CN115974967B

CN115974967B - Oligopeptide POP1, and preparation method and application thereof

Info

Publication number: CN115974967B
Application number: CN202210973822.2A
Authority: CN
Inventors: 宋芹; 许丽佳; 袁海梅; 王健; 肖鑫; 李红梅
Original assignee: Chengdu University
Current assignee: Chengdu University
Priority date: 2022-08-15
Filing date: 2022-08-15
Publication date: 2023-09-26
Anticipated expiration: 2042-08-15
Also published as: CN115974967A

Abstract

The application relates to the field of biological medicine, in particular to an oligopeptide POP1, a preparation method and application thereof; the primary structure amino acid sequence of the oligopeptide POP1 is shown as SEQ ID NO. 1; the polypeptide comprises oligopeptide POP1 or pharmaceutically acceptable salt thereof; the preparation method comprises the following steps: crushing periplaneta americana, extracting with an organic solvent, and decoloring to obtain a concentrated solution; carrying out multiple chromatography and separation and purification on the concentrated solution to obtain a compound; vacuum freeze drying is carried out on the compound to obtain oligopeptide POP1; the application comprises: the oligopeptide POP1 is used for preparing medicines for treating wounds, scalds and ulcers; short peptides with the length of only 5 amino acids are screened and separated from the periplaneta americana, and the structure is simpler, so that the synthesis and the preparation are simpler and more convenient than common long peptides, and the synthesis of active ingredients can be simplified.

Description

Oligopeptide POP1, and preparation method and application thereof

Technical Field

The application relates to the field of biological medicine, in particular to an oligopeptide POP1, a preparation method and application thereof.

Background

With the increasing demand for wound skin repair, various biological agents for wound repair have been developed on a large scale, wherein, since repair and healing of the wound is a complex process, not only proliferation, differentiation and migration processes of various repair cells, but also a series of inflammatory reactions are required to be considered, and thus, the current wound repair needs to be coordinated by adopting various agents.

Peptide drugs have excellent properties in promoting wound healing, reducing excessive inflammatory reactions and reducing scars, and have been widely reported in wound repair. At present, a rehabilitation new liquid prepared from periplaneta americana serving as a raw material is adopted for repairing a wound difficult to heal, and in clinical practice, a good treatment effect of the rehabilitation new liquid is proved, but only the screening and analysis of peptide substances in the components with the wound repairing effect are required for the current extraction and analysis of the effective components of the periplaneta americana, the screening and analysis are long peptide substances generally, the research on the short peptide substances is lacking, and the synthesis and preparation of the long peptide substances are complex, so that how to provide the short peptide substances for wound repairing so as to simplify the synthesis of the effective components is a technical problem which needs to be solved at present.

Disclosure of Invention

The application provides an oligopeptide POP1, a preparation method and application thereof, and aims to solve the technical problem of lack of short peptide substances for wound surface repair in the prior art.

In a first aspect, the application provides an oligopeptide POP1, wherein the amino acid sequence of the primary structure of the oligopeptide POP1 is shown in SEQ ID NO. 1.

Optionally, the oligopeptide POP1 has a molecular weight of < 550Da.

Optionally, the nucleotide sequence of the oligopeptide POP1 is X1-X2-X3-X4-X5,

wherein X1 is CCU or CCC or CCA or CCG;

x2 is AAA or AAG;

x3 is GCU or GCC or GCA or GCG;

x4 is GAA or GAG;

x5 is GCU or GCC or GCA or GCG.

In a second aspect, the present application provides a polypeptide comprising the oligopeptide POP1 of the first aspect, or a pharmaceutically acceptable salt thereof.

In a third aspect, the present application provides a method for preparing the oligopeptide POP1, wherein the method is used for obtaining the oligopeptide POP1 in the first aspect from periplaneta americana, and the method comprises:

crushing periplaneta americana, extracting with an organic solvent, and decoloring to obtain a concentrated solution;

carrying out multiple chromatography and separation and purification on the concentrated solution to obtain a compound;

and carrying out vacuum freeze drying on the compound to obtain the oligopeptide POP1.

Optionally, the organic solvent includes at least one of ethanol, methanol, and acetone.

Optionally, the multiple chromatography includes a first chromatography, a second chromatography, and a third chromatography, the target molecular weight of the first chromatography > the target molecular weight of the second chromatography, and the third chromatography is used for separating the compound of the second chromatography according to a polarity size.

Optionally, the separation and purification comprises separation and purification with a molecular sieve under conditions less than a reference molecular weight, wherein the reference molecular weight is 1000Da.

