CN101411871A - Method for preparing chicken thymosin long-acting dosage form - Google Patents
Method for preparing chicken thymosin long-acting dosage form Download PDFInfo
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- CN101411871A CN101411871A CNA2008100534075A CN200810053407A CN101411871A CN 101411871 A CN101411871 A CN 101411871A CN A2008100534075 A CNA2008100534075 A CN A2008100534075A CN 200810053407 A CN200810053407 A CN 200810053407A CN 101411871 A CN101411871 A CN 101411871A
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Abstract
The invention discloses a method for preparing a chicken extrasin long-term slow release formulation which is capable of being injected intramuscularly once each week. The method is characterized in that the chicken extrasin long-term slow release formulation is prepared by the double emulsion solvent evaporation method; lactide/glycolide multipolymer is taken as microsphere carrier; a chicken extrasin is taken as a coating object; and the release characteristic in vitro of the prepared chicken extrasin long-term slow release formulation meets the characteristic of long-term preparation. The method has the advantages that the chicken extrasin long-term slow release formulation for injection overcomes the defects that the prior chicken extrasin injection needs frequent injection for a long term and has inconvenient use, low biological utilization rate and large using amount of medicine, can have antiviral effect for a longer term after once injection, and is an ideal chicken extrasin long-term slow release formulation. The chicken extrasin long-term slow release formulation is easy to prepare, easy and convenient to apply, and easy to be absorbed, and has better effect of treating chicken diseases.
Description
Technical field
The invention belongs to the technology of preparing of bio-pharmaceutical, particularly a kind of preparation method of chicken thymosin long-acting dosage form.
Background technology
Thymosin be one group by the excretory peptide hormone of thymic epithelial cell.It can regulate the immunologic function of humans and animals, and disease resistance enhancemen is treated many constitutionales and secondary immunodeficiency disease, can also be used for the treatment of anaphylaxis and infectious disease simultaneously.1966, Goldstein etc. reported the cattle thymus gland element of preliminary purification biologically active from calf thymus, later on from pig, sheep, and had extracted thymosin in people's tire thymus.In recent years, thymosin has been widely used in clinical medicine as a kind of immunomodulator.
Chicken thymosin is the hormone of purification biologically active from the pigeon chest gland.Studies show that chicken thymosin is in the external function that can not only make the lymphocyte that takes off the E receptor recover its E receptor, and can promote the lymphocyte increment of In vitro culture; And can strengthen the lymphocyte competence for added value of chicken, increase the content of Y thymosin in the chicken peripheral blood.In recent years, one of popular main threat that has become livestock and poultry breeding industry of immunosuppressive disease large tracts of land.Therefore, immunoregulatory factor such as chicken thymosin has been subjected to the extensive concern of Chinese scholars.In veterinary clinic test, chicken thymosin has also been obtained satisfied effect at animal epidemic aspect preventing and treating.
Chicken thymosin all is with the administration of injecting better therapeutic effect to be arranged, and many persons need inject 1-2 times every day, logotype 2-5 days, and bring a lot of inconvenience for the use of culturing the owner, therefore being made into long-acting dosage form just becomes the target that people pursue.Defectives such as existing chicken thymosin exists needs long-term frequent injection, use inconvenience, bioavailability is low and the drug use amount is big.
Therefore, providing a kind of preparation method of chicken thymosin long-acting dosage form, effectively prevent and treat various chicken diseases, is one of this technical field scientific research personnel new problem of being badly in need of developing.
Summary of the invention
The objective of the invention is for overcoming the weak point of prior art, provide a kind of and prepare simply, use preparation method convenient, the obvious results chicken thymosin long-acting dosage form.
