CN107523599A - The method that fermentation method prepares peony pollen polypeptide - Google Patents
The method that fermentation method prepares peony pollen polypeptide Download PDFInfo
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- CN107523599A CN107523599A CN201710779679.2A CN201710779679A CN107523599A CN 107523599 A CN107523599 A CN 107523599A CN 201710779679 A CN201710779679 A CN 201710779679A CN 107523599 A CN107523599 A CN 107523599A
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- fermentation
- peony
- peony pollen
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- 241000736199 Paeonia Species 0.000 title claims abstract description 74
- 235000006484 Paeonia officinalis Nutrition 0.000 title claims abstract description 73
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 57
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 49
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 44
- 238000000855 fermentation Methods 0.000 title claims abstract description 40
- 238000000034 method Methods 0.000 title claims abstract description 38
- 230000004151 fermentation Effects 0.000 title claims abstract description 36
- 102000004882 Lipase Human genes 0.000 claims abstract description 30
- 108090001060 Lipase Proteins 0.000 claims abstract description 30
- 239000004367 Lipase Substances 0.000 claims abstract description 30
- 235000019421 lipase Nutrition 0.000 claims abstract description 30
- 239000000341 volatile oil Substances 0.000 claims abstract description 21
- 238000005238 degreasing Methods 0.000 claims abstract description 15
- 238000000605 extraction Methods 0.000 claims abstract description 15
- 239000012530 fluid Substances 0.000 claims abstract description 12
- 240000006024 Lactobacillus plantarum Species 0.000 claims abstract description 9
- 235000013965 Lactobacillus plantarum Nutrition 0.000 claims abstract description 9
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims abstract description 9
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 claims abstract description 9
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 7
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 7
- 239000000843 powder Substances 0.000 claims description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 25
- 229920001661 Chitosan Polymers 0.000 claims description 18
- 239000007788 liquid Substances 0.000 claims description 13
- 230000000975 bioactive effect Effects 0.000 claims description 12
- 102000004190 Enzymes Human genes 0.000 claims description 11
- 108090000790 Enzymes Proteins 0.000 claims description 11
- 239000012153 distilled water Substances 0.000 claims description 10
- 230000001954 sterilising effect Effects 0.000 claims description 10
- 238000003756 stirring Methods 0.000 claims description 10
- 239000000284 extract Substances 0.000 claims description 9
- 240000005001 Paeonia suffruticosa Species 0.000 claims description 8
- 235000003889 Paeonia suffruticosa Nutrition 0.000 claims description 8
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 claims description 7
- 235000013305 food Nutrition 0.000 claims description 7
- 238000002360 preparation method Methods 0.000 claims description 7
- 238000001035 drying Methods 0.000 claims description 6
- 238000001914 filtration Methods 0.000 claims description 6
- 239000004094 surface-active agent Substances 0.000 claims description 6
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 5
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 claims description 5
- 230000009471 action Effects 0.000 claims description 5
- 238000004140 cleaning Methods 0.000 claims description 5
- 230000009849 deactivation Effects 0.000 claims description 5
- 239000000706 filtrate Substances 0.000 claims description 5
- 238000005859 coupling reaction Methods 0.000 claims description 4
- 230000008014 freezing Effects 0.000 claims description 4
- 238000007710 freezing Methods 0.000 claims description 4
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- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 3
- 239000003921 oil Substances 0.000 claims 2
- 239000000243 solution Substances 0.000 claims 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims 1
- 229910019142 PO4 Inorganic materials 0.000 claims 1
- 230000003213 activating effect Effects 0.000 claims 1
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 239000007853 buffer solution Substances 0.000 claims 1
- 229910052799 carbon Inorganic materials 0.000 claims 1
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 150000002466 imines Chemical class 0.000 claims 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims 1
- 239000010452 phosphate Substances 0.000 claims 1
- 238000005507 spraying Methods 0.000 claims 1
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- 238000001694 spray drying Methods 0.000 abstract description 8
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- 101800004538 Bradykinin Proteins 0.000 description 5
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- QXZGBUJJYSLZLT-UHFFFAOYSA-N H-Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg-OH Natural products NC(N)=NCCCC(N)C(=O)N1CCCC1C(=O)N1C(C(=O)NCC(=O)NC(CC=2C=CC=CC=2)C(=O)NC(CO)C(=O)N2C(CCC2)C(=O)NC(CC=2C=CC=CC=2)C(=O)NC(CCCN=C(N)N)C(O)=O)CCC1 QXZGBUJJYSLZLT-UHFFFAOYSA-N 0.000 description 5
