CN1569896A - Preparation of composite specific yelk antibody igy against cutaneous infection and its use in treatment of cutaneous infection - Google Patents
Preparation of composite specific yelk antibody igy against cutaneous infection and its use in treatment of cutaneous infection Download PDFInfo
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- CN1569896A CN1569896A CN 03139875 CN03139875A CN1569896A CN 1569896 A CN1569896 A CN 1569896A CN 03139875 CN03139875 CN 03139875 CN 03139875 A CN03139875 A CN 03139875A CN 1569896 A CN1569896 A CN 1569896A
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Abstract
The invention discloses a skin infection resistant specific compound vitellus antibody IGY which can be prepared through, optimizing the main skin disease causing bacterium resulting in skin infection, preparing two or more single pathogenic agents causing skin infection, preparing immune eggs, preparing crude product of skin infection resistant pathogene specific yolk antibody, purifying the crude product, and mixing the single specific yolk antibody IGY proportionally.
Description
Technical field:
The present invention relates to a kind of anti-skin infections specific IgY and preparation method thereof, particularly relate to the preparation method and the combination preparation that are used to prevent and treat skin infections of suis property and mycotic infection of superficial part specificity composite IgY.
Background technology:
Treat skin infections of suis property and mycotic infection of superficial part tetter at present both at home and abroad, mainly use the medicine of microbiotic, chemical bactericide or hormones.But owing to cause that dermopathic Type B Streptococcus hemolyticus one class pathogenic bacterium resistance occurs to a lot of microbiotic, the particularly appearance of penicillin resistant and anti-tsiklomitsin and other Resistant strain, and be on the rise, add that microbiotic does not almost have killing action to superficial mycosis, so microbiotic is very limited to above-mentioned dermopathic curative effect.Chemical bactericide and hormone medicine are often cured the symptoms, not the disease, and toxic side effect is big, not only can't effect a radical cure tetter, increase the weight of the state of an illness sometimes on the contrary.The more important thing is and use in the above-mentioned treatment tetter process, can destroy the skin microecological balance, not only can not the kill harmful bacterium.Kill probiotics on the contrary, make the skin forfeiture resist the first road barrier that pathogenic bacterium are invaded, cause tetter to show effect repeatedly and be difficult to and cure.
Therefore, current still do not have ideal medicament ideal, environmental protection, that have no side effect to above-mentioned multiple cutaneous disease both at home and abroad.
Anti-skin infections specificity composite yolk antibody (immunoglobulin of yolk, IgY) belong to the IgG immunoglobulin like protein, main with five kinds of pathogenic bacterium immunity breeding hens that cause skin infections of suis property and mycotic infection of superficial part, make it to produce a large amount of corresponding immunoglobulin (Ig)s, make specific composite yolk antibody through extraction, separation, purifying.This compound antibody has specificity combination and restraining effect to the pathogenic bacterium that cause skin infections, and (tens of ng/ml) can bring into play useful effect under lower concentration.Its feature is that a pathogenic bacterium to focus plays restraining effect, and probiotics is then forbidden the slightest violation of the people's interests.The effect of this and microbiotic, hormones and pharmaceutical chemicals is distinct.With this compound antibody (IgY) treatment, both can protect and improve the microecological balance at skin focus position, can suppress pathogenic bacterium again, play and cure the disease and functions of prevention and health care.This prevents and treats the precedent that the tetter aspect is an initiative at home and abroad.
Technology contents:
The purpose of this invention is to provide the common suis property skin infections of a kind of direct inhibition and the specificity composite IgY of mycotic infection of superficial part pathogenic bacterium and the preparation method of preparation thereof; The present invention also aims to provide a kind of efficient, special efficacy biotechnological formulation of making by the specific IgY that can directly suppress Beta-hemolytic streptococcus, trichophyton, cotton-shaped surperficial tinea bacterium, Sabouraudites lanosus and microsporon gypseum, and preparation is made up of different formulations.
The objective of the invention is to be achieved through the following technical solutions:
1. preferably cause the representational pathogenic bacterium common, that multiple cutaneous is sick: according to the epidemiology survey result, (β-hemolyticstreptococcus), trichophyton (Trichophyton rubrum), acrothesium floccosum (E.floccosum), Sabouraudites lanosus (M.canis) and the little spore bacterium of gypsum sample (M.gypseum) are as representational tetter pathogenic agent to select the Type B Streptococcus hemolyticus.
2. antigen prepd: choose commercially available Type B Streptococcus hemolyticus, trichophyton, acrothesium floccosum, Sabouraudites lanosus and microsporon gypseum type strain, respectively by its cultural characters and optimal culture conditions, select the suitable culture base to cultivate and breeding so that ordinary method is a large amount of, collect culture, carry line, counting then, the dilution packing.Prepare single or complex antigen respectively.
