Saury Maillard peptide and its preparation method and application

Abstract

The present invention discloses a saury Maillard peptide with antihyperuricemic activity and its preparation method and application, with the method comprising the following steps: mincing a saury, adding water, heating, agitating, adjusting a pH value to 4.2, separating by centrifugation, and collecting the precipitate; adding water, proteases and amino acids to the precipitate, adjusting a pH value to 7.0, hydrolyzing, adding a reducing sugar to cause a reaction, centrifuging and collecting a supernatant which is saury Maillard peptide; and spray drying the peptide liquid to obtain a dry powder. The method of the present invention realizes the continuous action of enzymolysis and Maillard reaction to prepare the Maillard peptide, not only simplifying the production process, shortening the production cycle, and reducing the production costs, but also significantly enhancing the antihyperuricemic activity of the produced Maillard peptide. Animal experiments in rats showed that the obtained Maillard peptide prepared by the method of the present invention could significantly decrease the level of serum uric acid in rats, and display certain protective effect in their kidney.

Claims

1. A method of preparing a saury Maillard peptide with antihyperuricemic activity, comprising: (1) pretreatment of saury: removing head and internal organs of the saury, cleaning, mincing in a meat grinder, adding water 3 to 5 times the mass of the minced saury meat, heating and stirring at 40 C. to 50 C. for 1 to 2 h, adjusting a pH value of the mixture to 4.2, continuing to stir and heat for 1 to 1.5 h, separating by centrifugation, discarding the supernatant and the upper fat, and collecting a precipitate; and (2) enzymolysis-Maillard continuous reaction: adding water 1 to 1.5 times the mass of the saury precipitate to the saury precipitate, then adding protease and monomer amino acids, adjusting the pH value of the mixture to 7.0, hydrolyzing at 50 C. to 55 C. for 6 to 9 h, adding reducing sugar, heating at 100 C. to 121 C. for 1.0 to 2.0 h, and centrifuging, to obtain a saury Maillard peptide solution; and vacuum-concentrating and spray drying the peptide solution, thus obtaining saury Maillard peptide dry powder; calculated based on the mass of the saury precipitate, the amount of the added protease accounts for 1.5% to 3.0%, the amount of the added monomer amino acids accounts for 0.1% to 0.3%, and the amount of the added reducing sugar accounts for 0.5% to 2.5%; the monomer amino acids described in step (2) are tyrosine, phenylalanine or tryptophan; and the reducing sugar described in step (2) is glucose, xylose or ribose.

2. The method of preparing the saury Maillard peptide having antihyperuricemic activity according to claim 1, wherein the protease described in step (2) is an alkaline protease and a flavourzyme.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 displays the chromatogram of uric acid peaks for Example 2 and contrast products.

DETAILED DESCRIPTION OF THE EMBODIMENTS

(2) The present invention will be further described in detail below with reference examples and FIGURES; however, the embodiments of the present invention are not limited thereto.

(3) In the following examples, a method of determining the uric acid value by the reversed-phase high-performance liquid chromatography is as follows:

(4) (1) Preparation of solutions

(5) 0.2 mol/L (pH 7.5) phosphoric acid buffer (PBS): Accurately weighing 30.0838 g of Na.sub.2HPO.sub.4.12H.sub.2O and 2.4962 g of NaH.sub.2PO.sub.4.2H.sub.2O, and dissolving them with deionized water to a constant volume of 500 ml.

(6) Xanthine solution: Accurately weighing 6.4 mg of xanthine, first dissolving it with 1 ml of 1 M NaOH, then adding 100 ml of PBS, and adjusting the pH value to 7.5 with 1M HCl.

(7) Xanthine oxidase: Taking 120 l of an enzyme solution, and diluting it to 8 ml with PBS.

(8) Uric acid standard curve: Accurately weighing 10 mg of uric acid, adding 10 ml of water, and then diluting it to 0.1-0.9 mg/ml.

(9) Ammonium acetate-glacial acetic acid: Accurately weighing 3.85 g of ammonium acetate to a constant volume of 1000 ml, and then adding 4 ml of glacial acetic acid.