- 102000004877 Insulin Human genes 0.000 description 5
- 108090001061 Insulin Proteins 0.000 description 5
- QXZGBUJJYSLZLT-FDISYFBBSA-N bradykinin Chemical compound NC(=N)NCCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N1[C@H](C(=O)NCC(=O)N[C@@H](CC=2C=CC=CC=2)C(=O)N[C@@H](CO)C(=O)N2[C@@H](CCC2)C(=O)N[C@@H](CC=2C=CC=CC=2)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)CCC1 QXZGBUJJYSLZLT-FDISYFBBSA-N 0.000 description 5
- 239000000864 hyperglycemic agent Substances 0.000 description 5
- 229940125396 insulin Drugs 0.000 description 5
- 102000004169 proteins and genes Human genes 0.000 description 5
- 108090000623 proteins and genes Proteins 0.000 description 5
- 230000028327 secretion Effects 0.000 description 5
- 108010025252 Kassinin Proteins 0.000 description 4
- -1 Propyl group carbodiimide Chemical class 0.000 description 4
- 239000008363 phosphate buffer Substances 0.000 description 4
- 230000002255 enzymatic effect Effects 0.000 description 3
- 150000004676 glycans Chemical class 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 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
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000011149 active material Substances 0.000 description 2
- 150000001718 carbodiimides Chemical class 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
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- 239000000463 material Substances 0.000 description 2
- 235000008935 nutritious Nutrition 0.000 description 2
- 235000013406 prebiotics Nutrition 0.000 description 2
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- JBGSZRYCXBPWGX-BQBZGAKWSA-N Ala-Arg-Gly Chemical compound OC(=O)CNC(=O)[C@@H](NC(=O)[C@@H](N)C)CCCN=C(N)N JBGSZRYCXBPWGX-BQBZGAKWSA-N 0.000 description 1
- 108010064733 Angiotensins Proteins 0.000 description 1
- 102000015427 Angiotensins Human genes 0.000 description 1
- RWDVGVPHEWOZMO-GUBZILKMSA-N Arg-Cys-Val Chemical compound CC(C)[C@H](NC(=O)[C@H](CS)NC(=O)[C@@H](N)CCCNC(N)=N)C(O)=O RWDVGVPHEWOZMO-GUBZILKMSA-N 0.000 description 1
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- OCEHKDFAWQIBHH-FXQIFTODSA-N Cys-Arg-Cys Chemical compound C(C[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CS)N)CN=C(N)N OCEHKDFAWQIBHH-FXQIFTODSA-N 0.000 description 1
- ABLJDBFJPUWQQB-DCAQKATOSA-N Cys-Leu-Arg Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)NC(=O)[C@H](CS)N ABLJDBFJPUWQQB-DCAQKATOSA-N 0.000 description 1
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- 244000020551 Helianthus annuus Species 0.000 description 1
- 235000003222 Helianthus annuus Nutrition 0.000 description 1
- 102000001399 Kallikrein Human genes 0.000 description 1
- 108060005987 Kallikrein Proteins 0.000 description 1
- 108010093008 Kinins Proteins 0.000 description 1
- 102000002397 Kinins Human genes 0.000 description 1
- RVOMPSJXSRPFJT-DCAQKATOSA-N Lys-Ala-Arg Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O RVOMPSJXSRPFJT-DCAQKATOSA-N 0.000 description 1
- 241000143233 Mytilus coruscus Species 0.000 description 1
- 241000218201 Ranunculaceae Species 0.000 description 1
- KCFKKAQKRZBWJB-ZLUOBGJFSA-N Ser-Cys-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CS)C(=O)N[C@@H](C)C(O)=O KCFKKAQKRZBWJB-ZLUOBGJFSA-N 0.000 description 1
- SGZVZUCRAVSPKQ-FXQIFTODSA-N Ser-Val-Cys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CO)N SGZVZUCRAVSPKQ-FXQIFTODSA-N 0.000 description 1
- ODRUTDLAONAVDV-IHRRRGAJSA-N Ser-Val-Tyr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O ODRUTDLAONAVDV-IHRRRGAJSA-N 0.000 description 1
- 229930003270 Vitamin B Natural products 0.000 description 1
- 229930003268 Vitamin C Natural products 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 125000001931 aliphatic group Chemical group 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 239000002416 angiotensin derivative Substances 0.000 description 1
- 238000013475 authorization Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 235000019658 bitter taste Nutrition 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 238000007385 chemical modification Methods 0.000 description 1
- 230000006835 compression Effects 0.000 description 1
- 238000007906 compression Methods 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 210000002249 digestive system Anatomy 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002213 flavones Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 210000004051 gastric juice Anatomy 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
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- 238000010025 steaming Methods 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 235000019156 vitamin B Nutrition 0.000 description 1
- 239000011720 vitamin B Substances 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B9/00—Essential oils; Perfumes
- C11B9/02—Recovery or refining of essential oils from raw materials
- C11B9/027—Recovery of volatiles by distillation or stripping
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/06—Lysis of microorganisms
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/10—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a carbohydrate
