Antarctic krill active peptide and its application

Abstract

The present invention relates to the field of functional polypeptide preparation, and aims to provide an Antarctic krill active peptide and its application. The present invention provides an Antarctic krill active peptide, which is refined to obtain an Antarctic krill peptide liquid with a high protein concentration of 89.3 mg / g on a dry basis, and finally dried to prepare Antarctic krill peptide powder. Five highly active peptide segments contained therein are particularly provided, which are non-enzymatic peptide segments directly separated from Antarctic krill shelling processing liquid, have natural activity, and can be used in the preparation of uric acid-lowering products.

Description

Technical Field

[0001] The invention relates to the field of functional polypeptide preparation, and aims to provide an Antarctic krill active peptide and application thereof. Background Art

[0002] Uric acid is the final product of purine metabolism in the human body. The level of uric acid in human blood will continue to increase through nutritional metabolism as people consume seafood and other foods rich in purines. Theoretically, the maximum concentration of uric acid in normal human blood is about 416μmol / L. According to statistics, hyperuricemia in my country is increasing rapidly at a rate of 9.7% per year. In recent years, there has been great progress in the research of marine bioactive peptides. There are studies on the composite enzymatic hydrolysis of marine fish meat and seafood by-products. The marine fish enzymatic hydrolyzate prepared by selecting the best enzymatic hydrolysis process has a protective effect on the test experiment of the hyperuricemia model mice caused by potassium oxonate.

[0003] Antarctic krill biological resources are extremely abundant, but they are still underdeveloped. At the same time, krill is rich in protein and has a reasonable amino acid composition, making it the world's largest potential protein resource library. It has been found that krill peptides have a variety of biological activities, such as improving osteoarthritis, osteoporosis, diabetes, lowering blood pressure and antioxidant activity. At this stage, research on Antarctic krill peptides in improving hyperuricemia symptoms has also received great attention. Patent No. CN117568430A introduces a method of preparing Antarctic krill peptides that can be used as uric acid-lowering drugs or health products using enzymatic hydrolysis, but it requires the addition of exogenous enzymes to decompose the macromolecules. At present, there is a lack of methods for directly obtaining biologically active Antarctic krill peptides from processing wastewater, as well as related research on non-enzymatic peptide segments. Summary of the invention

[0004] The technical problem to be solved by the present invention is that at this stage, research on Antarctic krill peptides in improving the symptoms of hyperuricemia has received great attention, but the research direction is concentrated on enzymatic peptides that require the addition of exogenous enzymes to decompose large molecules. There is still a lack of methods for directly obtaining biologically active Antarctic krill peptides from processing wastewater, as well as research on related non-enzymatic peptide segments.

[0005] In order to solve the above problems, the present invention provides an Antarctic krill active peptide, which is refined to obtain an Antarctic krill peptide liquid with a high protein concentration of 89.3 mg / g on a dry basis, and finally dried to prepare Antarctic krill peptide powder. Five highly active peptide segments contained therein are particularly provided, which are non-enzymatic peptide segments directly separated from Antarctic krill shelling processing liquid, have natural activity, and can be used in the preparation of uric acid-lowering products.

[0006] To achieve the above object, the present invention is implemented by the following technical means: an Antarctic krill active peptide, including a polypeptide with an amino acid sequence as shown in SEQ ID NO.1-5.

[0007] SEQ ID NO.1:

[0008] LDGGF.

[0009] SEQ ID NO.2:

[0010] TVPVF.

[0011] SEQ ID NO.3:

[0012] IDDLF.

[0013] SEQ ID NO.4:

[0014] LVLTP.

[0015] SEQ ID NO.5:

[0016] NPVLE.

[0017] A method for preparing the above-mentioned Antarctic krill active peptide is artificial synthesis.

