CN110759970A - Antioxidant polypeptide derived from fermented milk, application thereof and additive - Google Patents
Antioxidant polypeptide derived from fermented milk, application thereof and additive Download PDFInfo
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 39
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 35
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 33
- 230000003078 antioxidant effect Effects 0.000 title claims abstract description 28
- 239000003963 antioxidant agent Substances 0.000 title claims abstract description 23
- 239000000654 additive Substances 0.000 title claims abstract description 13
- 230000000996 additive effect Effects 0.000 title claims description 9
- 235000015140 cultured milk Nutrition 0.000 title claims description 6
- 235000013305 food Nutrition 0.000 claims abstract description 17
- 239000000126 substance Substances 0.000 claims abstract description 9
- 241001465754 Metazoa Species 0.000 claims abstract description 8
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract 2
- 235000020247 cow milk Nutrition 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 2
- 239000004480 active ingredient Substances 0.000 claims 1
- 239000012752 auxiliary agent Substances 0.000 claims 1
- 150000001875 compounds Chemical class 0.000 abstract description 5
- 235000006708 antioxidants Nutrition 0.000 description 17
- 239000000047 product Substances 0.000 description 11
- 238000000034 method Methods 0.000 description 10
- OHDRQQURAXLVGJ-HLVWOLMTSA-N azane;(2e)-3-ethyl-2-[(e)-(3-ethyl-6-sulfo-1,3-benzothiazol-2-ylidene)hydrazinylidene]-1,3-benzothiazole-6-sulfonic acid Chemical compound [NH4+].[NH4+].S/1C2=CC(S([O-])(=O)=O)=CC=C2N(CC)C\1=N/N=C1/SC2=CC(S([O-])(=O)=O)=CC=C2N1CC OHDRQQURAXLVGJ-HLVWOLMTSA-N 0.000 description 9
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- 101800000068 Antioxidant peptide Proteins 0.000 description 3
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 3
- 230000002292 Radical scavenging effect Effects 0.000 description 3
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- -1 vitamin E Chemical class 0.000 description 3
- GVJHHUAWPYXKBD-UHFFFAOYSA-N (±)-α-Tocopherol Chemical compound OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
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- 150000002632 lipids Chemical class 0.000 description 2
- 125000003588 lysine group Chemical group [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 2
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 235000020183 skimmed milk Nutrition 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 238000004885 tandem mass spectrometry Methods 0.000 description 2
- 238000000108 ultra-filtration Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 108050001786 Alpha-s2 casein Proteins 0.000 description 1
- 208000024827 Alzheimer disease Diseases 0.000 description 1
- 201000001320 Atherosclerosis Diseases 0.000 description 1
- NLZUEZXRPGMBCV-UHFFFAOYSA-N Butylhydroxytoluene Chemical compound CC1=CC(C(C)(C)C)=C(O)C(C(C)(C)C)=C1 NLZUEZXRPGMBCV-UHFFFAOYSA-N 0.000 description 1
- MYMOFIZGZYHOMD-UHFFFAOYSA-N Dioxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- MKRDNSWGJWTBKZ-GVXVVHGQSA-N Gln-Val-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)N)N MKRDNSWGJWTBKZ-GVXVVHGQSA-N 0.000 description 1
- SSFWXSNOKDZNHY-QXEWZRGKSA-N Gly-Pro-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)CN SSFWXSNOKDZNHY-QXEWZRGKSA-N 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 229910002567 K2S2O8 Inorganic materials 0.000 description 1
- WGNOPSQMIQERPK-UHFFFAOYSA-N Leu-Asn-Pro Natural products CC(C)CC(N)C(=O)NC(CC(=O)N)C(=O)N1CCCC1C(=O)O WGNOPSQMIQERPK-UHFFFAOYSA-N 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- BNUKRHFCHHLIGR-JYJNAYRXSA-N Pro-Trp-Asp Chemical compound C1C[C@H](NC1)C(=O)N[C@@H](CC2=CNC3=CC=CC=C32)C(=O)N[C@@H](CC(=O)O)C(=O)O BNUKRHFCHHLIGR-JYJNAYRXSA-N 0.000 description 1
- RWRDLPDLKQPQOW-UHFFFAOYSA-N Pyrrolidine Chemical group C1CCNC1 RWRDLPDLKQPQOW-UHFFFAOYSA-N 0.000 description 1
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 1
- 241001122767 Theaceae Species 0.000 description 1
- GLEVLJDDWXEYCO-UHFFFAOYSA-N Trolox Chemical compound O1C(C)(C(O)=O)CCC2=C1C(C)=C(C)C(O)=C2C GLEVLJDDWXEYCO-UHFFFAOYSA-N 0.000 description 1
- FEXILLGKGGTLRI-NHCYSSNCSA-N Val-Leu-Asn Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](C(C)C)N FEXILLGKGGTLRI-NHCYSSNCSA-N 0.000 description 1
- 229930003427 Vitamin E Natural products 0.000 description 1
