CN115068592A - Recombinant human interferon alpha 1b eye drops stable at normal temperature after unsealing and preparation method thereof - Google Patents
Recombinant human interferon alpha 1b eye drops stable at normal temperature after unsealing and preparation method thereof Download PDFInfo
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- CN115068592A CN115068592A CN202210734636.3A CN202210734636A CN115068592A CN 115068592 A CN115068592 A CN 115068592A CN 202210734636 A CN202210734636 A CN 202210734636A CN 115068592 A CN115068592 A CN 115068592A
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- interferon alpha
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- human interferon
- eye drops
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Abstract
The invention discloses an eye drop of recombinant human interferon alpha 1b stable at normal temperature after unsealing and a preparation method thereof, relating to the field of pharmaceutical preparations, the eye drop comprises recombinant human interferon alpha 1b, and also comprises one or more of a protective agent, a stabilizing agent, an additional regulating agent and an osmotic pressure regulating agent, the eye drop can be stored for a long time under the condition of low temperature, can be stored for a long time under the condition of normal temperature without light protection after unsealing, can be stored for at least 24 months under the condition of light protection at 2-8 ℃ (conventional storage requirement), can be stored for at least 9 months under the condition of light protection at normal temperature (25 ℃ -35 ℃) without light protection after unsealing, can be stored for at least 9 days under the condition of normal temperature (25 ℃ -35 ℃) without light protection without resealing after unsealing, and can guarantee sterility during the storage period, and has no obvious change in the pH value and osmotic pressure of products, has no visible foreign matter and stable biological activity.
Description
Technical Field
The invention relates to the field of pharmaceutical preparations, in particular to an unsealed recombinant human interferon alpha 1b eye drop stable at normal temperature and a preparation method thereof.
Background
Interferon is a kind of protein medicine of cell factors which is originally produced by animal body and has broad-spectrum antiviral, antiproliferative and immunoregulatory actions, and can be divided into three large types of alpha, gamma and beta according to the difference of its production site and action mechanism, and each large type can be divided into several small subtypes, among several large types, alpha type is the most widely used one, and the interferon of this type which is clinically used at present mainly includes interferon alpha 2a, interferon alpha 2b, interferon alpha 1b, etc. In 1980, foreign scientists cloned interferon alpha 2a and interferon alpha 2b genes from European and American human bodies in sequence and expressed them in colibacillus recombinantly, making it possible to use them as drugs for clinical treatment. In 1981, the maryund academy cloned the alpha 1 interferon gene from umbilical cord blood leukocyte of healthy Chinese by modern molecular biology technology, and later named alpha 1b by International Commission on interferon naming. The recombinant human interferon alpha 1b and the recombinant human interferon alpha 2b have essential differences: although all are proteins, the recombinant human interferon alpha 1b is a single-chain polypeptide consisting of 166 amino acid residues, and the theoretical value of the molecular weight is 19.4KD +/-1.9 KD; the recombinant human interferon alpha 2b is a single-chain polypeptide consisting of 165 amino acid residues, and the theoretical value of the molecular weight is 19.2KD +/-1.9 KD; their primary structure differs by one amino acid, which also leads to their serial differences. Interferon alpha 1b, derived from the chinese gene, was later shown to be the most prominent antiviral type of chinese. In the research of various virus infections, the interferon alpha 1b is found to be the antiviral substance which appears earliest and produces the largest amount in the human body after Chinese people are infected by the virus. A large number of clinical practices show that the recombinant human interferon alpha 1b is most suitable for Chinese people to use, and the adverse reaction is obviously lower than that of recombinant human interferon alpha 2a and recombinant human interferon alpha 2b from western people.
The recombinant human interferon alpha 1b has broad-spectrum antiviral effect, is combined with a cell surface receptor, activates various cytoplasmic enzymes through a series of intermediate metabolism, and the enzymes can inhibit the propagation of viruses in cells and interfere the synthesis of virus nucleic acid and protein. The interferon can also inhibit functions of T cells and B cells, activate macrophages and NK cells, and promote the effect of killer T cells, thereby achieving the antiviral effect.
The ocular virus infectious diseases are inflammations caused by the infection of eyes by virus pathogens, and the most common pathogens are herpes simplex viruses and banded viruses. Clinically, the following diseases are mainly classified: ocular diseases with herpes simplex: is caused by Herpes Simplex Virus (HSV) infection of eyes, and comprises eyelid herpes simplex, single bleb conjunctivitis, keratitis (dendritic, cartographic, discoid, substantial keratitis), single bleb iridocyclitis, etc. ② herpes zoster ophthalmopathy: is caused by herpes zoster virus (VZV) infection of the eye, including herpes zoster of the eyelid, herpes zoster keratitis, scleritis, etc. ③ the adenovirus conjunctivitis keratitis. Fourthly, epidemic hemorrhagic conjunctivitis, which is commonly called as pinkeye.
