CN1463749A - Anti-sphagitis specificity composite IgY and preparation thereof - Google Patents
Anti-sphagitis specificity composite IgY and preparation thereof Download PDFInfo
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- CN1463749A CN1463749A CN02121244A CN02121244A CN1463749A CN 1463749 A CN1463749 A CN 1463749A CN 02121244 A CN02121244 A CN 02121244A CN 02121244 A CN02121244 A CN 02121244A CN 1463749 A CN1463749 A CN 1463749A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/02—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies from eggs
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P11/00—Drugs for disorders of the respiratory system
- A61P11/04—Drugs for disorders of the respiratory system for throat disorders
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/20—Immunoglobulins specific features characterized by taxonomic origin
- C07K2317/23—Immunoglobulins specific features characterized by taxonomic origin from birds
Abstract
The specific laryngopharyngitis resisting composite IgY preparation is prepared through following steps: selecting fine breed chicken with high immune response capacity, reinforced immunizing the selected chicken with composite antigen of complete components or thallus component, collecting the immune eggs laid by the immunized hens, preparing coarse powder of the laryngopharyngitis resisting composite IgY, purifying to obtain pure IgY and further preparing orally sprayed preparation. The preparation contains no harmful chemical matter and is safe and suitable for spraying orally.
Description
Technical field:
The present invention relates to a kind of composite IgY and preparation method thereof and preparation, particularly relate to a kind of anti-sphagitis specificity composite IgY and preparation method thereof and preparation
Background technology:
The method of diseases such as bacterial pharyngolaryngitis of current domestic and international treatment and tonsillitis is mainly used antibiotic, and the method is effective, but has several problems.The one, Resistant strain causes failing to respond to any medical treatment, and antibiotic lost efficacy has become global problem, and wherein Chinese Shanghai and two places, Hong Kong have the antibiotic more than 75% to lose efficacy.The 2nd, may bring out the generation of drug resistance strain, " antibacterial is drug resistance more and more, and antibiotic more and more lost efficacy " is the common problem of area that are fairly developed economically.The 3rd, the beneficial bacteria of oral cavity and throat has also been killed, cause dysbacteriosis, destroy microecological balance, produce adverse consequences; The 4th, can cause side effect such as diarrhoea, sensitivity response, fungal infection.Use Chinese herbal and crude drugs preparations to treat diseases such as bacterial pharyngolaryngitis and tonsillitis and also much see, as Herba Pileae Scriptae Tabellae, ice boron YANHOU PIAN, Waterelon Frost Lozenges, larynx treasured etc.The symptom effect of releiving of these medicaments is more outstanding than its antibiotic curative effect.Atmotherapy also is one of clinical common method, still to relax symptom as therapeutic purposes.Therefore at present the medicine that pharyngolaryngitis and tonsillitis are not still had ideal and have no side effect.
IgY (Immunoglobulin of yolk) genus IgG immunoglobulin like protein can combine with corresponding antigens generation specificity, thereby suppresses or changed the state or the activity of this antigen (as some antibacterial); Can promote segmented cell or macrophage engulfing by opsonic action to antibacterial; Can combine with viral, change the conformation of virus surface, stop virus to be adsorbed in permissive cell, simultaneously, the virus of formation---the IgY immune complex can be by macrophage phagocytic.
Technology contents:
The purpose of this invention is to provide a kind of direct inhibition or kill specific IgY of pharyngolaryngitis (pharyngolaryngitis) pathogenic bacterium and preparation method thereof; The present invention also aims to provide a kind of direct inhibition or kill the natural safety that the specific IgYs of pharyngolaryngitis (pharyngolaryngitis) pathogenic bacterium makes and have specific action, prevent and treat pharyngolaryngitis and tonsillitic preparation.