A kind of preparation method of chicken thymosin long-acting dosage form is characterized in that comprising the steps:
(1) a certain amount of chicken thymosin is dissolved in the PBS buffer solution that contains stabilizing agent, makes aqueous phase solution (W1), the content of chicken thymosin is 0.5-50 mg/ml;
(2) hand over fat/glycolide copolymer to make oil-phase solution (O) with the dichloromethane dissolving with third, the content of third friendship fat/glycolide copolymer is 0.1-0.5 grams per milliliter; The above-mentioned aqueous phase solution (W1) that contains chicken thymosin is joined in the oil-phase solution (O), oil-phase solution (O) is 20: 1 to 2: 1 with the ratio of aqueous phase solution (W1), these two kinds of solution are stirred down for 5-30 ℃ in temperature, mixing speed is 500-8000 rev/mins, mixing time is 0.02-0.2 hour, mix homogeneously is made colostrum W1/O, preserves down at 4-10 ℃;
(3) polyvinyl alcohol fully is dissolved in water and makes aqueous phase solution (W2), the weight concentration of polyvinyl alcohol is 0.5-50%; Above-mentioned colostrum W1/O is joined aqueous phase solution (W2) to stir, whipping temp is 5-30 ℃, mixing speed is 100-3000 rev/mins, mixing time is 0.2-0.5 hour, the ratio of colostrum W1/O and aqueous phase solution (W2) is 1: 50 to 1: 200, make emulsion W1/O/W2, preserve down at 4-10 ℃;
(4) emulsion W1/O/W2 is transferred in the NaCl aqueous solution, stir, whipping temp is 5-30 ℃, and mixing speed is 500-3000 rev/mins, and mixing time is 0.2-0.5 hour, makes the organic solvent volatilization, centrifugal then collection microsphere;
(5) in order to keep the stable of solution, add certain amount of stabilizer, stabilizing agent directly joins in described preparation process (1) solution.
Described stabilizing agent is selected from one or more of trehalose, bovine serum albumin and Polyethylene Glycol.
The addition of described stabilizing agent is 0.1%-10% of a final solution.
Described chicken thymosin is reorganization chicken thymosin and the natural thymosin of chicken.
The beneficial effect of the inventive method is: this injection chicken thymosin long-acting dosage form can reduce chicken thymosin medication number of times, is a kind of ideal chicken thymosin long-acting dosage form.It is simple to have preparation technology, realizes easily, and it is convenient to use, characteristics such as required cost is low, this method adopt toxicity little, materials such as the dispersant of injection safety and oil phase, make the uniform dispersing and dissolving of chicken thymosin in oil phase, finally become a kind of long-acting dosage form of chicken thymosin.Adopt the long-acting dosage form experiment in vitro of the chicken thymosin that this method makes to show, disturbing element with chicken is the parcel object, prepared chicken thymosin microsphere form rounding, even particle size distribution, particle size distribution is below 40 microns, and drug loading can reach more than 6.0%, and envelop rate is more than 30.0%, external slow release can reach 4 all releases, meets the durative action preparation feature.Can bring into play the antiviral drug effect in a long time after the shot.Chicken thymosin is made microball preparation, can prolong drug action time, improve curative effect of medication and economic benefit greatly.
The specific embodiment
Below in conjunction with embodiment, to details are as follows according to the specific embodiment provided by the invention:
Embodiment 1
A kind of preparation method of chicken thymosin long-acting dosage form is characterized in that concrete implementation step is as follows:
(1) 120 milligrams of natural thymosins of chicken is dissolved in 2.0 milliliters of PBS buffer solution that contain 2% bovine serum albumin, makes aqueous phase solution (W1);
(2) hand over fat/glycolide copolymer to be dissolved in 4 milliliters of dichloromethane with 900.0 milligram third and make oil-phase solution (O);
(3) the above-mentioned aqueous phase solution (W1) that contains chicken thymosin is joined in the oil-phase solution (O), make these two kinds of solution under 15 ℃ of temperature, mixing speed is that 8000 rev/mins, mixing time are 0.05 hour, mix, make colostrum W1/O, preserve down at 4-10 ℃;
(4) above-mentioned colostrum W1/O is joined under 2000 rev/mins of stirring states in 400 milliliter 3% the polyvinyl alcohol, whipping temp is 5-30 ℃, and mixing time is 0.2 hour, makes emulsion W1/O/W2, preserves down at 4-10 ℃;
(5) emulsion W1/O/W2 is joined in the 1%NaCl aqueous solution of 2 times of emulsion volumes, makes organic solvent volatilization by 1000 rev/mins of stirrings, then 1300 rev/mins centrifugal 20 minutes, collect microsphere, deposit in refrigerator and cooled and hide.
Prepared chicken thymosin microsphere form rounding, even particle size distribution, mean diameter are at 20.6 microns, and drug loading can reach 8.8%, and envelop rate is 32.3%, and extracorporeal releasing experiment shows that slow release can reach for 3 weeks, meets the durative action preparation feature.Zoopery proves this novel thymosin except that the anti-fowl disease toxic action with thymosin, and its half-life in vivo is 4.2 times of common chicken thymosin, thereby has prolonged its action time in vivo.
