third giant salamander peptide solution was then determined, and it was determined whether the protein concentration of the peptide solution was equal to the preset standard protein concentration of the peptide solution. If the protein concentration of the peptide solution is not equal to the protein concentration of the standard peptide solution, the third giant salamander peptide solution is adjusted to obtain an updated giant salamander peptide solution. The updated giant salamander peptide solution is used as the third giant salamander peptide solution. The process of performing protein detection on the third giant salamander peptide solution is repeated until the protein concentration of the peptide solution is equal to the protein concentration of the standard peptide solution. If the protein concentration of the peptide solution is equal to that of the standard peptide solution, then the third giant salamander peptide solution is taken as the target peptide segment.

[0039] It should be explained that a tangential flow ultrafiltration system is a device that uses ultrafiltration membranes for separation, used to separate peptides of different molecular weights from crude giant salamander peptide solutions. Setting up the pre-constructed tangential flow ultrafiltration system refers to configuring different ultrafiltration membranes within the system. A configured tangential flow ultrafiltration system refers to a tangential flow ultrafiltration system that has been set up and is ready for use, including a first-stage ultrafiltration membrane, a second-stage ultrafiltration membrane, and a third-stage ultrafiltration membrane. The first-stage ultrafiltration membrane is a membrane that retains larger molecules, used for the initial removal of large protein molecules and incompletely digested fragments. For example, the first-stage ultrafiltration membrane has a 10 kDa molecular weight cutoff. The second-stage ultrafiltration membrane is a membrane that retains medium molecular weight peptides, used for further separation of medium molecular weight peptides. For example, the second-stage ultrafiltration membrane has a 3 kDa molecular weight cutoff. The third-stage ultrafiltration membrane is a membrane that retains smaller molecules. For example, the third-stage ultrafiltration membrane has a 1 kDa molecular weight cutoff. The first giant salamander peptide solution refers to the peptide solution obtained after treatment with the first-stage ultrafiltration membrane. The second-stage giant salamander peptide solution refers to the peptide solution obtained after treatment with a second-stage ultrafiltration membrane. Macromolecular ultrafiltration refers to ultrafiltration using a first-stage ultrafiltration membrane, primarily removing large protein molecules and incompletely hydrolyzed fragments. The crude giant salamander peptide solution is passed through the first-stage ultrafiltration membrane, and the liquid that permeates through the first-stage membrane is collected. Medium-molecular ultrafiltration refers to ultrafiltration using a second-stage ultrafiltration membrane, primarily removing peptides with a 3kDa molecular weight cutoff.

[0040] Understandably, the third-stage giant salamander peptide solution refers to the peptide solution obtained after treatment through a third-stage ultrafiltration membrane. It contains small-molecule peptides with high peptide activity. The peptide protein concentration refers to the protein concentration in the third-stage giant salamander peptide solution. The standard peptide protein concentration refers to a pre-set target value for peptide protein concentration. Adjusting the third-stage giant salamander peptide solution means that if the peptide protein concentration is lower than the standard peptide protein concentration, the ultrafiltration time can be increased; if the peptide protein concentration is higher than the standard peptide protein concentration, the third-stage giant salamander peptide solution is diluted. The updated giant salamander peptide solution refers to the adjusted third-stage giant salamander peptide solution. The target peptide segment refers to the third-stage giant salamander peptide solution with a peptide protein concentration equal to the pre-set standard peptide protein concentration. The fact that the protein concentration in the peptide solution of this invention is not equal to the preset standard protein concentration indicates that some large protein molecules or incompletely hydrolyzed fragments may not have been completely retained, resulting in a lower protein concentration in the permeate than the standard protein concentration. Alternatively, a low flow rate or long duration during ultrafiltration may have led to excessive concentration of small peptides, resulting in a higher protein concentration than the standard protein concentration. When the protein concentration in the peptide solution equals the preset standard protein concentration, it indicates that the concentration of the target peptide meets the standard protein concentration, signifying a successful ultrafiltration process. This allows the process to proceed to the next step of purification and application, ensuring the smooth progress of subsequent steps and improving production efficiency and product quality.

[0041] Specifically, the process of detecting proteins in the third giant salamander peptide solution to obtain the protein concentration of the peptide solution includes: The first and second peptide solutions were extracted from the third giant salamander peptide solution. The first peptide solution was subjected to ultrafiltration and centrifugation to obtain the sample residue. The BCA method and Bradford method samples were extracted from the sample residue. The protein concentration of the sample determined by the BCA method was obtained by BCA method determination, and the protein concentration of the sample determined by the Bradford method was obtained by Bradford method determination. The concentration of the peptide solution was obtained based on the Bradford method and the BCA method. The pre-set exogenous standard protein was added to the second peptide solution sample to obtain the internal standard peptide solution sample. The internal standard peptide solution sample was subjected to ultrafiltration and centrifugation to obtain the internal standard sample residue. The internal standard concentration was obtained based on the internal standard sample residue. The exogenous internal standard concentration of the exogenous standard protein was obtained. The recovery rate of the internal standard was calculated based on the internal standard concentration and the exogenous internal standard concentration. If the recovery rate of the internal standard is not within the preset range, the protein concentration of the peptide solution is calculated based on the recovery rate of the internal standard and the concentration of the peptide solution. If the recovery rate of the internal standard is within the range of the internal standard recovery rate, then the concentration of the peptide solution measured shall be taken as the protein concentration of the peptide solution.