The present invention seeks to be achieved through the following technical solutions:
(1) the breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, difference immune hen, per injection 1ml antigen (about 2,000,000,000 above antibacterials).After injection for the first time, strengthen injection more once every two weeks, totally three times; In injecting the back 7th month for the first time, adopting the disclosed patented technology (number of patent application 00101270.3 of our company behind the egg labelling that these chickens produced, patent publication No. 1307061) extracts wherein IgY respectively, detect tiring of prepared IgY with ELISA method (enzyme-linked immunosorbent assay); Relatively it is tired again, selects wherein can make IgY and tire 〉=1: 128 the strong especially chicken of immunne response ability, and the egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of the preferred high immunne response ability of the present invention;
Above-mentioned hen can be selected from star chicken, leghorn chicken, brown blue chicken, Luo Man chicken;
(2) preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 40-60% by A bunch of Type B haemolysis type streptococcus, Diplococcus pneumoniae accounts for 35-25%, staphylococcus aureus accounts for 25-15%, the mixed homogenize or account for 45-30% by A bunch of Type B haemolysis type streptococcus, Diplococcus pneumoniae accounts for 15-25%, G bunch of Hemolytic streptococcus accounts for 10-15%, and staphylococcus aureus accounts for 10-20%, and Staphylococcus albus accounts for the mixed homogenize of 20-10%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, carry out the high speed pulverization homogenize, make the complex antigen of full bacterium and thalline composition with the 10000-30000rpm rotating speed;
(3) respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
(4) preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours-24 hours; Diluent is added in the high-speed centrifuge centrifugalize 20 minutes; Separating obtained supernatant added again carry out ultrafiltration in the ultrafilter and concentrate 10-20 doubly, the ratio in 0.1～0.3% adds sodium alginate in the slurry after concentrating, fully stirring; Add in the high speed centrifuge centrifugally again, get supernatant, remove lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product;
(5) purification of anti-sphagitis IgY: adopt the disclosed patented technology (number of patent application 00101270.3 of our company, patent publication No. 1307061) crosses ion exchange column and gel exchange column respectively the anti-sphagitis IgY crude extract of producing by above technology is carried out purification, promptly get pure IgY.
Use SDS-PAGE electrophoretic determination method, the anti-sphagitis IgY crude extract of producing by above technology is detected, recording wherein pure IgY content is more than 50%.
Adopting specifc immunity Activiation method of the present invention and the prepared anti-sphagitis composite IgY of improved Technology crude extract to carry out activity with reference to above-mentioned " ELISA method " detects, five kinds of pathogenic bacterium antibodies such as A bunch and G bunch of Hemolytic streptococcus, staphylococcus aureus, Diplococcus pneumoniae, Staphylococcus albus are tired to be reached more than 1: 256, has the antibodies of belonging to together of " common antigenic determinant " difference not of the same race or of the same race bunch and veriform antibacterial to tire to other with these 5 kinds of antibacterials and also reaches more than 1: 128.
The composite IgY crude extract of 0.02-80% or its purification thing are equipped with the adjuvant of 99.98-20%, can be made into clinical acceptable forms, as the product of dosage forms such as buccal tablet, nasal drop, nasal spray, mouth spraying agent.Be that Main Ingredients and Appearance is mixed with novel atomizing agent or buccal tablet with anti-sphagitis composite IgY crude extract or its purification thing particularly, the present invention directly imposes on the most concentrated throat of pathogenic bacterium with its design, and effect is direct, good effect, instant effect; Simultaneously, it has overcome conventional insecticide-applying way and will make medicine enter after the human circulation just generation effect and can bring all dysgenic disadvantages to human body.
The prepared anti-sphagitis composite IgY of the present invention is a kind of unique natural biological preparation, uses fool proof, without any side effects.Because it only directly produces had strong inhibitory effects at pharyngolaryngitis and tonsillitic infective pathogen body, and probiotics is but had good protective effect; Simultaneously, it can not bring out the generation of drug resistance strain fully, can be as " antibiotic " or other conventional medicines yet, but profitable strain is also all gone out together with pathogen and cause " dysbacteriosis "; Otherwise the anti-sphagitis composite IgY can make up a kind of normal flora ecology that does not produce inflammation in the throat, reaches the dual purpose of not only treating but also preventing.
Experimental example 1: mix with single A bunch Hemolytic streptococcus, single staphylococcus aureus, A bunch Hemolytic streptococcus and staphylococcus aureus, make three kinds of different antigen immune hens respectively, and produce three kinds of different I gY respectively as stated above.Detecting its antibodies with reference to " ELISA method " separately then tires.
| ??IgY | Detection antigen | Antibodies is tired |
| Anti-A bunch of streptococcus IgY | A bunch of Hemolytic streptococcus | ????1∶512 |
| Anti-staphylococcus aureus IgY | Staphylococcus aureus | ????1∶512 |
| Streptococcus, staphylococcus composite IgY | A bunch of Hemolytic streptococcus | ????1∶128 |
| Streptococcus, staphylococcus composite IgY | Staphylococcus | ????1∶128 |
| Streptococcus, staphylococcus composite IgY | Streptococcus and staphylococcus complex antigen | ????1∶256 |
After the above test of comparative analysis, we find: adopt several pathogenic bacterium to make complex antigen, the prepared composite IgY of this complex antigen immune hen of reuse, to complex antigen in conjunction with potency ratio to the single antigenic combination height of tiring.Because seldom single the having an effect of pathogenic bacterium of throat, all be just to produce symptoms such as inflammation by several pathogenic bacterium comprehensive functions, for example, staphylococcus aureus exists in the throat, the pathogenic bacterium of just having ready conditions, its individualism might not cause laryngopharyngitis, has only when Hemolytic streptococcus haunts, and it just can and then work.Therefore, with the complex antigen immune hen, prepared composite IgY, for the disease that is caused by various pathogens, effect is more far better than adopting the prepared IgY of single antigen immune hen.
