CN107722114A - A kind of polypeptide and its application - Google Patents
A kind of polypeptide and its application Download PDFInfo
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- CN107722114A CN107722114A CN201711004943.1A CN201711004943A CN107722114A CN 107722114 A CN107722114 A CN 107722114A CN 201711004943 A CN201711004943 A CN 201711004943A CN 107722114 A CN107722114 A CN 107722114A
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- 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
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- 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
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/30—Feeding-stuffs specially adapted for particular animals for swines
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0014—Skin, i.e. galenical aspects of topical compositions
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/08—Solutions
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/10—Dispersions; Emulsions
- A61K9/12—Aerosols; Foams
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- C—CHEMISTRY; METALLURGY
- 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
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
Abstract
A kind of polypeptide and its application, the polypeptide of the present invention, contain the continuous amino acid residue segment that sequence is Pro Gly Ala Asp or Asp Ala Gly Pro.Can be synthesized according to its sequential structure using Peptide synthesizer, also can from natural plants pseudo-ginseng, sophora flower, aloe and or the bletilla striata in extract and obtain.The polypeptide of the present invention has the function that to promote mouse stem cells and vascular progenitor migration and to vascular endothelial cell directed differentiation, so as to the formation for promoting the injury repair of vascular endothelial cell, promoting new vessels.Skin spray is prepared into, wound healing, preventing and treating inflammation can be promoted；Piglet by Oral liquid is prepared into, piglet intestinal villus can be promoted to regenerate and repair, fundamentally realize the prevention and radical cure of grice diarrhoea disease；Additive for animal feed is prepared into, can anti diar rhea and growth promotion；Gilt Uterus filling agent is prepared into, sow postpartum wound healing can be promoted, effectively prevents and treats sow inflammation.
Description
The application category divisional application.
The original application applying date：On 07 07th, 2016, application number：201610528672.9 denomination of invention：A kind of polypeptide
And its application.
Technical field
The present invention relates to a kind of polypeptide and its application.
Background technology
In Swine Production, disconnected navel is required for after all pig births, most of piglet can all dock, and the pig of half needs to go
Gesture, while gregarious fight, tail biting are faced, mechanical damage, hernia, prolapse of the anus suture operation etc..It is if right not in time in Swine Production
Caused wound is handled, and will cause the infection of the pathogen such as bacterium, virus, can trigger local inflammation, redness and concurrent
Illness, so as to influence pig growth performance, carcase quality declines or even causes death.Especially sow postpartum wound is handled not in time,
Often easily trigger the inflammation such as Gilt Uterus inflammation, vaginitis, cause Repeat breeding, miscarried after pregnant, suppurated during heat or before breeding
The normal production of phenomenon, directly harm kind swinery, increase the mortality of boar, reduce the utilization rate of kind of swinery.
At present, most of pig farms are not handled for minor cut or wound of pig, some pig farms even hog, castration, seam anus, docking,
Wound will not also be handled during disconnected navel, most of pig farm processing wound is namely disappeared with the tincture of iodine, alcohol even safflower oil
Poison.The tincture of iodine, alcohol, safflower oil play the role of sterilization, but have stimulation to wound simultaneously, cause pain, and have obstruction wound
The effect of healing.Pig farm also is sprinkling upon antibiotic on wound after piglet is castrated, and can play a part of sterilization, but not
Promote the effect of wound healing, often lead to swinery disease infection rate height, resistance is poor, growth retardation.
Control sow postpartum inflammation be one of key link in Swine Production, its be related to the production efficiency on whole pig farm and
Productivity.Sow postpartum uterus and birth canal have different degrees of damage, and pathogenic microorganism is easily invaded and bred, most pig farms
Sow postpartum treatment of wounds is carried out using injection of antibiotics or perfusion antibiotics, but with the generation of bacterial drug resistance,
The method is difficult to obtain very significant effect, and antibiotics can not rapidly promote wound healing, during wound healing
Between it is longer, the probability of its infection inflammation is then higher, and antibiotics has been difficult to fundamentally solve sow postpartum inflammation etc. at present
Problem.