[0042] Importantly, the first peptide solution sample refers to a portion extracted from the third giant salamander peptide solution, used for subsequent ultrafiltration centrifugation and other operations to determine the protein concentration in the peptide solution. The ultrafiltration centrifugation of this invention can efficiently remove small molecules such as free amino acids, nucleotides, and salt ions that may be present in the sample. These substances are common interfering components in the third giant salamander peptide solution and can affect the accuracy of the BCA method (affected by reducing substances) and the Bradford method (affected by acidic substances). Collecting the residual solution for subsequent determination yields a protein concentration result that more accurately reflects the peptide content. The second peptide solution sample refers to another portion extracted from the third giant salamander peptide solution, used to add exogenous standard proteins to prepare an internal standard peptide solution sample, used to correct and verify the accuracy of the determination results. Ultrafiltration centrifugation is a separation technique that uses an ultrafiltration membrane with a specific pore size to separate large protein molecules from small molecules in the first peptide solution sample under centrifugal force. Large protein molecules are retained above the ultrafiltration membrane, forming a sample residue, while small molecules are removed through the ultrafiltration membrane.

[0043] It should be explained that the methods for BCA method determination of samples and Bradford method determination of samples are both existing technologies and will not be described in detail here. BCA method protein concentration refers to the protein concentration value in the BCA method-determined sample obtained by the BCA method. Bradford method protein concentration refers to the protein concentration value in the Bradford method-determined sample obtained by the Bradford method. In the step of obtaining the peptide solution determination concentration based on the Bradford method protein concentration and the BCA method protein concentration, the peptide solution determination concentration = (Bradford method protein concentration + BCA method protein concentration) / 2. This invention, by comparing the concentration results obtained by the two methods, can preliminarily evaluate the amino acid composition characteristics of peptides in the third giant salamander peptide solution. Exogenous standard protein refers to a standard protein of known concentration, used to be added to the second peptide solution sample as an internal standard to correct and verify the accuracy of the determination results. For example, the exogenous standard protein is chicken oocyte albumin.

[0044] Further, the internal standard peptide solution sample refers to the sample obtained by adding exogenous standard protein to the second peptide solution sample. The method for ultrafiltration centrifugation of the internal standard peptide solution sample is the same as the method for ultrafiltration centrifugation of the first peptide solution sample, and will not be repeated here. The internal standard sample residue refers to the sample residue obtained after ultrafiltration centrifugation of the internal standard peptide solution sample. The method for obtaining the internal standard determination concentration based on the internal standard sample residue is the same as the method for obtaining the peptide solution determination concentration based on the sample residue, and will not be repeated here. The internal standard determination concentration refers to the average concentration of the internal standard sample residue determined by the Bradford method and the Bradford method, respectively. In the step of calculating the internal standard recovery rate based on the internal standard determination concentration and the exogenous internal standard concentration, the internal standard recovery rate = internal standard determination concentration / exogenous internal standard concentration. The internal standard recovery rate range refers to a pre-set range used to determine whether the internal standard recovery rate is within this range. If the internal standard recovery rate is within this range, the measurement result is relatively reliable; if it is not within this range, the peptide solution determination concentration needs to be corrected. This invention quantifies the presence of matrix inhibition or enhancement effects throughout the detection process by calculating the recovery rate of the internal standard, thereby obtaining the most accurate concentration value. In the step of calculating the peptide protein concentration based on the internal standard recovery rate and the peptide solution concentration, the peptide protein concentration = peptide solution concentration / internal standard recovery rate.

[0045] Specifically, obtaining the exogenous internal standard concentration of the exogenous standard protein includes: Determine the protein addition volume and concentration of the exogenous standard protein, and calculate the total amount of exogenous standard protein based on the protein addition volume and concentration. The total amount of exogenous standard protein is the product of the protein addition volume and the exogenous standard protein concentration. Obtain the total sample volume of the second peptide solution sample, and calculate the exogenous internal standard concentration based on the total sample volume and the total amount of exogenous standard protein. The exogenous internal standard concentration is the product of the total sample volume and the total amount of exogenous standard protein.

[0046] It should be explained that the protein addition volume refers to the volume of exogenous standard protein solution added to the second peptide solution sample. The exogenous standard protein concentration refers to the concentration of the exogenous standard protein solution added to the second peptide solution sample. The total sample volume refers to the total volume of the second peptide solution sample after adding the exogenous standard protein solution.

[0047] Specifically, the fine separation of the target peptide to obtain the target active peptide distillate includes: The target column set and SMB system are determined, and each target column in the target column set is connected in series with the SMB system to obtain the integrated SMB system; The target chromatographic column set in the integrated SMB system is divided into functional regions, which include multiple functional regions and correspond one-to-one with the target chromatographic column. Representative chromatographic columns were extracted from the integrated SMB system and connected to a pre-constructed high-performance liquid chromatography system to obtain a pre-configured high-performance liquid chromatography system. The target peptide is introduced into a prepared high-performance liquid chromatography system to obtain the target peptide to be separated. The target peptide to be separated is subjected to a phase B gradient elution operation to obtain the retention time of the target active peptide. Using the retention time of the target active peptide as the switching time, parameters are set for each functional region in the functional region set to obtain a parameterized functional region set, which includes multiple parameterized functional regions. The target peptide to be separated is separated by using a parameterized functional region set and switching time to obtain an initial target peptide. The initial target peptide is then detected in real time to obtain activity signals and ultraviolet signals. If the activity signal and ultraviolet signal meet the preset normal signal parameter conditions, the target active peptide distillate is collected using a pre-constructed fraction collector.