In a fourth aspect, the present application provides an application of the oligopeptide POP1, wherein the oligopeptide POP1 in the first aspect is used for preparing a medicament for treating wounds, scalds and ulcers.

Optionally, the medicament comprises an external preparation and an injection preparation.

Compared with the prior art, the technical scheme provided by the embodiment of the application has the following advantages:

according to the oligopeptide POP1 provided by the embodiment of the application, the short peptide with the length of only 5 amino acids is screened and separated from the periplaneta americana, and the short peptide only contains four amino acids of proline, lysine, alanine and glutamic acid, and meanwhile, the whole short peptide only has five amino acids, so that the structure is simpler, and the synthesis and preparation are simpler and more convenient than the common long peptide, so that the synthesis of active ingredients can be simplified.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the application and together with the description, serve to explain the principles of the application.

In order to more clearly illustrate the embodiments of the application or the technical solutions of the prior art, the drawings which are used in the description of the embodiments or the prior art will be briefly described, and it will be obvious to a person skilled in the art that other drawings can be obtained from these drawings without inventive effort.

FIG. 1 is a schematic flow chart of a method according to an embodiment of the present application;

FIG. 2 is a standard amino acid alignment chart provided in an embodiment of the present application;

FIG. 3 is a blank pattern of a blank control test provided by an embodiment of the present application;

FIG. 4 is a first amino acid pattern of the N-terminus of oligopeptide POP1 provided in the examples of the present application;

FIG. 5 is a second amino acid pattern of the N-terminus of oligopeptide POP1 according to an embodiment of the present application;

FIG. 6 is a third amino acid pattern of the N-terminus of oligopeptide POP1 according to an embodiment of the present application;

FIG. 7 is a fourth amino acid pattern of the N-terminus of oligopeptide POP1 according to an embodiment of the present application;

fig. 8 is a high performance liquid chromatogram of oligopeptide POP1 provided in an embodiment of the present application;

FIG. 9 is a first-order mass spectrum of oligopeptide POP1 provided by the embodiment of the application;

fig. 10 is a schematic diagram of the basic structure of an oligopeptide POP1 according to an embodiment of the present application;

FIG. 11 is a schematic diagram showing the secondary mass spectrum and the assignment of peaks of oligopeptide POP1 according to the embodiment of the present application;

FIG. 12 is a mass spectrum of the solid phase synthesized oligopeptide POP1 provided by the embodiment of the application;

FIG. 13 is a schematic diagram showing the results of the generation of collagen from a wound surface of a mouse according to an embodiment of the present application;

FIG. 14 is a graph showing the comparative results of human skin fibroblast proliferation activity assay provided in the examples of the present application;

fig. 15 is a graph showing the comparison result of hydroxyproline content of human skin fibroblasts according to the embodiment of the present application.

Detailed Description

The advantages and various effects of the present application will be more clearly apparent from the following detailed description and examples. It will be understood by those skilled in the art that these specific embodiments and examples are intended to illustrate the application, not to limit the application.

Throughout the specification, unless specifically indicated otherwise, the terms used herein should be understood as meaning as commonly used in the art. Accordingly, unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. In case of conflict, the present specification will control.

Unless otherwise specifically indicated, the various raw materials, reagents, instruments, equipment and the like used in the present application are commercially available or may be prepared by existing methods.

The technical scheme provided by the embodiment of the application aims to solve the technical problems, and the overall thought is as follows:

in one embodiment of the application, an oligopeptide POP1 is provided, wherein the amino acid sequence of the primary structure of the oligopeptide POP1 is shown as SEQ ID NO.1 and is Pro-Lys-Ala-Glu-Ala.

In some alternative embodiments, the oligopeptide POP1 has a molecular weight of < 550Da.

In the embodiment of the application, the molecular weight of the oligopeptide POP1 is controlled below 1000Da, so that the oligopeptide POP1 can be fully described as a short peptide.

In some alternative embodiments, the oligopeptide POP1 has a nucleotide sequence of X1-X2-X3-X4-X5,

wherein X1 is CCU or CCC or CCA or CCG;

x2 is AAA or AAG;

x3 is GCU or GCC or GCA or GCG;

x4 is GAA or GAG;

x5 is GCU or GCC or GCA or GCG.

