CN101070341A - Antibacterial peptide without mutagenic action - Google Patents
Antibacterial peptide without mutagenic action Download PDFInfo
- Publication number
- CN101070341A CN101070341A CN 200610017750 CN200610017750A CN101070341A CN 101070341 A CN101070341 A CN 101070341A CN 200610017750 CN200610017750 CN 200610017750 CN 200610017750 A CN200610017750 A CN 200610017750A CN 101070341 A CN101070341 A CN 101070341A
- Authority
- CN
- China
- Prior art keywords
- antimicrobial polypeptide
- mutagenesis
- polypeptide
- amino acid
- mixture
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Abstract
This invention relates to an antibiotic polypeptide without mutagenic action. It at least contain 22 amino acid, contain ( AFVKLLKK) or ( AFVKIMKK). The invention is useful for preparing antibiotic combination, has good antibiotic action.
Description
Technical field
The invention belongs to the polypeptide technical field, particularly can be used for preparing antimicrobial compound no mutagenesis antimicrobial polypeptide, its preparation method and contain the mixture of antimicrobial polypeptide.
Background technology
All biologies, from virus to the mankind, its protein all is made up of 20 seed amino acids.Polypeptide is that a class is made of amino acid but is different from proteinic intermediate material.Amino acid in the polypeptide is each other to claim peptide bond to connect.The amino acid that a kind of peptide contains is less than 10 peptide and is called oligopeptides, surpasses 10 amino acid just to be called polypeptide.Amino acid is called protein at the polypeptide more than more than 50.
Because more and more serious in existing antibiotic resistance problem, the new antibiotic of R and D has become an important topic of medical circle.In addition, people find that a class polypeptide has anti-microbial effect when the external defense mechanism of research biology.Therefore caused the very big interest of the world of medicine to the research and development of polypeptide antibiotic.Find first antibacterial peptide the forties, from then on, the anti-microbial effect of polypeptide has also caused investigator's great attention.Such as, Steiner et al (1981) finds can cause the cracking of bacterium from the isolated antimicrobial polypeptide in cecropia oral cavity.Zasloff (1987) has found Magainins.More (Mor et al., 1991) finds that dermaseptins also can make mammal lysis.This two peptide species all can extract from the skin histology of the frog.Polypeptide Venom melittin (1967) and neurotoxin pardaxin (1988) all can cause cell and bacterium for cracking.Initial antimicrobial polypeptide is mostly found in animal tissues.In these tissues, play defense reaction, and can suppress duplicate (Boman, 1995) of microorganism.
Polypeptide antibiotic is different from traditional antibiotic, and the antibacterial peptide effect is single-minded, can go to play a role from the angle of nucleic acid and gene.
1980, people such as Sweden scientist G.Boman found the anti-microbial effect of polypeptide.Afterwards, people found and isolated the polypeptide of anti-microbial activity from bacterium, fungi, batrachians, insect, higher plant, Mammals and even the mankind.Be referred to as " antibactetial peptides, ABP ", i.e. antibacterial peptide.But found again afterwards that some antibacterial peptide all had lethal effect to part fungi, protozoon, virus and cancer cells etc., just being called " peptide antibiotics " again is peptide antibiotics (Shai, et al., U.S.Patent, 7001983,2006; Paradies.U.S.Patent Documents, 4874850,1989; Jacob et al., U.S.Patent, 5635479,1997; Maloy, U.S.Patent, 1998; Lee et al, 5889148,1999; WO, 97-02286,1997).Natural antibacterial peptide normally is made up of more than 30 amino-acid residues, good water solubility, and molecular weight is approximately about 4000 dalton.Most of antibacterial peptide has thermostability, heats down at 100 ℃ still can keep its activity in 10～15 minutes.The iso-electric point of most antibacterial peptides can be greater than 7, show stronger positively charged ion feature (Peptide Antibiotics:Discovery, Modes of Action and Applications edited by Christopher J.Dutton et al1998-2005 Culinary and Hospitality Industry Publications Services).Simultaneously, antibacterial peptide all has stronger resistance to bigger ionic strength and higher or lower pH value.In addition, the part antibacterial peptide still possesses the ability of opposing trypsinase or pepsin hydrolysis.It is generally acknowledged that the antibacterial peptide germicidal action mainly is the integrity by the cell walls that destroys bacterium.And it is outer and dead to cause in the cell oar material to overflow born of the same parents.Antibacterial peptide is invested the bacterial film surface earlier by electrostatic attraction, hydrophobic C end inserts the interior hydrophobic region of film and changes the conformation of film, and a plurality of antibacterial peptides form ionic channel and cause ionic to overflow and death on film.Antibacterial peptide can make membranin cohesion, inactivation, causes that membrane permeability changes and causes bacterium death.The different classes of antibacterial peptide mechanism of action can be different.The antibacterial peptide majority has characteristics such as strong basicity, thermostability and broad-spectrum antimicrobial.Some antibacterial peptide all has strong lethal effect to part fungi, protozoon, virus and cancer cells etc.Antibacterial peptide all has the lethal effect of high-efficiency broad spectrum to Gram-negative and positive bacteria.Reported that antibacterial peptide has lethal effect to having the bacterium more than 100 kinds at least both at home and abroad.Linear polypeptide cecropins with spirane structure is first found animal antibacterial peptide, 1980, is separated to obtain by Boman etc. from U.S.'s sky silkworm chrysalis.Such peptide antibiotics generally contains 37～39 amino-acid residues, do not contain halfcystine, its N end regions has strong basicity, can form and be close to perfect parents' spirane structure, and can form hydrophobic spiral at the C end regions, the hinge area that has glycine and proline(Pro) to form between the two, the C end of most polypeptide is by amidation, and amidation has vital role to its anti-microbial activity.After this, people have been separated to cecropins class antibacterial peptide in succession from silkworm, tussah, fruit bat, sarcophagid.1989, people such as Lee were separated to cecropin P1 from chitterlings, had illustrated that cecropins may extensively exist in animal.Cecropins has very strong lethality to gram positive organism, negative bacterium group, and fungi and eukaryotic cell are not had toxicity.Cecropins is by synthetic and commercialization at present.Magainins also is a class antibacterial peptide of early finding.1987, Zasloff finds from the skin of the Amphibians frog (African clawed frogXenopus laevis) that at first magainins has antimycotic and anti-microbial effect.Contain 23 amino acid, and under water miscible condition, have anti-microbial effect.Therefore he set up a company and developed this product.It not only has lethal effect to gram-positive microorganism and gram-negative bacteria, and fungi and protozoon are also had lethal effect Zasloff, 1987; Hancock, 1998).Other one group of antimicrobial polypeptide is Defensins, it is a class zooblast endogenous sterilization polypeptide, from phagocytic cell, separate, has very wide antimicrobial spectrum, greater than the lethal effect to gram-negative bacteria, it also acts on fungi and part eukaryotic cell to the lethal effect of gram-positive microorganism.But the specific activity cecropins