[0018] A method for preparing Antarctic krill active peptides comprises the following steps:

[0019] (1) The Antarctic krill shelling liquid was heated to 100°C and then heated for 5-10 minutes. Flocculent aggregates were formed in the liquid and roughly separated using a 100-mesh filter cloth. After cooling, the liquid was centrifuged at 10,000 r / min and filtered using a 500-mesh filter cloth to obtain a clear peptide liquid on the upper layer. The final solid content of the Antarctic krill liquid was about 2g / 100g, of which nearly 60% was protein and 16% was lipid. Due to the presence of phospholipids, the lipids in the liquid were tightly bound to the proteins and difficult to separate, requiring further treatment. The heating and steaming process formed visible suspended lumps. The heating process destroyed the spatial structure of the macromolecular proteins in the aqueous phase, exposing the hydrophobic area to the outside to adsorb fat droplets, forming flocculent precipitation from the appearance, effectively adsorbing lipids, and facilitating further centrifugal degreasing. After centrifugation, a clear supernatant was obtained without obvious particles, meeting the membrane technology treatment conditions. Compared with traditional degreasing methods, including alkali refining and n-hexane extraction, this heating-centrifugation method does not involve any reagents, nor does it require the addition of flocculants, etc. There is no need to consider the removal of other reagents during subsequent processing, thereby minimizing the impact of the processing on nutrients.

[0020] (2) The upper clear peptide solution is subjected to membrane concentration and then freeze-dried to obtain Antarctic krill active peptides; the membrane concentration conditions are: 150Da nanofiltration, temperature 25°C, operating pressure 4.0MPa, and concentration multiple of 10-12 times. Too high a concentration multiple will lead to a gradual decrease in membrane flux, and the concentration of substances such as protein and chloride in the sample will continue to increase. The membrane flux decreases slowly in the early stage of concentration, and the adsorption gel layer has not yet formed. After a period of concentration, membrane scaling begins to occur, blocking the membrane pores, the mass transfer boundary layer thickens, and the concentration polarization and membrane pollution degree increase, resulting in a significant decrease in the retention rate of each component and the membrane flux. Therefore, controlling the concentration of the concentrated solution can effectively extend the service life of the membrane.

[0021] Furthermore, the Antarctic krill peptide obtained in step (2) is further screened for its uric acid-lowering active ingredient by gel column chromatography, comprising the following steps: preparing a 5 mg / ml aqueous solution of the recovered Antarctic krill peptide, slowly adding it into the chromatographic column along the wall of a 2×120 cm chromatographic column using a dextran gel Sephadex G-25, and balancing the chromatographic column with pure water twice the volume of the column; after balancing, adding the reserved 4 mL of the krill peptide aqueous solution into the chromatographic column; using pure water to carry the components forward, adjusting the flow rate to 0.6 mL / min, collecting the eluted solution in a group of 5 min, and then detecting at 220 nm, collecting and merging the eluate during the 75-85 min process according to the absorption peak.

[0022] With the continuous development of fishing and breeding technology and the increasing recognition of the consumer market, the seafood processing industry has always maintained a good development trend for a long time. However, the seafood processing process often produces a large amount of processing wastewater with high salt and high organic characteristics. Direct discharge will cause great damage to the environment. However, the cost of wastewater treatment brings great burden to seafood production and processing enterprises. Concentration separation is the main process to solve the separation of complex components of seafood processing wastewater and the recovery of pure water. While recycling water resources, it enriches soluble organic substances in the water phase as raw materials for nutrient recovery, as a new idea to solve processing wastewater. Compared with traditional concentration methods, membrane technology has the advantages of low energy consumption and high efficiency. The principle is to use external energy or pressure difference as the driving force, and use artificial or natural materials to synthesize membranes to separate, purify and concentrate solutes and solvents. There is almost no heat exchange in the process, which can completely retain the flavor and active substances of the concentrate. It is especially suitable for the separation of heat-sensitive substances. According to the needs, microfiltration, ultrafiltration, nanofiltration, electrodialysis and reverse osmosis technologies with different membrane pore sizes can be selected. At present, domestic and foreign research has carried out membrane technology to concentrate and recover organic nutrients in seafood processing fluids. At the same time, nanofiltration membrane technology also has a desalination effect during the concentration process, including nanofiltration and enzymatic hydrolysis to recover antioxidant peptides in anchovy wastewater; ultrafiltration and concentration of flavor nutrients in shrimp processing fluids to prepare seasonings, etc.