At present, a lot of eye disease patients are infected by viruses such as ophthalmic viral keratitis, viral conjunctivitis and the like, and eye drops in the market are of a lot of varieties at present, but most of the eye drops are antibiotic eye drops, and the antibiotic eye drops are only effective to bacterial infection. Aiming at viral infection eye diseases, the effective antiviral eye drops are few. The interferon has the functions of broad-spectrum antivirus, anti-tumor and regulating the immune function of the organism, and is a medicament which is well recognized by scholars and has good antivirus curative effect.
An aqueous solution of interferon has a problem of being unstable over a long period of time. CN1724567A discloses a stable recombinant human interferon alpha 1b aqueous solution, the auxiliary agents of which comprise sodium chloride, citric acid, disodium hydrogen phosphate dodecahydrate and human serum albumin, wherein the preferred range of the sodium chloride is 6.5-7.5 g/L. CN101214224A discloses a recombinant human interferon injection, which uses succinylated gelatin and Tween 80 to replace human serum albumin as a stabilizer. CN109010803A discloses a recombinant human interferon alpha 2b eye drop and a preparation method thereof, and auxiliary materials of the eye drop comprise sodium hyaluronate, histidine, edetate disodium, tween 80, a bacteriostatic agent and sodium chloride. CN111617031A discloses a stable recombinant human interferon alpha 1b eye drop and a production method thereof, wherein the eye drop is composed of recombinant human interferon alpha 1b, a protective agent, an osmotic pressure regulator and water. The proposal achieves the purpose of stably storing the interferon aqueous solution for a long time by adjusting the formula of the interferon preparation, particularly adding a stabilizer.
However, the above solutions, including other similar technologies, have the common problems that: the eye drops are special medicines, such as tablets, capsules, injection and the like, which are used immediately or in a short time after the medicines are unsealed, but the service life of the eye drops after the unsealing is short (about 1mL of small packages) within 24 hours and long (about 10-30mL of large packages) within 1 month, users usually do not put the eye drops back to the storage environment (2-8 ℃ away from light) required by the specification after the unsealing, and some users cannot mark the unsealing date, so that the eye drops are continuously used after the unsealing and quality guarantee period. The stability after unsealing is reduced, and the stability after unsealing mainly comprises two factors, wherein on one hand, reagent after unsealing is polluted by a user or the environment, and pollutants are caused by the influence of the reagent, and on the other hand, interferon self after unsealing is comprehensively influenced by various factors in the environment, so that the stability of the interferon self is reduced. In the prior art, the effects are usually reduced by adding antibacterial agents or adding a sealed bag after unsealing, but the effects are generally not obvious, and the increase of the antibacterial agents can also cause irritation to eyes and have negative effects on the performance of eye drops.
The eye is one of the most sensitive organs of human body, and the eye drops applied to the eyes should be added with chemical substances as little as possible on the premise of ensuring the stability of the solution at normal temperature after the solution is unsealed, so that the stimulation to the eyes is reduced.
Disclosure of Invention
Aiming at the problems, the invention adds one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator into the stock solution of the recombinant human interferon alpha 1b obtained by fermentation and purification, after being uniformly mixed, the mixture is sterilized and filtered by a filter membrane of 0.22 mu m, and finally, the eye drops of the recombinant human interferon alpha 1b are prepared by packaging after being qualified. The eye drops adopt reagents which avoid chemicals irritating eyes or are added with pure natural non-irritating components, and can achieve the stability for a long time under the conditions of normal temperature and no light shielding after unsealing. The specific scheme is as follows:
firstly, the invention provides the recombinant human interferon alpha 1b eye drops which are stable at normal temperature after being unsealed, and the eye drops comprise the recombinant human interferon alpha 1b and one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
Preferably, the protective agent comprises a mixture of arginine, sitosterol, human serum albumin.
Preferably, the concentration of arginine, sitosterol and human serum albumin in the eye drops of the recombinant human interferon alpha 1b in the protective agent is 1.0-1.5% (w/v), 0.3-0.5% (w/v) and 0.2-0.4% (w/v), respectively.
Preferably, the stabilizer comprises a mixture of water soluble beta-1, 3 glucan, lysine.
Preferably, the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the recombinant human interferon alpha 1b eye drops in the stabilizing agent are 0.5-1.2% (w/v) and 0.5-1.0% (w/v), respectively.
Preferably, assuming that the total concentration of the protective agent in the recombinant human interferon alpha 1b eye drops is i and the total concentration of the stabilizing agent is j, i is 2 betaxj, and beta is a constant and takes a value of 1-1.25.
Preferably, the additional modulator comprises a mixture of tribulus terrestris extract and okra extract.
Preferably, the concentration of the tribulus terrestris extract and the concentration of the okra extract in the eye drops of the recombinant human interferon alpha 1b in the additional regulator are respectively 0.6-0.9% (w/v) and 0.1-0.5% (w/v).
Preferably, the preparation method of the tribulus terrestris extract comprises the following steps: pulverizing fructus Atriplicis Sibiricae to above 80 mesh; adding distilled water according to the liquid-material ratio of 20:1-25:1 ml/g; heating to 40-90 deg.C, ultrasonic leaching for 90-180 min; filtering conventionally to obtain filtrate, centrifuging the filtrate at 2000-.