to gram-negative bacteria will hang down (Dorschner RA, et al., 2006) about 10 times.Apidaecins separates certain the amino acid whose linear polypeptide that contains that obtains from honeybee.Maloy and Kari UP (1995) find to contain the another kind of peptide antibiotics of proline(Pro).Generally contain 16～18 amino-acid residues, wherein proline content is up to 33%, and arginine content can reach 17%.Apidaecins has very strong activity to some gram-negative bacteria, and inoperative to gram positive organism.Apidaecins is to the High Fragmentation power of the pathogenic bacterium of some Gram negative phytopathogen and enterobacteriaceae, but can produce the neuron toxic action.Yet it only has good application prospects in plant antibacterium ospc gene engineering and foodstuffs industry.Also has a kind of Drosocin of being called antibacterial peptide (Otvos L Jr, 2002).It is the antibacterial peptide that derives from a kind of proline rich of fruit bat, structurally has certain similarity with apidaecins, but is connecting O-two sugar chains (N-acetylgalactosamine-semi-lactosi) on its Threonine hydroxyl of 11.It may influence proteinic synthetic and metabolism (Otvos, 2002).Coleoptericin and hemiptericin derive from Coleoptera and hemipteran respectively, are rich in glycine in the primary structure, molecular weight generally big (Imamura M, et al, 1999; CociancichS et al., 1994).But, because shortage does not still have the developer at present to the research of its anti-microbial effect.People such as Oppenheim separate from people's the parotid gland and lower jaw glandular secretion thing and have obtained one group of antibacterial peptide that is rich in Histidine, and length does not wait at 7～38 amino-acid residues, is called as histatins.Has activity (Raj PA and Dentino AR., 2005) for the multiple microorganism that causes oral cavity infection.Indolicidin is the peptide antibiotics that derives from the ox neutrophil leucocyte, gains the name because of containing 5 tryptophanes in its 13 amino acid.Its C end is amidated, intestinal bacteria and streptococcus aureus is all had very strong fungicidal activity (Shaw JE., et al., 2006), but can produce resistance (Samuelsen O., et al., 2005).The polypeptide that contains a disulfide linkage, this is a class quantity antibacterial peptide seldom, the 1st found this class polypeptide is bactenecin, derives from the ox neutrophil leucocyte.Contain 4 arginine in its 12 amino acid, between its 2nd and the 11st amino acids residue, form disulfide linkage.Bactenecin has activity (Raj PA and Dentino AR., 2005) to intestinal bacteria and streptococcus aureus.Comprise also in this class polypeptide that some derive from the peptide antibiotics of batrachia skin, generally have one by 7 amino acids formed " loop " and a long N end " tail " at the C end, as brevinin-1, brevinin-2.The polypeptide that contains two or more disulfide linkage, typical case's representative of this class polypeptide is defensins, the initial defensins that finds derives from the mammiferous tissue, generally contains 29～34 amino-acid residues, and wherein 6 conservative halfcystines form 3 intramolecular disulfide bonds; In addition, its 6th and the 15th 's arginine, the 24th glycine is also guarded.α-defensins can form 3 layers β laminated structure, is stabilized by the salt bridge between 3 disulfide linkage and Arg-6 and the Glu-24.At present, defensins has been synthesized and commercialization.Defensins has lethal effect to various bacteria and some fungi, and eukaryotic cell is had certain toxicity.Defensins is strong to the specific activity gram-negative bacteria of gram positive organism.A little less than the specific activity cecropins of defenssins, and under low ionic strength work usually.β-defensins is bigger than α-defensins, generally contains 38～42 amino-acid residues.All contain 3 disulfide linkage and 4～8 arginine.Insect defensins is similar to α-defensins at C-terminal, but has only two β laminated structures, centre to have one section α spiral to play stabilization, mainly gram positive organism is worked, and to not effect of fungi.Plant defensins generally has 45～54 amino-acid residues, can form 4 disulfide linkage, 3 β laminated structures and a αLuo Xuanjiegou.General of plant defensins works to fungi and bacterium is not acted on.Different plant defensins are to the antimicrobial spectrum difference of fungi.Thionins also is the peptide antibiotics that a class derives from plant, contains 45～47 amino-acid residues, and 6 or 83 or 4 disulfide linkage that halfcystine forms are arranged.Its secondary structure can form 2 antiparallel αLuo Xuanjiegou and 2 antiparallel β laminated structures.Thionins suppresses various plants malignant bacteria and fungi, but to the bacterium of Rhodopseudomonas and erwinia inoperative (Castro andFontes, 2005).Therefore, the research relevant with the human body bacterium is seldom arranged.Scientists has been illustrated another and has been called the natural micromolecular complicated antibacterial mechanisms of antibacterial peptide J25 (microcin MccJ25), and this antibacterial peptide can be blocked the enzyme of bacterium, might cause antibiotic appearance of new generation.The MccJ25 stable structure makes it to tolerate various extreme environments, and it might become a kind of sterilizing agent.Although thereby MccJ25 can suppress the RNAP killing bacteria of bacterium, it can not suppress human RNAP, therefore can the harmful to human cell.But, MccJ25 also has some shortcomings, and it only acts on intestinal bacteria and relevant Pseudomonas, and bacterium is easy to produce resistance.At present, corresponding resistance pathogenic strain system has all appearred in all conventional microbiotic, and the resistance problem of pathogenic bacterium has day by day seriously threatened people's health.The microbiotic of seeking brand-new type is a kind of effective way that solves the resistance problem.Antibacterial peptide is because the anti-microbial activity height, has a broad antifungal spectrum, and kind is many, and alternative scope is wide, and the target bacterial strain is difficult for producing reasons such as resistant mutation, and is considered to that wide application prospect will be arranged on medicine industry.At present, existing multiple polypeptides microbiotic is carrying out preclinical feasibility study.
The clinical application aspect also has many problems, and wherein topmost problem is a toxic action.For example, Colimycin, a kind of fat polypeptide, success is used for treating the Pseudomonasaeruginosa pulmonary infection clinically.As if patient can tolerate the treatment of colimycin.Mainly may be because the modification of chemical structure has reduced the toxic action of medicine.May be to have added fat (Robert E., et al., 1999) in the back of this polypeptide.Present most of clinical trial is to be used for topical therapeutic, and this treatment should be a safety and effective.Polypeptide that some toxicity are stronger and fat polypeptide, as Gramicidin S, PXB has been used to make skin ointments.These polypeptide also can be used for those conventional microbiotic and the invalid place of routine treatment.Utilizing the method treatment pulmonary infection of pulvis is a up-and-coming developing direction.Oral pharmaceutical may be used to treat intestinal tract infections, and Nisin is carrying out the clinical trial of anti-helicobacter.Although antibacterial peptide has been carried out a large amount of research, still there are many problems can enter the clinical treatment infectation of bacteria.Remain unsolved such as problems such as toxicity, absorption and synthetic costs.