[0048] It needs to be explained that a target column set refers to a group of chromatographic columns used to separate target peptides. An SMB system is a simulated moving bed chromatography system that simulates the movement of the stationary phase by periodically switching valves to achieve continuous separation. An integrated SMB system refers to a system obtained by connecting each target column in the target column set in series with a pre-constructed SMB system. A functional region set refers to dividing the target column set in the integrated SMB system into multiple functional regions, each corresponding to one target column. A target column refers to a single chromatographic column in the target column set used to separate the target peptide. A representative column is a column extracted from the target column set and used to connect to a high-performance liquid chromatography (HPLC) system. A prepared HPLC system refers to a system obtained by connecting a representative column to an HPLC system. HPLC is a chromatographic technique used for the separation, identification, and quantitative analysis of peptides. The target peptide to be separated refers to the target peptide introduced into the prepared HPLC system. The B-phase gradient elution operation for the target peptide to be separated refers to linearly increasing the proportion of the organic phase (B phase, such as acetonitrile or methanol) over time during the separation process in a high-performance liquid chromatography (HPLC) system, while correspondingly decreasing the proportion of the aqueous phase (A phase), so that the target peptides are eluted sequentially according to their differences in hydrophobicity. For example, a gradient program can be set to start from initial conditions (0% B phase and 100% A phase), increasing the concentration of B phase at a rate of 5% per minute until it reaches 100%, while simultaneously decreasing the proportion of A phase by 5%, until the target peptide is eluted.

[0049] Importantly, the retention time of the target active peptide refers to the time elapsed in a high-performance liquid chromatography (HPLC) system from injection to detection of the active peptide eluting from the representative column. Switching time refers to the time interval between valve switching cycles in the SMB system, used to simulate stationary phase movement. Parameter setting refers to the operation of setting the flow rate and gradient for each functional region, ensuring effective separation of the target peptide in each region by setting different flow rates and B-phase concentration gradients for each functional region. Parameterized functional region refers to the functional region after parameter setting. Initially separated target peptide refers to the initially separated peptide obtained after parameterized functional region processing in the SMB system. Real-time detection of the initially separated target peptide refers to real-time detection of the initially separated target peptide using an ultraviolet detector and an online microfluidic activity detector. Activity signal refers to the bioactivity signal of the target peptide measured by the online microfluidic activity detector. Ultraviolet signal refers to the absorbance signal of the target peptide measured by the ultraviolet detector. Target active peptide fraction refers to the fraction containing both high concentration and high bioactivity of the target peptide obtained through real-time detection and collection.

[0050] Furthermore, the condition that the activity signal and ultraviolet (UV) signal meet the preset normal signal parameter conditions means that the activity signal is greater than the preset normal activity signal threshold and the UV signal is greater than the preset normal UV signal threshold. The normal activity signal threshold and the normal UV signal threshold are both preset minimum values ​​for the activity signal, used to determine whether the bioactivity of the target peptide reaches an acceptable standard. When the activity signal measured by the online microfluidic activity detector is greater than or equal to this threshold, it indicates that the current peptide has sufficient bioactivity and can be collected. The normal UV signal threshold is a preset minimum value used to determine whether the concentration of the target peptide reaches an acceptable standard. When the UV signal measured by the UV detector is greater than or equal to this threshold, it indicates that the current peptide concentration is high enough and can be collected.

[0051] S5. The target active peptide is freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptide are determined based on the high-purity giant salamander peptide powder.

[0052] It should be explained that the freeze-drying of the target active peptide refers to freeze-drying the target active peptide using a freeze dryer. Many bioactive substances, such as peptides and proteins, are prone to denaturation or inactivation during high-temperature or prolonged drying processes. Freeze-drying, performed at low temperatures, effectively avoids the damage to bioactive substances caused by high temperatures, ensuring that the target active peptide retains its bioactivity during the drying process and does not lose its function due to high temperatures. Therefore, freeze-drying is performed. The determination of peptide molecular weight and amino acid sequence based on high-purity giant salamander peptide powder refers to using high-performance liquid chromatography-mass spectrometry (HPLC-MS) to analyze the peptide molecular weight and amino acid sequence in the high-purity giant salamander peptide powder. The HPLC-MS technique of this invention is existing technology and will not be described in detail here.

[0053] S6. The activity of high-purity giant salamander peptide powder was verified to obtain activity verification parameters. The peptide molecular weight, amino acid sequence and activity verification parameters were stored in a pre-constructed storage database to obtain an optimized storage database.

[0054] It should be explained that the storage database refers to a database used to store various data generated during the extraction and separation of giant salamander peptides, including peptide molecular weight, amino acid sequence, and activity verification parameters. In the production and purification of peptide products, molecular weight determination is used for quality control, ensuring product consistency and purity. Accurate molecular weight information helps in understanding the structure and function of peptides, providing fundamental data for subsequent bioactivity studies. During the production and purification of peptide products, amino acid sequence determination ensures product consistency and purity, preventing the introduction of impurities or incorrect peptides.

[0055] Specifically, the activity verification of high-purity giant salamander peptide powder, to obtain activity verification parameters, includes: Obtain the stop solution, enzyme solution, substrate solution and buffer solution. Prepare the reaction solution based on high-purity giant salamander peptide powder. Mix the enzyme solution, substrate solution, buffer solution and reaction solution to obtain the initial mixed reaction solution. The initial mixed reaction solution is subjected to a water bath reaction to obtain a mixed reaction solution. The termination solution is then added to the mixed reaction solution to obtain a termination reaction solution. The solution used to terminate the reaction was centrifuged to obtain the supernatant. The absorbance of the supernatant was measured to obtain the absorbance of the sample, as well as the absorbance of the substrate control and the blank control. The ACE inhibition rate was calculated based on the absorbance of the sample, the absorbance of the substrate control, and the absorbance of the blank control, and the ACE inhibition rate was used as the activity verification parameter.