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/04—Plant cells or tissues
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/18—Carboxylic ester hydrolases (3.1.1)
- C12N9/20—Triglyceride splitting, e.g. by means of lipase
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/02—Preparation of peptides or proteins having a known sequence of two or more amino acids, e.g. glutathione
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12Y301/00—Hydrolases acting on ester bonds (3.1)
- C12Y301/01—Carboxylic ester hydrolases (3.1.1)
- C12Y301/01003—Triacylglycerol lipase (3.1.1.3)
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- 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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Abstract
The invention discloses a kind of method that fermentation method prepares peony pollen polypeptide, including pollen broken wall, extraction of essential oil, fermentation, degreasing, spray drying.Peony pollen is freezed, mechanical crushing, broken wall is continued using Lactobacillus plantarum and saccharomyces cerevisiae anaerobic fermentation again, fermentation of bacillus subtilis is used again, peony pollen albumen is digested using neutral proteinase caused by its fermentation, the peony pollen polypeptide of activity intensity is obtained, degreasing is then carried out to zymotic fluid using chitosan-immobilized lipase.Have the beneficial effect that：The peony pollen polypeptide being prepared will not be hydrolyzed by erepsin, directly act on internal acceptor, and blood pressure lowering effect is notable.At the same time, peony pollen polypeptide also has effect of weight reducing for expectation.
Description
Technical field
The present invention relates to biological activity protein peptide art, specifically a kind of fermentation method prepares peony pollen polypeptide
Method.
Background technology
Tree peony（Paeonia suffruticosa Andr.）Belong to Ranunculaceae Paeonia, machaka, there is very high sight
Reward value and pharmaceutical value.For a long time, the utilization of tree peony deep processed product but seem serious hysteresis that production added value is very
It is low, do not form industrial chain.Peony pollen is not only nutritious but also contains various bioactivators.Amino acid content exists
More than 25.0%, rich in there is 17 kinds of amino acid, vitamin B, Vitamin C content enrich, also containing tree peony polysaccharide, tree peony flavones etc.
Bioactive substance.Protein content 39.3%, far above the protein content of general pollen, protein compression is referred to as by academia
The hat of contracting body, native protein.
Natural pollen has tough and tensile cell membrane, the sporopollenin in its structure have acidproof, alkaline-resisting, heatproof, it is pressure-resistant and
The physicochemical property highly stable to hydrochloric acid in gastric juice and other digestive system enzymes, hinders human body and pollen protein is absorbed.
Prior art such as Authorization Notice No. is CN103053893B Chinese invention patent, discloses a kind of natural pollen hair
Ferment product production technology, advanced air force crushing technology, biological fermentation equipment and technology are make use of, using natural pollen as original
Material, crushes broken wall, then access prebiotic fermentative microorganism and pollen protein is degraded into amino first with air force by natural pollen
Acid and polypeptide, so as to obtain natural pollen tunnings a kind of nutritious and containing a large amount of prebiotic substances, finally through overdrying
Product is made in dry and crushing.The said goods delicate mouthfeel, it is sour-sweet it is moderate, without bitter taste stink, but the recovery rate of tree peony polypeptide
It is not high, and the activity of the functional polypeptide extracted is not strong, application prospect is limited.
The content of the invention
It is an object of the invention to provide a kind of method that fermentation method prepares peony pollen polypeptide, preparation method operation
Simply, required instrument is few, and it is low to prepare cost；The peony pollen polypeptide of preparation has excellent blood pressure reduction effect, and has
Standby effect of weight reducing.
The present invention is directed to the problem of being mentioned in background technology, and the technical scheme taken is：Fermentation method prepares peony pollen egg
The method of white polypeptide, including pollen broken wall, extraction of essential oil, fermentation, degreasing, spray drying.The peony pollen albumen being prepared
Polypeptide will not be hydrolyzed by erepsin, directly act on internal acceptor, can promote the secretion of insulin and hyperglycemic factor and prolong
The duration of runs of the long food in enteron aisle.Meanwhile it can suppress Angiotensin-Converting（ACE）Activity, discharge kassinin kinin
Enzyme-kinin system（KKS）It is active, produces the bradykinin with the effect that reduces blood pressure, blood pressure lowering effect is notable, bright
It is aobvious to be better than prior art.At the same time, peony pollen polypeptide also has effect of weight reducing for expectation.