Also need to solve some problems at present.It at first is toxicity problem.This comprises that can antibacterial peptide single-mindedly cause the cracking of harmful germ and not influence normal tissue cell.Secondly, antibacterial peptide can or can not produce transgenation.
Consider that in addition the synthetic of cost problem antibacterial peptide will make things convenient for economy, the antibacterial peptide molecule will have high vigor.Our invention be one to be combined into polypeptide, remove its sequence and be different from existing antimicrobial polypeptide, and have stronger anti-microbial effect, do not have mutagenesis simultaneously, be more suitable for being used for treating the infection of infectation of bacteria, especially drug-resistant bacteria.
Summary of the invention
The object of the invention is to provide antimicrobial polypeptide, its preparation method of one group of no mutagenesis that can be used for preparing antimicrobial compound and contains the mixture of antimicrobial polypeptide.
For reaching above-mentioned purpose, the present invention adopts on the basis of sequence, anti-microbial activity and structural analysis to the natural antibacterial polypeptide, research, has designed and synthesized one group of antimicrobial polypeptide that does not have mutagenesis, and described antimicrobial polypeptide does not have mutagenesis.
Described antimicrobial polypeptide contains 22 amino acid at least.
Described antimicrobial polypeptide contains (AFVKLLKK) or (AFVKIMKK).
The aminoacid sequence of described antimicrobial polypeptide is CH
3CO-GIGKFLKKAKKFGKAFVKLLKK-NH
2(hereinafter to be referred as: DK-1) or CH
3CO-GIGKFLKKAKKFGKAFVKIMKK-NH
2(hereinafter to be referred as: DK-2).
The 1st, 3,13 amino acids are Gly in 22 amino acid of described antimicrobial polypeptide.
In 22 amino acid of described antimicrobial polypeptide the 4th, 7,10,11,14,18 amino acids are Lys.
It is Lys that described antimicrobial polypeptide amino acid A19 Lys connects 20A.
The 2nd amino acid is Ile in 22 amino acid of described antimicrobial polypeptide.
The 5th, 12,16 amino acids are Phe in 22 amino acid of described antimicrobial polypeptide.
The 6th, 20 amino acids is Leu in 22 amino acid of described antimicrobial polypeptide.
The 9th, 15 amino acids is Ala in 22 amino acid of described antimicrobial polypeptide.
The 17th amino acids is Val in 22 amino acid of described antimicrobial polypeptide.
Described antimicrobial polypeptide the 20th amino acids is L or M.
15 amino acids that described antimicrobial polypeptide contains AFVKLLK or AFVKIMKK are Ala.
The preparation method of the antimicrobial polypeptide of no mutagenesis adopts solid state chemistry synthetic.
The preparation method of the antimicrobial polypeptide of no mutagenesis adopts biosynthetic means.
The described mixture that contains antimicrobial polypeptide is characterized in that this mixture contains a kind of described antimicrobial polypeptide at least.
Described mixture contains two peptide species.
The a kind of of two peptide species is CH
3CO-GIGKFLKKAKKFGKAFVKLLKK-NH
2(DK-1); Another kind is CH
3CO-GIGKFLKKAKKFGKAFVKIMKK-NH
2(DK-2).
Described mixture is a creme, finish, or the external-use drug form of aqua.
Described mixture is oral, intramuscular injection, intravenous form.
A kind of method for the treatment of Gram-positive, gram negative bacterium and drug-fast bacteria infection uses any formulation of described mixture for the infectation of bacteria patient.
A kind of sterilizing agent contains any sterilizing agent formulation of described mixture.
Polypeptide among the present invention can adopt sophisticated chemical synthesis process to synthesize.The synthetic FMOC chemical process that adopts of polypeptide.Instrument is the Applied-432 Peptide synthesizer, is the Synergy Peptide synthesizer again.The concrete grammar that we adopt is the solid phase synthesis process that FMOC and HBTU activation combine.Synthetic from amino acid near C-terminal.Add an amino acid at every turn, be blended into till the N-terminal.Whenever add an amino acid and all will repeat following three steps: 1), from solid phase, remove the amino acid whose blocking group near N-terminal, 2), add next amino acid, 3), remove new close N-terminal amino acid blocking group.Between above-mentioned each process, cleaning step is arranged all.Synthesizer can be monitored removing blocking group and link coupled quality automatically.The printouts method is adopted in gross control.Quality controlling means synthetic and the synthetic product treatment measures are as follows: when end of synthesis, collect the polypeptide that has the protectiveness group and comprise leacheate.Then, the synthetic polypeptide is scaled off, and the blocking group on the removal side chain.Said process all carries out having under 90%TFA, 5%thioaisole and the ethanedithiol condition.Filter with glass wool (glass wool), with MTBE peptide group is precipitated then.And it is inferior to give a baby a bath on the third day after its birth with MTBE, so that remove residual blocking group.Residual acidic substance then neutralize.Antimicrobial polypeptide also can adopt biosynthetic means to obtain.Finally identify product purity or use the sequence measurement controlling quality with MS.
The present invention can provide a kind of new antibiotic method, it is characterized in that described mixture is oral, intramuscular injection, intravenous injection and various formulation.Be a kind of treatment Gram-positive, the method for gram negative bacterium and drug-fast bacteria infection.
The present invention can provide a kind of new antibiotic method, and antimicrobial polypeptide can be made into ointment, and drug dose 10-20ug/ml comprises ointment and paste.Concrete grammar is exemplified below: the antimicrobial polypeptide ointment is that antimicrobial polypeptide is added a kind of paste external preparation with d spiss of coating skin, mucous membrane or the surface of a wound easily of making in the suitable matrix.The specification of quality of ointment for evenly, fine and smooth, be applied to no coarse sensation on the skin; Suitable viscosity is arranged, easily the coating and do not melt; Do not have become sour rotten, oily water separation or layering do not take place, can keep intrinsic curative effect; Should non-stimulated and other reaction.The ointment that is used for the surface of a wound also should be aseptic.To matrix require as follows: lubricated non-stimulated, denseness is suitable, is easy to coating; Compatibility does not take place and changes in stable in properties; Have water-absorbent, can absorb wound exudate; Do not hinder the normal function of skin, have good Release Performance; Clothes is not polluted in easily eccysis.Be applicable to the infectious skin disease.Its kind has greasing base, emulsion-type matrix and contains three kinds of ointments of paste (Chinese Pharmacopoeia) of a large amount of (25-70%) moisture absorptions, convergency powder.