[0056] It should be explained that a stop solution is a solution used to stop a reaction, capable of rapidly terminating an enzymatic reaction. For example, a stop solution is 10% trichloroacetic acid. An enzyme solution is a solution containing a specific enzyme, used to catalyze a specific chemical reaction. A substrate solution is a solution containing a specific substrate, used to provide the substrate for the enzymatic reaction. For example, a substrate solution is Hippuryl-Histidyl-Leucine. A buffer solution is a solution that maintains a stable pH value, used to provide a suitable reaction environment. For example, a phosphate buffer. The preparation of the reaction solution based on high-purity giant salamander peptide powder refers to dissolving high-purity giant salamander peptide powder in a suitable solvent to prepare a reaction solution of a predetermined standard concentration. The standard concentration refers to a solution concentration set in advance. For example, 0.1 mg / mL. The initial mixed reaction solution is a solution obtained by mixing the enzyme solution, substrate solution, buffer solution, and reaction solution, used to initiate the reaction. A water bath reaction involves placing the reaction solution in a water bath and maintaining a constant temperature for the reaction. For example, the initial mixed reaction solution is placed in a 37°C water bath and reacted for 30 minutes. The mixed reaction solution is the solution after the water bath reaction. The termination reaction solution is the solution where the reaction stops after the addition of the termination solution. The supernatant is the liquid at the top of the centrifuge tube after centrifugation of the termination reaction solution. The absorbance measurement of the supernatant refers to the operation of measuring the absorbance of the supernatant at a specific wavelength (e.g., 228 nm) using a UV spectrophotometer. The sample absorbance is the absorbance value of the supernatant at a specific wavelength, reflecting the concentration of the reaction product. The substrate control absorbance is the absorbance of the substrate solution without the enzyme at a specific wavelength (e.g., 228 nm). The blank control absorbance is the absorbance value of the reaction system without the sample at a specific wavelength (e.g., 228 nm). In the step of calculating the ACE inhibition rate based on the sample absorbance, substrate control absorbance, and blank control absorbance, the ACE inhibition rate = (1 - (sample absorbance - substrate control absorbance) / (blank control absorbance - substrate control absorbance)). 100%. This invention, by calculating the ACE inhibition rate, can reveal the bioactivity of peptides, particularly their potential applications in the field of cardiovascular health.

[0057] S7. Extraction, separation and purification of giant salamander peptides were completed based on the optimized storage database and giant salamander oil.

[0058] It should be explained that an optimized storage database refers to a storage database that already stores peptide molecular weights, amino acid sequences, and activity verification parameters. Storing these data in the storage database facilitates data management and analysis.

[0059] To address the problems described in the background art, this invention identifies healthy giant salamanders and their processing equipment. The processing equipment includes a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. This invention selects healthy giant salamanders as raw materials because the proteins and other components in healthy salamanders are in a normal state, reducing abnormalities caused by disease or unhealthy factors, thus laying the foundation for extracting high-quality giant salamander peptides. The invention receives giant salamander peptide extraction instructions, obtains processing by-products based on healthy giant salamanders and the peptide extraction instructions, and performs pretreatment operations on the by-products to obtain the by-products to be processed. The pretreatment operations include washing and crushing. This invention... The processing byproducts of giant salamanders are pulverized into smaller particles, increasing the specific surface area of ​​the material, which is beneficial for subsequent degreasing, deodorization, and enzymatic hydrolysis, thus improving reaction efficiency. The byproducts are then degreased and deodorized to obtain degreased byproducts and giant salamander oil. Giant salamander protein powder is prepared from the degreased byproducts, and then a crude giant salamander peptide solution is prepared from the protein powder. This invention removes oil from the giant salamander processing byproducts, which on the one hand reduces the impact of oil on subsequent enzymatic hydrolysis and separation purification processes, improving the purity of the giant salamander peptides; on the other hand, the obtained giant salamander oil can be used as a raw material for other products, realizing further resource utilization. The invention also receives giant salamander peptide separation and purification instructions, and prepares the crude giant salamander peptide solution and giant salamander peptide separation... This invention obtains target peptides through purification commands, performs fine separation on these peptides to obtain target active peptide fractions. The fine separation further removes impurities and other unwanted peptides, yielding high-purity target active peptide fractions, thus improving the quality and bioactivity of the giant salamander peptides. The target active peptide fractions are then freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptides are confirmed based on the high-purity giant salamander peptide powder. This invention uses freeze-drying of the target active peptide fraction, which removes moisture at low temperatures, maximizing the preservation of the giant salamander peptide's bioactivity and structural integrity. Simultaneously, the dried powder is easy to store and use, extending the product's shelf life. This invention validates the activity of high-purity giant salamander peptide powder, obtaining activity validation parameters. A pre-constructed storage database is used to store the peptide molecular weight, amino acid sequence, and activity validation parameters, resulting in an optimized storage database. This invention enables the evaluation of the actual biological activity of giant salamander peptides, such as antioxidant and immunomodulatory functions, providing a scientific basis for product quality control and application. Based on the optimized storage database and giant salamander oil, the extraction, separation, and purification of giant salamander peptides are completed. This invention establishes a giant salamander peptide information database, facilitating subsequent research, development, and quality traceability, and also providing data support for the standardized production of giant salamander peptides. Therefore, this invention can improve the extraction and purification efficiency of giant salamander peptides.