- DGEZNRSVGBDHLK-UHFFFAOYSA-N [1,10]phenanthroline Chemical compound C1=CN=C2C3=NC=CC=C3C=CC2=C1 DGEZNRSVGBDHLK-UHFFFAOYSA-N 0.000 description 1
- XBJFCYDKBDVADW-UHFFFAOYSA-N acetonitrile;formic acid Chemical compound CC#N.OC=O XBJFCYDKBDVADW-UHFFFAOYSA-N 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 235000019282 butylated hydroxyanisole Nutrition 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
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- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000013480 data collection Methods 0.000 description 1
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- 238000001976 enzyme digestion Methods 0.000 description 1
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- 238000013467 fragmentation Methods 0.000 description 1
- 238000006062 fragmentation reaction Methods 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 235000013376 functional food Nutrition 0.000 description 1
- WIGCFUFOHFEKBI-UHFFFAOYSA-N gamma-tocopherol Natural products CC(C)CCCC(C)CCCC(C)CCCC1CCC2C(C)C(O)C(C)C(C)C2O1 WIGCFUFOHFEKBI-UHFFFAOYSA-N 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
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- 230000004054 inflammatory process Effects 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 108010078274 isoleucylvaline Proteins 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
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- 239000012528 membrane Substances 0.000 description 1
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- 210000000056 organ Anatomy 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 150000008442 polyphenolic compounds Chemical class 0.000 description 1
- 235000013824 polyphenols Nutrition 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 125000000719 pyrrolidinyl group Chemical group 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
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- 230000002000 scavenging effect Effects 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
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- 235000019165 vitamin E Nutrition 0.000 description 1
- 229940046009 vitamin E Drugs 0.000 description 1
- 239000011709 vitamin E Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/142—Amino acids; Derivatives thereof
- A23K20/147—Polymeric derivatives, e.g. peptides or proteins
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Polymers & Plastics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Animal Husbandry (AREA)
- Nutrition Science (AREA)
- Zoology (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Mycology (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention relates to a polypeptide compound Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg with antioxidant capacity, and the amino acid sequence of the polypeptide compound is Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg. The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg has antioxidant activity, not only has active function, but also can be used as additives of food, health care products, daily chemical products, animal food and the like, and has wide application range.
Description
Technical Field
The invention relates to polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg and application of the polypeptide as an antioxidant component in preparing additives of foods, health products, daily chemical products, animal foods and the like.
Background
The antioxidant is widely applied to the fields of food, daily chemical products and the like. On one hand, the antioxidant can prevent the above products from discoloring or deteriorating due to oxidation; on the other hand, some oxidants even play active functional roles in related products, such as playing roles in resisting oxidation, scavenging free radicals and the like in vivo. Chemically synthesized antioxidants such as Butylated Hydroxyanisole (BHA), 2, 6-di-tert-butyl-p-cresol (BHT) and the like are widely used in the food industry, but their use is increasingly limited due to the potential risk to human health. In recent years, natural and safe antioxidants have been actively sought, and various antioxidant compounds, such as vitamin E, tea polyphenol, food-derived antioxidant peptide, etc., are found in many natural animal and plant materials. In particular, antioxidant peptides are receiving attention because of their high safety, strong antioxidant properties, and absorbability.