Greasing base comprises three kinds of hydro carbons, lipoidis, lipid; The matrix of emulsifiable paste is emulsion-type matrix; Paste contains a large amount of moisture absorptions, convergency powder.
Vaseline in the hydrocarbon base (soft wax) fusing point is 38-60 ℃, and odorless is non-stimulated, does not become sour stable in properties.Be applicable to the medicine of meeting water unstable.Be not suitable for affected part acute and the volume transudate.Be characterized in suitable viscosity.Solid paraffin is a kind of of greasing base, and its fusing point is 50-65 ℃, can be used to regulate the denseness of ointment.Whiteruss also is a kind of of greasing base.It can mix with most fatty oils or volatile oil, regulates the denseness of ointment.Silicone also is a kind of of greasing base, and its viscosity increases with the increase of molecular weight, is the liquid of no color or smell, nontoxic, non-stimulated, easy coating, and stable in properties can be protected skin and as transdermal enhancer.Lipoidis has lanolin, is light brown yellow thickness semisolid, and fusing point is 36-42 ℃, and water-absorbent is strong, can absorb the water of about 2 times of its weight and form w/o type emulsion.Share with Vaseline, can improve the water-absorbent of Vaseline.Lipid matrix derives from animals and plants higher fatty acid glyceride and composition thereof.Be subject to the influence of temperature, light, oxygen and decompose, oxidation.
Emulsion-type matrix is formed: water+oil phase+emulsifying agent is applicable to that skin subacute, chronic, no transudate decreases.Avoid and be used for erosion, ulcer, blister and warts disease.Below be the example of emulsion formulations:
Monovalence soap prescription: stearic acid 100g, Viscotrol C 100g, whiteruss 100g, trolamine 8g, glycerine 40g, distilled water 452g.Be suitable for: emulsifiable paste.
Multivalence soap (w/o type emulsion-type matrix) prescription: calcium, magnesium, zinc and aluminum oxide+lipid acid multivalence soap.Characteristics: the poison that dissociates is little, and wetting ability is less than the monovalence soap, and lipophilicity is strong, and HLB＜6 easily form, and stability is high.Concrete prescription: stearic acid 12.5g, single stearic acid glycerine lipoprotein 17.0g, the cured 5.0g of honeybee, ceresine 75.0g, whiteruss 410.0ml, white vaseline 67.0g, double stearic acid aluminium 10.0g, calcium hydroxide 1.0g, ethylparoben 1.0g, distilled water 401.5ml.Method for making: oil phase fusing (85 ℃)+water (85 ℃) limit edged stirs.
Fatty alcohol sulphuric acid (ester) sodium class prescription: lauryl alcohol sulfuric acid (ester) sodium, character: anionic emulsifier, usual amounts 0.55-2% (weight percent), suitable pH value is 6-7, must not＜4,＞8.Compatibility: normal and other w/o type emulsifying agent share.Add 1.5%-2% (weight percent) sodium-chlor and can make it to lose emulsifying effect.Concrete prescription: stearyl alcohol 220g, sodium lauryl sulphate 15g, single stearic acid glycerine lipoprotein 17.0g, white vaseline 250g, methyl hydroxybenzoate 0.25g, Propyl Hydroxybenzoate 0.15g, propylene glycol 120g, adding distil water are to 1000g.Method for making: oil phase fusing 75 ℃+water (75 ℃) limit edged stirs.
High fatty alcohol and polyol ester class prescription: in the high fatty alcohol, 45-50 ℃ of hexadecanol fusing point, water insoluble, be used for O/W type emulsion matrix, can increase stability and viscosity; 56-60 ℃ of stearyl alcohol fusing point, water insoluble, can increase the Vaseline water-absorbent; Sodium laurylsulfate, O/W emulsion emulsifying agent, consumption 0.5%-2% (weight percent), pungency is less, HLB value 40, can with acid-basicity medicine, Ca, Mg ion compatibility, avoid with cationic drug and join thing.
The derivatives class prescription of Soxylat A 25-7: water-soluble base, characteristics: it is very fast to discharge medicine, and no greasy feeling easily is coated with exhibition, and nonirritant can the absorptive tissue exudate.Purposes: be used for moisteningly, the rotten to the corn surface of a wound helps the eliminating of secretory product, also can be used for the matrix of cavity mucous membrane or grease proofing protectiveness ointment.Shortcoming: lubrication is relatively poor, and wherein moisture easily evaporates, and easily goes mouldy.
Glycogelatin, method for making: gelatin 1-3%+ glycerine 10-30%+ water (surplus, weight percent).
Characteristics: form layer protecting film, more comfortable during use.
Derivatived cellulose: MC is dissolved in cold water, and CMC is cold and hot all molten, and its CMC-Na and heavy metal ion can not compatibilities.The PEG class, molecular weight: 300-6000 commonly used is liquid or waxy solid, characteristics: soluble in water, easily mix easy eccysis with transudate, character is more stable, is difficult for going mouldy.
The preparation of ointment:
1, the method for medicine adding: water soluble drug water
2, preparation method:
A. grind and method: semisolid+liquid
Medicine+part matrix (or liquid) is ground to fine and smooth pasty state, and all the other matrix of progressively increasing are checked till the no particle sensation.
B. fusion method: the matrix that fusing point is higher, matrix fusing, filtration+medicine stir evenly, condensation.
C. emulsion process: oil soluble matrix, fusing, filtration, filtrate+antimicrobial polypeptide+water, solution heating (80 ℃) is stirred, condensation.
The quality control of ointment:
1, the assay of antimicrobial polypeptide;
2, the mensuration of physical properties: fusing point, potential of hydrogen, physical appearance, viscosity;
3, pungency;
4, stable form;
5, drug release, the measuring method that penetrates and absorb.
The packing of ointment and storage:
1, packing: ointment tube (tin, aluminium, plastics); Plastics casing;
2, packing apparatus: automatic gear.Roll tail, dress box gear;
3, storage: the shady and cool dry place of lucifuge, temperature should not cross exceed low.
Two peptide species of the present invention are all water-soluble.Utilize polypeptide of the present invention to can be made into externally applied agents such as creme, finish or aqua, also can be made into oral preparation, injection.Can be used for treating Gram-positive, gram negative bacterium and drug-fast bacteria infection.Polypeptide of the present invention does not have mutagenesis.
The chemical structure of polypeptide of the present invention can identify that anti-microbial activity can be checked with bacteriostatic experiment by mass spectrum, and its toxic action can be assessed with acute toxicological experiment, long-term toxicological experiment, local toxicological experiment and mutagenicity test.