[0060] like Figure 2 The diagram shown is a functional block diagram of a system for the extraction, separation and purification of giant salamander peptides provided in an embodiment of the present invention.

[0061] The giant salamander peptide extraction, separation, and purification system 100 of the present invention can be installed in an electronic device. Depending on the functions implemented, the giant salamander peptide extraction, separation, and purification system 100 may include a raw material preparation module 101, a crude peptide solution preparation module 102, a peptide separation and purification module 103, and a peptide data management module 104. The module described in this invention can also be referred to as a unit, which refers to a series of computer program segments that can be executed by the processor of an electronic device and can perform a fixed function, and which are stored in the memory of the electronic device. The raw material preparation module 101 is used to determine healthy giant salamanders and giant salamander processing equipment. The giant salamander processing equipment includes a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. It receives giant salamander peptide extraction instructions, obtains giant salamander processing by-products according to the instructions, and performs pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed. The pretreatment operations include washing and crushing. The crude peptide liquid preparation module 102 is used to degrease and deodorize the by-products to be processed, to obtain degreased by-products and giant salamander oil, to prepare giant salamander protein powder based on the degreased by-products, and to prepare giant salamander crude peptide liquid based on the giant salamander protein powder. The peptide separation and purification module 103 is used to receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide liquid and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment to obtain the target active peptide distillate, freeze-dry the target active peptide distillate to obtain high-purity giant salamander peptide powder, and confirm the peptide molecular weight and amino acid sequence according to the high-purity giant salamander peptide powder. The peptide data management module 104 is used to verify the activity of high-purity giant salamander peptide powder, obtain activity verification parameters, and use a pre-constructed storage database to store the peptide molecular weight, amino acid sequence and activity verification parameters to obtain an optimized storage database. Based on the optimized storage database and giant salamander oil, the extraction, separation and purification of giant salamander peptides are completed.

[0062] In detail, the modules in the giant salamander peptide extraction, separation and purification system 100 described in this embodiment of the invention employ the same methods as described above. Figure 1 The extraction and purification methods for giant salamander peptides described herein are the same as those used in this study, and can produce the same technical effects, so they will not be repeated here.

[0063] like Figure 3 The diagram shown is a schematic diagram of an electronic device for implementing a method for extracting, separating, and purifying giant salamander peptides according to an embodiment of the present invention.

[0064] The electronic device 1 may include a processor 10, a memory 11 and a bus 12, and may also include a computer program stored in the memory 11 and executable on the processor 10, such as a method program for the extraction, separation and purification of giant salamander peptides.

[0065] The memory 11 includes at least one type of readable storage medium, including flash memory, portable hard drive, multimedia card, card-type memory (e.g., SD or DX memory), magnetic memory, disk, optical disk, etc. In some embodiments, the memory 11 can be an internal storage unit of the electronic device 1, such as the portable hard drive of the electronic device 1. In other embodiments, the memory 11 can be an external storage device of the electronic device 1, such as a plug-in portable hard drive, smart media card (SMC), secure digital card (SD), flash card, etc., equipped on the electronic device 1. Furthermore, the memory 11 includes both internal storage units and external storage devices of the electronic device 1. The memory 11 can be used not only to store application software and various types of data installed on the electronic device 1, such as the code of the extraction and purification method program for giant salamander peptide, but also to temporarily store data that has been output or will be output.

[0066] In some embodiments, the processor 10 may be composed of integrated circuits, such as a single packaged integrated circuit or multiple integrated circuits with the same or different functions, including combinations of one or more central processing units (CPUs), microprocessors, digital processing chips, graphics processors, and various control chips. The processor 10 is the control unit of the electronic device, connecting various components of the entire electronic device through various interfaces and lines. It executes programs or modules stored in the memory 11 (e.g., a method for extracting, separating, and purifying giant salamander peptides) and calls data stored in the memory 11 to perform various functions of the electronic device 1 and process data.

[0067] The bus 12 can be a peripheral component interconnect (PCI) bus or an extended industry standard architecture (EISA) bus, etc. The bus 12 can be divided into an address bus, a data bus, a control bus, etc. The bus 12 is configured to realize the connection and communication between the memory 11 and at least one processor 10, etc.

[0068] Figure 3 Only electronic devices with components are shown; it will be understood by those skilled in the art that... Figure 3The structure shown does not constitute a limitation on the electronic device 1, and may include fewer or more components than shown, or combine certain components, or have different component arrangements.

[0069] For example, although not shown, the electronic device 1 may also include a power supply (such as a battery) to power the various components. Preferably, the power supply can be logically connected to the at least one processor 10 through a power management device, thereby enabling functions such as charging management, discharging management, and power consumption management. The power supply may also include one or more DC or AC power supplies, recharging devices, power fault detection circuits, power converters or inverters, power status indicators, and other arbitrary components. The electronic device 1 may also include various sensors, Bluetooth modules, Wi-Fi modules, etc., which will not be described in detail here.

[0070] Furthermore, the electronic device 1 may also include a network interface. Optionally, the network interface may include a wired interface and / or a wireless interface (such as a Wi-Fi interface, a Bluetooth interface, etc.), which is typically used to establish communication connections between the electronic device 1 and other electronic devices.

[0071] Optionally, the electronic device 1 may further include a user interface, which may be a display, an input unit (such as a keyboard), and optionally, a standard wired interface or a wireless interface. Optionally, in some embodiments, the display may be an LED display, a liquid crystal display, a touch-sensitive liquid crystal display, or an OLED (Organic Light-Emitting Diode) touchscreen, etc. The display may also be appropriately referred to as a screen or display unit, used to display information processed in the electronic device 1 and to display a visual user interface.

