CN110759971A - Antioxidant polypeptide derived from cornu Cervi Pantotrichum, and its application and additive - Google Patents
Antioxidant polypeptide derived from cornu Cervi Pantotrichum, and its application and additive Download PDFInfo
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- CN110759971A CN110759971A CN201810843190.1A CN201810843190A CN110759971A CN 110759971 A CN110759971 A CN 110759971A CN 201810843190 A CN201810843190 A CN 201810843190A CN 110759971 A CN110759971 A CN 110759971A
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 35
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 27
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 27
- 230000003078 antioxidant effect Effects 0.000 title claims abstract description 21
- 239000000654 additive Substances 0.000 title claims abstract description 13
- 239000003963 antioxidant agent Substances 0.000 title claims description 16
- 230000000996 additive effect Effects 0.000 title claims description 9
- 235000013305 food Nutrition 0.000 claims abstract description 16
- 241001465754 Metazoa Species 0.000 claims abstract description 8
- 239000002537 cosmetic Substances 0.000 claims abstract description 7
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract 2
- 210000003056 antler Anatomy 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 2
- 239000004480 active ingredient Substances 0.000 claims 2
- 239000012752 auxiliary agent Substances 0.000 claims 1
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 abstract description 12
- 150000001875 compounds Chemical class 0.000 abstract description 5
- 229910052742 iron Inorganic materials 0.000 abstract description 5
- 239000000243 solution Substances 0.000 description 12
- 235000006708 antioxidants Nutrition 0.000 description 11
- 108090000623 proteins and genes Proteins 0.000 description 8
- 241000283690 Bos taurus Species 0.000 description 7
- 238000012986 modification Methods 0.000 description 7
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- 239000000047 product Substances 0.000 description 7
- 102000004169 proteins and genes Human genes 0.000 description 7
- 108010033276 Peptide Fragments Proteins 0.000 description 6
- 102000007079 Peptide Fragments Human genes 0.000 description 6
- 150000001413 amino acids Chemical group 0.000 description 6
- 238000000034 method Methods 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- KMVWNDHKTPHDMT-UHFFFAOYSA-N 2,4,6-tripyridin-2-yl-1,3,5-triazine Chemical compound N1=CC=CC=C1C1=NC(C=2N=CC=CC=2)=NC(C=2N=CC=CC=2)=N1 KMVWNDHKTPHDMT-UHFFFAOYSA-N 0.000 description 5
- 125000000539 amino acid group Chemical group 0.000 description 5
- 230000003647 oxidation Effects 0.000 description 5
- 238000007254 oxidation reaction Methods 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 4
- 230000007717 exclusion Effects 0.000 description 4
- 150000002500 ions Chemical class 0.000 description 4
- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 description 4
- 101800000068 Antioxidant peptide Proteins 0.000 description 3
- 230000003064 anti-oxidating effect Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000005556 structure-activity relationship Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 239000012224 working solution Substances 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
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 241000282994 Cervidae Species 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 2
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 2
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 210000004899 c-terminal region Anatomy 0.000 description 2
- 239000008367 deionised water Substances 0.000 description 2
- 229910021641 deionized water Inorganic materials 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- HQVFCQRVQFYGRJ-UHFFFAOYSA-N formic acid;hydrate Chemical compound O.OC=O HQVFCQRVQFYGRJ-UHFFFAOYSA-N 0.000 description 2
- 238000004108 freeze drying Methods 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 239000000852 hydrogen donor Substances 0.000 description 2
- 230000002209 hydrophobic effect Effects 0.000 description 2
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- 241001416181 Axis axis Species 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
- 241000283007 Cervus nippon Species 0.000 description 1
- 102100027442 Collagen alpha-1(XII) chain Human genes 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- PMVSDNDAUGGCCE-TYYBGVCCSA-L Ferrous fumarate Chemical compound [Fe+2].[O-]C(=O)\C=C\C([O-])=O PMVSDNDAUGGCCE-TYYBGVCCSA-L 0.000 description 1
- 101000861874 Homo sapiens Collagen alpha-1(XII) chain Proteins 0.000 description 1
- 229910021578 Iron(III) chloride Inorganic materials 0.000 description 1
- RWRDLPDLKQPQOW-UHFFFAOYSA-N Pyrrolidine Chemical group C1CCNC1 RWRDLPDLKQPQOW-UHFFFAOYSA-N 0.000 description 1
- 230000010757 Reduction Activity Effects 0.000 description 1
- 229920005654 Sephadex Polymers 0.000 description 1
- 239000012507 Sephadex™ Substances 0.000 description 1
- 241001122767 Theaceae Species 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 229930003427 Vitamin E Natural products 0.000 description 1
- 239000008351 acetate buffer Substances 0.000 description 1
- XBJFCYDKBDVADW-UHFFFAOYSA-N acetonitrile;formic acid Chemical compound CC#N.OC=O XBJFCYDKBDVADW-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 235000019282 butylated hydroxyanisole Nutrition 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000013480 data collection Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 230000002542 deteriorative effect Effects 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