Experimental example 2: the chicken kind of screening high immunne response
Make antigen with A bunch of Hemolytic streptococcus, with the immunity French " star chicken " respectively of same dosage, same injection system, Italian " leghorn chicken ", the U.S. " Luo Man chicken "; From these hens not of the same race are laid eggs, extract IgY respectively then, refer again to aforementioned " ELISA method " and detect tiring of these IgY one by one.
| The chicken kind | ??IgY | Antigen | Tire |
| France " star chicken " | Streptococcus IgY | A bunch of Hemolytic streptococcus | ????1∶64 |
| Italy's " leghorn chicken " | Streptococcus IgY | A bunch of Hemolytic streptococcus | ????1∶64 |
| The U.S. " Luo Man chicken " | Streptococcus IgY | A bunch of Hemolytic streptococcus | ????1∶128-256 |
From contrast test, can find out obviously that under the situation that condition is the same, other chicken kinds of potency ratio of the IgY that the immune U.S. " Luo Man chicken " is produced are much higher in every respect.
Experimental example 3: adopt the thalline composition to make antigenic antibodies potency test
Test and testing result see the following form:
| The used antigen of immunity | Immunity chicken kind | IgY | Detect used antigen | Antibodies is tired |
| The A bunch of full bacterium antigen of Type B Hemolytic streptococcus | Preferably " Luo Man chicken " | Anti-" Hemolytic streptococcus " IgY | A bunch of A type haemolysis type streptococcus | 1∶64 |
| A bunch of full bacterium of Type B Hemolytic streptococcus and thalline composition hybrid antigen | Preferably " Luo Man chicken " | Anti-" Hemolytic streptococcus " IgY | A bunch of A type haemolysis type streptococcus | 1∶256 |
Test and testing result confirm, adopt the thalline composition to make antigen, the made IgY that gets is tired to the antibodies of the antibacterial of non-immune other different shaped of the same race and hypotype increase substantially.This point is very important for the activity and the broad spectrum activity that improve prepared IgY.
Experimental example 4: adopt special multi-section position, large bolus injection mode to replace the reinforced immunological Activiation method antagonist of the muscle injection mode that generally adopts in conjunction with the influence of tiring.
Result of the test is as follows:
| ??IgY | Detect used antigen | Tire | |
| Before reinforced immunological activates | The anti-sphagitis composite IgY | Streptococcus, staphylococcal pneumonia diplococcus complex antigen | ??1∶128～1∶256 |
| After reinforced immunological activates | The anti-sphagitis composite IgY | Streptococcus, staphylococcal pneumonia diplococcus complex antigen | ??1∶256～1∶512 |
Adopt special multi-section position, large bolus injection mode to replace the prepared specificity composite IgY of reinforced immunological Activiation method of the muscle injection mode that generally adopts, the antibodies of a series of antibacterials with " common antigenic determinant " is tired all to be increased substantially.
Experimental example 5: the reinforced immunological Activiation method is to the influence of hen immune response time
After with the reinforced immunological Activiation method the extraordinary hen of being improved successively being carried out 3 immunity in 1 month, the immune response time of this kind hen extended to 13 months by 6 months.Promptly in 13 months time, in this hen is laid eggs, all maintain highly active antibody always, reduced the cost of IgY product significantly.
With 200 chickens being screened, the immunity of immune activation method is strengthened in above four of optional 10 employings; Simultaneously, select other 10 general " Luo Man chickens " in addition arbitrarily, adopt conventional method, make full bacterium antigen, with conventional these chickens of muscle injection mode immunity with five kinds of same antibacterials.Then, the different times after the inoculation first time extracts IgY with these two batches of different eggs that chicken was produced respectively, detects it more one by one and tires, and the result is as follows:
| The chicken kind | The used antigen of immunity | Injection system | The 1st injection back extracted IgY in 7th month and tired | The 1st injection back extracted IgY in 10th month and tired | The 1st injection back extracted IgY in 13rd month and tired |
| Generally " Luo Man chicken " | The complete average hybrid antigen of bacterium (various antibacterials respectively account for 20%) | Intramuscular injection | 1∶32 | 0 | 0 |
| Preferably " Luo Man chicken " | Full bacterium and thalline composition hybrid antigen | Subcutaneous and wing vein | 1∶256 | 1∶256 | 1∶128 |
Chang immune response time like this, the immune response time (be generally 4-5 individual month, the longest is 6 months) than the mechanism of other many research IgY of the whole world can realize when the hen of immunity is more than doubled.Reflected that a series of immune activation method of the present invention is advanced.