The mixture that two kinds of synthetic antimicrobial polypeptide of the present invention are formed wherein contains two peptide species: DK-1 and DK-2, and its ratio is 1: 1, the mixture DK-1+DK-2=DK (being designated hereinafter simply as DK) that these two kinds of synthetic antimicrobial polypeptide are formed.Every peptide species is made up of 22-L amino acid.This two peptide species is all water-soluble.It is characterized in that described antibacterial peptide has the aminoacid sequence of DK-1 or DK-2.Antibacterial peptide also comprises indivedual amino acid whose replacements and obtains functional equivalent or the stronger polypeptide of anti-microbial effect.Can kill gram-positive and gram negative bacterium.
Among the present invention, the polypeptide amino acid positively charged is big, can combine with the lipopolysaccharide (LPS) of leather orchid property negative bacteria or combine with the lipoteichoic acid of leather orchid property positive bacteria, increases the permeability of bacteria cell wall, and bacterium is produced lethal effect.For general anti-microbial effect and the antimicrobial agent effect that proves this class antimicrobial polypeptide, utilize U.S. GenScript Corporation biotech company to adopt the prepared one group of polypeptide (DK) of solid state chemistry synthetic method to study.
Utilize 96 well plate method to detect the germicidal action of one group of polypeptide (DK), and compare, to estimate the fungicidal activity of antimicrobial polypeptide DK among the present invention with synthetic natural antibacterial polypeptide (Pexiganan).The result shows that synthetic antimicrobial polypeptide DK fungicidal activity of the present invention is better than the above-mentioned natural antibacterial of synthetic in advance polypeptide (Pexiganan).Bacterial strain recovery back inoculation, 37 ℃ of overnight incubation; Then, choose bacterium in ordinary culture medium, 37 ℃ of overnight incubation; Bacterium liquid is diluted to the 104-105CFU/ milliliter, every hole inoculation 10ul bacterium liquid in 96 orifice plates, 37 ℃ of overnight incubation of 96 orifice plates are measured OD with microplate reader
620Value (Park et al, 1998).The measurement result of fungicidal activity sees Table 1.Growth concentration (the OD that contains the antimicrobial polypeptide bacterium
620Value) ratio with the growth concentration that does not add the antimicrobial polypeptide bacterium is minimal inhibitory concentration (minimal inhibitory concentration, MIC is meant the minimum concentration of remarkable bacteria growing inhibiting) greater than 90% o'clock antimicrobial polypeptide concentration.
Table 1. compares the minimal inhibitory concentration (MIC) of two kinds of antimicrobial polypeptides (DK and Pexiganan) to the anti-microbial activity of different bacterium.Visual inspection does not see that the drug level of bacterial growth is that the observations of minimum inhibitory concentration (MIC) sees the following form.
Minimum inhibitory concentration (MIC) microgram/ml
(2005.11 experimental result record)
| Bacterium is hidden numbering | Strain name | Quantity | MIC (the DK of microgram/ml) | MIC (the Pexigan an of microgram/ml) | Bacterium numbering | Remarks |
| 1.2465 | Streptococcus aureus | 1 | ?8 | ?16 | ?ATCC6538 | Reference culture |
| 1.879 | Streptococcus aureus | 1 | ?4 | ?8 | ?ATCC6358P | Penicillin resistant |
| 1.2387 | Pseudomonas aeruginosa | 1 | ?32 | ?32 | ?ATCC27853 | Reference culture |
| 1.2031 | Pseudomonas aeruginosa | 1 | ?32 | ?ATCC10145 | Anti-Ampicillin Trihydrate | |
| 1.3373 | Colon bacillus | 1 | ?16 | ?32 | ?ATCC8099 | Reference culture |
| 1.2429 | Staphylococcus epidermidis | 1 | ?1、 ?0.5 | ?ATCC12228 | Reference culture | |
| 1.2025 | Intestines (dung) coccus | 1 | ?4 | ?ATCC6057 | Reference culture | |
| 49619 | Streptococcus pneumoniae | 1 | ?ATCC49619 | Reference culture | ||
| 13883 | Klebsiella Pneumoniae | 1 | ?8 | ?ATCC13883 | Reference culture | |
| 25922 | Colon bacillus | 1 | ?16 | ?ATCC25922 | Reference culture | |
| 27853 | Pseudomonas aeruginosa | 1 | ?32 | ?ATCC27853 | Reference culture | |
| 29213 | Streptococcus aureus | 1 | ?4 | ?ATCC29213 | Reference culture |
| Streptococcus aureus | 1 | ?0.5 | To Azythromycin, the ammonia sterilization, Ciprofloxacin, gentamicin, left side oxygen fluorine, tsiklomitsin, the Ampicillin Trihydrate, paraxin, Oxazacillin, trimethoprim-sulfamethoxazole, penicillin, cefotaxime, the cefepime resistance | |||
| Escherichia coli | 1 | ?4 | Clinical isolates (sensitive strain) | |||
| Pseudomonas aeruginosa | 1 | ?4 | To ceftriaxone, cefepime, cefotaxime, ceftazime, cefoxitin, aztreonam, paraxin, left side oxygen fluorine, gentamicin, penbritin, amikacin, the trimethoprim-sulfamethoxazole resistance |
In addition, find that in DK and the mutagenic comparison of Pexiganan DK does not have mutagenesis.Adopt the Aims experimental technique that DK and Pexiganan mutagenesis are compared.Result of study finds that the Aims experiment of Pexiganan is positive, and the Aims of DK experiment is negative.The result shows that DK does not produce mutagenesis, and Pexiganan can produce mutagenesis.
At observation DK the time spent of doing of skin infections model is found that DK has therapeutic action.Get rabbit and weigh, and 1d loses hair or feathers at the back before experiment, depilation area 150cm
2, skin routine disinfection during experiment is with the skin that excises 2.0cm * 2.0cm area after the 2% PROCAINE HCL, PHARMA GRADE 2ml local infiltration anesthesia, thoroughly after the hemostasis, inject streptococcus aureus bacterium liquid 0.2ml in the subcutis, simultaneously wound surface smearing 0.2ml bacterium liquid, plastics film flap coverage, 2d continuously.3d gets surface of a wound fester and makes microbial culture, and random packet administration (king's Peng, Xu Shulan, Zhu Yan etc., 2000; Lan Shumin, Lu Huiqin, old filial piety etc., 2004; Di Ning, Di Fengtong, Wang Li etc., 2001).DK high dose group wound surface smearing, DK 32 micrograms/ml, low dose group wound surface smearing DK 16 micrograms/ml paste, each 1ml.Positive group is smeared the gentamicin paste, each 1ml.Model group is smeared the Natvosol paste, each 1ml.Below respectively organize equal every day 2 times, continuous 5 days, every day, the face situation respectively formed in record, put to death animal in 24 hours behind the last medicine, and the part is got skin and done pathological section, observes the wound healing situation.Discover the 3rd day and respectively form face edge redness, a large amount of purulent secretion, surface of a wound microbial culture turns out to be infection of staphylococcus aureus, the Modelling success.2d high dose group, the slight redness of positive group behind the medicine, secretory product obviously reduces; The a large amount of secretory product of the red and swollen obviously companion of model group; Low dose group is red and swollen obviously, moderate secretory product.Behind the medicine the 6th day, high and low dose group, the positive were formed the face incrustation, and the edge has on a small quantity and oozes out granulation tissue hyperplasia; The incrustation of the model group surface of a wound, the edge still has a small amount of secretory product, the slight hyperplasia of granulation tissue.Mirror sight is down seen more granulation tissue, inflammatory exudate, is not seen the regeneration epidermis.