Fermentation step is：After broken wall in peony pollen add 1 ~ 6 times of volume clear water, add the essential oils of 2 ~ 5wt ‰,
0.1 ~ 0.2 wt ‰ surfactant, the Lactobacillus plantarums of 1 ~ 4 wt ‰ and the saccharomyces cerevisiaes of 1 ~ 4 wt ‰ are inoculated with after sterilizing, in 32 ~
43 DEG C, 20 ~ 30r/min of stir speed (S.S.), constant-temperatureanaerobic anaerobic is fermented 24 ~ 72h under the conditions of natural pH, 0.5 ~ 2.0 wt ‰ is inoculated with after sterilizing
Bacillus subtilis, fermented 12 ~ 24h under the conditions of mutually synthermal, stir speed (S.S.), pH, and filtrated air is passed through during fermentation
0.001~0.01m3/ min, filter to take zymotic fluid, number of repetition 1 ~ 4 time.Wherein, Lactobacillus plantarum is purchased from Shandong sunflower life
Thing Science and Technology Ltd.；Saccharomyces cerevisiae is purchased from Angel Yeast Co., Ltd；Bacillus subtilis is purchased from the high dragon biology in Shandong
Science and Technology Ltd..Only exposore is destroyed by way of freezing and mechanical breaking-wall method, sporoderm-broken rate is not high, can directly result in
The recovery rate of peony pollen polypeptide is not high.Under peony essential oil existence condition, Lactobacillus plantarum and saccharomyces cerevisiae collaboration hair
Ferment, destroy the sporopollenin of peony pollen tapetum so that the active material outflow in pollen cell.Fermentation of bacillus subtilis mistake
Journey produces neutral proteinase, the peony pollen polypeptide by the peony pollen proteolysis of dissolution into relatively small molecular weight.
Pollen broken wall step is that peony pollen is placed in into 2 ~ 24h of freezing under -40 ~ -50 DEG C of environment, mechanical crushing broken wall, is obtained
Peony pollen after to broken wall.Exposore is tentatively destroyed using mechanical crushing method, is advantageous to postorder peony pollen
Fermentation operation.
Extraction of essential oil step is：Solid-liquid ratio 1g is pressed in peony petal powder：2 ~ 6 mL add absolute ethyl alcohol, are placed in microwave
Extracted in abstraction instrument, microwave power is 200 ~ 400W, and a microwave action time is 15 ~ 35s, and extraction time is 2 ~ 6 times, filtering
Take extract to stand, remove water layer after its layering is stable, add anhydrous sodium sulfate and stand overnight, filter to take filtrate, and rotate
Obtain peony essential oil.Compared to traditional SDE methods, the essential oil yield of the method extraction of said extracted peony essential oil is higher, extraction
Temperature is relatively low, and the active ingredient in essential oil will not be made to be thermally decomposed or Oxidative demage.
Defatting step is that 2 ~ 4 ‰ chitosan-immobilized lipase are added in zymotic fluid, and constant temperature digests at 25 ~ 50 DEG C
0.8 ~ 2.5h, enzymolysis terminate rear enzyme deactivation, are filtrated to get degreasing peony pollen polypeptide liquid.Lipase can be by the fat in zymotic fluid
Fat is progressively hydrolyzed into glycerine and aliphatic acid.Lipase after immobilization, its stability, enzymatic activity, heat resistance are significantly improved,
It is and reusable.
The preparation process of chitosan-immobilized lipase is：With drying Chitosan powder, with the 60 of pH6.0 ~ 6.8 ~
80mmol/L phosphate buffer soaked overnights, then add the 1- ethyls -3- of Chitosan powder quality 0.7 ~ 1.2%（3- first
Base amino）Propyl group 0.5 ~ 0.9h of carbodiimide activation, add the glutaraldehyde coupling reaction 1 of 1 ~ 2 times of volume of Chitosan powder ~
1.8h, cleaned 5 ~ 8 times with distilled water, 0.7 ~ 2.0mg/mL lipase solutions and 0.1 ~ 0.3 ‰ bioactive peptides are added, in ice bath
0.5 ~ 1.5h is slowly stirred, then places refrigerator overnight, secondary daily distilled water cleaning down, filtration drying obtains chitosan
Immobilized lipase.1- ethyls -3-（3- methylaminos）Propyl group carbodiimide carries out chemical modification, effective activation to chitosan
Chitosan surface hydroxyl groups.Chitosan surface amino groups after activation are coupled to obtain excellent lipase immobilization load with glutaraldehyde
Body.Lipase adsorption forms chitosan-immobilized lipase on carrier.