[0023] The present invention first optimizes the process parameters of the shelling machine and then performs roller extrusion shelling of Antarctic krill, recovers the processing liquid generated by water circulation during the shelling process (the material-liquid ratio is about 1:4), and the shelling processing liquid is recycled to enrich the solids in the aqueous phase. The peptides in the molecular interval with the highest uric acid-lowering activity obtained after gel column chromatography are subjected to amino acid sequence analysis, and their inhibitory effect on XOD activity is analyzed by molecular docking technology. Further molecular docking results show that the peptide segments with better binding ability with XOD in the Antarctic krill peptides obtained by the above method are the polypeptides shown in SEQ ID NO.1-5.

[0024] The application of the above-mentioned Antarctic krill active peptide in the preparation of uric acid-lowering products. The present invention uses in vitro activity experiments and animal experiments to prove that the krill peptide can improve hyperuricemia by inhibiting the production of uric acid in the liver, promoting the excretion of uric acid in the kidney, and maintaining kidney function. At the same time, the components with the highest uric acid-lowering activity in the recovered krill peptides are screened out by dextran gel column chromatography, showing that the polypeptides shown in SEQ ID NO.1-5 can effectively reduce the ability of XOD to produce uric acid.

[0025] The beneficial effects of the present invention are:

[0026] (1) The present invention provides an environmentally friendly Antarctic krill peptide preparation technology, and can provide high-quality protein nutrients and uric acid-lowering effects. Its advantages are that the raw material source is sufficient and the cost is low, the process flow is simplified, and it does not rely on the addition of organic reagents to separate lipid substances. The produced Antarctic krill peptide not only has uric acid-lowering activity, but also fully meets the requirements of the food production process and can be used as a protein source in food.

[0027] (2) The raw material used in the present invention is the waste liquid generated during the processing of Antarctic krill, which effectively improves the utilization rate of Antarctic krill protein during the production and processing process, reduces the loss of protein nutrients, and reduces energy consumption by at least one third compared with the traditional concentration method mediated by thermal energy. It has the advantages of low cost, high efficiency, and compliance with food safety standards.

[0028] (3) The present invention provides a group of peptide sequences of Antarctic krill active peptides, which are peptides naturally present in the Antarctic krill shelling processing fluid, have a wide source and are safer. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 The effect of membrane separation under different conditions on the protein content of freeze-dried powder.

[0030] Figure 2 It is the effect of Antarctic krill peptide on the basic indicators of mice.

[0031] Figure 3 It is the effect of Antarctic krill peptide on the blood indicators of mice.

[0032] Figure 4 The effect of Antarctic krill peptide on the expression of XOD in mouse liver and hURAT1 mRNA in kidney.

[0033] Figure 5 XOD inhibition rate of each peak of Antarctic krill peptide. DETAILED DESCRIPTION

[0034] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0035] In addition, all materials used in the embodiments of the present invention, unless otherwise specified, were purchased from the market.

[0036] Embodiment 1:

[0037] An Antarctic krill active peptide comprises a polypeptide with an amino acid sequence as shown in SEQ ID NO.1-5.

[0038] SEQ ID NO.1:

[0039] LDGGF.

[0040] SEQ ID NO.2:

[0041] TVPVF.

[0042] SEQ ID NO.3:

[0043] IDDLF.

[0044] SEQ ID NO.4:

[0045] LVLTP.

[0046] SEQ ID NO.5:

[0047] NPVLE.

[0048] Embodiment 2:

[0049] A method for preparing Antarctic krill active peptides comprises the following steps:

[0050] (1) The Antarctic krill shelling liquid is heated to 100° C. and then heated for 5-10 minutes to form flocculent aggregates in the liquid, which are roughly separated by a 100-mesh filter cloth. After cooling, the liquid is centrifuged at 10,000 r / min and filtered by a 500-mesh filter cloth to obtain a clear peptide liquid on the upper layer;

[0051] (2) The upper clear peptide solution is subjected to membrane concentration and then freeze-dried to obtain Antarctic krill active peptides; the membrane concentration conditions are: 150Da nanofiltration, temperature 25°C, operating pressure 4.0MPa, and concentration multiple of 10-12 times.