[0072] The extraction, separation, and purification method program for giant salamander peptides stored in the memory 11 of the electronic device 1 is a combination of multiple instructions. When run in the processor 10, it can achieve the following: The healthy giant salamander and the giant salamander processing equipment were identified. The giant salamander processing equipment includes: a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. Receive a giant salamander peptide extraction instruction, obtain giant salamander processing by-products according to healthy giant salamanders and the giant salamander peptide extraction instruction, perform pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed, wherein the pretreatment operations include: washing and crushing; The processing by-products were degreased and deodorized to obtain degreased by-products and giant salamander oil. Giant salamander protein powder was prepared based on the degreased by-products, and giant salamander crude peptide liquid was prepared based on the giant salamander protein powder. Receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment, and obtain the target active peptide distillate; The target active peptide was freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptide were determined based on the high-purity giant salamander peptide powder. The activity of high-purity giant salamander peptide powder was verified to obtain activity verification parameters. The peptide molecular weight, amino acid sequence and activity verification parameters were stored in a pre-constructed storage database to obtain an optimized storage database. The extraction, separation, and purification of giant salamander peptides were completed based on an optimized storage database and giant salamander oil.

[0073] Specifically, the processor 10's implementation method for the above instructions can be found in [reference needed]. Figures 1 to 3 The descriptions of the relevant steps in the corresponding embodiments are not repeated here.

[0074] Furthermore, if the modules / units integrated in the electronic device 1 are implemented as software functional units and sold or used as independent products, they can be stored in a computer-readable storage medium. The computer-readable storage medium can be volatile or non-volatile. For example, the computer-readable medium may include: any entity or device capable of carrying the computer program code, a recording medium, a USB flash drive, a portable hard drive, a magnetic disk, an optical disk, a computer memory, or a read-only memory (ROM).

[0075] The present invention also provides a computer-readable storage medium storing a computer program, which, when executed by a processor of an electronic device, can perform the following: The healthy giant salamander and the giant salamander processing equipment were identified. The giant salamander processing equipment includes: a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. Receive a giant salamander peptide extraction instruction, obtain giant salamander processing by-products according to healthy giant salamanders and the giant salamander peptide extraction instruction, perform pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed, wherein the pretreatment operations include: washing and crushing; The processing by-products were degreased and deodorized to obtain degreased by-products and giant salamander oil. Giant salamander protein powder was prepared based on the degreased by-products, and giant salamander crude peptide liquid was prepared based on the giant salamander protein powder. Receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment, and obtain the target active peptide distillate; The target active peptide was freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptide were determined based on the high-purity giant salamander peptide powder. The activity of high-purity giant salamander peptide powder was verified to obtain activity verification parameters. The peptide molecular weight, amino acid sequence and activity verification parameters were stored in a pre-constructed storage database to obtain an optimized storage database. The extraction, separation, and purification of giant salamander peptides were completed based on an optimized storage database and giant salamander oil.

[0076] In the embodiments provided by this invention, it should be understood that the disclosed devices, systems, and methods can be implemented in other ways. For example, the system embodiments described above are merely illustrative, and actual implementations may have other classification methods.

[0077] The modules described as separate components may or may not be physically separate. The components shown as modules may or may not be physical units; that is, they may be located in one place or distributed across multiple network units. Some or all of the modules can be selected to achieve the purpose of this embodiment according to actual needs.

[0078] Furthermore, the functional modules in the various embodiments of the present invention can be integrated into one processing unit, or each unit can exist physically separately, or two or more units can be integrated into one unit. The integrated unit can be implemented in hardware or in the form of hardware plus software functional modules.

[0079] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the present invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the present invention.

[0080] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can 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

1. A method for the extraction, separation, and purification of giant salamander peptides, characterized in that, The method includes: The healthy giant salamander and the giant salamander processing equipment were identified. The giant salamander processing equipment includes: a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. Receive a giant salamander peptide extraction instruction, obtain giant salamander processing by-products according to healthy giant salamanders and the giant salamander peptide extraction instruction, perform pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed, wherein the pretreatment operations include: washing and crushing; The processing by-products were degreased and deodorized to obtain degreased by-products and giant salamander oil. Giant salamander protein powder was prepared based on the degreased by-products, and giant salamander crude peptide liquid was prepared based on the giant salamander protein powder. Receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment, and obtain the target active peptide distillate; The target active peptide was freeze-dried to obtain high-purity giant salamander peptide powder. The molecular weight and amino acid sequence of the peptide were determined based on the high-purity giant salamander peptide powder. The activity of high-purity giant salamander peptide powder was verified to obtain activity verification parameters. The peptide molecular weight, amino acid sequence and activity verification parameters were stored in a pre-constructed storage database to obtain an optimized storage database. The extraction, separation, and purification of giant salamander peptides were completed based on an optimized storage database and giant salamander oil.