(10) (2) Sample pretreatment:

(11) Diluting the sample to 40 mg/ml, adding successively 50 l of the sample and 50 l of xanthine in a 96-well microplate, with 3 parallels for each sample, adding 150 l of xanthine oxidase after insulation at 37 C. for 10 min, adding 80 l of 1M HCl to stop the reaction after continuing insulation at 37 C. for 20 min, filtering through a 0.25 m aqueous filter till use.

(12) Chromatographic column: Zorbax Eclipse XDB-C18 column (5 m, 4.6250 mm, Agilent); and

(13) Mobile phase conditions: the eluent was 10% methanol+90% ammonium acetate-glacial acetic acid solution, the admission volume was 20 l, the flow rate was 1 ml/min, the detection wavelength was 290 nm, and the run duration was 10 min.

(14) (3) Calculation formula
Xanthine oxidase inhibition rate=(A.sub.0A)/A.sub.0100%

(15) where:

(16) A.sub.0the peak area of the uric acid peak of a sample without addition of peptides analyzed by high-performance liquid chromatography,

(17) Athe peak area of the uric acid peak of a sample with addition of peptides analyzed by high-performance liquid chromatography.

(18) As the index in the following examples, the experimental method of therapeutic effects of the peptide samples on the oteracil potassium-induced hyperuricemia in rats was as follow:

(19) (1) Experimental materials

(20) Animals: 140 SPF grade SD male rats, weight 20020 g, purchased from Laboratory Animal Center of Guangzhou University of Chinese Medicine.

(21) Medicines and reagents: Allophylline tablets (Guangdong Bidi Pharmaceutical Co., Ltd.); oteracil potassium (Shandong Zhongke Taidou Chemical Co., Ltd.); sodium carboxymethyl cellulose (Shanghai Celluloid Plant); and a uric acid kit (Nanjing Jiancheng Bioengineering Research Institute).

(22) Instruments: TGL-16G high-speed refrigerated centrifuge (Shanghai Anting Scientific Instrument Factory); and a multifunctional microplate reader for ELISA (BioTelc USA, Synergy HT).

(23) Animal feeding situations: Animals were bred in SPF grade Laboratory Animal Center of Jinan University, with rat feed and excipients all provided by the same. Housed under conditions of (202) C. and relative humidity of (60-70) % with a 12 h light-dark cycle and standard diet and water. (2) Experimental method

(24) Animal grouping and modeling: Taking a total of 140 healthy male SD rats, and randomly dividing them into a normal control group (20 rats) and a model group (120 rats); the model group rats were gavage with oteracil potassium (daily dose: 2 g/kg) for 7 days, then the rat were anesthetized via intraperitoneally injecting 3% pentobarbital sodium (30 mg/kg) after the last administration, their blood was taken from conjunctival (0.5 ml), then centrifuging at 4 C. and 3000 r/min for 15 min, and taking the upper serum to determine the uric acid content, and the rats in the normal control group were treated with a constant volume of solvent by intragastric administration. The rats with the uric acid content above 110 mol/L were identified to be successful in modeling.

(25) The rats successful in modeling, according to the uric acid content, was randomly dividing into a model control group (constant volume solvent), a tested peptide sample group (200 mg/kg) and an allopurinol group (50 mg/kg) at 14 rats per group, and the rats were gavage with oteracil potassium (daily dose: 2 g/kg) at a dose of 10 ml/kg, with the model rats treated with a constant volume of distilled water. Administering 3% pentobarbital sodium (30 mg/kg) for 50 min at the end of administering the above-mentioned peptide samples for 10 days and 20 days, drawing the conjunctival blood (0.5 ml), and determining the serum uric acid content; after the 30th day of treatment with anti-gout peptides, drawing the celiac artery blood 5 ml after anesthesia with 3% pentobarbital sodium, and determining the content of serum creatinine and urea nitrogen in addition to determining the serum uric acid content.

(26) Determination of serum uric acid: using the tungstic acid method and strictly following the determination method from the kit instructions.

(27) Determination of serum urea nitrogen and creatinine: Determining the serum urea nitrogen content by the diacetyl oxime method; and determining serum creatinine content by the picric acid method, with the specific operation strictly in accordance with the kit instructions.