[0052] Embodiment 3:

[0053] The Antarctic krill peptide uric acid-lowering active ingredient obtained in step (2) was further screened by gel column chromatography, and the rest was the same as in Example 2.

[0054] Verification results:

[0055] The present invention extracts protein peptides with uric acid lowering activity from high-organic Antarctic krill processing fluid. In existing studies, it is reported that small molecule peptides in marine protein peptides have the effect of inhibiting xanthine oxidase (XOD), thereby inhibiting the production of uric acid. The uric acid lowering effect of the recovered Antarctic krill peptides is demonstrated through animal experiments and compared with the activity of ordinary Antarctic krill peptides.

[0056] The method for measuring xanthine oxidase (XOD) inhibitory activity by colorimetry in the following examples is as follows:

[0057] (1) Solution configuration:

[0058] 0.2mol / LpH 7.5 phosphate buffer (PBS): Accurately weigh 60.1676g Na2HPO4·2H2O and dilute to 1L with deionized water.

[0059] Xanthine oxidase: accurately weigh 6.4 mg of xanthine, dissolve it in 1M ml NaOH, add 100 ml PBS, and adjust the pH to 7.5 with 1M HCl.

[0060] Allopurinol: Accurately weigh 1 mg of allopurinol and make up to 100 ml.

[0061] (2) Sample pretreatment:

[0062] 50 μL of the sample to be tested and 50 μL of 0.05U / mL XOD solution were added to a 96-well plate in sequence, shaken, and incubated at 25°C for 5 minutes. Then, 150 μL of 0.48mmol / L xanthine solution was added, shaken, and incubated at 25°C for 25 minutes. The absorbance of the solution was measured at a wavelength of 290nm and calculated according to the following formula:

[0063]

[0064] Wherein: A1 is the absorbance value of XOD, xanthine and sample solution; A2 is the absorbance value of xanthine and sample solution; A3 is the absorbance value of buffer solution, XOD and xanthine solution; A4 is the absorbance value of buffer solution and xanthine solution.

[0065] 1. The purification effect of membrane separation system on Antarctic krill protein peptides:

[0066] (1) 100 L of Antarctic krill shelling liquid (YY) with a solid content of about 2 g / 100 g was directly recovered from a krill shelling machine, placed in a pot and heated to 80° C. The temperature was maintained for 12 min. When flocculation and precipitation occurred, heating was stopped and the temperature was naturally cooled to 40-50° C.

[0067] (2) After coarse separation-centrifugation-filtration separation, a clarified peptide solution that meets the requirements of the membrane separation equipment is obtained, and the solid content is 1.1-1.3 g / 100 g.

[0068] (3) The peptide solution was concentrated using an organic membrane separation device, and the concentration ratios of the liquid during ultrafiltration and nanofiltration were compared. The operating environment settings were: nanofiltration (150Da) temperature 25°C, operating pressure 4.0MPa, concentration ratios 8, 10, and 12 times (N8, N10, and N12); ultrafiltration (3500Da) temperature 25°C, operating pressure 3.0MPa, and concentration ratios 8, 10, and 12 times (C8, C10, and C12). 100 ml of the concentrated liquid was taken and freeze-dried, and the dry protein content (attached) was determined by the Kjeldahl method. Figure 1 As shown in the figure, the protein peptide content of the freeze-dried powder obtained after nanofiltration membrane separation (all higher than 85g / 100g) is significantly higher than that of ultrafiltration. Considering that the separation efficiency decreases significantly when the concentration multiple exceeds 10 times, and the membrane pollution increases, the nanofiltration (150Da) temperature of 25°C, the operating pressure of 4.0MPa, and the concentration multiple of 10 times are selected as the preparation conditions.

[0069] The nanofiltration membrane separation technology in the present invention has the characteristics of high efficiency and low energy consumption. After the process parameters are optimized, the scaling problem in the process of concentrating high-organic processing fluid is effectively reduced, which meets the characteristics of environmentally friendly processing technology.