The amino acid sequence of bioactive peptide is SCASVCKARCLRARGCRCVSVYCRCLR.Above-mentioned bioactive peptide can be obvious
Strengthen the physics and chemisorption power between lipase and lipase immobilization carrier, improve immobilized lipase enzymatic conversion
Rate, and improve the reusable number of chitosan-immobilized lipase.
Surfactant is food grade surfactant.
Compared with prior art, the advantage of the invention is that：
1. the peony pollen polypeptide being prepared will not be hydrolyzed by erepsin, internal acceptor is directly acted on, can be promoted
The duration of runs of the secretion and extension food of insulin and hyperglycemic factor in enteron aisle.Meanwhile it can suppress angiotensins
Converting Enzyme（ACE）Activity, make kallikrein kinin system（KKS）It is active, generation has the effect of reducing blood pressure
Bradykinin, blood pressure lowering effect is notable, hence it is evident that better than prior art.At the same time, peony pollen polypeptide is also for meaning
Material has effect of weight reducing；
2. using 1- ethyls -3-（3- methylaminos）Propyl group carbodiimide activation chitosan, and be coupled to form fat with glutaraldehyde
Enzyme immobilization carrier, the bioactive peptide that amino acid sequence is SCASVCKARCLRARGCRCVSVYCRCLR is recycled to be carried to improve
Physics and chemisorbed power between body and lipase, improve stability, enzymatic activity, heat resistance and the access times of lipase；
3. under peony essential oil existence condition, Lactobacillus plantarum and saccharomyces cerevisiae cooperative fermentation, peony pollen tapetum is destroyed
Sporopollenin so that the active material outflow in pollen cell.
Embodiment
The present invention program is described further below by embodiment：
Embodiment 1：
The method that fermentation method prepares peony pollen polypeptide, comprises the following steps：
1）Pollen broken wall：Peony pollen is placed in 2 ~ 24h of freezing, mechanical crushing broken wall, after obtaining broken wall under -40 ~ -50 DEG C of environment
Peony pollen；
2）Extraction of essential oil：Solid-liquid ratio 1g is pressed in peony petal powder：2 ~ 6 mL add absolute ethyl alcohol, are placed in Microwave Extraction Apparatus
Extraction, microwave power are 200 ~ 400W, and a microwave action time is 15 ~ 35s, and extraction time is 2 ~ 6 times, filters to take extract
Stand, remove water layer after its layering is stable, add anhydrous sodium sulfate and stand overnight, filter to take filtrate, and rotate and obtain tree peony
Essential oil；
3）Fermentation：The clear water of 1 ~ 6 times of volume is added in peony pollen after broken wall, adds the essential oils of 2 ~ 5wt ‰, 0.1 ~ 0.2
Wt ‰ surfactant, the Lactobacillus plantarums of 1 ~ 4 wt ‰ and the saccharomyces cerevisiaes of 1 ~ 4 wt ‰ are inoculated with after sterilizing, in 32 ~ 43 DEG C, stir
Mix constant-temperatureanaerobic anaerobic under the conditions of 20 ~ 30r/min of speed, natural pH to ferment 24 ~ 72h, the withered grass buds of 0.5 ~ 2.0 wt ‰ are inoculated with after sterilizing
Spore bacillus, under the conditions of mutually synthermal, stir speed (S.S.), pH ferment 12 ~ 24h, be passed through during fermentation filtrated air 0.001~
0.01m3/ min, filter to take zymotic fluid, number of repetition 1 ~ 4 time；
4）Degreasing：2 ~ 4 ‰ chitosan-immobilized lipase are added in zymotic fluid, constant temperature digests 0.8 ~ 2.5h at 25 ~ 50 DEG C,
Enzymolysis terminates rear enzyme deactivation, is filtrated to get degreasing peony pollen polypeptide liquid.The preparation process of chitosan-immobilized lipase is：
With Chitosan powder is dried, with 60 ~ 80mmol/L phosphate buffer soaked overnights of pH6.0 ~ 6.8, chitosan is then added
1- ethyls-the 3- of powder quality 0.7 ~ 1.2%（3- methylaminos）Propyl group 0.5 ~ 0.9h of carbodiimide activation, adds chitosan
Glutaraldehyde 1 ~ the 1.8h of coupling reaction of 1 ~ 2 times of volume of powder, is cleaned 5 ~ 8 times with distilled water, adds 0.7 ~ 2.0mg/mL lipase
Solution and 0.1 ~ 0.3 ‰ bioactive peptides, are slowly stirred 0.5 ~ 1.5h in ice bath, then place refrigerator overnight, secondary daily steaming
Distilled water cleaning down, filtration drying obtain chitosan-immobilized lipase.The amino acid sequence of bioactive peptide is
SCASVCKARCLRARGCRCVSVYCRCLR；
5）Spray drying：Fermented liquid spray drying after degreasing is obtained into peony pollen polypeptide powder.Above-mentioned peony pollen
Polypeptide will not be hydrolyzed by erepsin, directly act on internal acceptor, can promote the secretion of insulin and hyperglycemic factor
With the duration of runs of the extension food in enteron aisle.Meanwhile it can suppress Angiotensin-Converting（ACE）Activity, make kassinin kinin
Discharge enzyme-kinin system（KKS）It is active, produces the bradykinin with the effect that reduces blood pressure, blood pressure lowering effect shows
Write, hence it is evident that better than prior art.At the same time, peony pollen polypeptide also has effect of weight reducing for expectation.