In the embodiment of the application, the arrangement of the nucleotide sequence of the oligopeptide POP1 is controlled, and the required nucleotide sequence can be amplified and converted into an expression vector by utilizing a molecular biology technology to obtain the oligopeptide POP1 shown in SEQ ID NO.1, so that the preparation of the oligopeptide POP1 can be realized in a biosynthesis mode.

Next, a polypeptide provided by an embodiment of the present application is described, where the polypeptide includes the oligopeptide POP1 described in the first aspect, or a pharmaceutically acceptable salt thereof.

Because the polypeptide described in the embodiments of the present application includes the oligopeptide POP1 provided in the foregoing embodiments of the present application, the composition of amino acids and the corresponding amino acid sequences in the oligopeptide POP1 are not described in detail herein. All polypeptides comprising the oligopeptide POP1 according to the embodiments of the present application are within the scope of the present application.

Next, as shown in fig. 1, a method for preparing an oligopeptide POP1 according to an embodiment of the present application is described, where the method is used to obtain the oligopeptide POP1 from periplaneta americana, and the method includes:

s1, crushing periplaneta americana, extracting with an organic solvent, and decoloring to obtain a concentrated solution;

s2, performing multiple times of chromatography on the concentrated solution, separating and purifying, and performing gradient elution to obtain a compound;

s3, performing vacuum freeze drying on the compound to obtain the oligopeptide POP1.

In the embodiment of the application, the oligopeptides can be gradually separated out by carrying out chromatography on the effective components in the periplaneta americana for multiple times, and then gradient elution is carried out on the oligopeptides, so that the purity of the screened oligopeptides is fully ensured.

Because the oligopeptide included in the preparation method described in the embodiment of the present application is the oligopeptide POP1 provided in the foregoing embodiment of the present application, the composition of amino acids and the corresponding amino acid sequences in the oligopeptide POP1 are not described in detail herein. All the preparation methods including the oligopeptide POP1 provided by the embodiment of the application belong to the scope of protection.

In some alternative embodiments, the organic solvent comprises at least one of ethanol, methanol, and acetone.

In the embodiment of the application, the types of the organic solvents are controlled, and the effective components in the periplaneta americana can be effectively extracted, so that the subsequent chromatography and separation force purification are convenient, and the purified oligopeptide is obtained.

In some alternative embodiments, the multiple chromatography comprises a first chromatography, a second chromatography, and a third chromatography, the multiple chromatography comprising a first chromatography, a second chromatography, and a third chromatography, the first chromatography having a target molecular weight > the target molecular weight of the second chromatography, the third chromatography being used to separate compounds of the second chromatography according to polarity size.

In the embodiment of the application, the particle size of the chromatographic column used for each chromatography is controlled to be gradually reduced, so that the chromatography effect can be improved, and the oligopeptide POP1 purified later can be obtained.

In some alternative embodiments, the separation and purification comprises separation and purification with a molecular sieve under conditions less than a baseline molecular weight, wherein the baseline molecular weight is 1000Da.

In the embodiment of the application, the reference molecular weight is controlled to be 1000Da, so that the oligopeptide meeting the small molecular weight can be screened out according to the standard.

Next, the application of the oligopeptide POP1 provided by the embodiment of the application will be described, and the oligopeptide POP1 is used for preparing a medicament for treating wounds, scalds and ulcers.

Because the application described in the embodiments of the present application includes the oligopeptide POP1 provided in the foregoing embodiments of the present application, the composition of amino acids and corresponding amino acid sequences in the oligopeptide POP1 are not described in detail herein. All applications including the oligopeptide POP1 in the embodiment of the application belong to the scope of the application to be protected.

In some alternative embodiments, the medicament includes an external preparation and an injection preparation.

In the embodiment of the application, the specific types of the medicines are controlled, and most forms of the medicines for wound repair, scald repair and ulcer repair can be covered.

Example 1

An oligopeptide POP1, wherein the amino acid sequence of the primary structure of the oligopeptide POP1 is shown as SEQ ID NO.1, and is Pro-Lys-Ala-Glu-Ala.

As shown in fig. 1, a preparation method of an oligopeptide POP1, the preparation method being used for obtaining the oligopeptide POP1 from periplaneta americana, the preparation method comprising:

extracting: extracting 5kg of periplaneta americana medicinal material with aqueous ethanol solution for 3 times, wherein the mass concentration of ethanol is 60% -90%, mixing the filtrates, recovering ethanol under reduced pressure until no ethanol smell exists, standing at room temperature for one night, and removing upper oily substance to obtain an extract.