2. The method for extraction, separation, and purification of giant salamander peptides as described in claim 1, characterized in that, The degreasing and deodorizing of the by-products to be processed yields degreased by-products and giant salamander oil, including: The by-products to be processed are spread in a material tray to obtain a loading tray. The loading tray is then placed in the extraction vessel to obtain the by-products to be extracted. Static extraction is performed on the extraction byproduct to obtain the first extraction byproduct. The first extraction byproduct is then dynamically circulated using a preset carbon dioxide gas to obtain a carbon dioxide fluid rich in oil. A separation vessel is used to separate a fluid rich in grease and carbon dioxide to obtain separated carbon dioxide gas and grease. The separated carbon dioxide gas is recovered using a carbon dioxide storage tank to obtain recovered carbon dioxide. The recovered carbon dioxide is then used as carbon dioxide gas and returned to the step of dynamically recycling the first extraction byproduct using the preset carbon dioxide gas until a pre-constructed extraction completion command is received. The defatting byproducts were obtained from the extraction vessel, and the oil was collected to obtain giant salamander oil.

3. The method for extraction, separation, and purification of giant salamander peptides as described in claim 2, characterized in that, The preparation of giant salamander protein powder based on defatting byproducts includes: The defatting byproduct was quick-frozen to obtain a pre-frozen defatting byproduct. The pre-frozen defatting byproduct was freeze-dried using a pre-constructed freeze-drying chamber to obtain an initial freeze-dried product. The temperature of the initial freeze-dried product was monitored to obtain the initial cold trap temperature. When the initial cold trap temperature reaches the preset standard cold trap temperature, the pre-built vacuum pump is started, and the freeze-drying chamber is evacuated using the started vacuum pump to obtain a vacuum freeze-drying chamber. The initial freeze-dried product in the vacuum freeze-drying chamber is heated using a preset initial heating temperature to obtain a preliminary dried product. The state data of the preliminary dried product is obtained, wherein the state data is either a porous state or a non-porous state. If the state data is non-porous, the preliminary dried product is used as the initial freeze-dried product, and the process returns to the step of heating the initial freeze-dried product in the vacuum freeze-drying chamber using a preset initial heating temperature until the state data is porous. If the state data is porous, the pre-dried product is continuously heated using a preset secondary drying time to obtain the secondary dried product. Giant salamander protein powder was obtained using secondary drying products and a pre-constructed inert gas.

4. The method for extraction, separation, and purification of giant salamander peptides as described in claim 3, characterized in that, The preparation of giant salamander crude peptide solution from giant salamander protein powder includes: To prepare a low cosolvent, the low cosolvent and giant salamander protein powder are mixed according to a preset mixing ratio to obtain an initial mixture. The initial mixture is then ultrasonically treated using a pre-constructed ultrasonic cell disruptor to obtain an ultrasonic mixture. The ultrasonic cell disruptor has preset ultrasonic parameters. The pH value of the ultrasonic mixture is measured to obtain the pH value of the mixture. The pH value of the mixture is adjusted to obtain the pH value of the standard mixture. The standard pH value mixture is then identified based on the pH value of the standard mixture. The standard pH mixture was subjected to temperature adjustment to obtain a standard temperature mixture, and appropriate amounts of alkaline protease and flavor protease were obtained. The standard temperature mixture was subjected to initial enzymatic hydrolysis using a preset first enzymatic hydrolysis time and an appropriate amount of alkaline protease to obtain the first enzymatic hydrolysis mixture. The first enzymatically hydrolyzed mixture was enzymatically hydrolyzed using a preset second enzymatic hydrolysis time and an appropriate amount of flavor protease to obtain an enzymatically hydrolyzed mixture; Enzyme inactivation was performed on the enzymatically hydrolyzed mixture using preset enzyme inactivation parameters to obtain an enzymatically hydrolyzed mixture. The enzymatically hydrolyzed mixture was then transferred to a pre-constructed centrifuge tube to obtain a mixture to be centrifuged. The mixture to be centrifuged was then centrifuged to obtain crude peptide solution of giant salamander.

5. The method for extraction, separation, and purification of giant salamander peptides as described in claim 4, characterized in that, The process of obtaining the target peptide fragment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instructions includes: The pre-constructed tangential flow ultrafiltration system was set up according to the giant salamander peptide separation and purification instructions to obtain a configured tangential flow ultrafiltration system, which includes: a first-stage ultrafiltration membrane, a second-stage ultrafiltration membrane and a third-stage ultrafiltration membrane; The crude peptide solution of giant salamander was subjected to macromolecular ultrafiltration using a first-stage ultrafiltration membrane to obtain the first giant salamander peptide solution. The first giant salamander peptide solution was subjected to medium-molecular ultrafiltration using a second-stage ultrafiltration membrane to obtain the second giant salamander peptide solution. The second giant salamander peptide solution was ultrafiltered using a third-stage ultrafiltration membrane to obtain a third giant salamander peptide solution. The protein concentration of the third giant salamander peptide solution was then determined, and it was determined whether the protein concentration of the peptide solution was equal to the preset standard protein concentration of the peptide solution. If the protein concentration of the peptide solution is not equal to the protein concentration of the standard peptide solution, the third giant salamander peptide solution is adjusted to obtain an updated giant salamander peptide solution. The updated giant salamander peptide solution is used as the third giant salamander peptide solution. The process of performing protein detection on the third giant salamander peptide solution is repeated until the protein concentration of the peptide solution is equal to the protein concentration of the standard peptide solution. If the protein concentration of the peptide solution is equal to that of the standard peptide solution, then the third giant salamander peptide solution is taken as the target peptide segment.