On the other hand, grice diarrhoea is as one of high morbidity in Swine Production, be influence piglet growth performance and piglet into
One of an important factor for motility rate.It has that cause of disease species is various, and the state of an illness is complicated, and high mortality, the morbidity duration is long, cures
The features such as rate is low, recurrent exerbation.From clinicopathological features, grice diarrhoea caused by different pathogeny with gut epithelium tissue by
The symptom such as damage, intestinal villus come off, small intestine epithelium malabsorption, so as to cause that large quantity of moisture and electrolyte, excrement are gathered in enteron aisle
Discharged in soft or liquid.Currently for the preventing and treating of grice diarrhoea, mainly start with terms of restraining with antibacterial two, it is difficult to reach
To effective control and the effect cured completely, especially it is also easy to produce drug resistance using antibiotic, reduces swinery growth performance etc..
The content of the invention
The technical problem to be solved in the present invention is overcome the deficiencies in the prior art, there is provided have promote wound healing,
Prevent inflammation generation caused by wound infection, fast quick-recovery and cure intestinal tract injury, prevent and treat polypeptide and its application of grice diarrhoea.
The present invention solves the technical scheme that its technical problem uses,
The polypeptide of the present invention, contain the continuous amino acid residue piece that sequence is Pro-Gly-Ala-Asp or Asp-Ala-Gly-Pro
Section.
The polypeptide is preferably made up of 7-15 amino acid residue.
The polypeptide, preferably include following（1）Any one amino acid sequence that ~ (10) represent：
（1）Ala-Met-His-Thr-Pro-Gly-Ala-Asp
（2）Met-Lys-Ser-His-Pro-Gly-Ala-Asp-Lys
（3）Lys-Pro-Gly-Ala-Asp-Met-His-Ser-Asp
（4）Met-Ser-His-Pro-Gly-Ala-Asp-Lys-Lys
（5）Lys-Met-Asp-Met-His-Pro-Gly-Ala-Asp-Ser
（6）Leu-Leu-Asp-Ala-Gly-Pro-Ser-His-Met-Ala
（7）Met- Arg-Ser- Pro-Gly-Ala-Asp-Lys-Leu-Lys-Lys
（8）Val-Ala-His-Ser-Pro-Gly-Ala-Asp-Ala-Leu-Glu-Gly-Lys-Leu
（9）Ala-Val-Met-His-Ser-Pro-Gly-Ala-Asp-Leu-Ile
(10) Asp-Phe-Leu-Ala-Met-His-Ser-Pro-Gly-Ala-Asp-Leu-Tyr-Trp-Ala。
Polypeptide of the present invention can be synthesized according to its sequential structure using Peptide synthesizer.
The polypeptide can from natural plants pseudo-ginseng, sophora flower, aloe and or the bletilla striata in extract and obtain.From natural plants pseudo-ginseng,
Sophora flower, aloe and or the bletilla striata in extract the method for the polypeptide, comprise the following steps:
（1）By the bud or flower piece or Chinese aloe leaf of plant material Roots of Panax Notoginseng or plant Chinese scholartree or bletilla striata rhizome or its combination, cut
Broken, homogenate, the Tris-Hcl that the pH for adding 0.04-0.06moL/L of 5-10 times equivalent to plant material volume is 7.5-8.5 delays
Solution is rushed, is ultrasonically treated 3 ~ 5min, centrifugation is removed slag, and takes supernatant, i.e. vegetable protein crude extract；
（2）By step（1）Gained vegetable protein crude extract ammonium sulfate precipitation, 1-4 DEG C of left overnight, is then centrifuged for, it is heavy to take
Form sediment, freeze-drying, obtain vegetable protein meals；
（3）Vegetable protein meals are taken, are dissolved with distilled water, it is 7.5-8.0 to adjust pH with 4-5 mol/L NaOH solution, is added
Equivalent to vegetable protein meals weight 0.5-1 ‰ complex enzyme composition, the complex enzyme composition is alkali protease with
Property protease in mass ratio 1:1-1:3 mixing；It is placed in 50-55 DEG C of thermostat water bath, digests 2.5-4h, enzymolysis terminates, by enzyme
Solution liquid boils 3 ~ 5min and carries out destroy the enzyme treatment；Room temperature is subsequently cooled to, is centrifuged, is taken supernatant, produce polypeptide crude extract；
（4）Polypeptide crude extract is taken, gel filtration chromatography is carried out with sephadex Sephadex G25, uses 0.04-0.05moL/L
PH be 7.5-8.0 Tris-Hcl cushioning liquid elute, Fractional Collections eluent, and to the bioactivity of each several part eluent
It is measured；
（5）Active eluent part is collected, is further purified with RPLC, obtains the more of high-purity
Peptide.