(1) single antigen prepd: respectively each 20～3,000,000,000 counting of β Streptococcus hemolyticus, trichophyton, acrothesium floccosum, Sabouraudites lanosus and microsporon gypseum are made 0.5ml cause of disease body fluid, add excellent dose of 0.5ml Fu Shi, fully be mixed and make single antigen.
(2) complex antigen preparation: the single antigen that above branch is installed is by following mixed:
β Streptococcus hemolyticus 20%
Trichophyton 30%
Acrothesium floccosum 10%
Acrothesium floccosum 10%
Sabouraudites lanosus 30%
Microsporon gypseum 10%
The antigen of aforementioned proportion mixes the back and adds excellent dose of Fu Shi in 1: 1 ratio, puts into the high-speed homogenization machine again with 10000～30000rpm homogenize, makes complex antigen.
3. select to have (SPE) that the healthy no-special pathogen of high immunne response ability the carries bird (comprise bird inlay, the duck of laying eggs, the female goose of laying eggs, the lay eggs turkey and the ostrich etc. of laying eggs) of laying eggs and carry out reinforced immunological.To every health lay eggs chest muscle or the above-mentioned antigen that makes of subcutaneous injection of bird, in injection the last for the first time 7 days at interval, more for the second time, injected for the third time with same dose, method in 7 days at interval again after second injection with same dosage, method injection.For the first time high immune egg is got to begin to choose in the 20th day in the injection back.
4. immune egg is collected and the specific IgY preparation
With 0.5% bromogeramine or 0.1%KmnO
4Solution soaks sterilization in 15～30 minutes with immune egg, sterile distilled water washes, dries, put in the beating machiae eggshell is smashed, screen out egg white with yolk and get yolk, add 4～8 times of distilled water dilutings and stir, transfer pH to 5.5～6.0 with the NaoH of 1mol/L concentration or the HCl of 1mol/L concentration, left standstill liquid under 4 ℃, the supernatant diluent is got the supernatant liquor ultrafiltration with 10000～30000rpm high speed centrifugation 30 minutes, and ultrafiltrated concentrates 10～20 times.Concentrated solution is done the specific IgY crude product that promptly gets antidermatosis with the freeze-drying spray.Crude product changes post through ion exchange column and gel and carries out purifying, promptly gets the composite IgY elaboration, and the content that records IgY with the SDS-PAGE electrophoretic method surpasses 50%.
Adopting enzyme ether immunization (ELISA method) to record the specific IgY biological activity tires and reaches more than 512.
With prepared specificity composite IgY crude product or its purifying thing 0.02～80%, be equipped with 99.98～20% auxiliary material, can make washing lotion, sprays, ointment etc.The present invention directly imposes on skin focus with above formulation, pathogenic bacterium directly can be killed, and is effective, and instant effect does not produce any toxic side effects.The present invention is the biotechnological formulation of a specific specificity, pure natural.10%, 30%, 10% make 0.5ml bacterium liquid and add the abundant mixing of 0.5ml freund adjuvant and make complex antigen, immune hen is chosen egg and is prepared specificity composite IgY then.Detect the special composite IgY antibodies of anti-skin infections with the ELISA method and tire result such as following table:
| ????IgY | Detection antigen | Antibodies is tired |
| Anti-skin infections specificity composite IgY | The β Streptococcus hemolyticus | ????1∶512 |
| Trichophyton | ????1∶512 | |
| Acrothesium floccosum | ????1∶256 | |
| Sabouraudites lanosus | ????1∶512 | |
| Microsporon gypseum | ????1∶128 | |
| Alpha fungus | ????1∶128 | |
| Trichophyton | ????1∶256 |
From The above results, adopt the anti-skin infections specificity composite IgY of the present invention's preparation to have the ideal broad spectrum, not only five kinds of pathogenic bacterium to direct immunization have very strong restraining effect, and other superficial part pathomycete that does not have direct immunization is had inhibition work too.
Experimental example 2: make ointment with anti-skin infections specificity composite IgY of the present invention, following dermatosis patient is carried out clinical observation.
(1) selects 20 of 10 of impetigo herpetiformis patients, the tinea manuum and tinea pedis patients at random.
(2) use this IgY ointment that skin focus is imposed to embrocate, each is once sooner or later.
(3), disappear with 8 impetigo herpetiformis patient inflammation bases after 5 days through administration, 16 masterhands, tinea pedis patient's symptom obviously alleviate.
(4) successive administration is 10 days, and 8 impetigo herpetiformis patients cure, and 2 class diseases disappear substantially; 16 masterhands, tinea pedis patient cure substantially, 4 symptoms obviously alleviate.