[0070] 2. Evaluation of the nutritional value of protein peptides during the preparation of Antarctic krill peptides:

[0071] (1) 100 L of Antarctic krill shelling liquid (YY) with a solid content of about 2 g / 100 g was directly recovered from a krill shelling machine, placed in a pot and heated to 80° C. The temperature was maintained for 12 min. When flocculation and precipitation occurred, heating was stopped and the temperature was naturally cooled to 40-50° C.

[0072] (2) Coarse filtration was performed using a 100-mesh filter cloth, and the separated supernatant was centrifuged at 10,000 r / min to separate small particles of visible suspended matter. All suspended precipitates (CD) were collected and freeze-dried to obtain an upper layer of clear peptide solution with a solid content of 0.8-1.0 g / 100 g.

[0073] (3) The peptide solution was concentrated using an organic membrane separation device under the following operating conditions: nanofiltration (150Da) temperature 25°C, operating pressure 4.0MPa, and concentration multiple 10 times. Freeze drying was performed to obtain the nanofiltration concentrate NL. The above samples, as well as common Antarctic krill peptide (LWT), raw shrimp (YX), and shelled shrimp meat (QK) were extracted and analyzed for amino acid composition and essential amino acid nutritional evaluation using an automatic amino acid analyzer.

[0074] AAS, EAAI and F values ​​were calculated according to the standard model proposed by FAO / WHO (1973):

[0075]

[0076] Where:

[0077] n—amino acid number;

[0078] t—amino acid number of the test protein;

[0079] s—The number of amino acids in egg protein.

[0080] The results are shown in Table 1 and Table 2:

[0081] Table 1 Amino acid composition:

[0082]

[0083] Table 2 Amino acid nutritional score:

[0084]

[0085]

[0086] The amino acid nutrition evaluation of peptide powder samples showed that the first limiting amino acid of various products was Ile, but except for NL-M (0.76) and LXT (0.65), the AAS values ​​were all above 0.8, indicating that the peptides recovered from the processing fluid as a source of human protein would not lead to the loss of each essential amino acid. During the entire processing process, the EAA / NEAA ratio of the final product and the intermediate product was above 0.9, and the essential amino acid content of all samples was above 45g / 100g, indicating that krill peptide products can provide sufficient essential amino acids for the human body.

[0087] In summary, the EAAI and F values ​​indicate that the krill peptides separated and recovered by the nanofiltration membrane of the present invention can be used as a protein nutritional supplement.

[0088] 3. Analysis of uric acid lowering activity of Antarctic krill peptide products in animal experiments:

[0089] Fifty 4-week-old Balb / c male mice were weighed and pre-raised for 2 days, and then 8 normal mice were selected and the rest were used as model mice. The model mice were injected with potassium oxonate (2g·kg-1) through the tail vein every day, and the dosage volume was (40mg / ml). Potassium oxonate requires 40-50℃ warm water to fully dissolve. After 7 consecutive days of injection, blood was collected from the tail vein to detect the uric acid content in the mouse serum, and the mice with uric acid content higher than 70umol / L were selected as successful modeling mice.

[0090] There were 8 mice in each of the normal group, model group, positive drug group (febuxostat), common krill peptide group, and sample group (krill peptide recovered by nanofiltration). The mice in the normal group were not injected with tail vein, while the mice in the other four groups were injected with potassium oxonate (2 g kg -1 ), the mice in the control group were fed with yeast extract powder feed to establish the hyperuricemia model mice, and the mice in the normal group were fed with yeast extract powder-free feed.

[0091] (1) Effects of Antarctic krill peptide on basic indicators of mice:

[0092] The weight of the high uric acid model mice before and after administration showed no significant difference between the weights of the mice in each group, indicating that krill peptides and processed liquid recovery peptides had no obvious toxic side effects. The weight gain of the mice in the normal group was significantly higher than that of the other groups. The possible reason is that the mice in the normal group were not treated with potassium oxonate, and the feed composition of the mice in the normal group was healthier. The main organs involved in uric acid metabolism are the liver and kidneys. The significant differences in the organ indexes of the groups indirectly verified that the drug intervention had no obvious toxic side effects on the organs of the mice (see Appendix). Figure 2 ).