Antioxidant Peptides (Antioxidant Peptides) have effects of inhibiting lipid oxidation and protecting human tissues and organs from free radical. Normally, the generation and elimination of free radicals in the body are in dynamic equilibrium, but when the free radicals are excessively generated in the body, an oxidation reaction occurs to attack membrane lipids, proteins, DNA and other biological macromolecules, so that the damage of tissue cells changes the cell structure and function, and further influences the health of the body, such as causing diseases of cancer, Alzheimer disease, inflammation, atherosclerosis and the like (reference 1: Appl Biochem Biotechnol 2015,176(7): 1815-1833). In the body, for example, hydroxyl radical (. OH), peroxy radical (. OOR), and superoxide radical (. O) are generally produced2) And peroxynitroso group (ONOO), which attack intracellular DNA and proteins when excessive, damage cells, accelerate aging, quench free radicals by supplying electrons or protons, and are known as antioxidants that can effectively promote human health by enhancing the body's antioxidant ability when taken in a diet (document 2: journal of Functional Foods 2011,3, 229-.
Disclosure of Invention
The invention aims to provide an application and a rapid screening method of polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg in an antioxidant component; the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg has antioxidant activity, can be used as additives of foods, health products, daily chemical products, animal foods and the like, and has good application prospect.
In order to achieve the aim, the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg is taken as an effective antioxidant component.
It has the sequence table of SEQ ID NO: 1, amino acid sequence; the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg is antioxidant component, and can be used as additive for food, health product, daily chemical product, animal food, etc.
The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg with antioxidant activity is derived from Alpha-S2-casein protein of a bovine (bovine) protein database, contains 13 amino acid residues, has an amino acid sequence of Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg, has a single-chain linear structure, is white powder, is easy to dissolve in water and has the molecular weight of 1521.8 Da; has better ABTS free radical and hydroxyl free radical scavenging activity and IC5013.44 ± 0.56 μ M (n ═ 3, Mean ± SD) and 77.6 ± 24.7 μ M (n ═ 3, Mean ± SD), respectively.
The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg has the characteristics required by antioxidant peptide: contains amino acid residues of benzene ring, pyrrolidine ring, and can be used as hydrogen donor required by free radical; containing hydrophobic amino acid residues, especially the N-terminal hydrophobic amino acid residues; the second amino acid residue at the C-terminal position readily forms hydrogen bonds. The second amino acid residue at the C terminal of the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg is lysine residue, and the hydroxyl group of the lysine residue is easy to form hydrogen bond with water molecules; contains 2 pyrrolidine rings and 1 benzene ring and can be used as a hydrogen donor required by free radicals; contains 5 hydrophobic amino acids. Therefore, the Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg conforms to the structure-activity relationship of antioxidation and has antioxidation function.
Compared with the prior art, the invention has the following beneficial effects:
the invention obtains and determines the structure of the active compound from the fermented cow milk for the first time, and the compound has better antioxidant activity, thereby having good potential and application prospect.
Detailed Description
Example 1
Preparation of polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg
The method combining LC-MS/MS and Shotgun proteomics technology is adopted. Cow milk is used as a raw material, and a peptide segment with antioxidation is screened by fermentation, centrifugation, ultrafiltration purification and LC-MS/MS analysis in combination with the structure-activity relationship characteristics.
The specific method comprises the following steps:
pure cultured Bulgaricus (Lb. bulgaricus, CICC6047) is inoculated into 200mL of 120g/L skimmed milk powder culture medium according to the proportion of 1% (V/V), cultured for 12h at 42 ℃, then inoculated into 200mL of 120g/L skimmed milk powder culture medium according to the proportion of 1% (V/V), cultured for 12h at 42 ℃ and used as a fermented milk sample. Centrifuging at 25000g and 4 deg.C for 15min, ultrafiltering the supernatant with ultrafiltration tube (with cut-off pore diameter of 10kDa) at 12000g and 20 deg.C, freeze drying the ultrafiltrate, and storing at-20 deg.C.
Peptide sequence library search: redissolving the freeze-dried fermented milk sample with 0.1% (V/V) formic acid aqueous solution to 0.4. mu.g/. mu.L, centrifuging for 3min at 15000g, performing mass spectrometry by LTQ Orbitrap XL, and taking the supernatant for LC-MS/MS analysis. The amount of sample per mass spectrometry was 8. mu.g. Chromatographic analysis conditions: the flow rate after the split was adjusted to 200nL/min for mobile phase A being 0.1% (V/V) formic acid aqueous solution and mobile phase B being 0.1% (V/V) formic acid acetonitrile solution. The linear gradient elution procedure was as follows: 0% B (0min) -5% B (5min) -35% B (95min) -80% B (110min) -0% B (120 min). The autoinjection system consisted of a 4cm homemade capillary trap column (200 μm i.d.) filled with C18AQ packing and a 12cm homemade capillary analytical column (75 μm i.d.).