6. The method for extraction, separation, and purification of giant salamander peptides as described in claim 5, characterized in that, The process of detecting proteins in the third giant salamander peptide solution to obtain the protein concentration of the peptide solution includes: The first and second peptide solutions were extracted from the third giant salamander peptide solution. The first peptide solution was subjected to ultrafiltration and centrifugation to obtain the sample residue. The BCA method and Bradford method samples were extracted from the sample residue. The protein concentration of the sample determined by the BCA method was obtained by BCA method determination, and the protein concentration of the sample determined by the Bradford method was obtained by Bradford method determination. The concentration of the peptide solution was obtained based on the Bradford method and the BCA method. The pre-set exogenous standard protein was added to the second peptide solution sample to obtain the internal standard peptide solution sample. The internal standard peptide solution sample was subjected to ultrafiltration and centrifugation to obtain the internal standard sample residue. The internal standard concentration was obtained based on the internal standard sample residue. The exogenous internal standard concentration of the exogenous standard protein was obtained. The recovery rate of the internal standard was calculated based on the internal standard concentration and the exogenous internal standard concentration. If the recovery rate of the internal standard is not within the preset range, the protein concentration of the peptide solution is calculated based on the recovery rate of the internal standard and the concentration of the peptide solution. If the recovery rate of the internal standard is within the range of the internal standard recovery rate, then the concentration of the peptide solution measured shall be taken as the protein concentration of the peptide solution.

7. The method for extraction, separation, and purification of giant salamander peptides as described in claim 6, characterized in that, The process of obtaining the exogenous internal standard concentration of the exogenous standard protein includes: Determine the protein addition volume and concentration of the exogenous standard protein, and calculate the total amount of exogenous standard protein based on the protein addition volume and concentration. The total amount of exogenous standard protein is the product of the protein addition volume and the exogenous standard protein concentration. Obtain the total sample volume of the second peptide solution sample, and calculate the exogenous internal standard concentration based on the total sample volume and the total amount of exogenous standard protein. The exogenous internal standard concentration is the product of the total sample volume and the total amount of exogenous standard protein.

8. The method for extraction, separation, and purification of giant salamander peptides as described in claim 7, characterized in that, The process of finely separating the target peptide to obtain the target active peptide distillate includes: The target column set and SMB system are determined, and each target column in the target column set is connected in series with the SMB system to obtain the integrated SMB system; The target chromatographic column set in the integrated SMB system is divided into functional regions, which include multiple functional regions and correspond one-to-one with the target chromatographic column. Representative chromatographic columns were extracted from the integrated SMB system and connected to a pre-constructed high-performance liquid chromatography system to obtain a pre-configured high-performance liquid chromatography system. The target peptide is introduced into a prepared high-performance liquid chromatography system to obtain the target peptide to be separated. The target peptide to be separated is subjected to a phase B gradient elution operation to obtain the retention time of the target active peptide. Using the retention time of the target active peptide as the switching time, parameters are set for each functional region in the functional region set to obtain a parameterized functional region set, which includes multiple parameterized functional regions. The target peptide to be separated is separated by using a parameterized functional region set and switching time to obtain an initial target peptide. The initial target peptide is then detected in real time to obtain activity signals and ultraviolet signals. If the activity signal and ultraviolet signal meet the preset normal signal parameter conditions, the target active peptide distillate is collected using a pre-constructed fraction collector.

9. The method for extraction, separation, and purification of giant salamander peptides as described in claim 8, characterized in that, The activity verification of high-purity giant salamander peptide powder was performed to obtain activity verification parameters, including: Obtain the stop solution, enzyme solution, substrate solution and buffer solution. Prepare the reaction solution based on high-purity giant salamander peptide powder. Mix the enzyme solution, substrate solution, buffer solution and reaction solution to obtain the initial mixed reaction solution. The initial mixed reaction solution is subjected to a water bath reaction to obtain a mixed reaction solution. The termination solution is then added to the mixed reaction solution to obtain a termination reaction solution. The solution used to terminate the reaction was centrifuged to obtain the supernatant. The absorbance of the supernatant was measured to obtain the absorbance of the sample, as well as the absorbance of the substrate control and the blank control. The ACE inhibition rate was calculated based on the absorbance of the sample, the absorbance of the substrate control, and the absorbance of the blank control, and the ACE inhibition rate was used as the activity verification parameter.

10. A system for the extraction, separation, and purification of giant salamander peptides, characterized in that, The system includes: The raw material preparation module is used to determine healthy giant salamanders and giant salamander processing equipment. The giant salamander processing equipment includes a material tray, an extraction vessel, a separation vessel, and a carbon dioxide storage tank. It receives giant salamander peptide extraction instructions, obtains giant salamander processing by-products according to the healthy giant salamanders and giant salamander peptide extraction instructions, and performs pretreatment operations on the giant salamander processing by-products to obtain by-products to be processed. The pretreatment operations include washing and crushing. The crude peptide solution preparation module is used to degrease and deodorize the by-products to be processed, to obtain degreased by-products and giant salamander oil, to prepare giant salamander protein powder based on the degreased by-products, and to prepare giant salamander crude peptide solution based on the giant salamander protein powder. The peptide separation and purification module is used to receive the giant salamander peptide separation and purification instruction, obtain the target peptide segment according to the giant salamander crude peptide solution and the giant salamander peptide separation and purification instruction, perform fine separation on the target peptide segment to obtain the target active peptide distillate, freeze-dry the target active peptide distillate to obtain high-purity giant salamander peptide powder, and confirm the peptide molecular weight and amino acid sequence based on the high-purity giant salamander peptide powder. The peptide data management module is used to verify the activity of high-purity giant salamander peptide powder, obtain activity verification parameters, and use a pre-constructed storage database to store peptide molecular weight, amino acid sequence and activity verification parameters to obtain an optimized storage database. Based on the optimized storage database and giant salamander oil, the extraction, separation and purification of giant salamander peptides are completed.

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