Experimental example 6: the immune effect of Different Chicken kind is observed
We choose " star chicken " 10, " leghorn chicken " 10, " brown blue chicken " 10, common " Luo Man chicken " 10, preferred " the Luo Man chicken " 10 of this test wantonly, adopt complex antigen of the present invention, use above-mentioned reinforced immunological Activiation method, respectively the above-listed five kinds of hens of immunity.Produce IgY respectively, detect it more separately and tire.
| The chicken kind | The used antigen of immunity | Injection system | Antibody titer |
| The star chicken | Five kinds of full bacterium of pathogenic bacterium such as A bunch of Hemolytic streptococcus and thalline composition complex antigen | Subcutaneous injection and wing intravenous injection combination | 1∶128 |
| Leghorn | Five kinds of full bacterium of pathogenic bacterium such as A bunch of Hemolytic streptococcus and thalline composition complex antigen | Subcutaneous injection and wing intravenous injection combination | 1∶128 |
| Brown blue chicken | Five kinds of full bacterium of pathogenic bacterium such as A bunch of Hemolytic streptococcus and thalline composition complex antigen | Subcutaneous injection and wing intravenous injection combination | 1∶128 |
| Common " Luo Man chicken " | Five kinds of full bacterium of pathogenic bacterium such as A bunch of Hemolytic streptococcus and thalline composition complex antigen | Subcutaneous injection and wing intravenous injection combination | 1∶128-256 |
| Preferably " Luo Man chicken " | Five kinds of full bacterium of pathogenic bacterium such as A bunch of Hemolytic streptococcus and thalline composition complex antigen | Subcutaneous injection and wing intravenous injection combination | 1∶256-512 |
Result of the test shows that immune activation method of the present invention can obviously activate and improve the immunne response function of some originally more blunt hens.
Experimental example 7: reduce antibacterial composition in the antigen, observe of the influence of immune activation method to the composite IgY broad spectrum activity.
To reduce by G bunch of Hemolytic streptococcus in five kinds of main pathogenic species in the antigen and Staphylococcus albus is rejected from antigen, again the special complex antigen of making full bacterium and thalline composition after three kinds of antigen antibacterial high speed pulverization homogenize remaining after simplifying, the same preferred breeding " Luo Man chicken " of above reinforced immunological Activiation method immunity that adopts extracts IgY then.At last, detect the composite IgY of the broad spectrum activity of gained respectively the antibodies of G bunch of Hemolytic streptococcus and Staphylococcus albus is tired, the result is as follows:
| The used antigen of immunity is formed | ??IgY | The antigen that is detected | Antibodies is tired |
| A bunch of Hemolytic streptococcus, Diplococcus pneumoniae, staphylococcus aureus | Anti-sphagitis wide spectrum composite IgY | G bunch of Hemolytic streptococcus | ??1∶128 |
| A bunch of Hemolytic streptococcus, Diplococcus pneumoniae, staphylococcus aureus | Anti-sphagitis wide spectrum composite IgY | Staphylococcus albus | ??1∶128 |
Find out from above test, thalline composition with the high speed pulverization gained is produced special antigen, other a series of immune activation methods have been adopted again, prepared anti-sphagitis composite IgY has ideal broad spectrum activity, not only to immune several pathogenic bacterium very strong inhibition effect is arranged, different shaped of the same race all there is ideal inhibitory action too with different bunches (as: A bunch of Hemolytic streptococcus and different bunches G bunch of Hemolytic streptococcus, the Staphylococcus albus of staphylococcus aureus and different genera).
Therefore, not only effect is remarkable for immune activation method of the present invention, and has very strong adaptability and motility.
Experimental example 8: anti-sphagitis IgY spray is treated 100 routine acute pharyngolaryngitises and tonsillitic facing
Bed observation of curative effect treatment fate pain symptom alleviates disappear routine number throat and the local red and swollen disappearance second day 17 58 58 routine local rednesses of tonsil sign first day 78 55 examples alleviate or the 3rd day 5 32 27 red and swollen disappearance of example that disappear of routine number pain symptom, and 5 examples alleviate the 4th day 055 red and swollen disappearance of example
The spray preparation is to causing the main pathogens of diseases such as pharyngolaryngitis and tonsillitis, as A family and G family streptococcus (streptococcus), golden yellow and Staphylococcus albus (staphylococci) and Diplococcus pneumoniae (S.pneumoniae), and not of the same race and genus, different shaped and hypotype all have streptococcus, staphylococcus and the Diplococcus pneumoniae of " common antigen ", all has special effect, so can fundamentally cure diseases such as the pharyngolaryngitis that causes by this bacterioid and tonsillitis.
Embodiment 1:
The preparation 100g anti-sphagitis IgY crude extract and the pure IgY technological process of production.
