CN104383537A - Circovirus egg yolk antibody oral liquid and preparation method thereof - Google Patents
Circovirus egg yolk antibody oral liquid and preparation method thereof Download PDFInfo
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- CN104383537A CN104383537A CN201410762711.2A CN201410762711A CN104383537A CN 104383537 A CN104383537 A CN 104383537A CN 201410762711 A CN201410762711 A CN 201410762711A CN 104383537 A CN104383537 A CN 104383537A
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Abstract
The invention discloses porcine a circovirus egg yolk antibody oral liquid and a preparation method thereof. The formula of the oral liquid is characterized by comprising the following raw materials in parts by volume: 95-105 parts of whole egg aqueous solution containing egg yolk antibodies, 0.2-0.3 part of caprylic acid, 0.2-0.3 part of formaldehyde and 0.01-0.02 part of thiomersal; the oral liquid further comprises mycillin, wherein the content of mycilline in the oral liquid is 800-1,000 unit/ml. During preparation, a clear egg yolk antibody oral liquid is prepared from whole hyperimmunity eggs. The egg yolk antibody oral liquid prepared by the method is less in loss in the production process, and has lots of immunologic active substances, high biological activity and no clinical side effect; meanwhile the oral liquid contains a great amount of various proteins and can reduce the adverse effects on antibodies caused by oral administration.
Description
Technical field
The invention belongs to pig biological product technical field, relate to a kind of pig circular ring virus 2 yolk antibody oral liquid and preparation method thereof.
Background technology
Pig circular ring virus, (Porcine circovirus, abbreviation: PCV) be a kind of minimum animal virus found so far.Existing known PCV has two serotypes, and namely PCV1 and PCV2, PCV2 are pathogenic virus, and it is the main pathogen of pmws (Postweaning Multisystemic Wasting Syndrome, PMWS).Pig has stronger susceptibility to PCV2, and infected pigs can discharge virus in the refuses such as nose liquid, feces, and direct oral cavity, respiratory tract approach infect the pig of all ages and classes.After farrowing sow infects PCV2, piglet can be infected through Placenta Hominis vertical transmission.
A kind of Mitochondrion IgG that yolk antibody IgY is, IgY is functionally equivalent to mammal IgG, but different in structure, has the space conformation of typical immunoglobulin.Yolk antibody has lot of superiority.(1) due to the gap of birds and mammal phylogenetics, birds are more suitable for the specific antibody of producing resisting mammal antigen.Yolk antibody can not excite complement system, does not react with the antibody of resisting mammal antigen, and not with the Fc receptors bind of mammal and antibacterial, this is significant in immunologic diagnosis.(2) yolk antibody has bioactive immunoglobulin as one, is storing, produces, processes, ingests and ensureing in digestion process that its stability is very crucial.Multitest shows, IgY has good stability, acidproof, alkaline-resisting, heat-resisting.Yolk antibody is more easily preserved, and deposits 6 months antagonist activity and have no significant effect under 4 DEG C of conditions under depositing 5 years or room temperature condition.
Summary of the invention
The invention provides a kind of pig circular ring virus 2 yolk antibody oral liquid and preparation method thereof, solve the pig circular ring virus 2 yolk antibody inconvenient problem with use of injection.
The object of the invention is to be realized by following form:
A kind of pig circular ring virus 2 yolk antibody oral liquid, comprises the following raw material calculated according to volume ratio: containing the shell egg aqueous solution 95-105 of yolk antibody, sad 0.2-0.3, formaldehyde 0.2-0.3, thimerosal 0.01-0.02; Also comprise the mycillin of 800-1000 units per ml oral liquid.
Its preparation process is:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological to chicken: to adopt three immune programme for children, every minor tick 10 days, every each vaccine consumption of chicken is 2ml, when antibody titer reaches 1:(128-256) time, carry out high-immunity egg collection;
2) 0.1-0.2% bromo geramine is used to carry out disinfection to high-immunity egg;
3) by 2) in the egg pulp of high-immunity egg carry out emulsifying, emulsification times 7-10min, then add pH be 4.2-4.3, temperature is the hcl acidifying water of 4-6 DEG C, the addition of hcl acidifying water is 5.5-6.5 times of egg pulp volume, after mixing, be cooled to 3-5 DEG C, leave standstill 18-24 hour;
4) get 3) in the supernatant, be warmed up to 25-32 DEG C, it is sad to add, stir 1-2h, leave standstill 10-12h, filter; Wherein sad volume: supernatant volume is (0.2-0.3): (95-105);
5) formaldehyde, thimerosal, mycillin is added, make the shell egg aqueous solution of yolk antibody: sad: formaldehyde: the volume ratio of thimerosal is (95-105): (0.2-0.3): (0.2-0.3): (0.01-0.02), and the content of mycillin is 800-1000 units per ml oral liquid;
6) subpackage, preserves.