Embodiment 1: prepare anti-skin infections specificity composite IgY
Respectively according to drawing cultural characters and the optimal culture conditions that holds Streptococcus sanguis, trichophyton, acrothesium floccosum, Sabouraudites lanosus, microsporon gypseum, select the suitable culture base, carry out bacterial strain and breed in a large number, collect culture, manage purifying, counting, dilution packing outward.At every turn with β Streptococcus hemolyticus and above-mentioned four kinds of fungies totally 50 hundred million, wherein β Streptococcus hemolyticus 20%, trichophyton 30%, acrothesium floccosum 10%, Sabouraudites lanosus 30%, microsporon gypseum 10%, make the 0.5ml mixed solution altogether, add Fu Shi again and help agent 0.5ml, fully mixing is made complex antigen.To 100 anosis Luo Si chicken immunes of laying eggs of health, every chest muscle injection 1ml injects the back and equally respectively injected the primary reinforcement immunity in the 7th day, the 14th day again with this complex antigen.The immunity back began to choose egg to the on the 20th day and stopped in 7 months, and superseded laying hen.
Immune egg is washed down with flowing water, 75% ethanol disinfection, dry, broken eggshell removes egg white with the yolk sieving, and leave and take yolk and add 6 times of distilled water and stir evenly, adjust pH to 6.0,4 ℃ of following standing over night were with centrifugal 30 minutes of this diluent 3000rpm.Getting supernatant liquor, is the ultra-filtration membrane ultrafiltration and concentration to 1/20 of 17～18KD with molecular weight cut-off, and concentrated solution is drying to obtain the specificity composite IgY crude product with cryospray.Crude product promptly gets the pure product of specificity composite IgY through ion exchange resin and gel exchange column layer folding.
Embodiment 2: the preparation of anti-immunoinfective specificity composite IgY ointment
Anti-skin infections specificity composite IgY 2.0%
Octadecanoic acid 6.0%
Whiteruss 9.0%
White vaseline 6.0%
Span-80????????????????????????????0.85％
Glycerine 11.0%
Potassium sorbate 0.1%
Tween-80 4.5%
Menthol 0.2%
Distilled water adds to 100%
Oil phase and water are heated to 80 ℃ respectively, and water is under agitation slowly added in the oil phase, stir to be condensed to about 40 ℃, add composite IgY solution, and fully grinding stirs makes water colour oil emulsifiable paste, promptly.
Embodiment 3: the preparation of anti-skin infections specificity composite IgY washing lotion
Get anti-skin infections specificity composite IgY lyophilized powder 150g, by 100 liters of anti-skin infections specificity composite IgY washing lotions of following formulated.
Anti-skin infections specificity composite IgY lyophilized powder 0.15%
Glycerine 1.5%
Press leaf oil 0.1%
Peppermint essence 0.2%
Borneol 0.1%
Ethanol 0.75%
Distilled water adds to 100 liters
Get 80 liters of distilled water earlier, add glycerine, composite IgY stirring and dissolving then; Dissolved adding in the ethanol of formula ratio by leaf oil, peppermint essence, borneol of formula ratio again, and under agitation slowly add in the aforementioned solution fully mixing, adjust pH to 6.5～7.0 are added distilled water to 100 liter at last, filter promptly.
Embodiment 4: the preparation of anti-skin infections specificity composite IgY sprays
Get anti-skin infections specificity composite IgY lyophilized powder 150g, allocate 100 liters of solution by following prescription:
Anti-skin infections specificity composite IgY 0.15%
Peppermint essence 0.1%
Fragrant citrus essence 0.1%
Glycerine 0.1%
Distilled water adds to 100 liters
Get 80 liters of distilled water earlier, add glycerine, composite IgY stirring and dissolving then; Peppermint essence, fragrant citrus essence are dissolved in an amount of ethanol, under agitation slowly add abundant mixing in the aforementioned solution, adjust pH value of solution value to 6.5～7.0; Add distilled water to 100 liter at last, filter the back can in the spray bottle promptly.
Claims (9)
1, anti-skin infections specificity composite yolk antibody (IGY) is characterized in that this antibody can be prepared by following method:
(A) preferably cause the main pathogenic bacterium of skin infections, preparation causes two or more the single pathogenic agent that causes skin infections of skin infections;
(B) use the pathogenic agent of step (A) to prepare immune egg respectively;
(C) prepare anti-skin infections pathogen specific yolk antibody (IGY) crude product, and crude product is carried out purifying;
(D) single special yolk antibody (IGY) is mixed in proportion, makes anti-skin infections specificity composite yolk antibody (IGY).