[0093] (2) Effects of Antarctic krill peptide on blood parameters of mice:

[0094] Analysis of the uric acid content in the serum at 0d, 15d, and 30d of intervention revealed that the uric acid content decreased significantly throughout the process. On the 30th day of product treatment, the mice were fasted for 12h and blood was collected from the eyeballs after ether anesthesia. In addition to measuring the serum uric acid content, serum creatinine, urea nitrogen content, and xanthine oxidase (XOD) data were also measured. The serum uric acid content of mice in the two krill peptide groups decreased significantly in the first 15d, but could not reach the therapeutic level of positive drugs. During the 15-30d treatment process, the serum content of the two groups of mice decreased significantly compared with the first 15d, and the serum urea nitrogen content and xanthine oxidase (XOD) showed a trend, and the protein expression results of the kidneys also confirmed this result.

[0095] (2) Effect of Antarctic krill peptide on the expression of XOD in mouse liver and hURAT1 mRNA in kidney uric acid transporter:

[0096] The experimental results showed that each drug group was significantly lower than the model group (p < 0.05), and the two krill peptide groups had an inhibitory effect on XOD that was even better than the positive drug. Considering that the XOD content in the serum of the positive drug was also high, it was indirectly confirmed that Febuxostat had a limited inhibitory effect on XOD. Compared with other groups, the expression of mURAT1 mRNA in the kidney of the model group was significantly increased. Compared with the model group, the positive drug group significantly inhibited the expression of mURAT1 mRNA, and the inhibitory effects of the krill peptide group and the sample group were obvious, but slightly higher than the positive drug group. Krill peptides recovered from the processing fluid and ordinary krill peptides promote uric acid excretion by controlling the downregulation of renal uric acid transporter mURAT1 mRNA, which indicates that Antarctic krill peptides can promote uric acid excretion by reducing the expression of mURAT1 mRNA in the kidney, and ultimately play a role in lowering uric acid (Appendix Figure 4 ).

[0097] 4. Sequence determination and uric acid-lowering activity screening of Antarctic krill peptide after purification:

[0098] Use the swollen Sephadex G-25 to slowly add it into the 2×120cm column along the wall of the column, and use twice the column volume of pure water to balance the column. After completion, add the spare 4mL sample solution into the column. Use pure water to carry the components to flow forward, adjust the flow rate to 0.6mL / min, collect the eluted solution every 3min, and then detect it at 220nm. According to the peak time, merge the eluents in the range of 20-30min, 40-50min, 65-75min, 75-85min, and 90-100min, and obtain five groups of samples, P1, P2, P3, P4, and P5. The colorimetric method is used to compare the XOD inhibition ability of each peak, and the component with the highest activity is selected. After testing, it is found that the sample with the highest activity is P4 recovered at 75-85min (attached Figure 5 ).

[0099] Dissolve the sample P4 in pure water to a concentration of 5 mg / ml, centrifuge at 10000 r / min for 20 min, take 0.4 ml of the supernatant, add 0.8 ml of chromatographic grade acetonitrile, place at 4 ° C for 2 h, centrifuge at 10000 r / min for 20 min, take 0.2 ml of the supernatant, blow nitrogen to dryness, add appropriate amount of ultrapure water and freeze-dry into powder; desalt with 0.1% trifluoroacetic acid using C18 small column, freeze-dry with nitrogen; finally re-dissolve with 50ul 0.1% formic acid, centrifuge at 10000 r / min and take 40ul of the upper layer for standby use. Detection was performed using ACQUITY UPLC Peptide CSHC18 (1mm*150mm, 1.7μm, 130A) column. Chromatographic conditions included gradient: 0-10min 5-20% B, 10-25min 20-35% B, 25-45min 35-90% B; flow rate: 0.15ml / min; column temperature: 40°C. Chromatographic conditions: positive ion mode, 100-1800m / z, 350-1800m / z. Search conditions: ProteomeDiscovery.