Embodiment 2：
Fermentation method prepares the most preferred method of peony pollen polypeptide, comprises the following steps：
1）Pollen broken wall：Peony pollen is placed under -45 DEG C of environment and freezes 12h, mechanical crushing broken wall, obtains peony after broken wall
Powder；
2）Extraction of essential oil：Solid-liquid ratio 1g is pressed in peony petal powder：4mL adds absolute ethyl alcohol, is placed in Microwave Extraction Apparatus and extracts
Take, microwave power 300W, a microwave action time is 25s, and extraction time is 3 times, filters to take extract standing, treats its point
Remove water layer after layer is stable, add anhydrous sodium sulfate and stand overnight, filter to take filtrate, and rotate and obtain peony essential oil；
3）Fermentation：The clear water of 3 times of volumes is added in peony pollen after broken wall, adds the essential oils of 4wt ‰, 0.2 wt ‰ is told
Temperature -80, the Lactobacillus plantarums of 3 wt ‰ and the saccharomyces cerevisiaes of 2wt ‰ are inoculated with after sterilizing, in 35 DEG C, stir speed (S.S.) 20r/min, natural pH
Under the conditions of constant-temperatureanaerobic anaerobic fermentation 36h, the bacillus subtilises of 1.0 wt ‰ are inoculated with after sterilizing, in mutually synthermal, stir speed (S.S.), pH bars
Fermented 24h under part, and filtrated air 0.008m is passed through during fermentation3/ min, filter to take zymotic fluid, number of repetition 3 times；
4）Degreasing：3 ‰ chitosan-immobilized lipase are added in zymotic fluid, constant temperature digests 1.5h at 32 DEG C, and enzymolysis terminates
Enzyme deactivation afterwards, it is filtrated to get degreasing peony pollen polypeptide liquid.The preparation process of chitosan-immobilized lipase is：With dry shell
Glycan powder, with pH6.5 70mmol/L phosphate buffer soaked overnights, then add Chitosan powder quality 1.0%
1- ethyls -3-（3- methylaminos）Propyl group carbodiimide activation 0.7h, the glutaraldehyde for adding 1.0 times of volumes of Chitosan powder are even
Connection reaction 1.2h, is cleaned 8 times with distilled water, adds 1.2mg/mL lipase solutions and 0.3 ‰ bioactive peptides, in ice bath slowly
0.9h is stirred, then places refrigerator overnight, secondary daily distilled water cleaning down, filtration drying obtains chitosan-immobilized fat
Fat enzyme.The amino acid sequence of bioactive peptide is SCASVCKARCLRARGCRCVSVYCRCLR；
5）Spray drying：Fermented liquid spray drying after degreasing is obtained into peony pollen polypeptide powder.Above-mentioned peony pollen
Polypeptide will not be hydrolyzed by erepsin, directly act on internal acceptor, can promote the secretion of insulin and hyperglycemic factor
With the duration of runs of the extension food in enteron aisle.Meanwhile it can suppress Angiotensin-Converting（ACE）Activity, make kassinin kinin
Discharge enzyme-kinin system（KKS）It is active, produces the bradykinin with the effect that reduces blood pressure, blood pressure lowering effect shows
Write, hence it is evident that better than prior art.At the same time, peony pollen polypeptide also has effect of weight reducing for expectation.