The polypeptide valid density is extremely low, is configured to the aqueous solution, polypeptide in skin spray, oral liquid and uterine perfusion agent
Valid density is 5-15 μ g/mL.
The polypeptide is prepared into skin spray, has the effect of promoting wound healing, effectively reduces wound infection
Risk；Oral liquid is prepared into, piglet intestinal villus can be promoted to regenerate and repair, realize effective preventing and treating of grice diarrhoea disease, can be
Rapid healing grice diarrhoea and no longer recurred in 3-5 days；Gilt Uterus filling agent is prepared into, sow postpartum wound can be promoted quick
Healing, effectively prevent and treat sow inflammation.Can be additionally used in application on human skin external application, for alleviate scytitis, fungal infection disease of skin and
Skin wound is promoted quickly to heal.
The polypeptide is added in feed as feed addictive to be applied, and is had and is promoted growth of animal and effective pre- anti-diarrhea
Effect.Applied in animal feed, polypeptide effective content is 1 ~ 30 μ g/kg in animal feed.
The present invention has found a variety of knots contained in pseudo-ginseng, sophora flower, aloe and the bletilla striata by being screened to thousands of kinds of plants
The peptide material of structure sequence, which has, promotes mouse stem cells and vascular progenitor migration and to vascular endothelial cell directed differentiation
Effect, so as to promote the injury repair of vascular endothelial cell, promotes the formation of new vessels.Skin spray is prepared into, is had
Play the role of to promote wound healing, preventing and treating inflammation, be prepared into Piglet by Oral liquid, have what is promoted the regeneration of piglet intestinal villus and repair
Effect, fundamentally realizes the prevention and radical cure of grice diarrhoea disease.
The present invention has the positive effect that：First, the polypeptide can be artificial synthesized, production cost is low, can extensive use and
Exploitation；Second, the polypeptide mechanism of action is clear and definite, there is huge medicinal potentiality to be exploited；Third, the polypeptide skin spray is used for
Swinery Wound healing, effect is very notable, can reduce swinery disease infection risk, greatly improve the pig farm level of production；Fourth, institute
Sow postpartum uterus wound healing can be rapidly promoted by stating polypeptide uterine perfusion agent, effectively reduce the generation of sow inflammation, improve kind
Pig utilization rate, improve the production efficiency on pig farm；Fifth, the polypeptide oral liquor, have and promote the reparation of piglet intestinal villus and regeneration
Effect, grice diarrhoea can be effectively prevented and treated, improve piglet survival ratio and growth performance, lift pig farm economic benefit；It is sixth, described more
Peptide valid density is extremely low, and use cost is low.And the polypeptide be natural plants in find, have it is green safe, efficient, without resistance
Property, noresidue, the characteristics of having no toxic side effect.
Brief description of the drawings
Fig. 1 and Fig. 2 is to observe to be calculated through in plant polypeptide promotion mouse vascular progenitor migration test in embodiment 3
The migrating cell number of 0.8 micron membranes；*:p<0.05,** :<0.01；
Fig. 3 and Fig. 4 is that plant polypeptide promotes VPC cells to break up situation map to vascular endothelial cell direction in embodiment 3；*:p<
0.05,** :<0.01；
Fig. 5 and Fig. 6 is the integrality of the Vascular Endothelial of embodiment 4 and the formational situation figure of new intima；Fig. 5 is representative diagram
Picture, Fig. 6 are the average relative thickness of the vascular wall theca interna+hypothallus of every group of six injured blood vessels；*:p<0.05,** :<
0.01；
Fig. 7 and Fig. 8 is the blood flow state figure of mouse bilateral hind leg in embodiment 4；Fig. 7 is presentation graphics, and Fig. 8 is every group six
The right side of mouse（Ligation）/ left side（Control）The percentage of blood flow ratio average；*:p<0.05,** :<0.01；
Fig. 9 is CD31 immunofluorescence dyeing results in embodiment 5；
Figure 10 is every group of three mouse in embodiment 5, the average value of capillary in every 10 400x visuals field；*:p<0.05；
Figure 11 is the influence figure of wound healing when polypeptide is to piglet castration in embodiment 6（When 1-7 rows are respectively piglet castration,
Control group 24h, test group 24h, the h of control group 48, the h of test group 48, the h of control group 72, the h of test group 72 situation map）.