2, anti-skin infections specificity composite yolk antibody according to claim 1 (IGY), it is characterized in that the antigen described in the step (A) is by the single pathogenic agent preparation that causes skin infections and get, described cause of disease has five kinds, is respectively Beta-hemolytic streptococcus, trichophyton, cotton-shaped surperficial tinea bacterium, Sabouraudites lanosus and microsporon gypseum.
3, anti-skin infections specificity composite yolk antibody according to claim 2 (IGY), it is characterized in that step (D) be with multiple single specific yolk antibody (IGY) in 1～10: 1～10: 1～10: 1～10 ratio uniform mixing, make anti-skin infections specificity composite yolk antibody (IGY).
4, anti-skin infections specificity composite yolk antibody according to claim 3 (IGY), it is characterized in that step (D) be with multiple single specific yolk antibody (IGY) in 1: 1: 1: 1 ratio uniform mixing, make anti-skin infections specificity composite yolk antibody (IGY).
5, the anti-skin infections specificity composite yolk antibody (IGY) of the arbitrary claim of claim 1～4 is in the medicine of preparation treatment skin infections and the application in the sterilized sanitary accessories.
6, anti-skin infections specificity composite yolk antibody according to claim 5 (IGY) is in the application of treatment in the skin infections, it is characterized in that prepared composite yolk antibody (IGY) is equipped with 99.98～20% auxiliary material by 0.02～80% quality percentage composition makes anti-skin infections combination preparation.
7, the application of anti-skin infections specificity composite yolk antibody according to claim 6 (IGY) in the treatment skin infections is characterized in that described combination preparation is anti-skin infections washing lotion.
8, the application of anti-skin infections specificity composite yolk according to claim 6 (IGY) antibody in the treatment skin infections is characterized in that described combination preparation is anti-skin infections sprays.
9, the application of anti-skin infections specificity composite yolk antibody according to claim 6 (IGY) in the treatment skin infections is characterized in that described combination preparation is anti-skin infections ointment.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103890173A (en) * | 2011-08-19 | 2014-06-25 | 鸵鸟制药株式会社 | Antibody and antibody-containing composition |
| CN106279349A (en) * | 2016-08-31 | 2017-01-04 | 广东工业大学 | For preparing extracting method and the preparation method of yolk antibody thereof of the dermatophytosis suppressor proteins antigen of yolk antibody |
| CN106397586A (en) * | 2016-08-31 | 2017-02-15 | 广东工业大学 | Anti-dermatophyte specific yolk antibody, preparation method and application thereof |
| CN109010823A (en) * | 2018-08-27 | 2018-12-18 | 广州汇高生物科技有限公司 | Anti- various pathogens composition and its preparation process and application, spray and its preparation process |
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2003
- 2003-07-18 CN CN 03139875 patent/CN1569896A/en active Pending
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103890173A (en) * | 2011-08-19 | 2014-06-25 | 鸵鸟制药株式会社 | Antibody and antibody-containing composition |
| CN103890173B (en) * | 2011-08-19 | 2016-02-03 | 鸵鸟制药株式会社 | Antibody and the composition containing antibody |
| US9828419B2 (en) | 2011-08-19 | 2017-11-28 | Ostrich Pharma Kk | Antibody and antibody-containing composition |
| US10106599B2 (en) | 2011-08-19 | 2018-10-23 | Ostrich Pharma Kk | Antibody and antibody-containing composition |
| US10428138B2 (en) | 2011-08-19 | 2019-10-01 | Ostrich Pharma Kk | Antibody and antibody-containing composition |
| US11041016B2 (en) | 2011-08-19 | 2021-06-22 | Ostrich Pharma Kk | Compositions containing anti-HIV ostrich antibodies |
| CN106279349A (en) * | 2016-08-31 | 2017-01-04 | 广东工业大学 | For preparing extracting method and the preparation method of yolk antibody thereof of the dermatophytosis suppressor proteins antigen of yolk antibody |
| CN106397586A (en) * | 2016-08-31 | 2017-02-15 | 广东工业大学 | Anti-dermatophyte specific yolk antibody, preparation method and application thereof |
| CN106397586B (en) * | 2016-08-31 | 2019-12-06 | 广东工业大学 | specific yolk antibody for resisting dermatophytes and preparation method and application thereof |
| CN106279349B (en) * | 2016-08-31 | 2019-12-10 | 广东工业大学 | extraction method of dermatophyte cytoplasm protein antigen for preparing yolk antibody and preparation method of yolk antibody |
| CN109010823A (en) * | 2018-08-27 | 2018-12-18 | 广州汇高生物科技有限公司 | Anti- various pathogens composition and its preparation process and application, spray and its preparation process |
| CN109010823B (en) * | 2018-08-27 | 2022-03-18 | 广州汇高生物科技有限公司 | Composition for resisting various pathogenic bacteria, preparation process and application thereof, spray and preparation process thereof |
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