Embodiment 3：
The method that fermentation method prepares peony pollen polypeptide, comprises the following steps：
1）Pollen broken wall：Peony pollen is placed under -50 DEG C of environment and freezes 12h, mechanical crushing broken wall, obtains peony after broken wall
Powder；
2）Extraction of essential oil：Solid-liquid ratio 1g is pressed in peony petal powder：6mL adds absolute ethyl alcohol, is placed in Microwave Extraction Apparatus and extracts
Take, microwave power 400W, a microwave action time is 15s, and extraction time is 3 times, filters to take extract standing, treats its point
Remove water layer after layer is stable, add anhydrous sodium sulfate and stand overnight, filter to take filtrate, and rotate and obtain peony essential oil；
3）Fermentation：The clear water of 3 times of volumes is added in peony pollen after broken wall, adds the essential oils of 5wt ‰, 0.2 wt ‰ is told
Temperature -20, the Lactobacillus plantarums of 4 wt ‰ and the saccharomyces cerevisiaes of 4 wt ‰ are inoculated with after sterilizing, in 35 DEG C, stir speed (S.S.) 30r/min, nature
Constant-temperatureanaerobic anaerobic fermentation 24h, is inoculated with the bacillus subtilises of 2.0 wt ‰, in mutually synthermal, stir speed (S.S.), pH under the conditions of pH after sterilizing
Under the conditions of ferment 12h, filtrated air 0.01m is passed through during fermentation3/ min, filter to take zymotic fluid, number of repetition 2 times；
4）Degreasing：4 ‰ chitosan-immobilized lipase are added in zymotic fluid, constant temperature digests 1.5h at 28 DEG C, and enzymolysis terminates
Enzyme deactivation afterwards, it is filtrated to get degreasing peony pollen polypeptide liquid.The preparation process of chitosan-immobilized lipase is：With dry shell
Glycan powder, with pH6.5 80mmol/L phosphate buffer soaked overnights, then add Chitosan powder quality 1.2%
1- ethyls -3-（3- methylaminos）Propyl group carbodiimide activation 0.7h, add the glutaraldehyde coupling of 2 times of volumes of Chitosan powder
1.8h is reacted, is cleaned 8 times with distilled water, 2.0mg/mL lipase solutions and 0.3 ‰ bioactive peptides is added, is slowly stirred in ice bath
0.5h is mixed, then places refrigerator overnight, secondary daily distilled water cleaning down, filtration drying obtains chitosan-immobilized fat
Enzyme.The amino acid sequence of bioactive peptide is SCASVCKARCLRARGCRCVSVYCRCLR；
5）Spray drying：Fermented liquid spray drying after degreasing is obtained into peony pollen polypeptide powder.Above-mentioned peony pollen
Polypeptide will not be hydrolyzed by erepsin, directly act on internal acceptor, can promote the secretion of insulin and hyperglycemic factor
With the duration of runs of the extension food in enteron aisle.Meanwhile it can suppress Angiotensin-Converting（ACE）Activity, make kassinin kinin
Discharge enzyme-kinin system（KKS）It is active, produces the bradykinin with the effect that reduces blood pressure, blood pressure lowering effect shows
Write, hence it is evident that better than prior art.At the same time, peony pollen polypeptide also has effect of weight reducing for expectation.
Routine operation in the operating procedure of the present invention is well known to those skilled in the art, herein without repeating.
Technical scheme is described in detail embodiment described above, it should be understood that it is described above only
For the specific embodiment of the present invention, it is not intended to limit the invention, all any modifications made in the spirit of the present invention,
Supplement or similar fashion replacement etc., should be included in the scope of the protection.
Sequence table
<110>Lanxi City silence bio tech ltd
<120>The method that fermentation method prepares peony pollen polypeptide
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 27
<212> PRT
<213>Artificial synthesized (Mytilus coruscus)
<400> 1
Ser Cys Ala Ser Val Cys Lys Ala Arg Cys Leu Arg Ala Arg Gly Cys
1 5 10 15
Arg Cys Val Ser Val Tyr Cys Arg Cys Leu Arg
20 25
Claims (7)
1. the method that fermentation method prepares peony pollen polypeptide, including pollen broken wall, extraction of essential oil, fermentation, degreasing, spraying are dry
It is dry, it is characterised in that：Described fermentation step is：The clear water of 1 ~ 6 times of volume is added in peony pollen after broken wall, add 2 ~
The surfactant of the essential oils of 5wt ‰, 0.1 ~ 0.2 wt ‰, it is inoculated with the Lactobacillus plantarums of 1 ~ 4 wt ‰ after sterilizing and 1 ~ 4 wt ‰ makes wine
Yeast, 24 ~ 72h of constant-temperatureanaerobic anaerobic fermentation, is inoculated with after sterilizing under the conditions of 32 ~ 43 DEG C, 20 ~ 30r/min of stir speed (S.S.), natural pH
The bacillus subtilises of 0.5 ~ 2.0 wt ‰, ferment 12 ~ 24h under the conditions of mutually synthermal, stir speed (S.S.), pH, is passed through during fermentation
0.001~0.01m of filtrated air3/ min, filter to take zymotic fluid, number of repetition 1 ~ 4 time.