Embodiment
The invention will be further described with reference to embodiments.
Embodiment 1
（1）Take natural plants Roots of Panax Notoginseng to shred, be homogenized, the pH for adding 0.06moL/L of 8 times equivalent to plant material volume is
8.0 Tris-Hcl cushioning liquid, is ultrasonically treated 4min, and centrifugation is removed slag, takes supernatant, produce Roots of Panax Notoginseng protein crude extract；
（2）By step（1）Gained Roots of Panax Notoginseng protein crude extract ammonium sulfate precipitation, 4 DEG C of left overnights, is then centrifuged for, it is heavy to take
Form sediment, freeze-drying, obtain Roots of Panax Notoginseng protein meals；
（3）Roots of Panax Notoginseng protein meals are taken, are dissolved with distilled water, is 8.0 with 5 mol/L NaOH solution regulation pH, adds suitable
0.5 ‰ complex enzyme composition is weighed in vegetable protein meals, the complex enzyme composition is alkali protease and neutral proteinase
In mass ratio 1:2 mixing；It is placed in 50 DEG C of thermostat water baths, digests 3h, enzymolysis terminates, and enzymolysis liquid is boiled into 3min is gone out
Ferment treatment；Room temperature is subsequently cooled to, is centrifuged, is taken supernatant, produce polypeptide crude extract；
（4）Polypeptide crude extract is taken, gel filtration chromatography is carried out with sephadex Sephadex G25, is with 0.05moL/L pH
8.0 Tris-Hcl cushioning liquid elution, Fractional Collections eluent, and the bioactivity of each several part eluent is measured；
（5）Active eluent part is collected, is further purified with RPLC, obtains the more of high-purity
Peptide.Through further screening active ingredients, its active peptides sequence is determined, obtains the active peptides of following amino acid sequence structure：
（4）Met-Ser-His-Pro-Gly-Ala-Asp-Lys-Lys
（5）Lys-Met-Asp-Met-His-Pro-Gly-Ala-Asp-Ser
（6）Leu-Leu-Asp-Ala-Gly-Pro-Ser-His-Met-Ala
（7）Met- Arg-Ser- Pro-Gly-Ala-Asp-Lys-Leu-Lys-Lys.
Embodiment 2
（1）Plant material Chinese aloe leaf is shredded, is homogenized, the pH for adding 0.06moL/L of 6 times equivalent to plant material volume is
8.5 Tris-Hcl cushioning liquid, 3.5min is ultrasonically treated, centrifugation is removed slag, and takes supernatant, i.e. Chinese aloe leaf protein crude extract；
（2）By step（1）Gained Chinese aloe leaf protein crude extract ammonium sulfate precipitation, 4 DEG C of left overnights, is then centrifuged for, it is heavy to take
Form sediment, freeze-drying, obtain Chinese aloe leaf vegetable protein meals；
（3）Take step（2）Gained Chinese aloe leaf protein meals, are dissolved with distilled water, are with 4mol/L NaOH solution regulation pH
8.0, the complex enzyme composition equivalent to Chinese aloe leaf protein meals weight 0.6 ‰ is added, the complex enzyme composition is alkaline egg
White enzyme and neutral proteinase in mass ratio 1:2 mixing；It is placed in 50 DEG C of thermostat water baths, digests 3.5h, enzymolysis terminates, by enzyme
Solution liquid boils 3min and carries out destroy the enzyme treatment；Room temperature is subsequently cooled to, is centrifuged, is taken supernatant, produce aloe polypeptide crude extract；
（4）Polypeptide crude extract is taken, gel filtration chromatography is carried out with sephadex Sephadex G25, is with 0.05moL/L pH
8.0 Tris-Hcl cushioning liquid elution, Fractional Collections eluent, and the bioactivity of each several part eluent is measured；
（5）Active eluent part is collected, is further purified with RPLC, obtains the more of high-purity
Peptide.Bioactivity screening is further carried out, active peptides are carried out with sequential structure measure, the final sequence for obtaining active aloe polypeptide
Array structure is as follows：
（4）Met-Ser-His-Pro-Gly-Ala-Asp-Lys-Lys
（5）Lys-Met-Asp-Met-His-Pro-Gly-Ala-Asp-Ser
（8）Val-Ala-His-Ser-Pro-Gly-Ala-Asp-Ala-Leu-Glu-Gly-Lys-Leu
（9）Ala-Val-Met-His-Ser-Pro-Gly-Ala-Asp-Leu-Ile
(10) Asp-Phe-Leu-Ala-Met-His-Ser-Pro-Gly-Ala-Asp-Leu-Tyr-Trp-Ala。
With above-mentioned same extracting method, the polypeptide fragment of following sequential structure is obtained respectively from the bletilla striata and sophora flower：
Obtained from the bletilla striata：
（1）Ala-Met-His-Thr-Pro-Gly-Ala-Asp
（2）Met-Lys-Ser-His-Pro-Gly-Ala-Asp-Lys.