Positive group is seen in the growth of part epidermis lining epithelium, the dermal papilla has a small amount of cell infiltration to carry out the transition to granulation tissue gradually, a small amount of inflammatory exudate is arranged.Low dose group sees that it is that granulation tissue, surface have promoting epidermization, regeneration section to lack hair follicle that the transition of part normal skin is arranged.
High dose group part normal skin extends for granulation tissue and surface-coated outgrowth epidermis lacks cutaneous appendages, surface-area a small amount of inflammatory exudate on every side.Therefore DK has therapeutic action to the skin infections model.
Because antimicrobial polypeptide mainly acts on the cell walls of bacterium, therefore, might have influence on Normocellular function, produce the general toxicity effect, comprise the side effect of cardiovascular aspect.DK is as externally applied agent, this experiment is carried out general pharmacology to DK and is learned research, observe by of the influence of percutaneous drug delivery approach the heart rate of laboratory animal, electrocardiogram(ECG, blood pressure, breathing, with other pharmacological action of understanding antimicrobial polypeptide or toxic action (the pharmacological research method is seen Zhou Xiumei etc., 2004; Xu Shuyun etc., 2003; Li Feng, the long pine 1995 of Jiang; Li Hua etc., 2004), for clinical use provides the research foundation.Observation experiment is carried out in side effect to the cardiovascular aspect of antimicrobial polypeptide.Experimental observation finds no the side effect of any cardiovascular aspect.
The long-time antimicrobial polypeptide that uses also might the toxigenicity effect.Therefore, the toxicity of antimicrobial polypeptide is carried out chronic toxicity observe, in chronic toxicological experiment, do not observe any toxic action.Concrete various chronic toxicological experiment results are as follows: the DK that gives rabbit skin partial smearing 10 μ g/ml, 50 μ g/ml, 100 μ g/ml, 1ml/ time, 1 time/day, continuous 28 days, observe local reaction and systemic reaction, routine blood test, cytometry and classification, blood biochemical check, organ index.The result does not find that DK has chronic toxicity reaction (local reaction and systemic reaction) to rabbit.
Laboratory animal is 30 of healthy adult white rabbit, and body weight is 2.00 ± 0.2kg, and male and female half and half are female for not producing and not having pregnantly, provided by lake region, the Wuhan City, Hubei Province animal cultivation factory that shines entirely.Conformity certification number: 0036417.Laboratory temperature 22-25 ℃, relative humidity 50%-60%, conventional feed divide cage to feed, free choice feeding, and amount of drinking water is not limit.Rabbit is divided into 5 groups at random: 6 every group, male and female half and half, 2 kinds of situations of every component intact skin and damaged skin, each 3.Normal group is only used physiological saline.Vehicle group is the DK of 100 μ g/ml with the vehicle of medicine, DK, the high dose group that low dose group is 50 μ g/ml with the DK of 10 μ g/ml, middle dosage group.Observation index is to observe local toxicity reaction (red, swollen situation) every day, and by the scoring of skin irritation standards of grading; Observe general toxic reaction (central nervous system, respiratory system, vegetative nervous system, stool and urine); Measure body weight weekly one time.At first heart extracting blood is inspected routine blood test, cytometry and classification respectively by ready samples during off-test, blood biochemical is checked (liver function, blood urea nitrogen, ionogen, blood fat, clotting time etc.); Put to death rabbit, take out important organ and claim organ weights, calculating organ index (brain, the heart, liver,spleen,kidney, lung, suprarenal gland), simultaneously important organ is carried out pathological examination (brain, the heart, lung, liver,kidney,spleen, stomach, small intestine, mesenteric lymph nodes, pancreas, uterus, ovary, testis, thymus gland, suprarenal gland, Tiroidina, administration local skin).Earlier normal group and high dose group are carried out pathological examination, if the no abnormal person of high dose group check result, then middle low dose group can not done.Histopathological examination: long-term contact is subjected to reagent thing DK after 28 days, and is as follows to the pathological examination result of brain, the heart, lung, liver,kidney,spleen, stomach, small intestine, mesenteric lymph nodes, pancreas, uterus, ovary, testis, thymus gland, suprarenal gland, Tiroidina and administration local skin:
High dose group:
1. skin: epidermis is normal, and squamous epithelium (1-2 layer) has angling, intradermal that more hair follicle is arranged.
2. brain: see that the neuronal cell structure has spongiocyte normally, on every side, do not see cell infiltration and focal necrosis.
3. the heart: cardiac muscle fibre marshalling, clear in structure, laterally, vertically section no abnormality seen, no cell infiltration
4. liver: liver rope marshalling, the liver cell structure is normal, the liver lobule clear in structure, does not see hepatocellular degeneration, necrosis, and the portal area structure is normal, and central vein does not have expansion.
5. spleen: acini lienalis and red pulp clear in structure, show no obvious abnormalities.
6. kidney: renal glomerulus clear in structure, uriniferous tubules show no obvious abnormalities.
7. lung: alveolar structure is normal, alveolus wall do not have thicken, no inflammatory exudate in the alveolar space.
8. thymus gland: cortex, medullary substance clear in structure, thymus corpuscle no abnormality seen change.
9. sialisterium: see a large amount of serous acinus, a small amount of mucous alveolus and conduit, no abnormal.
10. stomach: each layer structural integrity, stratum supravasculare partly come off, body of gland is complete substantially, normal.
11. small intestine: clear normal, each layer of the intestines wall no abnormality seen of fluff structures.
12. pancreas: acinus, conduit, pancreas islet are all visible, no abnormality seen.
13. mesenteric lymph nodes: mild reaction hyperplasia, some broadening of paracortex.
14. suprarenal gland: each band of cortex and medullary substance inner cell there is no unusually.
15. uterus: inner membrance and body of gland are all normal, no abnormality seen.
16. ovary: see ovarian follicles at different levels and interstitial gland.
17. testis: a large amount of convoluted seminiferous tubules have many androgones (at different levels), a small amount of sperm.