Filtered by the filter membrane of 1.0mm, 0.45mm, 0.22mm successively when step 4) is filtered.
The present invention is by clarifying yolk antibody oral liquid to obtaining after the extraction of high-immunity egg shell egg, low temperature acidification, sad process.Shell egg uses in the extraction of yolk antibody by the present invention, can directly orally use clinically, containing a large amount of different kinds of proteins in Ovum Gallus domesticus album, the value of high-immunity egg can be made full use of, improve immunity and the stress ability of animal body, bioavailability is high, has the impregnable advantage of biological activity.
Detailed description of the invention
Embodiment 1:
The formula of pig circular ring virus 2 yolk antibody oral liquid comprises following raw material: containing shell egg aqueous solution 95 L of yolk antibody, sad 0.2 L, formaldehyde 0.3 L, thimerosal 0.02 L, and the mycillin of 1000 units per ml oral liquids.
Its preparation method comprises the following steps:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological, every each vaccine consumption of chicken is 2ml, and every minor tick 10 days, carries out three immune programme for children, when antibody titer reaches 1:256, starts to collect high-immunity egg.
2) high-immunity egg will be collected arrange, 0.2% bromo geramine sterilization, for subsequent use.
3) water dilution method acidification: by 2) step high-immunity egg shell egg squeeze into carry out in Pasteur's tank body fully emulsified, emulsification times 7min, then the egg pulp after emulsifying is added the hcl acidifying water that pH is 4.2, temperature is 4 DEG C, hcl acidifying water addition is 6 times of egg pulp volume, mixing, place in 4 DEG C of environment and staticly settle for 18 hours, get supernatant; Wherein, hcl acidifying water in water, adds hydrochloric acid make its pH value reach 4.2.
4) sad process; By the supernatant of step 3), the shell egg aqueous solution namely containing yolk antibody weighs, and be warmed up to 25 DEG C, it is sad to add, and fully mixes 1h, then leaves standstill 10h; Wherein, by volume, the supernatant: sad=95:0.2;
5) step 4) gained clear liquor is used successively to the membrane filtration of 1.0mm, 0.45mm, 0.22mm, leave and take filtrate;
6) to step 5) gained filtrate adds formaldehyde, thimerosal, mycillin; By volume, containing the shell egg shell egg aqueous solution of yolk antibody: sad: formaldehyde: thimerosal=95:0.2:0.3:0.02; The content of mycillin in this oral liquid is 1000 units per ml;
7) subpackage, preserves.
For determining yolk antibody oral liquid placement effect duration at normal temperatures, do following test: this product be kept in 37 DEG C of incubators, preserve 24 months, 1 week, 2 weeks respectively, January, February, April, August, December, after 16 months, 20 months, 24 months, by agar diffusion (AGP) method, the yolk antibody measuring each point in time sampling product is tired change, the results are shown in Table 1.
Table 1. room temperature places the change that oral liquid different time sections is tired
result shows:yolk antibody can deposit 16 months under the preservation of room temperature.
Quality standard:
[character] this product is weak yellow liquid.
[steriling test] is undertaken by " Chinese veterinary pharmacopoeia ", asepsis growth.
[mycoplasma inspection] is undertaken by " Chinese veterinary pharmacopoeia ", grows without mycoplasma.
[exogenous virus inspection] is undertaken by " Chinese veterinary pharmacopoeia ", conforms with the regulations.
[safety verification] with 7 Japanese instar chickling 5, each oral 5 parts, observe 2 weeks, all should be good for and live.
[efficacy test]
1) protection detects: with the young pigs 30 of 7 ages in days, wherein 10, often only oral 5 part yolk antibodies, and another 20 contrast isolated rearings.24h, get 10 rear virus liquids of young pig intramuscular injection inoculation PCV dilution in test group and matched group, record 96h death condition, the protective rate of test group is more than 90%, and counteracting toxic substances matched group mortality rate is more than 75%, and young pig is not any change blank group.