Obtained from sophora flower：
（3）Lys-Pro-Gly-Ala-Asp-Met-His-Ser-Asp.
Embodiment 3
Plant polypeptide induction of vascular progenitor cells（VPC）Migrate and be directed differentiation to vascular endothelial cell
By vascular progenitor（VPC）The migration aperture in 0.8 micron of aperture is inoculated in, base is separately added into the control containing 1ng/mL
The sequential structure obtained in the common polypeptide of group, the 1ng/mL bletilla striatas of the present invention from natural plants（1）And sequential structure（2）（Matter
Amount is than being 1:2）Plant polypeptide composition, the culture medium of 10ng/mL blood vessel endothelial cell growth factor VEGFs, after 16 hours
Observation is calculated through the migrating cell number of 0.8 micron membranes.As a result Fig. 1 and Fig. 2 are seen.VEGF is that generally acknowledged having promotes blood vessel endothelium
So as to promote the growth factor that new vessels is formed, Fig. 1 and Fig. 2 show sequence of the present invention for cell differentiation, propagation and migration
Structure（1）Plant polypeptide promotes mouse vascular progenitor in the experiment of transwell migration models（VPC）Migration, its effect
VEGF is substantially better than, and compares the effect that polypeptide does not promote VPC cell migrations.
By vascular progenitor（VPC）The migration aperture in 0.8 micron of aperture is inoculated in, base is separately added into containing 0.1ng/mL
The common polypeptide of control group, 0.1ng/mL sequential structures of the present invention（1）And sequential structure（2）（Mass ratio is 1:2）Plant
After being cultivated 7 days in peptide composition, 10ng/mL VEGF cell differential mediums, the vascularization of analysis tool inner skin cell function
Situation, as a result see Fig. 3 and Fig. 4.It is the exclusive feature of vascular endothelial cell that tubular structure is formed on cell differential medium, often
It is usually used in functional selection of the stem cell into vascular endothelial cell differentiation research.Fig. 3 and Fig. 4 shows sequence knot of the present invention
Structure（1）And sequential structure（2）Plant polypeptide can be obviously promoted vascular progenitor and form tubular structure, its significant effect is better than
VEGF（P＜ 0.01）, prompt it to promote the directed differentiation of vascular endothelial cell.
Comprehensive two results showed thats are in vitro in culture experiment, sequential structure of the present invention（1）And sequential structure
（2）Plant polypeptide mouse vascular progenitor can be promoted to migrate and to vascular endothelial cell directed differentiation.
Embodiment 4
Plant polypeptide promotes the Reperfu- sion of blood vessel endothelium injury reparation and mouse hind leg ischemia model blood
With platinum wire ring, scraping damage right side of mice femoral artery blood vessel endothelium establishes mouse blood vessel endothelium injury model back and forth, in quilt
It is separately added into the common polypeptide of the control group containing 10ng/mL, the sophora flower of the present invention from natural plants and obtains on the outside of injured blood vessel
The sequential structure obtained（3）Plant polypeptide, VEGF, taken after 4 weeks and be damaged blood vessel, cut into slices, the complete of blood vessel endothelium is observed in HE dyeing
The formational situation of property and new intima, is as a result shown in Fig. 5 and Fig. 6.