Separating: taking the extracting solution, carrying out constant-temperature water bath to 60 ℃, adding 1% (mass/volume) active carbon, uniformly stirring by a magnetic stirrer, and filtering while the extracting solution is hot to obtain decolored concentrated solution.

Multiple column chromatography: first chromatography: taking a proper sample, separating by a Sephadex G25 chromatographic column (1.8 cm x 80 cm), eluting the sample by deionized water, collecting one part per 15mL, detecting peptide and amino acid by using thin layer chromatography on the collected part, combining similar parts containing the peptide according to detection results, respectively obtaining five groups of substances including SG1, SG2, SG3, SG4 and SG5 according to the molecular weight and the front-back sequence of the column, performing cell test on the five components, detecting fibroblast proliferation activity by using MTT (methyl thiazolyl tetrazolium), and the results show that: SG5> SG3> SG4> SG2> SG1, and adopting SG5 with the minimum molecular weight for further separation;

second chromatography: separating substances in SG5 group by using Sephadex G15 chromatographic column (1.8 cm. Times.80 cm), eluting sample of chromatographic column with deionized water, collecting one part per 10mL, detecting peptide and amino acid by using thin layer chromatography as a color developing agent and ninhydrin color developing agent as the collected fractions, combining similar fractions containing peptide according to detection result, respectively obtaining five groups of substances of SSG1, SSG2, SSG3, SSG4 and SSG5 according to molecular weight and front and back sequence of the column, performing cell test on all 5 components, and detecting fibroblast proliferation activity by MTT, wherein the result shows that: SSG5 is approximately equal to SSG4, SSG3, SSG2 and SSG1, the SSG5 is the most amount, SSG5 components are separated, and then vacuum drying is carried out to obtain SSG5 freeze-dried powder;

and (3) third chromatography: taking a sample SSG5 group of freeze-dried powder, dissolving in a small amount of water, separating by a C8 chromatographic column (3 cm x 40 cm), and performing gradient elution by using methanol water, wherein the method comprises the following steps of washing by using water, gradually increasing the dosage of methanol, collecting fractions, and performing specific gradient elution: 200mL of 100% water, 200mL of 80% water by volume: 20% methanol elution, 200mL volume ratio of 60% water: 40% methanol elution, 200mL 40% water by volume: 60% methanol elution, 200mL volume ratio of 20% water: 80% methanol, 200mL of 100% methanol by volume.

And (3) collecting the fractions by adopting thin-layer chromatography, wherein a developing agent is ninhydrin developing agent, detecting peptides and amino acids, merging similar fractions containing the peptides according to detection results, respectively obtaining seven components of substances of C1, C2, C3, C4, C5, C6 and C7 according to the polarity size and the sequence before and after column discharge, carrying out cell tests on the seven components, and adopting MTT to detect fibroblast proliferation activity, wherein the result shows that the C1 component is strongest under the same administration dosage.

Taking a C1 component sample, separating and purifying by reversed phase preparative HPLC, taking mobile phase water as A phase and acetonitrile as B phase, and performing gradient elution, wherein the gradient elution comprises the following procedures: washing with 5-17% of phase B for 0-9 min; washing the phase B of which the concentration is 17 to 30 percent in 9 to 17 minutes; washing with 30% -50% of phase B for 17-25 min; washing with 50-80% of phase B for 25-30 min; 30 min-35 min,80% -99%; washing with 99% phase B for 35-40 min), collecting the compound at 25deg.C, and vacuum freeze drying to obtain lyophilized powder of oligopeptide POP1.

Example 2

Example 2 and example 1 were compared, and the difference between example 2 and example 1 is that:

identifying the structure of the obtained oligopeptide:

1. the oligopeptide POP1 is subjected to amino acid sequencing, and the standard amino acid shown in the figure 2 is compared with a blank map shown in the figure 3, so that the first 4 amino acids of the oligopeptide POP1 are sequentially shown in figures 4 to 7,

2. under high resolution mass spectrum, as shown in FIG. 8, the oligopeptide POP1 has higher purity, and as shown in FIG. 9, the molecular weight of the oligopeptide is 514.28Da.

3. The secondary mass spectrum of oligopeptide POP1 was analyzed to obtain the assignment of the secondary mass spectrum fragment peaks as shown in FIG. 10.

The analysis of the above results shows that the amino acid sequence of the primary structure of oligopeptide POP1 is Pro-Lys-Ala-Glu-Ala as shown in FIG. 11.