(28) Determination of activity of serum xanthine oxidase (XOD) and adenosine deaminase (ADA): Taking 100 l of serum for determination of XOD, and taking 20 l of serum for determination of ADA, both in accordance with the kit instructions and operational requirements.

(29) Statistical treatment: all data is presented as meanstandard deviation, t test was performed to determine the significant difference between samples at the 95% confidence interval using SPSS 19.0 software.

EXAMPLE 1

(30) A method of preparing a saury Maillard peptide having antihyperuricemic activity is provided, comprising the following steps:

(31) (1) Pretreatment of saury: removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 3 times the mass of the minced saury meat, heating and stirring at 40 C. for 2 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting the precipitate;

(32) (2) enzymolysis-Maillard continuous reaction: adding water 1 time the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH solution, raising the temperature of the saury to 55 C., adding Alcalase 2.4 L of Novazymes, Flavourzyme 500 MG of Novozymes, and tyrosine as much as 0.5%, 1.0% and 0.10% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 55 C. for 6 h, adding xylose accounting for 0.5% of the mass of the saury precipitate, heating at 110 C. for 1.5 h, then taking the supernatant after centrifuging at 5000 r/min for 15-20 min, thus obtaining the saury Maillard peptide solution.

(33) (3) vacuum-concentrating the saury Maillard peptide solution to the 30% solid or more, and spray drying, thus obtaining the saury Maillard peptide product A.

(34) The therapeutical effects of the saury Maillard peptide product A on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

EXAMPLE 2

(35) A method of preparing a saury Maillard peptide having antihyperuricemic activity is provided, comprising the following steps:

(36) (1) Pretreatment of saury: removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 5 times the mass of the minced saury meat, heating and stirring at 50 C. for 1 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1.5 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting the precipitate;

(37) (2) enzymolysis-Maillard continuous reaction: adding water 1.5 times the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH solution, raising the temperature of the saury to 50 C., adding Alcalase 2.4 L of Novazymes, Flavourzyme 500 MG of Novozymes, and tryptophane as much as 0.8%, 1.2% and 0.20% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 50 C. for 9 h, adding glucose accounting for 1.5% of the mass of the saury precipitate, heating at 121 C. for 1.0 h, then taking the supernatant after centrifuging at 5000 r/min for 15-20 min, thus obtaining the saury Maillard peptide solution.

(38) (3) vacuum-concentrating the saury Maillard peptide solution to the 30% solid or more, and spray drying, thus obtaining the saury Maillard peptide product B.

(39) The therapeutical effects of the saury Maillard peptide product B on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

(40) The uric acid peak chromatogram of the saury Maillard peptide product B was shown in FIG. 1.

EXAMPLE 3

(41) A method of preparing a saury Maillard peptide having antihyperuricemic activity is provided, comprising the following steps:

(42) (1) Pretreatment of saury: removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 4 times the mass of the minced saury meat, heating and stirring at 45 C. for 1.5 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1.2 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting the precipitate;

(43) (2) enzymolysis-Maillard continuous reaction: adding water 1.2 times the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH, increasing the temperature of the saury to 53 C., adding Alcalase 2.4 L of Novazymes, Flavourzyme 500 MG of Novozymes, and phenylalanine as much as 0.5%, 1.5% and 0.3% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 53 C. for 9 h, adding glucose accounting for 2.5% of the mass of the saury precipitate, heating at 100 C. for 2.0 h, then centrifuging at 5000 r/min for 15-20 min, and taking the supernatant, thus obtaining the saury Maillard peptide solution.

(44) (3) vacuum-concentrating the saury Maillard peptide solution to the 30% solid or more, and spray drying, thus obtaining the saury Maillard peptide product C.

(45) The therapeutical effects of the saury Maillard peptide product C on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

Contrast Example 1

(46) A saury peptide, which was prepared as follows:

(47) (1) Removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 5 times the mass of the minced saury meat, heating and stirring at 50 C. for 1 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1.5 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting a precipitate;

(48) (2) adding water 1.5 times the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH solution, raising the temperature of the saury to 50 C., adding Alcalase 2.4 L of Novazymes and Flavourzyme 500 MG of Novozymes as much as 0.8% and 1.2% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 55 C. for 9 h, heating at 95 C. for 15 min to inactivate enzyme, then taking the supernatant after centrifuging at 5000 r/min for 15-20 min, thus obtaining the saury peptide solution.