Preferably, the preparation method of the okra extract comprises the following steps: cleaning okra pods, chopping and homogenizing; adding distilled water with the mass of 3 times, standing for 24h, centrifuging at 3000rpm for 5min, and taking the supernatant as a crude extract; adding 35% ethanol into the crude extract while stirring to generate precipitate, centrifuging at 3200rpm for 5min, collecting precipitate, and vacuum drying; dissolving the obtained product in 30-50 times of distilled water, dialyzing in 20 times of distilled water for 48 hr, changing the solution for 4 times per day, dialyzing, and lyophilizing to obtain okra extract.
Preferably, the osmotic pressure regulator comprises sodium chloride and borax.
Preferably, the concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.1-0.3% (w/v) and 0.4-0.6% (w/v), respectively.
The percentage (w/v) in the concentrations described herein refers to solute mass (g) ÷ solution volume (mL) × 100%.
Secondly, the invention also discloses a preparation method of the eye drops of the recombinant human interferon alpha 1b, which comprises the following steps:
s1, taking a stock solution of the recombinant human interferon alpha 1b, adding one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator, and uniformly mixing;
s2, filtering and sterilizing;
and S3, subpackaging the product.
Preferably, the protective agent, the stabilizing agent, the additional regulator and one or more of the osmotic pressure regulator are added in sequence.
Preferably, the concentration of the recombinant human interferon alpha 1b in the recombinant human interferon alpha 1b stock solution is 10-300 ten thousand IU/ml.
The invention can adopt phosphate buffer solution or PBS buffer solution as a buffer system for dissolving each component to prepare the eye drops, wherein the pH value of the phosphate buffer solution is 6-7.5, and the concentration is 15.0-40.0 mmol/L.
Advantageous effects
The beneficial technical effects of the invention are as follows:
(1) the recombinant human interferon alpha 1b eye drops can be used for treating ocular virus infectious diseases, including herpes simplex eye diseases: such as eyelid herpes simplex, herpes simplex conjunctivitis, keratitis (dendritic, cartographic, discoid, substantive keratitis), herpes simplex iridocyclitis), herpes zoster eye disease: such as blepharitis zoster, herpes zoster keratitis, scleritis, iridocyclitis, adenovirus conjunctivitis keratitis, epidemic hemorrhagic conjunctivitis;
(2) the protective agent adopted by the invention is a mixture of arginine, sitosterol and human serum albumin, and the stabilizing agent adopted is a mixture of water-soluble beta-1, 3 glucan and lazy amino acid, so that the stability of the eye drops in the environment of normal-temperature illumination after unsealing is effectively enhanced by the additive;
(3) the invention also adopts an additional regulator which is a mixture of the tribulus terrestris extract and the okra extract, the tribulus terrestris extract and the okra extract have application records in the past work, but the addition of the tribulus terrestris extract in the eye drops in the prior work has the effects of improving eyesight, relieving itching, relieving allergic conjunctivitis and the like, the addition of the okra extract in the eye drops has the effects of relieving eye dryness and fatigue, but in the invention, the core effective component of the invention is recombinant human interferon alpha 1b, the only active substance playing a role in treatment is recombinant human interferon alpha 1b, the core purpose of the invention is to improve the stability of the eye drops of the recombinant human interferon alpha 1b, particularly the stability under the condition of normal temperature illumination after unsealing, the addition amount of the two plant extracts is very small, in fact, at such an addition amount, the conventional effect cannot be achieved basically, but the purpose of adding the two plant extracts in the present application is not just that of performing conventional treatment, and such an addition amount has no obvious influence on the overall properties (including therapeutic embodiment and stability) of the eye drops when any one of the two extracts is added alone, and when the two extracts are added simultaneously and added according to the range defined by the present invention, the overall stability of the eye drops is remarkably improved, the therapeutic property is not remarkably changed, and we innovatively adopt the two plant extracts as special stabilizers;
(4) the eye drop can be stored for a long time under the condition of low temperature, can be stored for a long time under the condition of normal temperature and no light shielding after being unsealed, can be stored for at least 24 months under the condition of light shielding at 2-8 ℃ (the conventional storage requirement), can be stored for at least 9 months under the condition of normal temperature (25 ℃ -35 ℃) and no light shielding for at least 9 days under the condition of normal temperature (25 ℃ -35 ℃) and no light shielding after being unsealed (not being resealed), and the above preservation means that the sterility is ensured during the storage period, the pH value and the osmotic pressure of the product are not obviously changed, no visible foreign matters exist, and the biological activity is stable.
Detailed Description
The embodiments of the present invention are described below with reference to specific embodiments, and other advantages and effects of the present invention will be easily understood by those skilled in the art from the disclosure of the present specification. The invention is capable of other and different embodiments and of being practiced or of being carried out in various ways, and its several details are capable of modification in various respects, all without departing from the spirit and scope of the present invention.
The following examples used phosphate buffer pH 7.0 at 20.0 mmol/L.