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- 239000012634 fragment Substances 0.000 description 1
- 238000013467 fragmentation Methods 0.000 description 1
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- 238000002546 full scan Methods 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
- 238000010438 heat treatment Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 125000002883 imidazolyl group Chemical group 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 125000001360 methionine group Chemical group N[C@@H](CCSC)C(=O)* 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 1
- 150000008442 polyphenolic compounds Chemical class 0.000 description 1
- 235000013824 polyphenols Nutrition 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 125000001500 prolyl group Chemical group [H]N1C([H])(C(=O)[*])C([H])([H])C([H])([H])C1([H])[H] 0.000 description 1
- 238000010298 pulverizing process Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 125000000168 pyrrolyl group Chemical group 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 108091008146 restriction endonucleases Proteins 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- -1 vitamin E Chemical class 0.000 description 1
- 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
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- 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
- 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
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
-
- 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)
- General Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Molecular Biology (AREA)
- Medicinal Chemistry (AREA)
- Epidemiology (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- Dermatology (AREA)
- Gerontology & Geriatric Medicine (AREA)
- Birds (AREA)
- Genetics & Genomics (AREA)
- Mycology (AREA)
- Nutrition Science (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Peptides Or Proteins (AREA)
- Cosmetics (AREA)
Abstract
The invention relates to a polypeptide compound Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg with iron reduction capacity, and the amino acid sequence of the polypeptide compound is Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg. The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg has antioxidant activity, not only has active function, but also can be used as additives of food, cosmetics, health products, animal food and the like, and has wide application range.
Description
Technical Field
The invention relates to polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg and application of the polypeptide as an antioxidant component in additives of foods, health care products, cosmetics, 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.
Disclosure of Invention
The invention aims to provide an application and a rapid screening method of polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg in antioxidant components; the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg has antioxidant activity, can be used as additives of food, health products, cosmetics, animal food and the like, and has good application prospect.
In order to realize the purpose, the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg is taken as an effective antioxidant component.
It has the sequence table of SEQ ID NO: 1, amino acid sequence; the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg is used as antioxidant component, and can be used as additive for food, health product, cosmetic, animal food, etc.
The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg with antioxidant activity is derived from Cervus Nippon Temminck. Since the protein pool of deer is not perfect enough and the total protein amount is small, while the homology of cattle and deer gene is over 90% (Sui Z G, Yuan H M, Liang Z, et al. Talanta,2013,107,189-194.), the bovine (bovine) was selected as the protein database in this experiment. The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg is derived from Collagen type XII alpha 1chain protein of a bovine (bovine) protein library, contains 13 amino acid residues, has an amino acid sequence of Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg, is a single-chain linear structure, is white powder, is easy to dissolve in water, and has a molecular weight of 1296 Da; has better iron reducing ability activity, and the iron reducing ability per 100 mu M is equivalent to that of 9.20 +/-0.54 mu M ascorbic acid (n is 3, Mean +/-SD).
The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg has the characteristics required by antioxidant peptide: amino acid residues containing benzene ring, pyrrolidine ring and imidazole ring can be used as hydrogen donors required by free radicals; containing hydrophobic amino acid residues, especially the N-terminal hydrophobic amino acid; 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-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg is serine, and the hydroxyl group of the serine is easy to form hydrogen bond with water molecules; contains 4 pyrrole rings and can be used as a hydrogen donor required by free radicals, so the Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-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 pilose antler 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-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg
The method combining LC-MS/MS and Shotgun proteomics technology is adopted. The spotted deer antler is used as a raw material, and a peptide segment with antioxidation is screened by carrying out enzymolysis on protein, centrifugation, purification and LC-MS/MS analysis and combining the structure-activity relationship characteristics.