Mouse hind leg ischemia model is established by ligaturing and cutting right side of mice femoral artery, is being damaged on the outside of blood vessel respectively
Add the control polypeptide containing 10ng/mL, sequential structure of the present invention（3）Plant polypeptide, VEGF, using colored more after 2 weeks
The general blood flow state for strangling scanner detection mouse bilateral hind leg.As a result Fig. 7 and Fig. 8 are seen.
As shown in Figure 5 and Figure 6, in In-vivo test in mice, sequential structure of the present invention（3）Plant polypeptide can promote
The injury repair of mouse femoral artery damage model blood vessel and the formation of angiogenesis inhibiting inner membrance and plaque.Institute of the present invention
State sequential structure（3）Plant polypeptide promotes the integrality of blood vessel endothelium reparation suitable with VEGF, meanwhile, in the angiogenesis of formation
The thickness of film is far below control group and VEGF groups.From Fig. 7 and Fig. 8, the plant polypeptide group mouse ischemic acra blood of the present invention
Stream ratio average be far above control group and VEFG groups, shows in its promotion mouse hind leg ischemia model new green blood in ischemic tissue
The formation of pipe is and more more effective than VEGF so as to promote the Reperfu- sion of ischemic acra blood.
The experiment of this group shows that plant polypeptide of the present invention has in vivo and promotes blood vessel endothelium injury reparation and new green blood
The effect that pipe is formed.
Embodiment 5
Plant polypeptide promotes the formation of new vessels in heart infarction slough.
Mouse heart infarction model is established by ligaturing mouse left ventricular coronary artery.It is injected separately into mouse heart infarction region
The sequence obtained is extracted from natural plants pseudo-ginseng and aloe in 10ng/mL control polypeptide, Example 1 and Example 2 of the present invention
Array structure（4）And sequential structure（5）（Mass ratio is 1:3）Plant polypeptide composition and VEGF, after 30 days, to mouse left ventricle
Cardiac muscle sections, and carry out immunofluorescence dyeing observation CD31 positive cells（Predominantly vascular endothelial cell）.As a result figure is seen
9。
CD31 immunofluorescence dyeing results as shown in Figure 9（Artwork green is CD31 positive cells, and blueness is DAPI dyeing
Nucleus）, plant polypeptide group CD31 positive cell quantities are significantly more than control group and VEGF groups, show sequence knot of the present invention
Structure（4）And sequential structure（5）（Mass ratio is 1:3）Plant polypeptide composition have promote heart infarction slough Vascular Endothelial
The formation of cell differentiation and new vessels.
Embodiment 6
Polypeptide spray is used for effect observation during piglet castration
After piglet castration, of the present invention structure sequence of the spray containing 10 μ g/mL on its wound（6）The skin spray of polypeptide
1mL, recovery situation of the wound in 24,48,72 hours is observed.Blank physiological saline of the control group spray without polypeptide is molten
Liquid.As a result Figure 11 is seen.
Sprayed after test group castration in wound containing of the present invention by Peptide synthesizer composite structure sequence（6）Polypeptide
Spray, blank physiological saline solution of the control group spray without polypeptide, is carried out piglet castration 24,48,72 hours to wound respectively
Take pictures, as can be seen from Figure 11, compared with control group, using after 24 hours, wound location redness is bright after polypeptide spray of the present invention
It is aobvious to disappear；After 48 hours, speed of wound healing is substantially accelerated；Using 72 hours, wound healed completely.Its speed of wound healing
It significantly faster than control group, can completely be healed a wound in 2 ~ 3 days, show that polypeptide of the present invention has and rapidly promote wound and be cured
The effect of conjunction.