Preparation example:
1. preparation of recombinant human interferon alpha 1b stock solution:
the recombinant human interferon alpha 1b stock solution is prepared by the company (Changchun biological products research institute, Limited liability company), and can also be prepared to the required concentration by adopting the commercial recombinant human interferon alpha 1b for use, or prepared by adopting other preparation methods disclosed in the prior art, and the effect and the trend of the combined use of other components can not generate obvious difference.
Constructing recombinant plasmid containing recombinant human interferon alpha 1b liquid gene by using an escherichia coli expression technology, converting the recombinant plasmid into escherichia coli to obtain stable engineering bacteria, fermenting the engineering bacteria, removing impurities such as polymers in the interferon through primary extraction and several purification process operations to obtain recombinant human interferon alpha 1b stock solution, preparing the recombinant human interferon alpha 1b stock solution to the concentration of 10 ten thousand IU/ml, and ensuring the purity to accord with the regulation of the three parts (2015 edition) of pharmacopoeia of the people's republic of China. The operations of plasmid preparation, transformation, fermentation, purification and the like are all conventional operations in the prior art.
2. Preparation of tribulus terrestris extract: pulverizing fructus Atriplicis Sibiricae to 120 mesh; adding distilled water according to the liquid-material ratio of 20:1 ml/g; heating to 70 deg.C, ultrasonic extracting for 120 min; filtering at 3000rpm for 5min, collecting supernatant, filtering with 0.22 μm filter membrane, and lyophilizing to obtain fructus Atriplicis Sibiricae extract.
3. Preparing an okra extract: cleaning okra pods, chopping, and homogenizing; adding distilled water with the mass of 3 times, standing for 24h, centrifuging at 3000rpm for 5min, and taking the supernatant as a crude extract; adding 35% ethanol into the crude extract while stirring to generate precipitate, centrifuging at 3200rpm for 5min, collecting precipitate, and vacuum drying; dissolving the obtained product in 50 times of distilled water, dialyzing in 20 times of distilled water for 48 hr, changing the solution 4 times per day, dialyzing, and lyophilizing to obtain okra extract.
In addition to the above components, other components including buffers are commercially available.
Example 1 Kaifeng Normal temperature Stable recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, a protective agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
Comparative example 1 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the recombinant human interferon alpha 1b eye drops are 0.2% (w/v) and 0.5% (w/v) respectively.
The protective agent comprises arginine.
The concentration of arginine in the protective agent in the eye drops of the recombinant human interferon alpha 1b is 1.5% (w/v).
Comparative example 2 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises sitosterol.
The concentration of sitosterol in the protective agent in the eye drops of the recombinant human interferon alpha 1b is 0.5% (w/v).
Comparative example 3 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine and sitosterol.
The concentrations of arginine and sitosterol in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v) and 0.5% (w/v).
Comparative example 4 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine and human serum albumin.
The concentrations of arginine and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v) and 0.4% (w/v).
Comparative example 5 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of sitosterol and human serum albumin.
The concentrations of sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 0.5% (w/v) and 0.4% (w/v).
Example 2 Kaifeng Normal temperature Stable recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of water-soluble beta-1, 3 glucan and lysine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 1.2% (w/v) and 1.0% (w/v), respectively.
Example 3 Kaifeng Normal temperature Stable recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.0% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of water-soluble beta-1, 3 glucan and lysine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 1.0% (w/v) and 0.8% (w/v), respectively.
Comparative example 6A recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the recombinant human interferon alpha 1b eye drops are 0.2% (w/v) and 0.5% (w/v) respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises water-soluble beta-1, 3 glucan.
The concentration of the water-soluble beta-1, 3 glucan in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b is 1.2% (w/v).
Comparative example 7 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises lazine.
The concentration of the lazine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b is 1.0 percent (w/v).
Example 4-7 an unsealed recombinant human interferon alpha 1b eye drop stable at normal temperature
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
If the total concentration of the protective agent and the total concentration of the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are i and j, i is 2 beta multiplied by j, and beta is a constant and takes a value of 1-1.25.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.0-1.5% (w/v), 0.3-0.5% (w/v) and 0.2-0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of the water-soluble beta-1, 3 glucan and the lazine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 0.5-1.2% (w/v) and 0.5-1.0% (w/v), respectively.
Examples 4 to 7 differ in the concentration of the individual components of the protective agent, stabilizer:
example 4: the concentrations of arginine, sitosterol and human albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v); the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the eye drops of the recombinant human interferon alpha 1b in the stabilizing agent are 0.5 percent (w/v) and 0.5 percent (w/v), respectively.
Example 5: the concentrations of arginine, sitosterol and human albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.4% (w/v), 0.4% (w/v) and 0.4% (w/v); the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the eye drops of the recombinant human interferon alpha 1b in the stabilizing agent are 0.5(w/v) and 0.5(w/v), respectively.
Example 6: the concentrations of arginine, sitosterol and human albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v); the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the eye drops of the recombinant human interferon alpha 1b in the stabilizing agent are 0.5 percent (w/v) and 0.7 percent (w/v), respectively.