2. the method that fermentation method according to claim 1 prepares peony pollen polypeptide, it is characterised in that：Described flower
Powder broken wall step is that peony pollen is placed under -40 ~ -50 DEG C of environment into 2 ~ 24h of freezing, mechanical crushing broken wall, is obtained male after broken wall
Red pollen.
3. the method that fermentation method according to claim 1 prepares peony pollen polypeptide, it is characterised in that：Described essence
Oil extract step is：Solid-liquid ratio 1g is pressed in peony petal powder：2 ~ 6 mL add absolute ethyl alcohol, are placed in Microwave Extraction Apparatus and extract
Take, microwave power is 200 ~ 400W, and a microwave action time is 15 ~ 35s, and extraction time is 2 ~ 6 times, and it is quiet to filter to take extract
Put, remove water layer after its layering is stable, add anhydrous sodium sulfate and stand overnight, filter to take filtrate, and rotate and obtain tree peony essence
Oil.
4. the method that fermentation method according to claim 1 prepares peony pollen polypeptide, it is characterised in that：Described is de-
Fat step is that 2 ~ 4 ‰ chitosan-immobilized lipase are added in zymotic fluid, and constant temperature digests 0.8 ~ 2.5h, enzyme at 25 ~ 50 DEG C
Solution terminates rear enzyme deactivation, is filtrated to get degreasing peony pollen polypeptide liquid.
5. the method that fermentation method according to claim 4 prepares peony pollen polypeptide, it is characterised in that：Described shell
The preparation process of chitosan-immobilized lipase is：With Chitosan powder is dried, with 60 ~ 80mmol/L phosphate of pH6.0 ~ 6.8
Buffer solution soaked overnight, then add the 1- ethyls -3- of Chitosan powder quality 0.7 ~ 1.2%（3- methylaminos）Propyl group carbon two
Imines activates 0.5 ~ 0.9h, adds the glutaraldehyde 1 ~ 1.8h of coupling reaction of 1 ~ 2 times of volume of Chitosan powder, is cleaned with distilled water
5 ~ 8 times, 0.7 ~ 2.0mg/mL lipase solutions and 0.1 ~ 0.3 ‰ bioactive peptides are added, 0.5 ~ 1.5h is slowly stirred in ice bath
, refrigerator overnight, secondary daily distilled water cleaning down are then placed, filtration drying obtains chitosan-immobilized lipase.
6. the method that fermentation method according to claim 5 prepares peony pollen polypeptide, it is characterised in that：Described work
The amino acid sequence of property small peptide is SCASVCKARCLRARGCRCVSVYCRCLR.
7. the method that fermentation method according to claim 1 prepares peony pollen polypeptide, it is characterised in that：Described table
Face activating agent is food grade surfactant.
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| CN109207550A (en) * | 2018-11-22 | 2019-01-15 | 哈尔滨氧态健康科技股份有限公司 | A kind of preparation method of peony seeds polypeptide |
| CN110951809A (en) * | 2019-09-04 | 2020-04-03 | 内蒙古立泰锐晟智能科技有限公司 | Process for preparing quinoa wheat polypeptide powder |
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| CN114058665A (en) * | 2021-12-02 | 2022-02-18 | 浙江宜格企业管理集团有限公司 | Preparation method of composite flower peptide and application of composite flower peptide in preparation of cosmetics |
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| CN114767566A (en) * | 2022-04-12 | 2022-07-22 | 广州优理氏生物科技有限公司 | Peony seed protein powder, preparation method, yeast repair essence cream and application |
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| CN114410724B (en) * | 2022-03-02 | 2023-09-15 | 厦门元之道生物科技有限公司 | Paeonia ostii peptide with tyrosinase activity inhibiting function and preparation method and application thereof |
| CN114767566A (en) * | 2022-04-12 | 2022-07-22 | 广州优理氏生物科技有限公司 | Peony seed protein powder, preparation method, yeast repair essence cream and application |
| CN114767566B (en) * | 2022-04-12 | 2022-11-29 | 广州优理氏生物科技有限公司 | Peony seed protein powder, preparation method, yeast repair essence cream and application |
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Application publication date: 20171229 |