CN112933216B

CN112933216B - Stable pharmaceutical composition containing both insulin and dexamethasone sodium phosphate

Info

Publication number: CN112933216B
Application number: CN201911344584.3A
Authority: CN
Inventors: 睢诚; 苗春徽; 赵干; 程鑫; 彭海长; 俞庆龄
Original assignee: Advaccine Suzhou Biopharmaceutical Co ltd
Current assignee: Advaccine Suzhou Biopharmaceutical Co ltd
Priority date: 2019-12-24
Filing date: 2019-12-24
Publication date: 2023-04-28
Anticipated expiration: 2039-12-24
Also published as: CN112933216A

Abstract

The invention provides a stable pharmaceutical composition containing insulin and dexamethasone sodium phosphate at the same time, and relates to a pharmaceutical composition, which comprises insulin, dexamethasone sodium phosphate, zinc, phenols, a surfactant, an isotonic agent and a buffer solution. Meanwhile, the invention also provides an HPLC method for simultaneously detecting insulin and dexamethasone sodium phosphate in the pharmaceutical composition.

Description

Stable pharmaceutical composition containing both insulin and dexamethasone sodium phosphate

Technical Field

The invention relates to a pharmaceutical composition, in particular to a stable pharmaceutical composition containing insulin and dexamethasone sodium phosphate.

Background

Type I diabetes is a type of autoimmune disease, accounting for about 5-10% of all diabetics. The patient's own immune cells infiltrate the islets, causing the insulin-producing cells to be destroyed. At present, the treatment of type I diabetes is mainly exogenous insulin substitution treatment, patients need to inject insulin daily, the method not only brings great inconvenience to the patients every day, but also can only relieve clinical symptoms such as hyperglycemia, can not change the condition that the pathological immune cells of the patients attack pancreas, can not recover the damaged pancreas function, and can not control long-term complications.

Chinese patent CN103372214 reports that a pharmaceutical composition composed of type I diabetes protein antigen (insulin, etc.) and immunosuppressant (dexamethasone, etc.) can inhibit the killing effect of autoimmune cells on pancreas, and effectively treat or prevent type I diabetes. However, the report only relates to the way of mixing the two components and does not provide a method for preparing the two components into a stable pharmaceutical composition.

The instability of the protein comprises physical and chemical aspects, wherein the physical instability is the conformational change of the protein molecule caused by factors such as temperature, pH and the like, so that the protein is denatured, adsorbed, aggregated, precipitated and the like, and the chemical instability is the phenomenon such as hydrolysis, oxidation, reduction, racemization, isomerism, peptide bond cleavage, disulfide bond exchange and the like due to modification or change of amino acid residues. These instability factors can lead to reduced or lost activity of the protein. Insulin is formed by the disulfide linkage of two peptide chains, containing three asparagine and three glutamine residues, which are easily hydrolytically denatured in solution, or undergo aggregation and deagglomeration. Insulin belongs to biological products, also called macromolecular drugs, and compound injection containing insulin or insulin analogues is marketed at present, and other active pharmaceutical ingredients are also macromolecular drugs, because the pharmaceutical ingredients which are proteins (or polypeptides) are similar in instability, and stability of a plurality of ingredients can be better controlled when the compound preparation is developed. For example, xultophy 100/3.6 (insulin/liraglutide compound prefilled injection pen) of Norand Norod company is compound injection containing insulin deluge and macromolecular polypeptide drug liraglutide. Dexamethasone sodium phosphate is a chemically synthesized drug, also known as a small molecule drug. Because of the difference in the properties of the medicines, no compound injection which contains macromolecular and micromolecular medicines and can keep all active ingredients stable at the same time is successful at present. Dexamethasone sodium phosphate is an adrenocortical hormone, is widely applied to the treatment of autoimmune diseases, and is a small molecular medicine, so that physical instability such as protein denaturation and precipitation can not occur, but chemical instability still exists. In the aqueous solution, dexamethasone sodium phosphate is easy to hydrolyze to generate dexamethasone which is almost insoluble in water, the dexamethasone Mi Songyi is oxidized, and phenomena such as white spots, precipitation, color change and the like of unstable injection products can occur. Therefore, an auxiliary material with reducibility is usually added into the dexamethasone sodium phosphate injection, and the hydrolysis is inhibited by adjusting the pH. In addition, oxygen needs to be removed by filling nitrogen or the like when filling the injection, and oxidation and hydrolysis are further suppressed.