2) healing power measures: with the young pigs 120 of 7 ages in days, make negative control for 40, the virus liquid of 80 eye dripping inoculation PCV dilution, and after infection 12h, wherein 40, often only oral this product 5 parts, every day 1 time, continuous 3 days, another 40 were done positive control, isolated rearing.72h after treatment, record each group of death condition, counteracting toxic substances positive controls mortality rate is more than 70%, and treatment group survival rate is more than 100%, and healthy negative control group 40 is not only any change.
Embodiment 2:
The formula of pig circular ring virus 2 yolk antibody oral liquid comprises following raw material: containing shell egg aqueous solution 105 L of yolk antibody, sad 0.2 L, formaldehyde 0.2 L, thimerosal 0.01 L, and the mycillin of 800 units per ml oral liquids.
Its preparation method comprises the following steps:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological, every each vaccine consumption of chicken is 2ml, and every minor tick 10 days, carries out three immune programme for children, when antibody titer reaches 1:128, starts to collect high-immunity egg.
2) high-immunity egg will be collected arrange, 0.1% bromo geramine sterilization, for subsequent use.
3) water dilution method acidification: by 2) step high-immunity egg shell egg squeeze into carry out in Pasteur's tank body fully emulsified, emulsification times 10min, then the egg pulp after emulsifying is added the hcl acidifying water that pH is 4.3, temperature is 6 DEG C, hcl acidifying water addition is 5.5 times of egg pulp volume, mixing, place in 3 DEG C of environment and staticly settle for 24 hours, get supernatant; Wherein, hcl acidifying water in water, adds hydrochloric acid make its pH value reach 4.3.
4) sad process; By the supernatant of step 3), the shell egg aqueous solution namely containing yolk antibody weighs, and be warmed up to 32 DEG C, it is sad to add, and fully mixes 2h, then leaves standstill 12h; Wherein, by volume, the supernatant: sad=105:0.2;
5) step 4) gained clear liquor is used successively to the membrane filtration of 1.0mm, 0.45mm, 0.22mm, leave and take filtrate;
6) to step 5) gained filtrate adds formaldehyde, thimerosal, mycillin; By volume, containing the shell egg shell egg aqueous solution of yolk antibody: sad: formaldehyde: thimerosal=105:0.2:0.2:0.01; The content of mycillin in this oral liquid is 800 units per ml;
7) subpackage, preserves.
For determining yolk antibody oral liquid placement effect duration at normal temperatures, do following test: this product be kept in 37 DEG C of incubators, preserve 24 months, 1 week, 2 weeks respectively, January, February, April, August, December, after 16 months, 20 months, 24 months, by agar diffusion (AGP) method, the yolk antibody measuring each point in time sampling product is tired change, the results are shown in Table 1.The results are shown in Table 2.
Table 2. room temperature places the change that injection different time sections is tired
result shows:yolk antibody can deposit 16 months under the preservation of room temperature.
Quality standard:
[character] this product is weak yellow liquid.
[steriling test] is undertaken by " Chinese veterinary pharmacopoeia ", asepsis growth.
[mycoplasma inspection] is undertaken by " Chinese veterinary pharmacopoeia ", grows without mycoplasma.
[exogenous virus inspection] is undertaken by " Chinese veterinary pharmacopoeia ", conforms with the regulations.
[safety verification] with 7 Japanese instar chickling 5, each oral 5 parts, observe 2 weeks, all should be good for and live.
[efficacy test]
1) protection detects: with the young pigs 30 of 7 ages in days, wherein 10, often only oral 4 part yolk antibodies, and another 20 contrast isolated rearings.24h, get 10 rear virus liquids of young pig intramuscular injection inoculation PCV dilution in test group and matched group, record 96h death condition, the protective rate of test group is more than 95%, and counteracting toxic substances matched group mortality rate is more than 80%, and young pig is not any change blank group.
2) healing power measures: with the young pigs 120 of 7 ages in days, make negative control for 40, the virus liquid of 80 eye dripping inoculation PCV dilution, and after infection 12h, wherein 40, often only oral this product 4 parts, every day 1 time, continuous 3 days, another 40 were done positive control, isolated rearing.72h after treatment, record each group of death condition, counteracting toxic substances positive controls mortality rate is more than 78%, and treatment group survival rate is more than 90%, and healthy negative control group 40 is not only any change.