Example 7: the concentrations of arginine, sitosterol and human albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.3% (w/v), 0.3% (w/v) and 0.4% (w/v); the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the eye drops of the recombinant human interferon alpha 1b in the stabilizing agent are 0.5 percent (w/v) and 0.5 percent (w/v), respectively.
Example 8 Kaifeng Normal temperature Stable recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of water-soluble beta-1, 3 glucan and lysine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 1.2% (w/v) and 1.0% (w/v) respectively.
The additional regulator comprises a mixture of tribulus terrestris extract and okra extract.
The concentrations of the tribulus terrestris extract and the okra extract in the recombinant human interferon alpha 1b eye drops in the additional regulator are 0.9% (w/v) and 0.5% (w/v), respectively.
Comparative example 8 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of water-soluble beta-1, 3 glucan and lysine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 1.2% (w/v) and 1.0% (w/v), respectively.
The additional modifier comprises Tribulus terrestris extract.
The concentration of the tribulus terrestris extract in the eye drops of the recombinant human interferon alpha 1b in the additional regulator is 0.9% (w/v).
Comparative example 9 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.2% (w/v) and 0.5% (w/v), respectively.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of water-soluble beta-1, 3 glucan and lysine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 1.2% (w/v) and 1.0% (w/v), respectively.
The additional conditioning agent comprises a mixture of okra extracts.
The concentration of the okra extract in the recombinant human interferon alpha 1b eye drops as the additional regulator is 0.5% (w/v).
Example 9 Kaifeng Normal temperature Stable recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, and also comprises a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
The osmotic pressure regulator comprises sodium chloride and borax.
The concentrations of sodium chloride and borax in the osmotic pressure regulator in the recombinant human interferon alpha 1b eye drops are 0.2% (w/v) and 0.5% (w/v) respectively.
If the total concentration of the protective agent and the total concentration of the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are i and j, i is 2 beta multiplied by j, and beta is a constant and takes a value of 1-1.25.
The protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
The concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.5% (w/v), 0.5% (w/v) and 0.4% (w/v).
The stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
The concentrations of the water-soluble beta-1, 3 glucan and the lazine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 0.5% (w/v) and 0.5% (w/v), respectively.
The additional regulator comprises a mixture of Tribulus terrestris extract and Abelmoschus esculentus extract.
The concentrations of the tribulus terrestris extract and the okra extract in the recombinant human interferon alpha 1b eye drops in the additional regulator are 0.9% (w/v) and 0.5% (w/v) respectively.
Comparative example 10 recombinant human interferon alpha 1b eye drops
Comprises recombinant human interferon alpha 1b, human serum albumin, sodium chloride (and buffer solution).
Wherein the concentration of human serum albumin in the eye drops of the recombinant human interferon alpha 1b is 2% (w/v); the concentration of sodium chloride in the eye drops of the recombinant human interferon alpha 1b is 0.85% (w/v).
The method for preparing the recombinant human interferon alpha 1b eye drops by the formula in the above embodiment/comparative example comprises the following steps:
s1, taking a stock solution of the recombinant human interferon alpha 1b, adding one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator, and uniformly mixing;
s2, filtering and sterilizing;
and S3, subpackaging the product.
The protective agent, the stabilizing agent, the additional regulator and the osmotic pressure regulator are added in sequence, when one component is not included in the formula, the component is removed, and the protective agent, the stabilizing agent, the additional regulator and the osmotic pressure regulator are added in the same sequence as the components.
The components can be prepared into a semi-finished product with higher concentration by using a buffer solution, and then the semi-finished product is added into a system for preparing the eye drops by a certain amount to obtain the required concentration through concentration and volume calculation.
The sterilizing filtration comprises the following steps:
1) two 293 filters are connected in series, the screw caps on the filters are screwed, and the liquid inlet pipeline of the filters is connected with the liquid supply pipeline of the sterilization tank. The filtrate pipeline is sent into the buffer cabin of the aseptic operation isolator through the pipeline hole of the aseptic operation isolator, and the pipeline hole is sealed. And (3) feeding a liquid inlet pipeline connected to the 50L-100L semi-finished product tank into a buffer cabin of the aseptic operation isolator through a pipeline hole of the aseptic operation isolator, and sealing the pipeline hole. And putting articles such as sample bottles, bottle stoppers and the like into the buffer cabin of the sterile operation isolator through the buffer cabin door. The sterile manipulation isolator VHP was opened and sterilized for 30-40 minutes.
2) And after the filtrate pipeline connected to the filter and the liquid inlet pipeline connected to the 50L-100L semi-finished product tank are subjected to outer package, the filtrate pipeline and the liquid inlet pipeline are conveyed into the main cabin of the aseptic operation isolator through the pipeline hole between the main cabin and the buffer cabin, and the pipeline hole is sealed. The sample bottle, the bottle plug and other articles are put into the main cabin of the aseptic operation isolator through the door between the two cabins. Removing the innermost package, and connecting the two pipelines in a buckling manner.
3) And (4) placing the 50L-100L semi-finished product tank on a weighbridge, resetting, opening a tank bottom valve and a liquid supply valve of the sterilization tank, and starting sterilization and filtration. And (4) observing the positions of the upper filter plate and the lower filter plate of the filter and the exhaust valve, and carrying out the next operation if no liquid overflows. And (4) adjusting the peristaltic pump to 3 grades, and filtering the liquid in the sterilized tank to a sterilized 50L-100L semi-finished product tank to reach the required weight (converting the required preparation volume into the weight according to the specific gravity). And (3) closing the peristaltic pump, clamping the filter filtrate silica gel tube by using a hemostatic forceps, and obtaining the filtrate in the barrel, namely the semi-finished product of the recombinant human interferon alpha 1b eye drops. And then, carrying out semi-finished product verification, and carrying out next-step subpackaging after the semi-finished product is qualified.
And (3) subpackaging products:
the split charging specification of the eye drops finished product of the recombinant human interferon alpha 1b is 20 ten thousand IU/2.0 ml/bottle. The filling quantity requirement of the product is as follows: 2.15 ml/bottle is filled, and the filling amount of each bottle is not less than the amount marked on the label.
When filling begins, the environment of a filling room is confirmed, the temperature, the humidity and the pressure difference of the filling room are confirmed, the temperature is controlled to be 15-26 ℃, the relative humidity is controlled to be 30-70%, and a certain pressure difference (not less than 10Pa) is kept between the filling room and other areas; the confirmation device: confirming that the eye drops filling machine and the electronic balance operate normally; confirming that the inner packaging material is sterilized by ethylene oxide; confirming that the filling appliance, the oscillating hopper and the runway are sterilized under high pressure; confirming a semi-finished product to be filled: before filling, carefully checking the name and the batch number of a product to be filled and the quantity of a semi-finished product to be filled, checking whether a tank opening and a tank body are sealed completely, and spraying and sterilizing the semi-finished product to be filled by using a sporicide;
after the preparation work confirms that the bottles are correct, the well-connected filling pipelines are sequentially connected with the peristaltic pump and the semi-finished product tank to be filled. And fixing the filling needle head on the needle head clamp. Fixing the sterilized bottle oscillating hopper, the inner plug oscillating hopper and the outer cover oscillating hopper on an oscillator, and then pouring the bottle, the bottle plug and the bottle cap into the oscillating hopper. The compressed air valve is opened. And opening a key of the filling host machine, and electrifying the filling host machine. The inner plug oscillator and the outer cap oscillator are opened and adjusted to the appropriate speed. And turning on a bottle arranging power supply, turning on a bottle arranging oscillator, adjusting the bottle arranging oscillator to a proper speed, and turning on a bottle arranging switch. After the eye drop bottle, the plug and the cover inlet on the filling and sealing machine are sufficiently used, the machine can be prepared to start to fill.
The filling quantity is checked at the beginning of the filling process by an electronic balance, the filling quantity should be observed frequently during the filling process, and the filling quantity must be checked once every 1 hour of filling. The loading is accurate and uneven loading is prevented. The filling speed is less than or equal to 75 bottles/minute. The operation is kept at 55-60 bottles/min under the condition of stable operation of the equipment. The filling is finished within 24 hours, and after the filling is finished, the semi-finished product tank to be filled is opened to check whether precipitates or other foreign matters exist. Aseptic handling should be strictly observed during filling.
And (3) product property inspection: (0 month and 0 day in the reference period means the measurement was performed on the day after production or after unsealing)
Firstly, the stability of the recombinant human interferon alpha 1b eye drops was tested by the method described in Chinese patent CN202010591909.4 (our earlier work) on the samples prepared in the above examples and comparative examples, and the method was characterized in that the eye drops were packaged without being unsealed after aseptic manipulation and were performed under specific conditions.
In order to further determine the stability of the formula and the preparation method, whether the medicine is stable at the storage temperature of 2-8 ℃ is judged through summarizing and analyzing detection data, a theoretical basis is provided for determining the packaging, transportation, storage conditions and the effective period of the product, and the effective period of the product is reset in the stable period.
1) The recombinant human interferon alpha 1b eye drops are subjected to accelerated stability investigation at 23-27 ℃, and comprise the following steps:
according to the physicochemical property of the recombinant human interferon alpha 1b and the self-specificity of the dosage form of the eye drops, the accelerated stability investigation test at 23-27 ℃ is carried out on the eye drop. 3 batches of recombinant human interferon alpha 1b eye drops (production batches: S20220201, S20220202, S20220220203) prepared according to the optimized formulation and process of example 2 were used as subjects for accelerated stability test at 23-27 ℃.
The examination items refer to the pharmacopoeia of the people's republic of China (2015 edition), the appearance, visible foreign matters, loading, pH value, osmolality, biological activity and sterility test are selected as examination indexes, the stability examination for 6 months is carried out, and the specific examination items and examination time are shown in table 1.
TABLE 1 accelerated stability study of recombinant human interferon alpha 1b eye drops
Note: in the table, the symbol "√" is for detection; "/" is not detected.
The result of the accelerated stability test at 23-27 ℃ is as follows: through a 23-27 ℃ accelerated stability investigation test for 6 months, the physical and chemical properties such as appearance, visible foreign matters, loading, pH value and the like are not changed; the sterility test is qualified; the osmotic pressure molar concentration slightly fluctuates but is stable, and the biological activity basically has no fluctuation. The specific examination and detection results are shown in Table 2.
Table 2 investigation result of accelerated stability of recombinant human interferon alpha 1b eye drops at 23-27 DEG C
2) The long-term stability investigation test of the recombinant human interferon alpha 1b eye drops at 2-8 ℃ is as follows:
according to the physicochemical property of the recombinant human interferon alpha 1b and the self-specificity of the dosage form of the eye drop, a long-term stability investigation test at 2-8 ℃ is carried out on the invention. 3 batches of recombinant human interferon alpha 1b eye drops (production batches: 20200101, 20200102 and 20200103) produced in 2020 and prepared according to the optimized formula and process of the embodiment 2 are subjected to a long-term stability investigation test at 2-8 ℃. The examination items refer to pharmacopoeia of the people's republic of China (2015 edition), identification, appearance, visible foreign matters, loading, pH value, osmolality, biological activity and sterility test are selected as examination indexes, stability examination is carried out for 24 months, and specific examination items and examination time are shown in table 3.
Table 3 recombinant human interferon alpha 1b eye drops 2 ~ 8 ℃ long-term stability investigation
Note: in the table, the symbol "√" is for detection; "/" is not detected.
2-8 ℃ long-term stability investigation result: through a long-term stability investigation test at 2-8 ℃ for 24 months, physical and chemical properties such as identification, appearance, visible foreign matters, loading and the like are not changed, sterility inspection is not changed, the pH value is stable, the osmotic pressure molar concentration has a micro fluctuation phenomenon but is stable, and the biological activity has a fluctuation phenomenon but is within the quality standard range within 24 months. The specific examination and detection results are shown in Table 4.
Table 4 investigation result of long-term stability of recombinant human interferon alpha 1b eye drops at 2-8 DEG C
Analysis of the stability investigation test result of the recombinant human interferon alpha 1b eye drops: stability of three batches of recombinant human interferon alpha 1b eye drop finished products (20200101, 20200102 and 20200103) is examined, and the stability of the recombinant human interferon alpha 1b eye drop finished products is better as can be observed from a summary table of data of various detections. All detection results of the finished eye drop product in the monitoring period are within the qualified range specified in pharmacopoeia of the people's republic of China (2015 edition). The detection data show that the finished product of the eye drop meets the quality standards in terms of identification, appearance, visible foreign matters, loading, sterility inspection and the like. The pH value is stable, and the osmotic pressure molar concentration is slightly fluctuated, but is more stable in the detection period, and the biological activity is stable. The biological activity of the three batches of eye drops fluctuates in the monitoring period, but the three batches of eye drops meet the quality standard within 24 months.
And (4) conclusion: through the accelerated stability investigation at 23-27 ℃ and the long-term stability investigation at 2-8 ℃ of the eye drops of the recombinant human interferon alpha 1b, all indexes of the eye drops of the recombinant human interferon alpha 1b have no obvious change in a detection period. Therefore, the recombinant human interferon alpha 1b eye drops can be proved to be stable in dark storage at the temperature of 2-8 ℃, and the effective service life can be set to be 0-24 months.
Quality control evaluation
1) Identification test
The positive result is determined by the immuno-spot method (rule 3402) in three parts (2015) of pharmacopoeia of the people's republic of China.
2) Visual inspection
It should be a colorless liquid.
3) Visible foreign body
According to the pharmacopoeia of the people's republic of China, three parts (2015 edition) (general rule 0904), the product should be qualified.
4) Loading capacity
According to the pharmacopoeia of the people's republic of China, three parts (2015 edition) (0105 in general), the amount of the active ingredients should not be less than the labeled amount.
5) pH value
According to the pharmacopoeia of the people's republic of China (2015 edition) (general rule 0631), it should be regarded as
6) Osmolality
According to the pharmacopoeia of the people's republic of China (2015 edition) (general rule 0632), it should be regarded as
7) Biological activity
In accordance with the pharmacopoeia of the people's republic of China (2015 edition) (Tonghuang)3523), inhibition of cytopathic effect of the first method, should be in the amounts indicated
8) Sterility testing
According to the pharmacopoeia of the people's republic of China, three parts (2015 edition) (general rule 1101), the strain should be grown aseptically.
Based on the special purpose of the invention, 1 batch of samples of the above examples and comparative examples are respectively subjected to a long-term stability test under the conditions of unsealing, sealing/unsealing and normal temperature (25 ℃ -35 ℃) non-light-shielding storage, and the specific examination items and examination time are shown in tables 5 and 6. The results are shown in tables 7 and 8.
TABLE 5 resealing of the recombinant human interferon alpha 1b eye drops after unsealing, Long-term stability investigation under ambient temperature illumination
Note: in the table, the symbol "√" is for detection; "/" is not detected.