Example 3

Example 3 and example 2 are compared, and the difference between example 3 and example 2 is that:

the synthesis of the active oligopeptide POP1 is completed by adopting a conventional solid-phase synthesis method of a polypeptide synthesizer, the synthesized product is checked by adopting a liquid chromatography-mass spectrometry method, the mass spectrometry analysis result is shown in figure 13, and the mass spectrum is compared with the separated oligopeptide POP1 shown in figure 9, so that the mass spectrum of the synthesized product is similar to that of the separated product.

Example 4

Example 4 and example 3 were compared, and the difference between example 4 and example 3 is that:

taking 40 male Kunming mice about 8 weeks, feeding in separate cages, randomly dividing into blank control group, positive drug group (rehabilitation new liquid), POP1 low dose group (0.1 mg/mL) and POP1 high dose group (0.2 mg/mL), anaesthetizing the mice with diethyl ether, dehairing and sterilizing the backs, placing 20g weight in boiling water for 10min, placing weight on dehaired skin of the mice for 8s, and scalding area of 4cm ² Becomes a shallow II degree scald model.

The next day, 0.1mL of POP1, normal saline and rehabilitation new solution with different concentrations are added dropwise, and the administration is carried out 1 time a day. And observing the wound healing condition at 6d, 12d and 18d, and calculating the wound healing rate according to the formula of the wound healing rate= (S1-Sn)/S1, wherein S1 is the first day wound area, and Sn is the nth day wound area. Tissue sections were prepared, the synthesis of collagen in wound tissues was observed by a polarized light microscope, and the results were photographed and shown in table 1.

TABLE 1

Note that: compared to the blank model group, the data and P <0.05 were expressed as significantly different.

According to the tissue slice conditions shown in Table 1 and FIG. 13, the active oligopeptide POP1 can obviously promote the generation of type I collagen, and the collagen is orderly and tightly arranged after 18 days, which is obviously superior to a blank control group and a rehabilitation new liquid group.

Example 5

Example 5 and example 4 were compared, and the difference between example 5 and example 4 is that:

the MTT assay was used to determine human skin fibroblast proliferation activity. Human skin fibroblasts were seeded in 24-well plates (104/well) and cultured, POP1 oligopeptides were added to the medium, PBS was added to the blank, rehabilitation solution was added to the positive, and cell proliferation activity was detected using MTT kit.

As shown in FIG. 14, the oligopeptide has strong activity of promoting proliferation of human skin fibroblasts, and is superior to the rehabilitation solution.

Example 6

Example 6 and example 5 were compared, and the difference between example 6 and example 5 is that:

because hydroxyproline is a special material for synthesizing collagen, the content of the collagen is relatively stable, and therefore, in clinic and experiment, the measurement of the total collagen amount usually adopts a hydroxyproline method, and the total collagen content is indirectly reflected.

Human skin fibroblasts were seeded in 24-well plates (10 ⁴ And/or hole), adding POP1 oligopeptide into the culture medium, adding PBS into a blank group, adding a rehabilitation solution into a positive group, incubating for 24 hours, sucking the cell culture solution, and determining the content of hydroxyproline by a digestion method.

As shown in FIG. 15, the oligopeptide has the activity of promoting the synthesis of collagen by human skin fibroblasts and is superior to the rehabilitation solution.

Example 7

Example 7 and example 6 were compared, and the difference between example 7 and example 6 was:

weighing 0.1g of oligopeptide POP1 and 50g of propylene glycol, mixing, grinding, adding 100mL of water for injection for dilution, uniformly mixing, adding 9g of sodium chloride, dissolving, adding water for injection to 1000mL, adjusting the pH value to 5.5-6.5, filtering, filling and sealing, and sterilizing to obtain 1000 injections.

Example 8

Example 8 and example 7 were compared, and the difference between example 8 and example 7 was:

5g of polyvinyl alcohol, 0.5g of carbomer and 1g of carboxymethyl cellulose are weighed, respectively swelled, then mixed, added with 0.1g of oligopeptide POP1, fully stirred and mixed, and stood for defoaming, thus obtaining the gel containing the oligopeptide POP1.

Example 9

Example 9 and example 8 were compared, and example 9 and example 8 differ in that:

0.1g of oligopeptide POP1 is weighed and dissolved in 45mL of distilled water, then the mixture is filtered, 5mL of glycerol and 1mL of azone are added into the filtrate, and the distilled water is added to the volume of 50mL, so that the external liniment containing 0.2% of active oligopeptide POP1 is prepared.