(1) the breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, respectively immune 1000 climing hens of sieve, per injection 1ml antigen after injection for the first time, is strengthened injection once every two weeks, totally three times again; In injection the last for the first time 7th month, egg labelling that these chickens produced is extracted wherein IgY respectively by the back conventional method, detect tiring of prepared IgY with ELISA method (enzyme-linked immunosorbent assay); Relatively it is tired again, selects wherein can make IgY and tire 〉=200 of strong especially chickens of 1: 128 immunne response ability.The egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of preferred high immunne response ability and select 40;
(2) preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 40% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15%, and G bunch of Hemolytic streptococcus accounts for 15%, and staphylococcus aureus accounts for 20%, and Staphylococcus albus accounts for 10% mixed homogenize; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, carry out the high speed pulverization homogenize, make the complex antigen of full bacterium and thalline composition with the 10000-30000rpm rotating speed;
(3) respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced 1000 on immune egg after 20 days;
(4) preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours; Diluent is added in the high-speed centrifuge, separated 20 minutes with 8000-12000rpm rotating speed high speed centrifugation; Separating obtained supernatant reentered carry out ultrafiltration in the ultrafilter and concentrate 15 times, the ratio in 0.2% adds alginic acid in the slurry after concentrating, fully stirring; It is centrifugal to add high speed centrifuge again, gets supernatant, removes lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product; Cross ion exchange column and gel exchange column anti-sphagitis IgY crude extract is carried out purification, promptly get pure IgY.
Embodiment 2:
The preparation 100g anti-sphagitis IgY crude extract and the pure IgY technological process of production.
(1) the breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, respectively immune 1000 climing hens of sieve, per injection 1ml antigen after injection for the first time, is strengthened injection once every two weeks, totally three times again; In injection the last for the first time 7th month, egg labelling that these chickens produced is extracted wherein IgY respectively by the back conventional method, detect tiring of prepared IgY with ELISA method (enzyme-linked immunosorbent assay); Relatively it is tired again, selects wherein can make IgY and tire 〉=200 of strong especially chickens of 1: 128 immunne response ability.The egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of preferred high immunne response ability and select 40;
(2) preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, staphylococcus aureus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above three kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; By accounting for 40-60% by A bunch of Type B haemolysis type streptococcus, Diplococcus pneumoniae accounts for 35-25% with cultured antibacterial, and staphylococcus aureus accounts for the mixed homogenize of 25-15%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, carry out the high speed pulverization homogenize, make the complex antigen of full bacterium and thalline composition with the 10000-30000rpm rotating speed;
(3) respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced 1000 on immune egg after 20 days;
(4) preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours; Diluent is added in the high-speed centrifuge, separated 20 minutes with 8000-12000rpm rotating speed high speed centrifugation; Separating obtained supernatant reentered carry out ultrafiltration in the ultrafilter and concentrate 15 times, the ratio in 0.2% adds alginic acid in the slurry after concentrating, fully stirring; It is centrifugal to add high speed centrifuge again, gets supernatant, removes lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product; Cross ion exchange column and gel exchange column anti-sphagitis IgY crude extract is carried out purification, promptly get pure IgY.
Embodiment 3:
Anti-sphagitis composite IgY crude extract or purification thing prepare mouth spray liquid
Compound anti-sphagitis pathogen IgY crude extract or purification thing 0.15%
Aspartame 0.12%
Herba Menthae essence 0.35%
Fragrant citrus essence 0.30%
Honey peach essence 0.10%
Distilled water adds to 100ml
Get the about 80.0ml of distilled water earlier, the order by the prescription composition successively adds and stirs then; Add water to 100ml at last, stir; By 0.22 μ membrane filtration degerming device filtration sterilization, obtain sterile liquid again, promptly prevent and treat pharyngolaryngitis IgY spray, adopt aseptic spray bottle fill.
Embodiment 4:
Compound anti-sphagitis pathogen IgY crude extract or purification thing 0.30%
Aspartame 0.24%
Herba Menthae essence 0.70%
Fragrant citrus essence 0.60%
Honey peach essence 0.20%
Distilled water adds to 100ml
Get the about 80.0ml of distilled water earlier, the order by the prescription composition successively adds and stirs then; Add water to 100ml at last, stir; By 0.22 μ membrane filtration degerming device filtration sterilization, obtain sterile liquid again, promptly prevent and treat pharyngolaryngitis IgY spray, adopt aseptic spray bottle fill.
Embodiment 5:
Compound anti-sphagitis pathogen IgY crude extract or purification thing 90g add the conventional dressing of 10g, make 500 of buccal tablets.