(49) (3) vacuum-concentrating the saury peptide solution to the 30% solid or more, and spray drying, thus obtaining the contrast product 1.

(50) The therapeutical effects of the contrast product 1 on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

(51) The uric acid peak chromatogram of the contrast product 1 was shown in FIG. 1.

Contrast Example 2

(52) A saury peptide, which was prepared as follows:

(53) (1) Removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 5 times the mass of the minced saury meat, heating and stirring at 50 C. for 1 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1.5 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting a precipitate;

(54) (2) adding water 1.5 times the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH, raising the temperature of the saury to 50 C., adding Alcalase 2.4 L of Novazymes, Flavourzyme 500 MG of Novozymes, and tryptophane as much as 0.8%, 1.2% and 0.15% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 55 C. for 9 h, heating at 95 C. for 15 min to inactivate enzyme, then taking the supernatant after centrifuging at 5000 r/min for 15-20 min, thus obtaining the saury peptide solution;

(55) (3) vacuum-concentrating the saury peptide solution to the 30% solid or more, and spray drying, thus obtaining the contrast product 2.

(56) The therapeutical effects of the contrast product 2 on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

(57) The uric acid peak chromatogram of the contrast product 2 was shown in FIG. 1.

Contrast Example 3

(58) A saury peptide, which was prepared as follows:

(59) (1) Removing the head and internal organs of the saury, cleaning, mincing over a meat grinder, adding water 5 times the mass of the minced saury meat, heating and stirring at 50 C. for 1 h, then adjusting the pH value of the system to 4.2 with HCl (0.5 mol/L), continuing to stir and heat for 1.5 h, separating by centrifugation (5000 r/min, 15-20 min), discarding the supernatant and the upper fat, and collecting a precipitate;

(60) (2) adding water 1.5 times the mass of the saury precipitate to the saury precipitate, adjusting the pH value of the system to 7.0 with 0.5 mol/L NaOH, raising the temperature of the saury to 50 C., adding Alcalase 2.4 L of Novazymes and 500 MG of Flavourzyme Novozymes as much as 0.8% and 1.2% of the mass of the saury precipitate, respectively, insulatingly hydrolyzing at 55 C. for 9 h, adding glucose as much as 0.5% of the mass of the saury precipitate, heating at 121 C. for 1.0 h, then taking the supernatant after centrifuging at 5000 r/min for 15-20 min, thus obtaining the saury Maillard peptide solution;

(61) (3) vacuum-concentrating the saury Maillard peptide solution to the 30% solid or more, and spray drying, thus obtaining the contrast product 3.

(62) The therapeutical effects of the contrast product 3 on the hyperuricemia in rats induced by oteracil potassium were shown in Tables 1, 2 and 3.

(63) The uric acid peak chromatogram of the contrast product 3 was shown in FIG. 1.

(64) TABLE-US-00001 TABLE 1 Effects of tuna extracts acting for different duration on serum uric acid content in rats with hyperuricemia induced by oteracil potassium Before Treatment for Treatment for Treatment for administration 10 days 20 days 30 days Uric acid Uric acid Uric acid Uric acid value value value value Group n (mol/L) n (mol/L) n (mol/L) n (mol/L) Normal 12 81.3 13.2 12 78.2 10.7 10 82.7 7.9 8 76.9 9.8 group Model 15 222.2 21.4 15 220.3 31.2.sup.a 15 216.2 17.0.sup.a 14 218.0 34.5.sup.a group Allopurinol 14 230.4 19.1 14 89.5 22.4.sup.b 14 94.5 13.9.sup.b 14 92.8 20.9.sup.b group Saury 14 220.6 20.2 14 185.8 35.3.sup.c 14 171.5 23.8.sup.b 14 169.2 20.6.sup.b Maillard peptide product A Saury 14 227.5 19.4 14 186.7 35.4.sup.c 14 168.2 14.3.sup.b 11 163.5 33.8.sup.b Maillard peptide product B Saury 14 220.2 18.0 14 183.1 27.1.sup.c 14 173.7 23.8.sup.b 12 169.3 23.8.sup.b Maillard peptide product C Contrast 14 220.4 27.2 14 208.6 24.5.sup.c 13 190.0 21.3.sup.c 12 190.8 19.9.sup.c product 1 Contrast 14 216.0 24.5 14 200.2 30.1.sup.c 12 182.2 23.8.sup.c 10 185.2 16.6.sup.c product 2 Contrast 14 229.1 20.8 14 188.1 31.0.sup.c 12 174.7 18.3.sup.b 10 172.2 19.3.sup.b product 3 Note: Compared with the normal group: .sup.ap < 0.01, .sup.cp > 0.05; compared with the model group: .sup.bp < 0.01, .sup.cp < 0.05, .sup.fp > 0.05.