Embodiment 3:
The formula of pig circular ring virus 2 yolk antibody oral liquid comprises following raw material: containing shell egg aqueous solution 100 L of yolk antibody, sad 0.3 L, formaldehyde 0.3 L, thimerosal 0.02 L, and the mycillin of 1000 units per ml oral liquids.
Its preparation method comprises the following steps:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological, every each vaccine consumption of chicken is 2ml, and every minor tick 10 days, carries out three immune programme for children, when antibody titer reaches 1:256, starts to collect high-immunity egg.
2) high-immunity egg will be collected arrange, 0.2% bromo geramine sterilization, for subsequent use.
3) water dilution method acidification: by 2) step high-immunity egg shell egg squeeze into carry out in Pasteur's tank body fully emulsified, emulsification times 8min, then the egg pulp after emulsifying is added the hcl acidifying water that pH is 4.3, temperature is 5 DEG C, hcl acidifying water addition is 6.5 times of egg pulp volume, mixing, place in 5 DEG C of environment and staticly settle for 20 hours, get supernatant; Wherein, hcl acidifying water in water, adds hydrochloric acid make its pH value reach 4.3.
4) sad process; By the supernatant of step 3), the shell egg aqueous solution namely containing yolk antibody weighs, and be warmed up to 30 DEG C, it is sad to add, and fully mixes 2h, then leaves standstill 11h; Wherein, by volume, the supernatant=100:0.3;
5) step 4) gained clear liquor is used successively to the membrane filtration of 1.0mm, 0.45mm, 0.22mm, leave and take filtrate;
6) to step 5) gained filtrate adds formaldehyde, thimerosal, mycillin; By volume, containing the shell egg shell egg aqueous solution of yolk antibody: sad: formaldehyde: thimerosal=100:0.3:0.3:0.02; The content of mycillin in this oral liquid is 1000 units per ml;
7) subpackage, preserves.
Embodiment 4:
The formula of pig circular ring virus 2 yolk antibody oral liquid comprises following raw material: containing shell egg aqueous solution 98 L of yolk antibody, sad 0.3 L, formaldehyde 0.2 L, thimerosal 0.01 L, and the mycillin of 1000 units per ml oral liquids.
Its preparation method comprises the following steps:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological, every each vaccine consumption of chicken is 2ml, and every minor tick 10 days, carries out three immune programme for children, when antibody titer reaches 1:128, starts to collect high-immunity egg.
2) high-immunity egg will be collected arrange, 0.1% bromo geramine sterilization, for subsequent use.
3) water dilution method acidification: by 2) step high-immunity egg shell egg squeeze into carry out in Pasteur's tank body fully emulsified, emulsification times 9min, then the egg pulp after emulsifying is added the hcl acidifying water that pH is 4.3, temperature is 6 DEG C, hcl acidifying water addition is 5.5 times of egg pulp volume, mixing, place in 5 DEG C of environment and staticly settle for 22 hours, get supernatant; Wherein, hcl acidifying water in water, adds hydrochloric acid make its pH value reach 4.3.
4) sad process; By the supernatant of step 3), the shell egg aqueous solution namely containing yolk antibody weighs, and be warmed up to 28 DEG C, it is sad to add, and fully mixes 2h, then leaves standstill 12h; Wherein, by volume, the supernatant: sad=98:0.3;
5) step 4) gained clear liquor is used successively to the membrane filtration of 1.0mm, 0.45mm, 0.22mm, leave and take filtrate;
6) to step 5) gained filtrate adds formaldehyde, thimerosal, mycillin; By volume, containing the shell egg shell egg aqueous solution of yolk antibody: sad: formaldehyde: thimerosal=98:0.3:0.2:0.01; The content of mycillin in this oral liquid is 1000 units per ml;
7) subpackage, preserves.
With the young pig 90 of 7 ages in days, only feed embodiment 3 set product for first group 30, only feed embodiment 4 set product for second group 30,3rd group 30 only as positive controls, isolated rearing, intramuscular inoculation PCV dilute virus liquid, when showing clinical symptoms, start treatment, one group, two groups every draught animals take embodiment product 5ml, every day 1 time, continuous 3 days, 3rd group of normal saline 2 ml, every day 1 time, continuous 3 days.72h after treatment, records each group of morbidity and treatment situation.Table 3.
Table 3 experimental conditions
Embodiment 5:
The formula of pig circular ring virus 2 yolk antibody oral liquid comprises following raw material: containing shell egg aqueous solution 102 L of yolk antibody, sad 0.2 L, formaldehyde 0.3 L, thimerosal 0.01 L, and the mycillin of 800 units per ml oral liquids.