Mass spectrometry conditions: the mass spectrum fragmentation mode is Collision Induced Dissociation (CID), the temperature of the capillary is 200 ℃, the electrospray voltage is 1.8kV, the normalized collision energy is 35.0 percent, and the primary scanning range is 400-2000 Da. The MS and the MS/MS are subjected to spectrum acquisition by using a data-dependent mode, and the scanning mode is full scanning. MS/MS scan was performed for the 6 ion peaks with the highest abundance in the full scan, with dynamic exclusion set as: the number of repetitions was 2, the repeat tolerance time was 30s, and the dynamic exclusion time was 90 s. The system control and data collection was performed using Xcalibur (v2.1, Thermo corporation) software.
RAW files collected by Xcalibur were converted to MGF format using Thermo protein discover Daemon (v1.4) and then retrieved in the Bovine (Bovine, protein number 17890) protein database (http:// www.uniprot.org /) using the software Mascot (version 2.3.0, Matrix Science, London, UK). The variable modification was set to oxidation of methionine (+15.9949 Da); no enzyme digestion, maximum cut-missing number and fixed modification are set, the mass tolerance deviation of parent ions is 20ppm, and fragment ions are 0.8 Da. Score >20 was set when peptide fragments were derived, with significant difference P <0.01, while controlling false positive rate (FDR) < 1%.
The peptide sequences obtained are shown in the attached table. And (4) screening by combining the structure-activity relationship to obtain the polypeptide with the sequence of Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg.
Example 2
ABTS free radical scavenging activity detection of polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg
Principle of
ABTS generates stable blue-green cationic free radical ABTS after being oxidized by active oxygen+Anti-oxidative component with ABTS+The reaction is carried out to fade the reaction system, so that the change of absorbance can be detected under the maximum absorption wavelength of 734nm to examine the strength of the antioxidant activity of the sample. Comparing the sample with a control standard system containing trolox or Vc, and calculating the antioxidant capacity of the measured substance.
Method of producing a composite material
The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg used in the experiment is synthesized by Shanghai Qiang Yao biological technology limited company, and the purity of the polypeptide is>95 percent. The formulation contained 8mM ABTS (M. 548.68), 3mM K2S2O8The solution of (4) was left standing at room temperature in the dark, and after 16 hours, the OD value of the solution at 734nm was diluted to 1.5 with 0.1M PBS (pH 7.4, containing 0.15mM NaCl) to obtain an ABTS solution. And (3) diluting Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg into solutions with different concentrations by PBS, immediately preparing a reaction solution according to the table, carrying out shading reaction at room temperature for 30min, and calculating the ABTS free radical clearance rate of the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg. Clearance ═ 1- (ODsample-ODblank)/ODcontrol) × 100%. Vc is used as a positive control drug to replace polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg by the same method.
TABLE-grouping and sample addition for ABTS free radical scavenging experiments
Note: "-" is the corresponding volume of PBS
The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg is subjected to ABTS free radical scavenging activity detection by the method according to different concentrations, and the result is shown in the table III, under the same condition, the clearance rate of 28.4 mu M Vc is 46.0 +/-3.1 percent, and the clearance rate of 56.8 mu M Vc is 100.0 +/-0.2 percent. The half clearance concentration IC of the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg can be calculated from the table II50＝13.4±0.6μM(Mean±SD，n＝3)。
ABTS clearance of epididate concentrations for polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg
Example 3
Detection of hydroxyl radical scavenging activity of polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg
Principle of
According to the Fenton reaction principle, Fe2+Can form a red complex with phenanthroline, has a maximum absorption peak at 536nm (510 nm), and has a high absorption peak when phenanthroline-Fe2+Oxidized to phenanthrene-Fe by hydroxyl radical3+When an antioxidant is added, the absorption intensity at 536nm becomes small, and the hydroxyl radical is inhibited. Therefore, the removal rate of OH by the sample can be calculated according to the change of the absorbance values before and after the sample is added.