[0100] In the P4 group, most of the Antarctic krill protein substances are oligopeptides. Peptides with high hydrophobic amino acid content are more likely to bind to xanthine oxidase (XOD) through important domains, and reduce the activity by changing the structure of the enzyme, thereby reducing the production of uric acid. According to the data in Table 1, the hydrophobic amino acid content in the recovered peptide powder accounts for 39.01% of the total amino acid content, which is slightly higher than the 38.14% of the commercially available Antarctic krill peptide. At the same time, studies have shown that there is a certain correlation between basic amino acids and the uric acid-lowering activity of active peptides. The content of basic amino acids in wastewater recovered peptide powder reaches 17.62%, which is higher than the commercially available krill peptide powder. The above 11 peptide sequences were semi-flexibly docked with XOD using the Auto dock molecular docking software. The XOD crystal structure ID comes from the PDB protein database, number: 1FIQ. According to the binding energy, LDGGF, TVPVF, IDDLF, LVLTP, and NPVLE show excellent functions in binding to XOD. The N-terminal of the sequence is mostly hydrophobic amino acids, with Lys being the most, which can effectively inhibit the synthesis of uric acid.

[0101] Table 3P4 amino acid sequence:

[0102]

[0103] In summary, Antarctic krill processing liquid is used as raw material, and oil and insoluble substances are separated by heating-centrifugation pretreatment. After concentration by membrane technology, Antarctic krill peptide powder is obtained by drying. Animal experiments have shown that it has good uric acid lowering activity, and by affecting the effect of xanthine oxidase (XOD) in mice, the serum uric acid level of mice is effectively improved. In the present invention, the Antarctic krill processing liquid, which has a large output but complex composition, is first pre-treated by relatively simple process operations such as heating and centrifugation to meet the requirements of using membrane technology and optimize the membrane concentration process, reduce the loss of the nanofiltration membrane itself during the concentration and separation process, delay membrane scaling, greatly improve the service life of the membrane, and produce krill peptides with reasonable amino acid composition and uric acid lowering activity without relying on enzymes or other additives.

[0104] Finally, it should be noted that although the above embodiments describe the specific implementation of the present invention, they are not intended to limit the present invention; those skilled in the art should understand that these are only examples, and the protection scope of the present invention is defined by the appended claims. All modifications or equivalent replacements should be included in the protection scope of the present invention.

Claims

1. An Antarctic krill active peptide, characterized in that: Its amino acid sequence is shown in any one of SEQ ID NO.1-5.

2. The method for preparing the Antarctic krill active peptide according to claim 1, characterized in that: It is artificially synthesized.

3. A method for preparing the Antarctic krill active peptide according to claim 1, characterized in that The following steps are involved: (1) The Antarctic krill shelling liquid was heated to 100°C and then heated for 5-10 min to form flocculent aggregates in the liquid, which were roughly separated by a 100-mesh filter cloth. After cooling, the liquid was centrifuged at 10,000 r / min and filtered with a 500-mesh filter cloth to obtain a clear peptide liquid on the upper layer; (2) The upper clear peptide solution is subjected to membrane concentration and then freeze-dried to obtain Antarctic krill active peptides; the membrane concentration conditions are: 150Da nanofiltration, temperature 25°C, operating pressure 4.0MPa, and concentration multiple of 10-12 times; (3) The Antarctic krill peptide obtained in step (2) was further screened for its uric acid-lowering active ingredient by gel column chromatography, comprising the following steps: preparing a 5 mg / ml aqueous solution of the recovered Antarctic krill peptide, adding it to a chromatography column along a 2×120 cm column using a dextran gel Sephadex G-25, using pure water to carry the components forward at a flow rate of 0.6 mL / min, collecting the eluted solution once every 3 min, and then detecting at 220 nm, collecting and merging the eluate during 75-85 min according to the absorption peak.

4. Use of the Antarctic krill active peptide according to claim 1 in the preparation of uric acid-lowering drugs.

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