Up to now, there is no report on a pharmaceutical composition containing both insulin and dexamethasone sodium phosphate in the same preparation. In clinical studies using a combination of both, the mode of use is mostly to inject the two drugs separately. In preclinical studies, two drug injections were mixed for immediate use by researchers, but the activity and stability of the drug in the mixed formulation were not examined. The auxiliary materials used by the insulin injection on the market are not in the prescription of dexamethasone sodium phosphate injection in all countries; also, although some of the common auxiliary materials of dexamethasone sodium phosphate injection, such as propylene glycol, are inactive ingredients which can be used in macromolecular drug injection, none of the common auxiliary materials are components in common insulin injection, and whether the auxiliary materials have an effect on the stability of insulin is not known.

Aiming at the problems of the prior art, a stable pharmaceutical composition containing insulin and dexamethasone sodium phosphate simultaneously is sought, so that the pharmaceutical composition can be used for preparing compound injection, and the compound injection can enable both insulin and dexamethasone sodium phosphate to have activity in the injection and maintain higher stability for a long time.

Disclosure of Invention

Aiming at the problems existing in the prior art, the invention provides a stable pharmaceutical composition containing insulin and dexamethasone sodium phosphate, which can be used for preparing compound injection, and the compound injection can enable both insulin and dexamethasone sodium phosphate to have activity in the injection and maintain higher stability for a long time.

In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:

the invention provides a stable pharmaceutical composition containing insulin and dexamethasone sodium phosphate at the same time, wherein the pharmaceutical composition comprises insulin, dexamethasone sodium phosphate, zinc and buffer.

Further, the pharmaceutical composition comprises 10-50 μg/mL insulin, 10-50 μg/mL dexamethasone sodium phosphate, 0.01-0.5 μg/mL zinc, and 5-50mmol/L buffer.

Further, the pharmaceutical composition further comprises 5-50 μg/mL of phenols, 0-100 μg/mL of surfactant, and 0-9mg/mL of isotonic agent.

Preferably, the pharmaceutical composition comprises: 20. Mu.g/mL insulin, 20. Mu.g/mL dexamethasone sodium phosphate, 0.12. Mu.g/mL zinc, 20. Mu.g/mL phenol, 10. Mu.g/mL surfactant, 8mg/mL isotonic agent and 10mmol/L buffer.

Further, the zinc is added in the form of zinc salts, including one or more of zinc chloride, zinc bromide, zinc iodide, zinc fluoride, zinc sulfate, and zinc acetate; zinc chloride is preferred.

Further, the phenols include one or more of cresol, chlorocresol, and phenol; the cresol includes one or more of m-cresol, o-cresol and p-cresol.

Preferably, the phenols are m-cresol and/or phenol.

Further, the surfactant includes one or more of tween 20 (polyoxyethylene sorbitan monolaurate), tween 40 (polyoxyethylene sorbitan monopalmitate), tween 80 (polyoxyethylene sorbitan monooleate), poloxamer 188 (polyoxyethylene polyoxypropylene block copolymer), polyoxyethylene 35 castor oil, polyoxyethylene 40 hydrogenated castor oil, 15-hydroxystearic acid polyethylene glycol ester, polyethylene glycol 300, polyethylene glycol 400, and polyethylene glycol 600.

Preferably, the surfactant is tween 20 or tween 80.

Further, the isotonic agent includes one or more of mannitol, sorbitol, lactose, dextrose, trehalose, and sodium chloride.

Preferably, the isotonic agent is sodium chloride.

Further, the buffer comprises one or more of phosphate, acetate, citrate, arginine, glycylglycine and Tris buffer (2-amino-2-hydroxymethyl-1, 3-propanediol), buffer ph=6-8.

Preferably, the buffer is a phosphate buffer.

Further, the weight ratio of insulin to dexamethasone sodium phosphate is 5:1-1:5.

Preferably, the weight ratio of insulin to dexamethasone sodium phosphate is 1:1.

The pharmaceutical composition can be applied to preparing compound injection.

In addition, the invention also provides an HPLC method for simultaneously detecting insulin and dexamethasone sodium phosphate in the pharmaceutical composition.

The invention has the technical effects that:

1. the invention provides a compound injection containing the two medicines simultaneously through development and optimization of a preparation prescription, which can ensure that the two medicines have activity in the injection and can keep higher stability for a long time. After the mixture is preserved for 1 year at the temperature of 2-8 ℃, the concentration and purity of insulin and dexamethasone sodium phosphate are not obviously changed, and the properties (clarity, pH and the like) of the injection are not obviously changed;

2. the common auxiliary materials used in the commercial insulin injection are zinc, phenol/m-cresol, glycerol and trace amounts of hydrochloric acid and sodium hydroxide for regulating the pH value. The invention discovers that if glycerin is used as an isotonic agent, dexamethasone sodium phosphate in the compound injection is unstable, and the hydrolysis speed of dexamethasone sodium phosphate in the compound injection is accelerated. In addition, the buffer solution is added instead of purely using hydrochloric acid and sodium hydroxide to adjust the pH value, so that the hydrolysis of dexamethasone sodium phosphate can be inhibited. The addition of zinc and phenols can increase the stability of insulin without affecting dexamethasone sodium phosphate. In order to ensure that the insulin has optimal stability, the zinc content and the insulin content in the commercial insulin injection have fixed proportion, and the phenolic content and the insulin content have minimum proportion, and the invention also uses the same proportion;

3. the formula of dexamethasone sodium phosphate injection in different countries is different due to the influence of pharmacological properties of the raw materials, and common auxiliary materials comprise sodium citrate, sodium bisulphite, disodium edetate, benzyl alcohol, propylene glycol and the like. The invention discovers that the addition of auxiliary materials such as sodium bisulphite, edetate disodium and the like can cause unstable insulin, especially unstable chemistry; meanwhile, the auxiliary materials can not obviously improve the stability of dexamethasone sodium phosphate in the compound injection, and can not inhibit the hydrolysis of dexamethasone sodium phosphate into dexamethasone. When the prescription provided by the invention is used, the dexamethasone sodium phosphate in the compound preparation can still be kept stable even if an oxygen removal process which is generally used for preparing the dexamethasone sodium phosphate injection is not used when the injection is filled.

Drawings

FIG. 1 is a HPLC spectrum for simultaneous detection of insulin, dexamethasone sodium phosphate, dexamethasone;

FIG. 2 is a graph showing the change in insulin content at 25℃in example 3;

FIG. 3 is a graph showing the change in insulin content at 40℃in example 3;

FIG. 4 is a graph showing the variation of dexamethasone sodium phosphate content upon storage at 25℃in example 3;

FIG. 5 is a graph showing the variation of dexamethasone sodium phosphate content upon storage at 40℃in example 3;

FIG. 6 is a graph showing the change in insulin content at 4℃in example 4;

FIG. 7 is a graph showing the variation of dexamethasone sodium phosphate content upon storage at 4deg.C in example 4;

FIG. 8 is a graph showing the change in insulin content at 40℃in example 4;

FIG. 9 is a graph showing the variation of dexamethasone sodium phosphate content upon storage at 40℃in example 4;

FIG. 10 is a graph showing the change in insulin content in example 5;

FIG. 11 is a graph showing the variation of dexamethasone sodium phosphate content in example 5;

FIG. 12 is a graph showing the change in insulin content in example 6;

FIG. 13 is a graph showing the variation of dexamethasone sodium phosphate content in example 6;

FIG. 14 is a graph showing the change in insulin content in example 7;

FIG. 15 is a graph showing the variation of dexamethasone sodium phosphate content in example 7;

FIG. 16 is a graph showing the change in blood glucose in the mice of example 9;

FIG. 17 is a graph showing the change in blood glucose in mice in example 10.

Detailed Description

It is worth noting that insulin in the present application is purchased from Jiangsu wanbang Biochemical medicine Co., ltd., lot number 201801B01; dexamethasone sodium phosphate is purchased from the company of the pharmaceutical Co., ltd., the batch number is 20171105, and the rest of the raw materials are common commercial products, so the sources thereof are not specifically described.

Example 1: development of an HPLC method for simultaneous detection of insulin and dexamethasone sodium phosphate in the pharmaceutical composition according to the invention.

Through development, the detection of insulin, dexamethasone sodium phosphate and dexamethasone can be accomplished simultaneously under the following HPLC conditions:

HPLC conditions: the detection wavelength is 214nm, the sample injection amount is 20 mu L, the column temperature range is 40-47 ℃, and the sample cell temperature is 4 ℃.

The column was subjected to the conditions shown in tables 1 to 4 to obtain the column having the optimum conditions shown in Table 5.

TABLE 1

TABLE 2

TABLE 3 Table 3

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Table 4 condition 10 mobile phase gradient in table 3

Time/min	Mobile phase A%	Mobile phase B%
			0	100	0
20	0	100
			20.1	100	0
30	0	0

TABLE 5 optimal conditions

Chromatographic column model	Sepax Bio-C8 250mm*4.6mm 5μm
		Mobile phase	Buffer (200 mM Na) ₂ SO ₄ ph=2.3)/acetonitrile=74/26
Flow rate	0.8mL/min
		Column temperature	45℃
Run time	50min

FIG. 1 is an HPLC spectrum diagram for simultaneously detecting insulin, dexamethasone sodium phosphate and dexamethasone under the condition of an optimal chromatographic column, and the compounds corresponding to each peak in FIG. 1 are shown in Table 6.

TABLE 6

Example 2: effect of Glycerol, sodium bisulfite and buffer salt solutions on stability of insulin and dexamethasone sodium phosphate

Human insulin drug substance, dexamethasone sodium phosphate drug substance, zinc chloride, sodium bisulphite, glycerol, disodium hydrogen phosphate and sodium dihydrogen phosphate are dissolved. Compound formulations of different prescriptions were formulated as in table 7 and final pH was adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 7 prescription design of Compound preparation

The compound preparation is prepared into sterile preparations, which are packaged in penicillin bottles, covered with rubber plugs and tightly covered with aluminum-plastic composite covers, and then are respectively placed in thermostated containers at 4 ℃, 25 ℃ and 40 ℃. After 40 days of standing, the content of insulin, dexamethasone sodium phosphate and dexamethasone was detected by HPLC, the percentage content of insulin and dexamethasone sodium phosphate compared to the theoretical value was recorded, and the peak area of dexamethasone was recorded, and the results are shown in Table 8.

Table 8 stability of insulin and dexamethasone sodium phosphate when Compound formulations of different prescriptions are placed at different temperatures

The results show that:

1. after the glycerol is added, the stability of dexamethasone sodium phosphate is reduced, and the content is greatly reduced after 40 days.

2. Without the addition of glycerol, and without the use of phosphate buffer, the insulin content was lower than in the other batches, and the formation of dexamethasone was not inhibited.

3. No glycerol was added and phosphate buffer was used, the insulin content was higher than in the other batches, the dexamethasone sodium phosphate content was not significantly changed, no dexamethasone was formed after 40 days at 4 and 25 ℃, and the dexamethasone content generated by hydrolysis at 40 ℃ was lower than in the other batches.

4. When glycerol is added, sodium bisulfite can reduce the decrease of the content of dexamethasone sodium phosphate, but can not inhibit the hydrolysis of dexamethasone sodium phosphate; when no glycerol is added, sodium bisulphite has no obvious effect on the content of dexamethasone sodium phosphate, and the formation of dexamethasone cannot be inhibited; the addition of sodium bisulphite has a great influence on the stability of insulin, and the content of insulin can be greatly reduced.

Example 3: influence of phenols and surfactants on the stability of insulin and dexamethasone sodium phosphate

Human insulin drug substance, dexamethasone sodium phosphate drug substance, zinc chloride, m-cresol, tween 20, disodium hydrogen phosphate and sodium dihydrogen phosphate are dissolved. Compound formulations of different prescriptions were formulated as in table 9 and final pH was adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 9 prescription design of compound preparation

The compound preparation is prepared into sterile preparations, which are packaged in penicillin bottles, covered with rubber plugs and tightly covered with aluminum-plastic composite covers, and then are respectively placed in thermostated containers at 25 ℃ and 40 ℃. Samples were taken periodically, the content of insulin, dexamethasone sodium phosphate and dexamethasone was checked by HPLC, and the percentage of insulin and dexamethasone sodium phosphate content compared to the theoretical values was recorded, as well as the peak area of dexamethasone.

After 35 days of standing, no dexamethasone was detected for all batches. The results of insulin and dexamethasone sodium phosphate content are shown in figures 2 to 5.

The results show that:

1. the addition of m-cresol can slightly increase the stability of insulin; the addition of tween 20 increased the initial insulin content on day 0, possibly reducing the insulin adsorption of the container, and also significantly increased the stability of the insulin, reducing the rate of decrease in insulin content.

2. All batches of dexamethasone sodium phosphate remained stable after 35 days at 25℃and 40℃with no significant changes in content.

Example 4: effect of different drug concentrations on insulin and dexamethasone sodium phosphate stability

Dissolving human insulin crude drug, dexamethasone sodium phosphate crude drug, zinc chloride, disodium hydrogen phosphate and sodium dihydrogen phosphate. Compound formulations of different prescriptions were formulated as in table 10 and final pH was adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 10 compound preparation prescription design

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The compound preparation is a sterile preparation, which is packaged in sterilized penicillin bottles, covered by rubber plugs and tightly rolled by aluminum-plastic combined covers, and then is placed in an incubator at 4 ℃ and 40 ℃ respectively. Samples were taken periodically, the contents of insulin, dexamethasone sodium phosphate and dexamethasone were monitored by HPLC, the percentage of insulin and dexamethasone sodium phosphate content compared to the theoretical values were recorded, and the peak area of dexamethasone was sampled 3 times per batch in parallel and the results averaged.

No dexamethasone was detected for all batches after 28 days at 40 ℃ or 276 days at 4 ℃. The results of insulin and dexamethasone sodium phosphate content are shown in figures 6 to 9.

The results show that:

1. all batches of insulin remained stable after 276 days at 4 ℃ without obvious change in content; after 28 days of storage at 40 ℃, the content of all batches of insulin was reduced, the reduction was close.

2. All batches of dexamethasone sodium phosphate remained stable after either 276 days at 4℃or 28 days at 40℃with no significant changes in content.

3. The compound preparation with the drug concentration of 20 mug/mL and 50 mug/mL has basically the same change trend after the content of insulin and dexamethasone sodium phosphate is placed at 4 ℃ for 276 days or at 40 ℃ for 28 days.

Example 5: effect of different pH on insulin and dexamethasone sodium phosphate stability

Human insulin drug substance, dexamethasone sodium phosphate drug substance, zinc chloride, m-cresol, tween 20, disodium hydrogen phosphate and sodium dihydrogen phosphate are dissolved. Compound formulations of different prescriptions were formulated as in table 11, with final pH adjusted to 6.8, 7.0, 7.2, 7.6 and 8.0 with hydrochloric acid and sodium hydroxide, respectively.

Table 11 compound preparation prescription design

The compound preparation is a sterile preparation, which is packaged in sterilized penicillin bottles, covered by rubber plugs and tightly rolled by aluminum-plastic combined covers, and then placed in a constant temperature cabinet at 4 ℃. Samples were taken periodically, the contents of insulin, dexamethasone sodium phosphate and dexamethasone were monitored by HPLC, the percentage of insulin and dexamethasone sodium phosphate content compared to the theoretical values were recorded, and the peak area of dexamethasone was sampled 3 times per batch in parallel and the results averaged.

After 90 days of standing, no dexamethasone was detected for all batches. The results of insulin and dexamethasone sodium phosphate content are shown in figures 10 and 11.

The results show that: all batches of insulin and dexamethasone sodium phosphate remained stable after 90 days at 4 ℃ with no significant changes in content.

Example 6: effect of different surfactants on insulin and dexamethasone sodium phosphate stability

Dissolving human insulin raw material, dexamethasone sodium phosphate raw material, zinc chloride, m-cresol, tween 20, tween 80, 15-hydroxystearic acid polyethylene glycol ester, disodium hydrogen phosphate and sodium dihydrogen phosphate. Compound formulations of different prescriptions were formulated as in table 12, with final pH adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 12 compound preparation prescription design

The compound preparation is a sterile preparation, which is packaged in sterilized penicillin bottles, covered by rubber plugs and tightly rolled by aluminum-plastic combined covers, and then placed in an incubator at 25 ℃. Samples were taken periodically, the contents of insulin, dexamethasone sodium phosphate and dexamethasone were monitored by HPLC, the percentage of insulin and dexamethasone sodium phosphate content compared to the theoretical values were recorded, and the peak area of dexamethasone was sampled 3 times per batch in parallel and the results averaged.

After 38 days of standing, no dexamethasone was detected for all batches. The results of insulin and dexamethasone sodium phosphate content are shown in figures 12 and 13.

The results show that: all batches of insulin and dexamethasone sodium phosphate remained stable after 38 days at 25℃ with no significant changes in content.