The specific method comprises the following steps:
freeze drying fresh cornu Cervi Pantotrichum, pulverizing, adding deionized water to make cornu Cervi Pantotrichum concentration 33.3g/L, stirring, adding trypsin with mass of 0.5% (W/W) of cornu Cervi Pantotrichum, and performing enzymolysis at 40 deg.C for 3 hr; after enzymolysis, the enzymolysis liquid is sieved by a sieve of 80 meshes, and residues are extracted once by the same method. Heating the enzymolysis liquid of the two times to 90 ℃, preserving the temperature for 15 minutes, respectively filtering the enzymolysis liquid through 8 layers of gauze and 200-mesh sieves, centrifuging the obtained filtrate for 10 minutes at the speed of 10000g, and collecting supernatant; measuring the peptide concentration of the supernatant through Nanodrop Onec at 205nm, and adding water to dilute the supernatant to 20mg/mL of peptide concentration for later use; and filling the swelled Sephadex G-25medium filler into a gel column with the diameter of 2cm and the column height of 30 cm. Loading the enzymolysis solution on a gel column, eluting with deionized water as eluent at the flow rate of 3.5mL/min, eluting with 1/20 of bed volume as one flow portion for 2 bed volumes, collecting all 40 flow portions, and determining the peptide concentration at 205nm by using a Nanodrop Onec. Combining the 16 th to 25 th fractions according to the peptide concentration, and freeze-drying to obtain cornu Cervi Pantotrichum extract.
Subjecting the extract of cornu Cervi Pantotrichum to mass spectrometry with LTQ Orbitrap Velos: re-dissolving cornu Cervi Pantotrichum extract with 0.1% (V/V) formic acid water solution to obtain 0.4mg/mL solution, and performing LC-MS/MS analysis. One end of the capillary was drawn to a tip with an inner diameter of about 5 μm and a C18AQ packing was pressed into the column by air pressure, the length of the column being about 15 cm. The tip of the capillary was connected to a mass spectrometer. The mobile phase A used is 0.1% (V/V) formic acid water solution, the mobile phase B is 0.1% formic acid acetonitrile solution, and the linear gradient elution process is as follows: 0% B (0min) -2% B (2min) -25% B (87min) -35% B (97min) -90% B (99min) -90% B (109min) -2% B (110min) -2% B (120 min). The flow rate was 60. mu.L/min.
The temperature of the ion transmission capillary tube is set to be 200 ℃, the electrospray voltage is 1.8KV, and the normalized collision energy is 35.0%. Both MS and MS/MS were mapped using a data-dependent mode. The mass spectrometry scan conditions were set as: selecting 10 highest abundance ion peaks from full scans with each m/z of 400-2000 to perform MS/MS scanning, wherein dynamic exclusion (dynamic exclusion) is set as: the number of repetitions (repeat count) was 2, the repetition duration (repeat duration) was 30s, and the dynamic exclusion duration (exclusion duration) was 90 s. The system control and data collection was performed using Xcalibur software (version2.2, Thermo).
The collected raw file data was converted to mgf format using Thermo protein discover Daemon (v1.4) and retrieved using Mascot (version 2.3.0, Matrix Science, London, UK) in a bovine database (bovine, protein number 17890, downloaded from http:// www.uniprot.org /), with the following parameters: no restriction enzyme site, maximum number of missed cuts and fixed modification are set; a variable modification of a methionine residue, a proline residue plus 15.9949 Da; the mass tolerance (peptide tolerance) of the parent ion was 20ppm and the mass tolerance (fragmentation tolerance) of the fragment ion was 0.8 Da. When the peptide fragment result is derived, score >25 is set, and the significance difference P is adjusted to control the false positive rate (FDR) of the peptide fragment within 1 percent. The identification result is shown in the attached table I. And (4) screening by combining the structure-activity relationship to obtain the polypeptide with the sequence of Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg.