(65) TABLE-US-00002 TABLE 2 Effects of tuna extracts acting for 30 days on serum creatinine and urea nitrogen content in rats with hyperuricemia induced by oteracil potassium Number of Creatinine Urea nitrogen Group animals (mol/L) (mmol/L) Normal group 8 37.5 6.6 112.6 30.2 Model group 14 58.3 11.4.sup.a 192.7 48.2.sup.a Allopurinol group 14 41.8 13.2.sup.b 159.2 24.6.sup.b Saury Maillard peptide 14 50.4 8.6.sup.b 149.0 30.6.sup.b product A Saury Maillard peptide 11 47.8 10.4.sup.b 141.2 38.7.sup.b product B Saury Maillard peptide 11 49.9 20.5.sup.b 141.5 28.7.sup.b product C Contrast product 1 12 54.5 10.2.sup.c 167.2 26.5.sup.c Contrast product 2 10 55.3 8.1.sup.c 159.1 22.6.sup.b Contrast product 3 10 51.4 12.8.sup.b 156.4 32.2.sup.b Note: Compared with the normal control group: .sup.ap < 0.01; compared with the model group: .sup.bp < 0.01, .sup.cp < 0.05, .sup.fp > 0.05.

(66) TABLE-US-00003 TABLE 3 Effects of example and contrast products acting for 30 days on serum ADA and XOD contents in rats with hyperuricemia induced by oteracil potassium Number of ADA XOD Group animals (U/ml) (U/L) Normal group 8 7.31 5.25 15.48 2.65 Model group 14 17.71 7.03.sup.a 35.02 5.18.sup.a Allopurinol group 14 12.28 7.88.sup.b 8.61 2.46.sup.b Saury Maillard peptide 14 12.08 7.79.sup.b 17.28 6.36.sup.b product A Saury Maillard peptide 11 10.71 6.55.sup.b 15.89 5.93.sup.b product B Saury Maillard peptide 11 11.66 6.31.sup.b 19.63 4.24.sup.b product C Contrast product 1 12 15.38 6.07.sup.c 25.88 5.82.sup.c Contrast product 2 10 15.68 5.33.sup.c 26.63 5.62.sup.c Contrast product 3 10 13.51 7.64.sup.b 23.53 5.22.sup.c Note: Compared with the normal control group: .sup.ap < 0.01; compared with the model group: .sup.bp < 0.01, .sup.cp < 0.05, .sup.fp > 0.05.

(67) During the metabolism of purine in the human body, hypoxanthine and xanthine is generated from ATP or other substances through a series of metabolic reactions, both of which will be oxidized into uric acid by xanthine oxidase (XOD) in the human body, thus the generation of uric acid would be inhibited by inhibiting the activity of xanthine oxidase, results in reduce of uric acid level in the body.

(68) The present invention adopted the method of in vitro high-performance liquid chromatography to determine the inhibition of xanthine oxidase, which was as follows: First, mixing the sample with xanthine oxidase to make them interact; The amount of uric acid produced in the xanthine hydrolysis system would be reduced if the sample could interact with xanthine oxidase and inhibit the activity of the enzyme, Thus the inhibition rate of xanthine oxidase could be calculated by detecting the amount of uric acid produced.