Its preparation method comprises the following steps:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological, every each vaccine consumption of chicken is 2ml, and every minor tick 10 days, carries out three immune programme for children, when antibody titer reaches 1:256, starts to collect high-immunity egg.
2) high-immunity egg will be collected arrange, 0.2% bromo geramine sterilization, for subsequent use.
3) water dilution method acidification: by 2) step high-immunity egg shell egg squeeze into carry out in Pasteur's tank body fully emulsified, emulsification times 10min, then the egg pulp after emulsifying is added the hcl acidifying water that pH is 4.2, temperature is 4 DEG C, hcl acidifying water addition is 6 times of egg pulp volume, mixing, place in 5 DEG C of environment and staticly settle for 24 hours, get supernatant; Wherein, hcl acidifying water in water, adds hydrochloric acid make its pH value reach 4.2.
4) sad process; By the supernatant of step 3), the shell egg aqueous solution namely containing yolk antibody weighs, and be warmed up to 30 DEG C, it is sad to add, and fully mixes 2h, then leaves standstill 12h; Wherein, by volume, the supernatant: sad=102:0.2;
5) step 4) gained clear liquor is used successively to the membrane filtration of 1.0mm, 0.45mm, 0.22mm, leave and take filtrate;
6) to step 5) gained filtrate adds formaldehyde, thimerosal, mycillin; By volume, containing the shell egg shell egg aqueous solution of yolk antibody: sad: formaldehyde: thimerosal=102:0.2:0.3:0.01; The content of mycillin in this oral liquid is 800 units per ml;
7) subpackage, preserves.
Claims (3)
1. a pig circular ring virus 2 yolk antibody oral liquid, is characterized in that: comprise the following raw material calculated according to volume ratio: containing the shell egg aqueous solution 95-105 of yolk antibody, sad 0.2-0.3, formaldehyde 0.2-0.3, thimerosal 0.01-0.02; Also comprise the mycillin of 800-1000 units per ml oral liquid.
2. a preparation method for pig circular ring virus 2 yolk antibody oral liquid as claimed in claim 1, is characterized in that concrete steps are:
1) adopt commercially available pig circular ring virus inactivated vaccine to carry out reinforced immunological to chicken: to adopt three immune programme for children, every minor tick 10 days, every each vaccine consumption of chicken is 2ml, when antibody titer reaches 1:(128-256) time, carry out high-immunity egg collection;
2) 0.1-0.2% bromo geramine is used to carry out disinfection to high-immunity egg;
3) by 2) in the egg pulp of high-immunity egg carry out emulsifying, emulsification times 7-10min, then add pH be 4.2-4.3, temperature is the hcl acidifying water of 4-6 DEG C, the addition of hcl acidifying water is 5.5-6.5 times of egg pulp volume, after mixing, be cooled to 3-5 DEG C, leave standstill 18-24 hour;
4) get 3) in the supernatant, be warmed up to 25-32 DEG C, it is sad to add, stir 1-2h, leave standstill 10-12h, filter; Wherein sad volume: supernatant volume is (0.2-0.3): (95-105);
5) formaldehyde, thimerosal, mycillin is added, make the shell egg aqueous solution of yolk antibody: sad: formaldehyde: the volume ratio of thimerosal is (95-105): (0.2-0.3): (0.2-0.3): (0.01-0.02), and the content of mycillin is 800-1000 units per ml oral liquid;
6) subpackage, preserves.
3. the preparation method of pig circular ring virus 2 yolk antibody oral liquid according to claim 2, is characterized in that: filtered by the filter membrane of 1.0mm, 0.45mm, 0.22mm successively when step 4) is filtered.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102716476A (en) * | 2012-05-31 | 2012-10-10 | 郑州后羿制药有限公司 | Cell inactivated vaccine, egg yolk antibody injection, and preparation method of cell inactivated vaccine |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102716476A (en) * | 2012-05-31 | 2012-10-10 | 郑州后羿制药有限公司 | Cell inactivated vaccine, egg yolk antibody injection, and preparation method of cell inactivated vaccine |
Non-Patent Citations (1)
| Title |
|---|
| 张小莺 等: "《卵黄抗体技术》", 31 May 2011, 科学出版社 * |
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Application publication date: 20150304 |