Claims (15)
1, a kind of anti-sphagitis composite IgY crude extract is characterized in that this IgY crude extract is prepared by following method:
The breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen is strengthened injection once every two weeks, totally three times again after injection for the first time; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=1: 128 the strong especially chicken of immunne response ability, and the egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of the preferred high immunne response ability of the present invention;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 45-30% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15-25%, and G bunch of Hemolytic streptococcus accounts for 10-15%, and staphylococcus aureus accounts for 10-20%, and Staphylococcus albus accounts for the mixed homogenize of 20-10%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, and the high speed pulverization homogenize makes the complex antigen of full bacterium and thalline composition;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours-24 hours; Diluent is added in the high-speed centrifuge centrifugalize 20 minutes; Separating obtained supernatant added again carry out ultrafiltration in the ultrafilter and concentrate 10-20 doubly, the ratio in 0.1～0.3% adds sodium alginate in the slurry after concentrating, fully stirring; Add in the high speed centrifuge centrifugally again, get supernatant, remove lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product.
2, a kind of anti-sphagitis composite IgY crude extract as claimed in claim 1, it is characterized in that this IgY crude extract is prepared by following method: the breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen, after injection for the first time, strengthen injection more once every two weeks, totally three times; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=200 of strong especially chickens of 1: 128 immunne response ability.The egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of preferred high immunne response ability;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 40% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15%, and G bunch of Hemolytic streptococcus accounts for 15%, and staphylococcus aureus accounts for 20%, and Staphylococcus albus accounts for 10% mixed homogenize; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, carry out the high speed pulverization homogenize, make the complex antigen of full bacterium and thalline composition with the 10000-30000rpm rotating speed;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours; Diluent is added in the high-speed centrifuge, separated 20 minutes with 8000-12000rpm rotating speed high speed centrifugation; Separating obtained supernatant reentered carry out ultrafiltration in the ultrafilter and concentrate 15 times, the ratio in 0.2% adds alginic acid in the slurry after concentrating, fully stirring; It is centrifugal to add high speed centrifuge again, gets supernatant, removes lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product.
3, anti-sphagitis composite IgY crude extract as claimed in claim 1 or 2, it is characterized in that complex antigen preparation in this IgY crude extract preparation method can also be chosen by A bunch of Type B haemolysis type streptococcus accounts for 40-60%, Diplococcus pneumoniae accounts for 35-25%, and staphylococcus aureus accounts for the mixed homogenize of 25-15%.
4, a kind of anti-sphagitis composite IgY as claimed in claim 1 or 2 is characterized in that the breeding hen in this IgY preparation method can be selected from star chicken, leghorn chicken, brown blue chicken or Luo Man chicken.
5, a kind of anti-sphagitis composite IgY as claimed in claim 3 is characterized in that the breeding hen in this IgY preparation method can be selected from star chicken, leghorn chicken, brown blue chicken or Luo Man chicken.
6, the pure IgY of a kind of anti-sphagitis is characterized in that this IgY is prepared by following method:
The breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen after injection for the first time, is strengthened injection once every two weeks, totally three times again; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=1: 128 the strong especially chicken of immunne response ability, and the egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of the preferred high immunne response ability of the present invention;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 45-30% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15-25%, and G bunch of Hemolytic streptococcus accounts for 10-15%, and staphylococcus aureus accounts for 10-20%, and Staphylococcus albus accounts for the mixed homogenize of 20-10%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, and the high speed pulverization homogenize makes the complex antigen of full bacterium and thalline composition;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours-24 hours; Diluent is added in the high-speed centrifuge centrifugalize 20 minutes; Separating obtained supernatant added again carry out ultrafiltration in the ultrafilter and concentrate 10-20 doubly, the ratio in 0.1～0.3% adds sodium alginate in the slurry after concentrating, fully stirring; Add in the high speed centrifuge centrifugally again, get supernatant, remove lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product;
Cross ion exchange column and gel exchange column anti-sphagitis IgY crude extract is carried out purification, promptly get pure IgY.
7, a kind of anti-sphagitis IgY as claimed in claim 6, it is characterized in that this IgY is prepared by following method: the breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen, after injection for the first time, strengthen injection more once every two weeks, totally three times; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=200 of strong especially chickens of 1: 128 immunne response ability.The egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of preferred high immunne response ability;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 40% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15%, and G bunch of Hemolytic streptococcus accounts for 15%, and staphylococcus aureus accounts for 20%, and Staphylococcus albus accounts for 10% mixed homogenize; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, carry out the high speed pulverization homogenize, make the complex antigen of full bacterium and thalline composition with the 10000-30000rpm rotating speed;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours; Diluent is added in the high-speed centrifuge, separated 20 minutes with 8000-12000rpm rotating speed high speed centrifugation; Separating obtained supernatant reentered carry out ultrafiltration in the ultrafilter and concentrate 15 times, the ratio in 0.2% adds alginic acid in the slurry after concentrating, fully stirring; It is centrifugal to add high speed centrifuge again, gets supernatant, removes lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product;
Cross ion exchange column and gel exchange column anti-sphagitis IgY crude extract is carried out purification, promptly get pure IgY.