Long-term rabbit skin partial smearing 10 μ g/ml, 50 μ g/ml, the 100 μ g/ml DK of giving, 1ml * 1 time/d * 28d, intact skin group and damaged skin group local skin do not have the obvious stimulation performance (concrete grammar are seen Liao Xiaoyan, Li Yajie, Cai Wenzhi etc., 2004; Liu Yaqian, Chen Hua, Li Chunhai etc., 2002; Zou Yi, Yang Fangju, Han Xu etc., 2004; Tang Jikun, Liu Min, Pan Zhengxing, 2002; ), when contact was subjected to reagent, rabbit whole body general state diet, muscular strength, activity etc. were all normal; Compare between index administration group such as blood test and biochemistry detection and organ weights and index and the control group, the P value does not have showing property difference all greater than 0.05; The pathological examination high dose group does not see that tissue abnormalities changes; Pointed out by The above results: the long term toxicity test of DK is not found local skin and general toxic reaction.
Single eye stimulation test experimental result is found not see that irritant reaction is arranged behind low dosage and the middle dosage DK-1 eye drip; But behind the high dosage eye drip, the visible cornea diffusivity of 6-48h muddiness, iris conjunctiva are all congested, intraocular secretory product is many, and eyelid and eyelashes humidity stimulate score value between 3.0-9.5, disappear substantially behind the 48h, and this medicine has light moderate to stimulate to eye when high dosage.
Observe 10 μ g/ml and 100 μ g/ml DK solution do not have hemolytic action by the naked eyes method.For eliminating macroscopic error, colorimetric method for determining (Yu Lian, Su Jin, Tian Haixia etc. 2003 have been carried out again; Wu Jian, Yao Jian, Zhu Shunxing etc., 2003), absorbancy is better after the visible dosing of result, and haemolysis degree (%) is starkly lower than 5% of national standard respectively 0.93 ± 0.72,1.10 ± 0.78, no hemolytic action.
Give that situations such as animal whole body general state, four limbs activity, feed, defecation, fur, eyes mucous membrane, breathing, behavior there is no unusually in the DK process (research method is seen Wang Beiying, Li Yikui chief editor. new Chinese medicine development technology and method, the p809 of Shanghai science tech publishing house; Liu Yaqian, Chen Hua, Li Chunhai etc. 2002), observe in experiment that to smear DK solution treated animal anti-infection ability strong, the incrustation healing is fast.Repeatedly to the DK test, intact skin is smeared vehicle and DK, there is no erythema and oedema, and stimulating score value is 0, shows that this medicine is non-stimulated to intact skin; The damaged skin group is smeared vehicle and DK vehicle group one to seven day all inadequate as seen erythema and oedema, stimulates score value 0.5, and reluctantly as seen DK 1-3 days erythema and oedema disappeared after the 3rd day substantially, showed that this medicine is non-stimulated substantially to damaged skin.
Embodiment
Embodiment 1, oil-in-water antimicrobial polypeptide emulsifiable paste:
Prescription: stearic acid 150g, Viscotrol C 50ml, Liquid Paraffin 32ml, glycerine 80ml, ethyl hydroxybenzoate 1g, trolamine 120ml, external application essence are an amount of, and distilled water adds to 1000g (the about 650ml of distilled water), antimicrobial polypeptide DK 65-100ug.
Compound method: adopt profit phase Hybrid Heating preparation method at present, antimicrobial polypeptide directly adds in the distilled water, distilled water is put in the heating kettle, adding stearic acid, Viscotrol C, Liquid Paraffin, glycerine, ethyl hydroxybenzoate heating make it fusion, put slightly and be chilled to about 70 ℃, slowly add trolamine, the limit edged stirs, cooling is stirred and is made it into cream, adds essence then and stirs promptly.The preparation of the more preceding method of this method is simple, temperature easy to control, and oil phase, water temperature are definitely identical, do not need water-bath to heat, and the finished product quality is also better, and this method is applicable to the preparation of most oil-in-water-type emulsifiable pastes.
Embodiment 2, antimicrobial polypeptide paste
Prescription: starch 150g, oxide powder and zinc 100g, ethanol are an amount of, Vaseline adds to 1000g, antimicrobial polypeptide DK 10-50ug.
Compound method: the Vaseline paste matrix mixing and stirring to about 60 ℃ of heating, by above-mentioned formula rate, zinc oxide and starch are inserted in the large-scale plastic tub, mix and rub thin mistake with the hands and sieve for No. 3.Gradation adds antimicrobial polypeptide, treats promptly cold.
Claims (23)
1. the antimicrobial polypeptide that does not have mutagenesis, described antimicrobial polypeptide is no mutagenesis.
2. the antimicrobial polypeptide of no mutagenesis according to claim 1 is characterized in that described antimicrobial polypeptide contains 22 amino acid at least.
3. the antimicrobial polypeptide of no mutagenesis according to claim 1 is characterized in that described antimicrobial polypeptide contains (AFVKLLKK) or (AFVKIMKK).
4. the antimicrobial polypeptide of no mutagenesis according to claim 1, the aminoacid sequence that it is characterized in that described antimicrobial polypeptide is CH
3CO-GIGKFLKKAKKFGKAFVKLLKK-NH
2Or CH
3CO-GIGKFLKKAKKFGKAFVKIMKK-NH
2
5. the antimicrobial polypeptide of no mutagenesis according to claim 3 is characterized in that the 1st, 3,13 amino acids are Gly in 22 amino acid of described antimicrobial polypeptide.
6. the antimicrobial polypeptide of no mutagenesis according to claim 3 is characterized in that in 22 amino acid of described antimicrobial polypeptide the 4th, 7,10,11, and 14,18 amino acids are Lys.
7. the antimicrobial polypeptide of no mutagenesis according to claim 3 is characterized in that amino acid A19 Lys connection 20A is Lys in the described antimicrobial polypeptide.
8. according to the antimicrobial polypeptide of the no mutagenesis of claim 3, it is characterized in that the 2nd amino acid is Ile in 22 amino acid of described antimicrobial polypeptide.
9. the antimicrobial polypeptide of no mutagenesis according to claim 3 is characterized in that the 5th, 12,16 amino acids are Phe in 22 amino acid of described antimicrobial polypeptide.
10. the antimicrobial polypeptide of no mutagenesis according to claim 3 is characterized in that the 6th, 20 amino acids is Leu in 22 amino acid of described antimicrobial polypeptide.
11. the antimicrobial polypeptide according to the no mutagenesis of claim 3 is characterized in that the 9th, 15 amino acids is Ala in 22 amino acid of described antimicrobial polypeptide.
12. the antimicrobial polypeptide according to the no mutagenesis of claim 3 is characterized in that the 17th amino acids is Val in 22 amino acid of described antimicrobial polypeptide.