Example 2 detection of iron-reducing Activity of polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg
Principle of
Fe3+-tripyridyl-triazazine (TPTZ) reduced to Fe by antioxidant ingredients2+The sample is blue, the maximum light absorption value is at 593nm, and the antioxidant activity of the sample is calculated according to the light absorption value.
Experimental methods
The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg used in the experiment is synthesized by Nanjing Jie peptide biotechnology limited, and has purity>95 percent. TPTZ working solution was prepared from 25mL of 0.3M pH 3.6 acetate buffer, 2.5mL of 10mM TPTZ solution (containing 40mM hydrochloric acid), and 2.5mL of 20mM FeCl3And (4) solution composition. 1.8mL of fresh TPTZ working solution is mixed with 0.2mL of Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg solution, the mixture reacts at 37 ℃ for 10min, and the absorbance is measured at 593nm by using a spectrophotometer. Preparing Vc solution of 27 mu g/mL, transferring 3,6,9,12, 15 and 20 mu L, mixing with TPTZ working solution of 1.8mL, adding distilled water to 2.0mL, reacting at 37 deg.C for 10min, measuring absorbance at 593nm with spectrophotometer, calculating Vc regression curve with Vc concentration on abscissa and absorbance on ordinate, where y is 0.0149x +0.0912, and R is regression equation20.9988. Calculating according to the absorbance value of Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg and a regression equation: the iron reducing ability of a Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg solution with a certain concentration is equivalent to the amount of a Vc solution substance (μ M).
Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg solutions with the concentrations of 50, 100, 250 and 500 mu M are respectively prepared, and the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg is subjected to iron reduction activity detection according to the method. The results are shown in the table I:
TABLE one iron reduction Capacity of Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg at different concentrations
The polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg has antioxidant activity, can be used as additives of food, health products, cosmetics, animal feed and the like, and has wide application.
Attached table: peptide fragment of pilose antler extract identified by LC-MS/MS
Take TGTPGLPGPPGPMGPPGDR as an example: the modified type of the peptide segment is oxidation (M); 3Pro (O) (P), indicating that the peptide fragment has 1 methionine oxidation modification and 3 proline hydroxyl modification. The modification position of the peptide fragment is 0.0002002000001002000.0, which indicates that the proline hydroxyl group modification at the 3 position of the peptide fragment is positioned at the 4 th, 7 th and 16 th residues, and the methionine oxidation modification is positioned at the 13 th 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.
The names, abbreviations and abbreviations of diamino acids in the attached tables
Claims (5)
1. An antioxidant polypeptide derived from velvet antler, comprising: the polypeptide is Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg and has a sequence table SEQ ID NO: 1; the amino acid sequence of the polypeptide is Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg.
2. The use of the antioxidant polypeptide derived from velvet antler as claimed in claim 1 in the preparation of an additive having an antioxidant function for foods, cosmetics, health products or animal foods.
3. Use according to claim 2, characterized in that: the additive takes the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg in the claim 1 as an active ingredient.
4. An additive for food, cosmetics, health products or animal food with an antioxidant function, which is characterized in that: the additive takes the polypeptide Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Hyp-Gln-Gly-Glu-Ser-Arg in the claim 1 as an active ingredient.
5. The additive of claim 4, wherein: the additive also contains a carrier and/or an auxiliary agent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810843190.1A CN110759971B (en) | 2018-07-27 | 2018-07-27 | Cornu Cervi Pantotrichum-derived antioxidant polypeptide, and its application and additive |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810843190.1A CN110759971B (en) | 2018-07-27 | 2018-07-27 | Cornu Cervi Pantotrichum-derived antioxidant polypeptide, and its application and additive |
Publications (2)
| Publication Number | Publication Date |
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| CN110759971A true CN110759971A (en) | 2020-02-07 |
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| CN114652633A (en) * | 2022-05-20 | 2022-06-24 | 广州优理氏生物科技有限公司 | Preparation method of polypeptide enzymolysis and fermentation composition and application of polypeptide enzymolysis and fermentation composition in skin care products |
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| CN114652633B (en) * | 2022-05-20 | 2022-08-02 | 广州优理氏生物科技有限公司 | Preparation method of polypeptide enzymolysis and fermentation composition and application of polypeptide enzymolysis and fermentation composition in skin care products |
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