Embodiment 7
Influence of the polypeptide to sow postpartum inflammation and reproductive performance
Experiment is using 108 first-born production sows as subjects, to of the present invention by Peptide synthesizer composition sequence structure（7）With
（8）（Mass ratio 1:1）Peptide composition uterine perfusion agent is studied the prevention and control of sow inflammation and the influence of production performance.Selection
108 sows within expected date of childbirth interval 10 days are randomly divided into two groups of test group and control group, tested as subjects
Group sow postpartum uterine perfusion polypeptide uterine perfusion agent of the present invention（Content of peptides is 8 μ g/mL）, each 100mL, daily 1
It is secondary, it is used continuously 3 days.Control group takes routine healthcare method, and wound is smeared or perfusion Conventional antibiotic class uterine perfusion agent
Continuous use 3 days.Experiment terminated since the 1st day sow postpartum after weaning 7 days to sow.
, mammitis incidence scorching to Gilt Uterus during experiment and wean heat situation count, and the results are shown in Table 1.
Mechanical damage easily causes bacterium infection in sow reproductive process, and triggers inflammation to cause treating sow metritis, mammitis etc..
Treating sow metritis will influence sow life post-partum estrus wean interval time, Oestrus rate and rutting rate, when serious even
Sow is caused to be eliminated in advance.As shown in Table 1, compared with control group, test group hysteritis and incidence of mammits are greatly reduced,
Wean diestrum shorten, wean 7 days in Oestrus rate significantly improve, show polypeptide of the present invention can effectively reduce sow production
The incidence of inflammation afterwards, further improve the heat situation of sow.
Embodiment 8
Effect of the polypeptide oral liquor to preventing and treating grice diarrhoea
It is of the present invention by Peptide synthesizer composition sequence structure（9）With（10）Polypeptide（Mass ratio is 1:2）Composition prepare and
Into Piglet by Oral liquid, contrast test is carried out to the treatment of diarrhea of weaned piglets.Grice diarrhoea sample is to include wean to answer
Swash, turn group, reload, grice diarrhoea caused by factor, the test group illness piglet such as change in weather and bacterium infection gavage this hair
The bright polypeptide oral liquor（Content of peptides is 15 μ g/mL）, each 2-3mL, 1 time a day, it is used continuously 3-5 days, control group is adopted
With conventional antibiotic treatment, test data statistical result is shown in Table 2.
Weanling pig, which reloads, turns group, Changes in weather and bacteriovirus infection easily causes grice diarrhoea, and grice diarrhoea, which has, to be infected
Property and be difficult to radical-ability, and piglet growth performance is had a strong impact on, if treating not in time, it is easy to cause piglet dead, to pig
Field and raiser bring huge economic loss.As shown in Table 2, using polypeptide oral liquor of the present invention, can quickly control and
Grice diarrhoea is cured, cure rate more than 97% is realized using 3 days interior energies, and recurrence rate is 0, and it is cured speed and cured completely
Rate is significantly larger than normal antibiotics treatment, and the state of an illness does not recur, and grice diarrhoea is thoroughly effected a radical cure.
The additive of polypeptide feed growth promotion of embodiment 9 and anti-diarrhea effect
Selection 1100 wean turn colony weight similar in child care pig stochastic averagina be divided into test group and control group.Experimental period is from son
Pig is transferred to nursery house and starts to producing nursery house to terminate, totally 70 days.The whole feed addition per ton of test group is containing described in embodiment 1
The feed addictive of polypeptide, addition concentration is that content of peptides is 10mg in feed per ton.Other rearing conditions are consistent with control group.
Additive of polypeptide feed influences to be shown in Table 3 on child care pig growth performance.
As shown in Table 3, test group child care daily gain in pigs, survival rate, qualification rate are all improved significantly, child care pig child care stage material
Meat ratio, diarrhea rate significantly reduce, and 550 piglet cradling stages of test group only have 2 hairs life diarrhoea, and child care pig health degree substantially changes
Kind, experiment shows, polypeptide described in embodiment 1, which has, promotes piglet growth, the effect of prevention of diarrhea in piglets.