13. the antimicrobial polypeptide according to the no mutagenesis of claim 3 is characterized in that described antimicrobial polypeptide the 20th amino acids is L or M.
14. the antimicrobial polypeptide of no mutagenesis according to claim 4 is characterized in that 15 amino acids that described antimicrobial polypeptide contains AFVKLLK or AFVKIMKK are Ala.
15. do not have the preparation method of the antimicrobial polypeptide of mutagenesis, it is characterized in that adopting solid state chemistry synthetic.
16. do not have the preparation method of the antimicrobial polypeptide of mutagenesis, it is characterized in that adopting biosynthetic means.
17. a mixture that contains the antimicrobial polypeptide that has or not mutagenesis is characterized in that this mixture contains the described antimicrobial polypeptide of a kind of 1-14 at least.
18. the mixture that contains the antimicrobial polypeptide that has or not mutagenesis according to claim 17 is characterized in that containing two peptide species.
19. the mixture that contains the antimicrobial polypeptide that has or not mutagenesis according to claim 18, what it is characterized in that two peptide species a kind ofly is CH
3CO-GIGKFLKKAKKFGKAFVKLLKK-NH
2Another kind is CH
3CO-GIGKFLKKAKKFGKAFVKIMKK-NH
2
20., it is characterized in that described mixture is the external-use drug form of creme, finish or aqua according to each described mixture that contains the antimicrobial polypeptide that has or not mutagenesis of claim 17-19.
21., it is characterized in that described mixture is oral, intramuscular injection or intravenous form according to each described mixture that contains the antimicrobial polypeptide that has or not mutagenesis of claim 17-19.
22. a method for the treatment of Gram-positive, gram negative bacterium and drug-fast bacteria infection is characterized in that using any formulation that contains the described mixture of with good grounds claim 1-16 to the infectation of bacteria patient.
23. a sterilizing agent is characterized in that containing any sterilizing agent formulation of the described mixture of with good grounds claim 1-16.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610017750 CN101070341A (en) | 2006-05-08 | 2006-05-08 | Antibacterial peptide without mutagenic action |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610017750 CN101070341A (en) | 2006-05-08 | 2006-05-08 | Antibacterial peptide without mutagenic action |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN101070341A true CN101070341A (en) | 2007-11-14 |
Family
ID=38897770
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 200610017750 Pending CN101070341A (en) | 2006-05-08 | 2006-05-08 | Antibacterial peptide without mutagenic action |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101070341A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102906105A (en) * | 2010-03-31 | 2013-01-30 | 诺瓦生命科学有限公司 | Compounds and their use |
| CN111494603A (en) * | 2020-05-23 | 2020-08-07 | 河南科技学院 | Antibacterial peptide nano ointment and preparation method and application thereof |
| CN112480234A (en) * | 2020-12-14 | 2021-03-12 | 上海交通大学 | Bioactive peptide AAGGYDVEKNNSRIKLGLK, and preparation method and application thereof |
| CN113214377A (en) * | 2020-01-21 | 2021-08-06 | 四川大学 | Antibacterial peptide AP-64 and preparation method and application thereof |
-
2006
- 2006-05-08 CN CN 200610017750 patent/CN101070341A/en active Pending
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102906105A (en) * | 2010-03-31 | 2013-01-30 | 诺瓦生命科学有限公司 | Compounds and their use |
| CN102906105B (en) * | 2010-03-31 | 2017-05-10 | 诺瓦生命科学有限公司 | Compounds and their use |
| CN113214377A (en) * | 2020-01-21 | 2021-08-06 | 四川大学 | Antibacterial peptide AP-64 and preparation method and application thereof |
| CN113214377B (en) * | 2020-01-21 | 2022-10-28 | 四川大学 | Antibacterial peptide AP-64 and preparation method and application thereof |
| CN111494603A (en) * | 2020-05-23 | 2020-08-07 | 河南科技学院 | Antibacterial peptide nano ointment and preparation method and application thereof |
| CN111494603B (en) * | 2020-05-23 | 2022-11-01 | 河南科技学院 | Antibacterial peptide nano ointment and preparation method and application thereof |
| CN112480234A (en) * | 2020-12-14 | 2021-03-12 | 上海交通大学 | Bioactive peptide AAGGYDVEKNNSRIKLGLK, and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106236708B (en) | Antimicrobial compositions containing free fatty acids | |
| DE69835279T2 (en) | COMPOSITIONS AND METHODS FOR THE TREATMENT OF INFECTIONS USING CATIONIC PEPTIDES ALONE OR IN COMBINATION WITH ANTIBIOTICS | |
| AU2002324481B2 (en) | Emu-based formulations for wound treatment related application information | |
| CA2850908C (en) | Aqueous antimicrobial composition containing coniferous resin acids | |
| JPH07330505A (en) | Antimicrobial composition | |
| EP1049378B1 (en) | An antimicrobial composition | |
| US5731007A (en) | Pharmaceutical composition for skin diseases | |
| CN101070341A (en) | Antibacterial peptide without mutagenic action | |
| KR102272901B1 (en) | Composition for external application to the skin containing meso-2,3-butanediol as a preservative | |
| CN102100735B (en) | Medicinal composition for treating skin diseases | |
| CN110279844A (en) | Artificial synthetic antimicrobial peptide is preparing application and inhibiting bacteria and diminishing inflammation acne-removing composition in inhibiting bacteria and diminishing inflammation acne-eliminating cosmetic or external medicine preparation | |
| RU2636530C2 (en) | Pharmaceutical compositions for treatment of wounds and burns | |
| ITRM20130657A1 (en) | TREATMENT OF DERMATOPATHIES BY VEGETAL EXTRACT BY DAPHNE LAUREOLA | |
| CN114366692B (en) | Oil-control acne-removing composition and application thereof | |
| RU2481834C2 (en) | Antimicrobial composition for treatment of wounds and burns | |
| CN109432201A (en) | Gynaecologic antibiotic gel and preparation method thereof | |
| WO2008028497A1 (en) | A composition of egg white copper complexes with medicinal benifits | |
| CN113230200A (en) | Ointment for treating bacterial skin diseases and preparation method thereof | |
| HU186767B (en) | Process for producing compositions for treating dermatologicalanomalys | |
| RU2708069C1 (en) | Agent for treating mastitis in cows | |
| RU2517065C1 (en) | Wound-healing drug | |
| CN102349920B (en) | Sulfanilamide medicament composition for wound surface | |
| KR101454874B1 (en) | Composition of anti-bacterial cosmetics containing msm | |
| CN113134075A (en) | Antibacterial peptide emulsifiable paste and preparation method thereof | |
| JPWO2008032847A1 (en) | Topical skin preparation |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |