CN101612388A - One boar is used alpha-interferon high-efficient injection liquid and preparation method thereof - Google Patents
One boar is used alpha-interferon high-efficient injection liquid and preparation method thereof Download PDFInfo
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- CN101612388A CN101612388A CN200810154201A CN200810154201A CN101612388A CN 101612388 A CN101612388 A CN 101612388A CN 200810154201 A CN200810154201 A CN 200810154201A CN 200810154201 A CN200810154201 A CN 200810154201A CN 101612388 A CN101612388 A CN 101612388A
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Abstract
The invention discloses prescription of a kind of alpha-interferon high-efficient injection liquid and preparation method thereof.The present invention at first makes alpha-interferon the macromolecular compound of Polyethylene Glycol, add based on the curative effect reinforcing agent of polyinosini simultaneously and select special stabilizing agent for use, both kept the blood drug level of alpha-interferon, prolonged the plasma half-life of alpha-interferon, guaranteed the higher antiviral activity of alpha-interferon performance again, therefore, the clinical practice to alpha-interferon has crucial meaning.
Description
Technical field
The present invention relates to high-efficient injection liquid of alpha-interferon and preparation method thereof, relate to gama-interferon high-efficient injection liquid of the alpha-interferon that pig uses and preparation method thereof specifically, belong to field of pharmaceutical preparations.
Background technology
Interferon (Interferon, be called for short IFN) is to have bioactive glycoproteins such as antiviral, antitumor and immunoloregulation function by a class of emiocytosis under specific antigenic stimulus, can be divided into three types of α, β, γ.Wherein alpha-interferon is mainly produced by mononuclear cell, also is to study and use maximum interferoids at present.
The main biologic activity of IFN-α has: (1) suppresses virus replication, mainly is to synthesize plurality of enzymes and produce paracrine action by inducing cell; (2) suppress the increment of cell, as tumor cell etc.; (3) ability of reinforcement NK cell killing virus infected cell, the NK cell has interferon receptors; (4) change the expression of MHC molecule, strengthen the expression of MHC I quasi-molecule and suppress the expression of MHC II quasi-molecule.
Various natural and recombinants all have therapeutical effect, in case with their purification, separate and be prepared into medicine, just they can be carried out administration through parenteral route, are used for different medical indications.Yet, the alpha-interferon of parenteral route administration has immunogenicity, and plasma half-life is shorter, therefore, be difficult to messenger drug and keep medicative blood drug level in animal body with alpha-interferon, this also is the common problem that interferon exists in clinical practice.By with alpha-interferon and high molecular polymer synthesising complex, can effectively overcome these difficult problems.
Polyethylene Glycol (PEG) be one with-CH
2CH
2O-is the macromolecular linear polymer of base structure, is commonly used to carry out proteinic modification.By controlling PEG and the proteinic ratio degree of may command Pegylation to a certain extent.Pegylated protein has following characteristics: (1) increases dissolubility and stability; (2) change pharmacokinetics: the most significant characteristic of PEG-protein conjugate is the time that has prolonged body-internal-circulation, promptly with the time lengthening of organ, cells contacting.PEG also can protected protein, reduces proteinic decomposition.(3) change immunogenicity: many enzymes that therapeutic value arranged are not because of being that human protein has immunogenicity to the people.These enzymes can make immunogenicity reduce or disappearance after PEG modifies.In vitro study confirms that the bonded peg molecule of interferon is many more, and molecular weight is big more, and its physicochemical property is stable more, and the half-life is long more, but the loss of bioactivity of interferon is many more.Ideal to can be applicable to clinical Polyethylene Glycol interferon should be both balances, promptly keep strengthening the molecular weight of Polyethylene Glycol on the original bioactive basis of interferon as far as possible, prolongs its half-life.Pharmacokinetic confirms that the more common interferon of Polyethylene Glycol interferon infiltration rate is slow after administration, but behind the 3-4h, also can reach effective blood concentration that suppresses virus.
But, because the high molecular polymer of alpha-interferon can make the antiviral activity of interferon itself decrease, therefore need a kind of blood drug level that improves alpha-interferon that contains to help component of preserving steady in a long-term simultaneously, prepare the injection of the IFN-α (being PEG-IFN-α) of Pegylation.
Summary of the invention
The present invention is based on following inventive principle: the macromolecular compound of at first alpha-interferon being made Polyethylene Glycol, add curative effect reinforcing agent simultaneously based on polyinosini, both kept the blood drug level of alpha-interferon, prolong the plasma half-life of alpha-interferon, guaranteed the higher antiviral activity of alpha-interferon performance again.In addition, when preparation alpha-interferon high-efficient injection liquid, selected for use tween 80, made that prepared injection can preservation steady in a long-term as stabilizing agent.
Therefore, first purpose of the present invention provides a kind of alpha-interferon high-efficient injection liquid, comprises: PEG-IFN-α, polyinosini, osmotic pressure regulator, stabilizing agent, dispersant, antiseptic.Wherein, although polyinosini does not have obviously to improve the content of the IFN-α (being PEG-IFN-α) of Pegylation in the body, but it can keep the blood drug level of alpha-interferon, prolonged the plasma half-life of alpha-interferon, guarantee the higher antiviral activity of alpha-interferon performance again, therefore brought into play excellent synergism.
In a specific embodiments, the component of described injection is as follows:
Polyethylene Glycol alpha-interferon polymer 10000-10000000IU/mL
Polyinosini 1-2.5g
Osmotic pressure regulator 4-16g
Stabilizing agent 20-120mL
Dispersant 20-100mL
Antiseptic 0.1-1.0mL
Water for injection is to 1000mL
PH transfers to 6.0-8.0
In another embodiment, osmotic pressure regulator is a sodium chloride, and dispersant is a glycerol, and stabilizing agent is that polyoxyethylene 20 sorbitan monooleate (tween 80), antiseptic are m-cresol or phenol.Wherein, described stabilizer function is to make the preservation of interferon long-term stability and biological activity can not reduce.At present, more stabilizing agent is chosen or is raiseeed serum albumin (as bSA).Although end user or poultry serum albumin have certain advantage as active stabilizer, but people or poultry serum albumin are as the endogenous material from human body or animal, may carry disease virus such as multiple disease (for example hepatitis, acquired immune deficiency syndrome (AIDS), rabies virus etc.) and other infectious disease, thereby have highly potential danger in using clinically.Therefore, the used active stabilizer of the present invention is selected from tween 80 (polyoxyethylene 20 sorbitan monooleate), lysine, 2-HP-or soybean lecithin etc.Wherein, tween 80 has safe through using in the pharmaceutical preparation of a large amount of per os approach or non-digestive tract approach, advantages such as good dispersion discover that recently tween 80 has various biological and pharmacological activity, comprises anti-tumor activity etc., therefore, stabilizing agent tween 80 preferably.
Also in another embodiment, the component of described injection is as follows:
Polyethylene Glycol porcine alpha-interferon polymer 10
6IU/mL
Polyinosini 2g
NaCl?????????????????????????????????8g
Tween 80 50mL
Glycerol 80mL
Between phenyl methylcarbamate 0.7mL
Water for injection is to 1000mL
PH transfers to 7.0
Second purpose of the present invention is provided for preparing the method for above-mentioned high-efficient injection liquid, comprising:
(1) alpha-interferon of preparation Pegylation;
(2) add polyinosini, improve the antiviral activity of alpha-interferon;
(3) by after the following content mixing, degerming is handled, and promptly gets described high-efficient injection liquid
Polyethylene Glycol alpha-interferon polymer 10000-10000000IU/mL
Polyinosini 1-2.5g
Osmotic pressure regulator 4-16g
Stabilizing agent 20-120mL
Dispersant 20-100mL
Antiseptic 0.1-1.0mL
Water for injection is to 1000mL
PH transfers to 6.0-8.0
In a specific embodiments, the preparation process of the alpha-interferon of Pegylation is as follows: molecular weight polyethylene glycol is about 30KD, the adding proportion of Polyethylene Glycol and alpha-interferon is 5: 1 (mol ratio), by control buffer system pH value with the interferon Pegylation, pH of buffer is 3.0,0 ℃ of reaction 5h obtains Polyethylene Glycol alpha-interferon polymer.
In another embodiment, the stabilizing agent of interpolation is a tween 80, and dispersant is a glycerol, and antiseptic is a phenyl methylcarbamate.
Also in another embodiment, the component of described injection is preferably as follows:
Polyethylene Glycol porcine alpha-interferon polymer 10
6IU/mL
Polyinosini 2g
NaCl?????????????????????????????????8g
Tween 80 50mL
Glycerol 80mL
Between phenyl methylcarbamate 0.7mL
Water for injection is to 1000mL
PH transfers to 7.0
The 3rd purpose of the present invention provides the purposes of above-mentioned alpha-interferon high-efficient injection liquid in being used for the treatment of disease.
In a specific embodiments, described alpha-interferon is a porcine alpha-interferon, described purposes, and wherein said disease is epidemic diarrhea, swine fever and transmissible gastroenteritis, or is used for the treatment of epidemic diarrhea, swine fever and the transmissible gastroenteritis disease of pig.
The invention effect:
1. the alpha-interferon high-efficient injection liquid has prolonged the plasma half-life of alpha-interferon itself, simultaneously, has improved the antiviral activity of alpha-interferon;
2. the alpha-interferon high-efficient injection liquid provides a kind of more efficiently method for the clinical practice of alpha-interferon;
3. zest solvent such as injection is soft is to pathological changes mucosa nonirritant;
4. dosage is big, is particularly useful for the administration of polypeptide and protein medicaments.
5. use tween 80 as stabilizing agent, tween 80 is used in the pharmaceutical preparation of a large amount of per os approach or non-digestive tract approach, has advantages such as safe, good dispersion, and has various biological and pharmacological activity, comprises anti-tumor activity etc.
The specific embodiment:
Be described in further detail the present invention below by embodiment
Embodiment 1: the preparation of porcine alpha-interferon high-efficient injection liquid
With the porcine alpha-interferon is example: get the Polyethylene Glycol that molecular weight is 30KD, being added into pH with 5: 1 in molar ratio ratio of porcine alpha-interferon is in 3.0 the Tris buffer, and 0 ℃ of reaction 5h obtains Polyethylene Glycol porcine alpha-interferon polymer.
Press following set of dispense system injection A:
Polyethylene Glycol porcine alpha-interferon polymer 10
6IU/mL
Polyinosini 2g
NaCl?????????????????????????????8g
Tween 80 50mL
Glycerol 80mL
Between phenyl methylcarbamate 0.7mL
Water for injection is to 1000mL
PH transfers to 7.0
Press following set of dispense system injection B:
Polyethylene Glycol porcine alpha-interferon polymer 10
6IU/Ml
Polyinosini 1g
NaCl?????????????????????????????4g
Tween 80 20mL
Glycerol 20mL
Between phenyl methylcarbamate 0.1mL
Water for injection is to 1000mL
PH transfers to 7.0
Press following set of dispense system injection C:
Porcine alpha-interferon 10
6IU/mL
Polyinosini 2.5g
NaCl????????????????????????16g
Tween 80 120mL
Glycerol 100mL
Between phenyl methylcarbamate 1.0mL
Water for injection is to 1000mL
PH transfers to 7.0
Embodiment 2: the activity level test of porcine alpha-interferon high-efficient injection liquid
According to above-mentioned dilution proportion porcine alpha-IFN stock solution and Polyethylene Glycol porcine alpha-interferon polymer solution, adopt the half cell to suppress the pathological changes method, use the antiviral activity of interferon in each solution of VSV-MDBK system measurement.(method is with reference to Pharmacopoeia of the People's Republic of China version in 2005) experimental result is as follows:
| Sample | Antiviral activity (IU/mL) |
| Porcine alpha-interferon | ??2.23×10 6 |
| The PEG-porcine alpha-interferon | ??0.67×10 6 |
| Injection A:PEG-porcine alpha-interferon+reinforcing agent | ??2.57×10 6 |
| Injection B:PEG-porcine alpha-interferon+reinforcing agent | ??2.27×10 6 |
| Injection C:PEG-porcine alpha-interferon+reinforcing agent | ??2.39×10 6 |
Annotate: reinforcing agent is a polyinosini
Data show that porcine alpha-interferon injection of the present invention increases than the antiviral activity of porcine alpha-interferon itself.
Embodiment 3: the pharmacokinetics of porcine alpha-interferon high-efficient injection liquid
With the porcine alpha-interferon is example, carries out pharmacokinetic: adopt
125The I labelling method.Specific operation process is seen Figure of description.Experimental result such as following table:
| Sample | Half-life (h) |
| Porcine alpha-interferon | ??0.9-1.8 |
| The PEG-porcine alpha-interferon | ??2.5-4.3 |
| PEG-porcine alpha-interferon+reinforcing agent | ??2.8-5.0 |
Annotate: reinforcing agent is a polyinosini
Data show that porcine alpha-interferon injection of the present invention all prolongs to some extent than the plasma half-life of porcine alpha-interferon stock solution and PEG-porcine alpha-interferon.
Analysis-by-synthesis embodiment 2 and embodiment 3, porcine alpha-interferon high-efficient injection liquid of the present invention had both prolonged the plasma half-life of porcine alpha-interferon itself, simultaneously, improved the antiviral activity of porcine alpha-interferon again, for the clinical practice of porcine alpha-interferon provides more efficiently method.
Description of drawings
Accompanying drawing is for using
125The I labelling method carries out the flow chart of pharmacokinetic to porcine alpha-interferon.
Claims (10)
1, a kind of alpha-interferon high-efficient injection liquid, its component is as follows: PEG-IFN-α, reinforcing agent, osmotic pressure regulator, stabilizing agent, dispersant, antiseptic and water for injection, wherein reinforcing agent is selected from polyinosini, and stabilizing agent is selected from polyoxyethylene 20 sorbitan monooleate (tween 80), lysine, 2-HP-or soybean lecithin.
2, alpha-interferon high-efficient injection liquid according to claim 1, wherein osmotic pressure regulator is a sodium chloride, dispersant is that glycerol, stabilizing agent are that polyoxyethylene 20 sorbitan monooleate (tween 80), antiseptic are m-cresol or phenol.
3, alpha-interferon high-efficient injection liquid according to claim 2, wherein component is as follows
Polyethylene Glycol alpha-interferon polymer 10000-10000000IU/mL
Polyinosini 1-2.5g
Osmotic pressure regulator 4-16g
Stabilizing agent 20-120mL
Dispersant 20-100mL
Antiseptic 0.1-1.0mL
Water for injection is to 1000mL
PH transfers to 6.0-8.0
4, alpha-interferon high-efficient injection liquid according to claim 3, wherein component is as follows:
Polyethylene Glycol porcine alpha-interferon polymer 10
6IU/mL
Polyinosini 2g
NaCl?????????????????????????8g
Tween 80 50mL
Glycerol 80mL
Between phenyl methylcarbamate 0.7mL
Water for injection is to 1000mL
PH transfers to 7.0
5, according to each described alpha-interferon high-efficient injection liquid among the claim 1-4, wherein alpha-interferon is a porcine alpha-interferon.
6, a kind of method that is used to prepare the described alpha-interferon high-efficient injection liquid of claim 1-5, it is characterized in that: at first prepare the Polyethylene Glycol alpha-interferon, then by add the antiviral activity that polyinosini improves alpha-interferon in injection, in addition corresponding dispersant, osmotic pressure regulator, stabilizing agent and antiseptic are used as the highly efficient intensifying agent of alpha-interferon injection simultaneously.
7, the described method of claim 6, wherein stabilizing agent is selected from polyoxyethylene 20 sorbitan monooleate (tween 80), lysine, 2-HP-or soybean lecithin, osmotic pressure regulator is selected from sodium chloride, and dispersant is selected from glycerol, and antiseptic is selected from a phenyl methylcarbamate.
8, claim 6 or 7 described methods, wherein the preparation process of the alpha-interferon of Pegylation is as follows: molecular weight polyethylene glycol is about 30KD, the adding proportion of Polyethylene Glycol and alpha-interferon is 5: 1 (mol ratio), by control buffer system pH value with the interferon Pegylation, pH of buffer is 3.0,0 ℃ of reaction 5h obtains Polyethylene Glycol alpha-interferon polymer.
9, the prepared alpha-interferon high-efficient injection liquid of the described alpha-interferon high-efficient injection liquid of claim 1-5, or the method for claim 6-8 is used for preparing the purposes of disease treatment.
10, the described purposes of claim 9, wherein said disease is epidemic diarrhea, swine fever and transmissible gastroenteritis, or is used for the treatment of epidemic diarrhea, swine fever and the transmissible gastroenteritis disease of pig.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112244025A (en) * | 2020-11-27 | 2021-01-22 | 上海创宏生物科技有限公司 | Sterile treatment agent for macromolecular protein and/or nucleic acid and preparation method and application thereof |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112244025A (en) * | 2020-11-27 | 2021-01-22 | 上海创宏生物科技有限公司 | Sterile treatment agent for macromolecular protein and/or nucleic acid and preparation method and application thereof |
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Open date: 20091230 |