Embodiment 2
A kind of preparation method of chicken thymosin long-acting dosage form is characterized in that concrete implementation step is as follows:
(1) 90.0 milligrams of chicken genetic engineering thymosins is dissolved in 2 milliliters of PBS buffer solution that contain 2% trehalose, makes aqueous phase solution (W1);
(2) hand over fat/glycolide copolymer to be dissolved in 4 milliliters of dichloromethane with 850.0 milligram third and make oil-phase solution (O);
(3) the above-mentioned aqueous phase solution (W1) that contains chicken thymosin is joined in the oil-phase solution (O), make these two kinds of solution under 15 ℃ of temperature, mixing speed is that 8000 rev/mins, mixing time are 0.05 hour, mix, make colostrum W1/O, preserve down at 4-10 ℃;
(4) above-mentioned colostrum W1/O is joined under 1500 rev/mins of stirring states in 300 milliliter 2% the polyvinyl alcohol, whipping temp is 5-30 ℃, and mixing time is 0.2 hour, makes emulsion W1/O/W2, preserves down at 4-10 ℃;
(5) emulsion W1/O/W2 is joined in the 3.5%NaCl aqueous solution of 3 times of emulsion volumes, make the organic solvent volatilization by 1500 rev/mins of stirrings, 2000 rev/mins then, centrifugal 15 minutes, collect microsphere, deposit in refrigerator and cooled and hide.
Prepared chicken thymosin microsphere form rounding, even particle size distribution, mean diameter are at 25.2 microns, and drug loading can reach 8.6%, and envelop rate is 39.8%, and extracorporeal releasing experiment shows that slow release can reach for 3 weeks, meets the durative action preparation feature.Zoopery proves this novel thymosin except that the anti-fowl disease toxic action with thymosin, and its half-life in vivo is 3 times of common chicken thymosin, thereby has prolonged its action time in vivo.
Embodiment 3
A kind of preparation method of chicken thymosin long-acting dosage form is characterized in that concrete implementation step is as follows:
(1) 45.0 milligrams of chicken thymosins is dissolved in 0.5 milliliter of PBS buffer solution that contains 2% Macrogol 4000, makes aqueous phase solution (W1);
(2) hand over fat/glycolide copolymer to be dissolved in 2.5 milliliters of dichloromethane with 1100.0 milligram third and make oil-phase solution (O);
(3) the above-mentioned aqueous phase solution (W1) that contains chicken thymosin is joined in the oil-phase solution (O), make these two kinds of solution under 15 ℃ of temperature, mixing speed is that 8000 rev/mins, mixing time are 0.05 hour, mix, make colostrum W1/O, preserve down at 4-10 ℃;
(4) above-mentioned colostrum W1/O is joined under 2000 rev/mins of stirring states in 400 milliliter 3% the polyvinyl alcohol, whipping temp is 5-30 ℃, and mixing time is 0.2 hour, makes emulsion W1/O/W2, preserves down at 4-10 ℃;
(5) emulsion W1/O/W2 is joined in the 2%NaCl aqueous solution of 3 times of emulsion volumes, makes organic solvent volatilization by 800 rev/mins of stirrings, then 1800 rev/mins centrifugal 10 minutes, collect microsphere, deposit in refrigerator and cooled and hide.
Prepared chicken thymosin microsphere form rounding, even particle size distribution, mean diameter are at 28.9 microns, and drug loading can reach 7.2%, and envelop rate is 35.8%, and extracorporeal releasing experiment shows that slow release can reach for 4 weeks, meets the durative action preparation feature.Zoopery proves this novel thymosin except that the anti-fowl disease toxic action with thymosin, and its half-life in vivo is 5.6 times of common chicken thymosin, thereby has prolonged its action time in vivo.
Above-mentioned detailed description of the preparation method of this chicken thymosin long-acting dosage form being carried out with reference to embodiment; be illustrative rather than determinate; can list several embodiment according to institute's limited range; therefore in the variation and the modification that do not break away under the general plotting of the present invention, should belong within protection scope of the present invention.