8, as claim 6 or 7 described anti-sphagitis IgY, it is characterized in that complex antigen preparation in this IgY preparation method can also be chosen by A bunch of Type B haemolysis type streptococcus accounts for 40-60%, Diplococcus pneumoniae accounts for 35-25%, and staphylococcus aureus accounts for the mixed homogenize of 25-15%.
9, require 6 or 7 described a kind of anti-sphagitis composite IgYs as profit, it is characterized in that the breeding hen in this IgY preparation method can be selected from star chicken, leghorn chicken, brown blue chicken or Luo Man chicken.
10, a kind of preparation method of anti-sphagitis composite IgY crude extract is characterized in that this method is:
The breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen is strengthened injection once every two weeks, totally three times again after injection for the first time; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=1: 128 the strong especially chicken of immunne response ability, and the egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of the preferred high immunne response ability of the present invention;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 45-30% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15-25%, and G bunch of Hemolytic streptococcus accounts for 10-15%, and staphylococcus aureus accounts for 10-20%, and Staphylococcus albus accounts for the mixed homogenize of 20-10%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, and the high speed pulverization homogenize makes the complex antigen of full bacterium and thalline composition;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours-24 hours; Diluent is added in the high-speed centrifuge centrifugalize 20 minutes; Separating obtained supernatant added again carry out ultrafiltration in the ultrafilter and concentrate 10-20 doubly, the ratio in 0.1～0.3% adds sodium alginate in the slurry after concentrating, fully stirring; Add in the high speed centrifuge centrifugally again, get supernatant, remove lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product.
11, the preparation method of the pure IgY of a kind of anti-sphagitis is characterized in that this method is:
The breeding chicken of the preferred high immunne response ability of tool: make antigen with A bunch of hemolytic chain bacterium, the difference immune hen, per injection 1ml antigen after injection for the first time, is strengthened injection once every two weeks, totally three times again; In injecting the back 7th month for the first time, the IgY extracting respectively behind the egg labelling that these chickens produced wherein detects tiring of prepared IgY with the ELISA method; Relatively it is tired again, selects wherein can make IgY and tire 〉=1: 128 the strong especially chicken of immunne response ability, and the egg that this batch of reuse chicken is produced is hatched good chicken kind, treats that it grows up to 2-3 month, as the special chicken of the preferred high immunne response ability of the present invention;
The preparation of complex antigen: choosing A bunch of Type B Hemolytic streptococcus, Diplococcus pneumoniae, G bunch Hemolytic streptococcus, staphylococcus aureus, Staphylococcus albus is pharyngolaryngitis and the main pathogenic bacterium of tonsillitis; Buy commercially available above five kinds of main pathogenic bacterium bacterial strains, with bacteria culture media conventional cultivation in three gas incubators respectively; Cultured antibacterial is accounted for 45-30% by A bunch of Type B haemolysis type streptococcus, and Diplococcus pneumoniae accounts for 15-25%, and G bunch of Hemolytic streptococcus accounts for 10-15%, and staphylococcus aureus accounts for 10-20%, and Staphylococcus albus accounts for the mixed homogenize of 20-10%; In 1: 1 ratio freund adjuvant is added in the mixed body of the bacterial strain mix and to stir, make the full bacterium complex antigen of emulsus; Full bacterium complex antigen is put into the high-speed homogenization machine, and the high speed pulverization homogenize makes the complex antigen of full bacterium and thalline composition;
Respectively preferred special chicken is carried out reinforced immunological: use the complex antigen of being produced, the reinforced immunological method that adopts subcutaneous injection and wing intravenous injection to combine, respectively preferred special chicken is carried out immunity, every chicken per injection amount reaches 1-2ml antigen, is equivalent to hundred million antibacterials of 30-60; Strengthen injection more once every two weeks, be total to immunity three times in one month; Inoculation was for the first time searched chicken and is produced immune egg after 20 days;
The preparation of anti-sphagitis composite IgY crude extract: smash with the immune egg that eggbeater will be searched, remove Ovum Gallus domesticus album, stay egg yolk, stir with the sieving of egg yolk sieve; Measure the volume of the egg yolk of gained, extraordinarily go into distilled water, dilute and mix homogeneously by the 4-6 of this volume; Adjust between the pH to 5.5-6.0 with NaOH solution;
The diluent of adjusting pH value is further stirred, it is cooled to 2-6 ℃ then, left standstill 12 hours-24 hours; Diluent is added in the high-speed centrifuge centrifugalize 20 minutes; Separating obtained supernatant added again carry out ultrafiltration in the ultrafilter and concentrate 10-20 doubly, the ratio in 0.1～0.3% adds sodium alginate in the slurry after concentrating, fully stirring; Add in the high speed centrifuge centrifugally again, get supernatant, remove lipoprotein; Slurry after the centrifugalize is added ultrafilter, cross the ultra micro film, carry out filtration sterilization; Product after the filtration sterilization is carried out lyophilization with freezer dryer, promptly make anti-sphagitis composite IgY crude extract dry powder finished product;
Cross ion exchange column and gel exchange column anti-sphagitis IgY crude extract is carried out purification, promptly get pure IgY.