Method of producing a composite material
The reaction system was as shown in table three, and after adding the solutions of the respective volumes in order, incubation was performed at 37 ℃ for 1h, and then absorbance was measured at 536nm to calculate the hydroxyl radical clearance, I% ((a) = I ═ c)i-A0)/(A-A0)×100％(AiAbsorbance of the sample for the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg, A0Absorbance for the injured group, and a for the intact group). The clearance rate is determined by the same method by taking Vc as a positive control drug to replace a sample of Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg.
Grouping and sample adding amount of experiment for removing free radicals in epitrihydroxy group
Note: "-" indicates 1.0mL of PBS was substituted for
Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg with different concentrations was prepared, and the hydroxyl radical scavenging activity was determined by the above method, and the results are shown in Table four.
Hydroxyl radical scavenging activity of epi-tetrapolypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg
The polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg has antioxidant activity, can be used as additives of foods, health products, daily chemical products, animal foods and the like, and has wide application.
Attached table:
peptide sequence contained in fermentation liquid attached to table
Note: take EMPFPKYPVEPFTESQSLTLTDVENLHLPLPL as an example: the modification type of the peptide fragment is oxidation (M), which indicates that the peptide fragment has 1 methionine oxidation modification. The modification position of the peptide fragment is 0.01000000000000000000000000000000.0, which indicates that the methionine oxidation modification of the peptide fragment is located at the 2 nd residue.
The amino acids related to the sequences in the attached table I are all abbreviated as amino acids, and the abbreviations, abbreviations and names of the amino acids are shown in the attached table II.
Attached table of two amino acid names, abbreviations and abbreviations
Sequence listing
<110> institute of chemistry and physics, large connection of Chinese academy of sciences
<120> an antioxidant polypeptide derived from fermented milk, and its application and additive
<160>1
<170>SIPOSequenceListing 1.0
<210>1
<211>13
<212>PRT
<213> Artificial Sequence (Artificial Sequence)
<400>1
Gly Pro Ile Val Leu Asn Pro Trp Asp Gln Val Lys Arg
1 5 10
Claims (4)
1. An antioxidant polypeptide derived from fermented milk, which is characterized in that: the polypeptide is Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg and has a sequence table SEQ ID NO: 1; the amino acid sequence of the polypeptide is Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg.
2. The use of the antioxidant polypeptide derived from fermented cow's milk according to claim 1 in the preparation of an additive having antioxidant function for foods, health products, daily chemical products, animal foods, etc.
3. An additive of food, health care products, daily chemical products or animal food with an antioxidant function is characterized in that: the additive comprises the polypeptide Gly-Pro-Ile-Val-Leu-Asn-Pro-Trp-Asp-Gln-Val-Lys-Arg as an active ingredient.
4. The additive of claim 3, wherein: the additive also contains a carrier and/or an auxiliary agent.
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Cited By (1)
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| WO2024041604A1 (en) * | 2022-08-24 | 2024-02-29 | 四川大学华西第二医院 | Use of yogurt-derived polypeptide in preparation of drug for delaying telomere shortening and anti-aging drug |
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| CN105254740A (en) * | 2015-10-16 | 2016-01-20 | 上海交通大学 | Bioactive polypeptide NQFYQKF as well as preparation and application thereof |
| CN107573413A (en) * | 2014-12-19 | 2018-01-12 | 浙江辉肽生命健康科技有限公司 | Cow's milk αs2The preparation and application of casein derived biologically active peptide |
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Patent Citations (3)
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| CN107573413A (en) * | 2014-12-19 | 2018-01-12 | 浙江辉肽生命健康科技有限公司 | Cow's milk αs2The preparation and application of casein derived biologically active peptide |
| CN107602688A (en) * | 2014-12-19 | 2018-01-19 | 浙江辉肽生命健康科技有限公司 | Cow's milk αs2The preparation and application of casein derived biologically active peptide |
| CN105254740A (en) * | 2015-10-16 | 2016-01-20 | 上海交通大学 | Bioactive polypeptide NQFYQKF as well as preparation and application thereof |
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| Y. JIN等: "Peptide profiling and the bioactivity character of yogurt in the simulated gastrointestinal digestion", 《JOURNAL OF PROTEOMICS》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2024041604A1 (en) * | 2022-08-24 | 2024-02-29 | 四川大学华西第二医院 | Use of yogurt-derived polypeptide in preparation of drug for delaying telomere shortening and anti-aging drug |
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