(69) As FIG. 1 shows, the products obtained from examples and contrast examples could inhibit the activity of xanthine oxidase compared with the blank group (PBS), but different products exhibited distinctly different inhibition rates. The xanthine oxidase inhibition rate of the contrast product 1 is 10.40%, which indicated that the protein peptide product with certain antihyperuricemic activity could be obtained by hydrolyzing saury protein through enzymolysis; the xanthine oxidase inhibition rate of the contrast product 2 was increased to 12.34%, which was increased about 18.7%. The main difference between the contrast products 1 and 2 is the addition of monomer amino acids before enzymolysis of the product 2; with progress of the enzymolysis, monomer amino acids can display substrate inhibition on the enzymolytic system, during the enzymolytic process alter the amino acid composition of the final hydrolysate, and thereby improve the antihyperuricemic activity. The xanthine oxidase inhibition rate of the contrast product 3 was significantly improved compared to the products 1 and 2; the contrast product 3 was added reducing sugar to have the Maillard reaction compare to the contrast product 1, indicated that some substances with high antihyperuricemic activity was generated from saury hydrolysates during Maillard reaction. Above knowable, the saury hydrolysate has a certain antihyperuricemic effect, and both the addition of monomer amino acids for synergistic hydrolysis and the Maillard reaction of the hydrolysate can improve its antihyperuricemic activity. Therefore, the xanthine oxidase inhibition rate of the saury Maillard peptide B (Example 2) prepared by the combination of the three methods was as high as 25.59%.

(70) As Table 1 shows, the serum uric acid content was significantly increased (p<0.01) in the model rats treated with oteracil potassium for nearly 40 days, but was significantly decreased (p<0.01) after the rats were treated with allopurinol, which was mainly because allopurinol is a medicine that reduces uric acid production and serum uric acid concentration by inhibiting the xanthine oxidase activity. While the other administration groups (example products and contrast products) all had the significant effect of reducing serum uric acid in rats. Generally, the example products exhibited higher antihyperuricemic activity than the contrast products.

(71) Table 2 shows that the serum creatinine and urea nitrogen content of rats after administration. The results indicated that each example product could significantly reduce the serum creatinine and urea nitrogen content in the rats (p<0.01). Serum creatinine is the product of human muscle metabolism and urea nitrogen is the main end product of human protein metabolism. Under normal circumstances, both of them are excreted through kidneys by glomerular filtration, and their level in plasma indicated renal function. The above results indicated that the saury Maillard peptide prepared by the process of the present invention had an effect in significantly decreasing the serum creatinine level, and had a certain protective effect in the renal function of the hyperuricemia model rats.

(72) Xanthine oxidase (XOD) and adenosine deaminase (ADA) are key enzymes in uric acid metabolism. Wherein XOD widely exists in a variety of animals and human body. Liver has the highest content of XOD, followed by the small intestine, while the content of XOD in the rest of tissues is less than 3% of the content in the liver and small intestine. XOD can directly regulate uric acid levels in the body by subsequently oxidizing Xanthine and hypoxanthine into uric acid. The adenosine is catalyzed by ADA into hypoxanthine nucleotide and is finally oxidized by XOD into uric acid. The increase in activity of XOD and ADA contributes to promotion of the nucleic acid catabolism and the production of uric acid. As Table 3 shows, when the model rats were treated with oteracil potassium for nearly 40 days, the activity of serum xanthine oxidase (XOD) and adenosine deaminase (ADA) in the model rats was significantly increased (p<0.01). The example products (the saury Maillard peptides A, B and C) showed a strong effect on reducing serum ADA and XOD activity in rats; the contrast products, although also having a certain effect on reducing these two enzymes, had an overall effect inferior to that of the example products.

(73) In conclusion, the antihyperuricemic peptide prepared according to the present invention has the effect in reducing the levels of urea nitrogen and creatinine which are high in the serum of rats suffered from hyperuricemia-induced kidney injury, suggesting that it has certain protective effect on renal function. Besides, it can reduce the XOD and ADA enzyme activity in serum, suggesting that it reduces the activity of key enzymes so as to reduce the catabolism of nucleic acids and to decrease the uric acid production. Therefore, the antihyperuricemic peptide prepared by the method of the present invention has a good application prospect.

(74) The above examples are preferred embodiments of the present invention; however, the embodiments of the present invention are not limited by the above examples, and any other alteration, modification, substitution, combination and simplification made without departing from the spiritual essence and principle of the present invention are equivalent replacements and fall within the scope of protection of the present invention.