12,, it is characterized in that the breeding hen in this method can be selected from star chicken, leghorn chicken, brown blue chicken or Luo Man chicken as the preparation method of claim 10 or 11 described a kind of anti-sphagitis composite IgYs.
13, as the preparation method of claim 10 or 11 described anti-sphagitis IgY, it is characterized in that complex antigen preparation in this method can also be chosen by A bunch of Type B haemolysis type streptococcus accounts for 40-60%, Diplococcus pneumoniae accounts for 35-25%, and staphylococcus aureus accounts for the mixed homogenize of 25-15%.
14, various anti-sphagitis preparations, said preparation can be buccal tablet, nasal drop, nasal spray, pan work etc.It is characterized in that containing in the said preparation anti-sphagitis pathogen IgY crude extract or the purification thing of 0.02-80%.
15, a kind of anti-sphagitis mouth spraying agent is characterized in that this is characterized in that this mouth spraying agent is to be made by following component:
Compound anti-sphagitis pathogen IgY crude extract or purification thing 0.15-0.3%
Aspartame 0.12-0.24%
Herba Menthae essence 0.35-0.70%
Fragrant citrus essence 0.30-0.60%
Honey peach essence 0.10-0.20%
Distilled water adds to 100%
Priority Applications (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB021212449A CN1207057C (en) | 2002-06-11 | 2002-06-11 | Anti-sphagitis specificity composite IgY and preparation thereof |
| AU2003242070A AU2003242070A1 (en) | 2002-06-11 | 2003-05-08 | Anti-pharyngolaryngitis composition with igy and its agent |
| PCT/CN2003/000332 WO2003104280A1 (en) | 2002-06-11 | 2003-05-08 | Anti-pharyngolaryngitis composition with igy and its agent |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB021212449A CN1207057C (en) | 2002-06-11 | 2002-06-11 | Anti-sphagitis specificity composite IgY and preparation thereof |
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| Publication Number | Publication Date |
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| CN1463749A true CN1463749A (en) | 2003-12-31 |
| CN1207057C CN1207057C (en) | 2005-06-22 |
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| Application Number | Title | Priority Date | Filing Date |
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| CNB021212449A Expired - Fee Related CN1207057C (en) | 2002-06-11 | 2002-06-11 | Anti-sphagitis specificity composite IgY and preparation thereof |
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| Country | Link |
|---|---|
| CN (1) | CN1207057C (en) |
| AU (1) | AU2003242070A1 (en) |
| WO (1) | WO2003104280A1 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013004020A1 (en) * | 2011-07-07 | 2013-01-10 | 深圳市宝舜泰生物医药股份有限公司 | Pseudomonas aeruginosa (pa-msha) strain igy and preparation method and use thereof |
| CN104892756A (en) * | 2015-06-18 | 2015-09-09 | 成都安蒂康生物科技有限公司 | Antibacterial chicken egg yolk complex antibody as well as preparation method and application thereof |
| CN112957464A (en) * | 2021-02-24 | 2021-06-15 | 江秀坤 | Application of chicken egg yolk immunoglobulin in preparing throat-moistening health-care product |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2746394A4 (en) | 2011-08-19 | 2015-07-15 | Ostrich Pharma Kk | Antibody and antibody-containing composition |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1261454C (en) * | 2000-01-25 | 2006-06-28 | 雅臣药业集团（远东）有限公司 | Praparation of anticaries bacteria IgY, and anticaries composition |
-
2002
- 2002-06-11 CN CNB021212449A patent/CN1207057C/en not_active Expired - Fee Related
-
2003
- 2003-05-08 AU AU2003242070A patent/AU2003242070A1/en not_active Abandoned
- 2003-05-08 WO PCT/CN2003/000332 patent/WO2003104280A1/en not_active Application Discontinuation
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013004020A1 (en) * | 2011-07-07 | 2013-01-10 | 深圳市宝舜泰生物医药股份有限公司 | Pseudomonas aeruginosa (pa-msha) strain igy and preparation method and use thereof |
| CN104892756A (en) * | 2015-06-18 | 2015-09-09 | 成都安蒂康生物科技有限公司 | Antibacterial chicken egg yolk complex antibody as well as preparation method and application thereof |
| CN112957464A (en) * | 2021-02-24 | 2021-06-15 | 江秀坤 | Application of chicken egg yolk immunoglobulin in preparing throat-moistening health-care product |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1207057C (en) | 2005-06-22 |
| AU2003242070A1 (en) | 2003-12-22 |
| WO2003104280A1 (en) | 2003-12-18 |
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