Claims (5)
1, a kind of preparation method of chicken thymosin long-acting dosage form is characterized in that comprising the steps:
(1) a certain amount of chicken thymosin is dissolved in the PBS buffer solution that contains stabilizing agent, makes aqueous phase solution (W1), the content of chicken thymosin is 0.5-50 mg/ml;
(2) hand over fat/glycolide copolymer to make oil-phase solution (0) with the dichloromethane dissolving with third, the content of third friendship fat/glycolide copolymer is 0.1-0.5 grams per milliliter; The above-mentioned aqueous phase solution (W1) that contains chicken thymosin is joined in the oil-phase solution (0), oil-phase solution (0) is 20: 1 to 2: 1 with the ratio of aqueous phase solution (W1), these two kinds of solution are stirred down for 5-30 ℃ in temperature, mixing speed is 500-8000 rev/mins, mixing time is 0.02-0.2 hour, mix homogeneously is made colostrum W1/0, preserves down at 4-10 ℃;
(3) polyvinyl alcohol fully is dissolved in water and makes aqueous phase solution (W2), the weight concentration of polyvinyl alcohol is 0.5-50%; Above-mentioned colostrum W1/0 is joined aqueous phase solution (W2) to stir, whipping temp is 5-30 ℃, mixing speed is 100-3000 rev/mins, mixing time is 0.2-0.5 hour, the ratio of colostrum W1/0 and aqueous phase solution (W2) is 1: 50 to 1: 200, make emulsion W1/0/W2, preserve down at 4-10 ℃;
(4) emulsion W1/0/W2 is transferred in the NaCl aqueous solution of 1-5%, stir, whipping temp is 5-30 ℃, and mixing speed is 500-3000 rev/mins, and mixing time is 0.2-0.5 hour, makes the organic solvent volatilization, centrifugal then collection microsphere;
(5) in order to keep the stable of solution, add certain amount of stabilizer, stabilizing agent directly joins in described preparation process (1) solution.
2, the preparation method of chicken thymosin long-acting dosage form according to claim 1 is characterized in that described third friendship fat/glycolide molecular weight of copolymer is 2000-5000Da.
3, the preparation method of chicken thymosin long-acting dosage form according to claim 1 is characterized in that described stabilizing agent is selected from one or more of trehalose, bovine serum albumin and Polyethylene Glycol.
4, the preparation method of chicken thymosin long-acting dosage form according to claim 1, the addition that it is characterized in that described stabilizing agent is 0.1%-10% of a final solution.
5, the preparation method of chicken thymosin long-acting dosage form according to claim 1 is characterized in that described chicken thymosin is reorganization chicken genetic engineering thymosin and the natural thymosin of chicken.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008100534075A CN101411871A (en) | 2008-06-03 | 2008-06-03 | Method for preparing chicken thymosin long-acting dosage form |
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| CNA2008100534075A CN101411871A (en) | 2008-06-03 | 2008-06-03 | Method for preparing chicken thymosin long-acting dosage form |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101669905B (en) * | 2009-09-08 | 2011-06-01 | 中国人民解放军第二军医大学 | Thymosin alpha1 injection-type subcutaneous implant |
| CN110430866A (en) * | 2017-03-03 | 2019-11-08 | Gtreebnt科技有限公司 | Stabilization external preparation containing extrasin beta -4 as effective component |
-
2008
- 2008-06-03 CN CNA2008100534075A patent/CN101411871A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101669905B (en) * | 2009-09-08 | 2011-06-01 | 中国人民解放军第二军医大学 | Thymosin alpha1 injection-type subcutaneous implant |
| CN110430866A (en) * | 2017-03-03 | 2019-11-08 | Gtreebnt科技有限公司 | Stabilization external preparation containing extrasin beta -4 as effective component |
| JP2020508970A (en) * | 2017-03-03 | 2020-03-26 | ジー−トゥリー・ビーエヌティ・カンパニー・リミテッドＧ−Ｔｒｅｅｂｎｔ Ｃｏ．， Ｌｔｄ． | Stabilized external preparation containing thymosin beta-4 as active ingredient |
| EP3590497A4 (en) * | 2017-03-03 | 2020-12-30 | G-Treebnt Co., Ltd. | Stabilized external use agent comprising thymosin beta-4 as effective ingredient |
| US11179443B2 (en) | 2017-03-03 | 2021-11-23 | G-Treebnt Co., Ltd. | Stabilized external preparation comprising thymosin beta 4 as an active ingredient |
| CN110430866B (en) * | 2017-03-03 | 2023-04-07 | Hlb医疗有限公司 | Stable external preparation containing thymosin beta-4 as active ingredient |
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Open date: 20090422 |