Sequence table
<110>Hu Guotian, Zeng Lingfang
<120>A kind of polypeptide and its application
<160> 10
<170> SIPOSequenceListing 1.0
<210> 1
<211> 8
<212> PRT
<213>Artificial sequence ()
<400> 1
Ala Met His Thr Pro Gly Ala Ala
1 5
<210> 2
<211> 9
<212> PRT
<213>Artificial sequence ()
<400> 2
Met Leu Ser His Pro Gly Ala Ala Leu
1 5
<210> 3
<211> 9
<212> PRT
<213>Artificial sequence ()
<400> 3
Leu Pro Gly Ala Ala Met His Ser Ala
1 5
<210> 4
<211> 9
<212> PRT
<213>Artificial sequence ()
<400> 4
Met Ser His Pro Gly Ala Ala Leu Leu
1 5
<210> 5
<211> 10
<212> PRT
<213>Artificial sequence ()
<400> 5
Leu Met Ala Met His Pro Gly Ala Ala Ser
1 5 10
<210> 6
<211> 10
<212> PRT
<213>Artificial sequence ()
<400> 6
Leu Leu Ala Ala Gly Pro Ser His Met Ala
1 5 10
<210> 7
<211> 11
<212> PRT
<213>Artificial sequence ()
<400> 7
Met Ala Ser Pro Gly Ala Ala Leu Leu Leu Leu
1 5 10
<210> 8
<211> 14
<212> PRT
<213>Artificial sequence ()
<400> 8
Val Ala His Ser Pro Gly Ala Ala Ala Leu Gly Gly Leu Leu
1 5 10
<210> 9
<211> 11
<212> PRT
<213>Artificial sequence ()
<400> 9
Ala Val Met His Ser Pro Gly Ala Ala Leu Ile
1 5 10
<210> 10
<211> 15
<212> PRT
<213>Artificial sequence ()
<400> 10
Ala Pro Leu Ala Met His Ser Pro Gly Ala Ala Leu Thr Thr Ala
1 5 10 15
Claims (1)
1. a kind of polypeptide, it is characterised in that its amino acid sequence is：
Lys-Pro-Gly-Ala-Asp-Met-His-Ser-Asp。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201711004943.1A CN107722114A (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201610528672.9A CN106046130B (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
| CN201711004943.1A CN107722114A (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201610528672.9A Division CN106046130B (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
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| CN107722114A true CN107722114A (en) | 2018-02-23 |
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| CN201610528672.9A Expired - Fee Related CN106046130B (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
| CN201711004942.7A Pending CN107602682A (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
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| CN201711004942.7A Pending CN107602682A (en) | 2016-07-07 | 2016-07-07 | A kind of polypeptide and its application |
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| CN107279460A (en) * | 2017-06-27 | 2017-10-24 | 兰溪市酉泽饲料技术服务有限公司 | Penaeus Vannmei fermented feed |
| CN109321622B (en) * | 2017-08-01 | 2021-10-12 | 北京善肽堂生物科技有限公司 | Preparation method and application of pseudo-ginseng polypeptide |
| CN111110595B (en) * | 2020-01-21 | 2023-12-12 | 常明泉 | Vegetable protein emulsifiable paste, preparation method and application thereof |
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| US20130101628A1 (en) * | 2011-04-29 | 2013-04-25 | Northwestern University | Novel vegf mimetic peptide-based scaffolds for therapeutic angiogenesis and methods for their use |
| CN103082136B (en) * | 2013-01-15 | 2014-01-08 | 中北大学 | Pig feed additive |
| EP2983682A4 (en) * | 2013-04-12 | 2017-03-08 | Mangogen Pharma Inc. | Therapeutic viral microparticles for promoting stent biofunctionality and wound healing in vertebrate individuals |
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- 2016-07-07 CN CN201711004943.1A patent/CN107722114A/en active Pending
- 2016-07-07 CN CN201610528672.9A patent/CN106046130B/en not_active Expired - Fee Related
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| WO2001085193A2 (en) * | 2000-05-08 | 2001-11-15 | Biogen, Inc. | Method for promoting neovascularization using a tweak agonist and an angiogenic factor |
| CN101516902A (en) * | 2006-07-31 | 2009-08-26 | 脉管生物生长有限公司 | Polypeptides and polynucleotides encoding same and use thereof in the treatment of medical conditions associated with ischemia |
| CN105254717A (en) * | 2015-08-18 | 2016-01-20 | 广州一代医药科技有限公司 | Polypeptide specifically binding to CD34 molecule and application thereof |
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Also Published As
| Publication number | Publication date |
|---|---|
| CN107602682A (en) | 2018-01-19 |
| CN106046130A (en) | 2016-10-26 |
| CN106046130B (en) | 2017-12-12 |
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