TABLE 6 examination of the stability of the recombinant human interferon alpha 1b eye drops under normal temperature illumination for a long period of time without resealing after unsealing
Note: in the table, the symbol "√" is for detection; "/" is not detected.
TABLE 7 results of long-term stability investigation of the recombinant human interferon alpha 1b eye drops after unsealing and resealing under normal temperature illumination
TABLE 8 results of long-term stability investigation under normal temperature illumination without resealing of recombinant human interferon alpha 1b eye drops after unsealing
Analysis of investigation test results of the unsealed normal-temperature stability of the recombinant human interferon alpha 1b eye drops: the property change is mainly reflected by biological activity and a sterile inspection result, and therefore, in the formula provided by the invention, the stabilizing effect of arginine on sitosterol and human serum albumin in the protective agent is not promoted, the effect superposition or promotion is not realized after the sitosterol and the human serum albumin are mixed, and the effect of the invention can be achieved only by adding the arginine and simultaneously existing the sitosterol and the human serum albumin. The two components in the stabilizer do not achieve obvious effect when used independently, and the synergistic effect of the two components is obvious. The defined proportional relationship between the stabilizer and the protective agent can effectively increase the stability of the system. The two plant extracts are used as additional regulators respectively and are added into a system independently, the two plant extracts do not contribute to stability, the stability can be obviously improved after the two plant extracts are added simultaneously, a breakthrough discovery has not been made in principle, but the two plant extracts are suspected to generate substances which have antibacterial activity and pertinence after certain substances in the two plant extracts act with each other, so that the stability of the system is improved, and the two plant extracts are pure natural components which are not irritant to eyes, so that the discovery can play a role in promoting a plurality of work of people in the future. The resealing after unsealing refers to unsealing the packaged reagent in the air in a non-sterile environment, and resealing after opening and standing for 30s (simulating the standard use scene of the reagent in the hands of a user); unsealing after unsealing means unsealing the packaged reagent in the air in a non-sterile environment, and standing the reagent in an open state.
While the preferred embodiments and examples of the present invention have been described in detail, the present invention is not limited to the embodiments and examples, and various changes can be made without departing from the spirit of the present invention within the knowledge of those skilled in the art.
Claims (10)
1. An unsealed recombinant human interferon alpha 1b eye drop stable at normal temperature is characterized in that: comprises recombinant human interferon alpha 1b, and also comprises one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator.
2. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 1, which is characterized in that: the protective agent comprises a mixture of arginine, sitosterol and human serum albumin.
3. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 2, which is characterized in that: the concentrations of arginine, sitosterol and human serum albumin in the protective agent in the eye drops of the recombinant human interferon alpha 1b are respectively 1.0-1.5% (w/v), 0.3-0.5% (w/v) and 0.2-0.4% (w/v).
4. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 1, which is characterized in that: the stabilizer comprises a mixture of water-soluble beta-1, 3-glucan and lysine.
5. The kaifeng ambient temperature stable recombinant human interferon alpha 1b eye drops according to claim 4, which is characterized in that: the concentrations of the water-soluble beta-1, 3 glucan and the lazine in the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are 0.5-1.2% (w/v) and 0.5-1.0% (w/v), respectively.
6. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 3 or 5, which is characterized in that: if the total concentration of the protective agent and the total concentration of the stabilizing agent in the eye drops of the recombinant human interferon alpha 1b are i and j, i is 2 beta multiplied by j, and beta is a constant and takes a value of 1-1.25.
7. The kaifeng recombinant human interferon alpha 1b eye drops which are stable at normal temperature according to claim 1, which is characterized in that: the additional regulator comprises a mixture of tribulus terrestris extract and okra extract.
8. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 1, which is characterized in that: the concentrations of the tribulus terrestris extract and the okra extract in the eye drops of the recombinant human interferon alpha 1b in the additional regulator are 0.6-0.9% (w/v) and 0.1-0.5% (w/v), respectively.
9. The kaifeng recombinant human interferon alpha 1b eye drops stable at normal temperature according to claim 1, which is characterized in that: the osmotic pressure regulator comprises sodium chloride and borax; the concentrations of sodium chloride and borax in the osmotic pressure regulator in the eye drops of the recombinant human interferon alpha 1b are 0.1-0.3% (w/v) and 0.4-0.6% (w/v), respectively.
10. A preparation method of recombinant human interferon alpha 1b eye drops stable at normal temperature after being unsealed as claimed in any one of claims 1 to 9, which is characterized in that: the method comprises the following steps:
s1, taking a stock solution of the recombinant human interferon alpha 1b, adding one or more than two of a protective agent, a stabilizing agent, an additional regulator and an osmotic pressure regulator, and uniformly mixing;
s2, filtering and sterilizing;
s3, subpackaging the product;
and the protective agent, the stabilizing agent, the additional regulator and the osmotic pressure regulator are added in sequence.
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| CN117959251A (en) * | 2024-03-29 | 2024-05-03 | 长春生物制品研究所有限责任公司 | Recombinant human interferon alpha 1b eye drops and preparation method thereof |
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