Example 10

Comparing example 10 with example 9, example 10 differs from example 9 in that:

weighing 0.1g of oligopeptide POP1, dissolving in 10mL of deionized water, adding 0.4g of dissolved carbomer, mixing, adding triethanolamine, adjusting the pH value to 6-7, uniformly coating on a die which is pre-coated with liquid paraffin, drying at 45 ℃ for 1h, taking out the film, and cutting into the required size to obtain the oligopeptide-containing film.

The above examples also satisfy the following conditions:

chromatographic conditions: using Thermo Scientific Accucore ^TM C18 column (3 mm. Times.100 mm,2.6 μm), mobile phase 0.1% aqueous formic acid (phase A) and 0.1% formazanGradient elution is carried out on the acid acetonitrile solution (B phase) (0-9 min, 5-17% of B phase, the rest is A phase, 9 min-17 min, 17-30% of B phase, the rest is A phase, 17 min-25 min, 30-50% of B phase, the rest is A phase, 25 min-30 min, 50-80% of B phase, the rest is A phase, 30 min-35 min, 80-99% of B phase, the rest is A phase, 35 min-40 min,99% of B) at the column temperature of 30 ℃ and the sample injection amount of 3 mu L. Flow rate 0.3 mL/min ^-1 Column temperature was 30 ℃.

Mass spectrometry conditions: the high-resolution mass spectrum adopts an electrospray ion source (ESI), positive ion mode scanning, a spray voltage of 3.2kV, an ion source temperature of 350 ℃, a sheath air flow rate of 35arb, an auxiliary air flow rate of 10arb and an ion transmission tube temperature of 320 ℃. The scanning mode is full scanning/data dependent secondary scanning (FullMS/dd-MS 2), the primary resolution is 70000, the secondary resolution is 17500, the scanning range m/z is 100-1500, and the collision energy gradient is 20eV,40eV and 60eV.

One or more technical solutions in the embodiments of the present application at least have the following technical effects or advantages:

(1) The oligopeptide provided by the embodiment of the application has 5 amino acids, is convenient for biosynthesis and chemical synthesis, is favorable for mass production and preparation, and has good prospect in industrial production.

(2) The oligopeptide provided by the embodiment of the application has the effects of promoting the synthesis of human skin fibroblasts and wound collagen, promoting the proliferation of the human skin fibroblasts and promoting wound repair.

(3) The method provided by the embodiment of the application is simple to operate through separation and screening based on drug effect screening, and meanwhile, the screening standard is clear, so that the method is suitable for extracting oligopeptides from American cockroaches in batches.

(4) The application provided by the embodiment of the application can be used for preparing the extracted oligopeptide into injection for repairing wound surfaces and also into common medicines such as gel, film, liniment and the like.

It should be noted that in this document, relational terms such as "first" and "second" and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising … …" does not exclude the presence of other like elements in a process, method, article or apparatus that comprises the element.

The endpoints and any values of the ranges disclosed herein are not limited to the precise range or value, and are understood to encompass values approaching those ranges or values. For numerical ranges, one or more new numerical ranges may be found between the endpoints of each range, between the endpoint of each range and the individual point value, and between the individual point value, in combination with each other, and are to be considered as specifically disclosed herein.

The foregoing is only a specific embodiment of the application to enable those skilled in the art to understand or practice the application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the application. Thus, the present application is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. The oligopeptide POP1 is characterized in that the amino acid sequence of the primary structure of the oligopeptide POP1 is shown in SEQ ID NO. 1.

2. A pharmaceutically acceptable salt of a polypeptide, wherein the polypeptide is the oligopeptide POP1 of claim 1.

3. A method for preparing the oligopeptide POP1, wherein the method is used for obtaining the oligopeptide POP1 of claim 1 from periplaneta americana, and the method comprises the following steps:

4. The method according to claim 3, wherein the organic solvent comprises at least one of ethanol, methanol, and acetone.

5. The method according to claim 3, wherein the plurality of chromatographies includes a first chromatograph, a second chromatograph, and a third chromatograph, the first chromatograph having a target molecular weight > the second chromatograph, the third chromatograph being configured to separate the compound of the second chromatograph according to a polarity size.

6. The method of claim 3, wherein the separation and purification comprises separation and purification with a molecular sieve under conditions less than a baseline molecular weight, wherein the baseline molecular weight is 1000Da.

7. Use of the oligopeptide POP1 according to claim 1 for the preparation of a medicament for the treatment of wounds, scalds and ulcers.

8. The use according to claim 7, wherein the medicament comprises an external preparation and an injection preparation.