Example 7: effect of isotonic agent on insulin and dexamethasone sodium phosphate stability

The human insulin drug substance, dexamethasone sodium phosphate drug substance, zinc chloride, m-cresol, tween 20, sodium chloride, disodium hydrogen phosphate and sodium dihydrogen phosphate are dissolved by ultrapure water. Compound formulations of different prescriptions were formulated as in table 13, with final pH adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 13 prescription design of compound preparation

After 181 days of standing, no dexamethasone was detected for all batches. The results of insulin and dexamethasone sodium phosphate content are shown in figures 14 and 15.

The results show that: all batches of insulin and dexamethasone sodium phosphate remained stable after 181 days at 4 ℃ with no significant change in content.

Example 8:

the human insulin drug substance, dexamethasone sodium phosphate drug substance, zinc chloride, m-cresol, tween 20, sodium chloride, disodium hydrogen phosphate and sodium dihydrogen phosphate are dissolved by ultrapure water. Compound formulations of different formulations were formulated as in table 14, with final pH adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Table 14 compound preparation prescription design

In addition, a compound preparation was prepared according to the formulation shown in the following comparative example, and the final pH was adjusted to 7.4 with hydrochloric acid and sodium hydroxide.

Comparative example 1

The only difference from the E2 batch in example 8 is that the compound formulation has the following groups and corresponding concentrations: 50. Mu.g/mL insulin, 50. Mu.g/mL dexamethasone sodium phosphate, 1.25. Mu.g/mL zinc chloride, and 60mmol/L phosphate buffer.

Comparative example 2

The only difference from the R1 lot in example 8 is that the compound formulation has the following groups and corresponding concentrations: 10. Mu.g/mL insulin, 10. Mu.g/mL dexamethasone sodium phosphate, 0.0125. Mu.g/mL zinc chloride, 3. Mu.g/mL m-cresol, 3. Mu.g/mL Tween 20, 10mg/mL sodium chloride and 3mmol/L phosphate buffer.

Comparative example 3

The only difference from the R1 lot in example 8 is that the compound formulation has the following groups and corresponding concentrations: 10. Mu.g/mL insulin, 10. Mu.g/mL dexamethasone sodium phosphate, 0.02. Mu.g/mL zinc chloride, 5. Mu.g/mL m-cresol, 5. Mu.g/mL Tween 20, 9mg/mL sodium chloride and 3mmol/L phosphate buffer.

The compound preparation is a sterile preparation, which is packaged in penicillin bottles, covered by rubber plugs and tightly covered by aluminum-plastic combination covers, placed in a constant temperature box at 40 ℃, placed for 30 days and sampled, and the contents of insulin, dexamethasone sodium phosphate and dexamethasone are detected by HPLC.

Comparing the HPLC detection data of E, R, E, E2, R1 and R2 batches in example 8 and the compound preparation prepared in comparative examples 1-3 after being placed at 40 ℃ for 30 days, the results show that:

no change in the sodium phosphate content was observed for batches E, R, E1, E2, R1 and R2, no more than 30% decrease in the insulin content was observed for dexamethasone, and it was assumed that these batches remained stable for a long period of time when placed at 4 c, as indicated by the G, I batch in example 4.

2. The dexamethasone sodium phosphate content in comparative example 1 was substantially unchanged, but dexamethasone could be detected, while the insulin content was reduced by more than 40%. It is speculated that excessive zinc reduces the stability of dexamethasone sodium phosphate, whereas the hydrolysis of dexamethasone further reduces the stability of insulin in the system.

3. In comparative example 2, the content of dexamethasone sodium phosphate is reduced, dexamethasone is formed, the content of insulin is also greatly reduced, and the stability of the preparation is poor when the dosage of other components is outside the protection range of the invention.

4. The dexamethasone sodium phosphate content in comparative example 3 is basically unchanged, but the dexamethasone and insulin content can be detected to be reduced by about 30%, and the concentration of the buffer solution in the prescription is presumed to be low, the buffer capacity is weak, the dexamethasone sodium phosphate can not be inhibited from being hydrolyzed to form dexamethasone, but the dosages of zinc, m-cresol and tween 20 are reasonable, and the stability of insulin is maintained.

Example 9: insulin and dexamethasone sodium phosphate compound preparation for delaying early-stage T1D progression

STZ formulation concentration was 10mg/mL. The injection quantity of the mice is 50mg/kg. I.e. 20g mice were injected at a volume of 100 μl. STZ was injected intraperitoneally, five times for five consecutive days. Immediately after five continuous STZ injections, the administration of commercial insulin injections combined with commercial dexamethasone sodium phosphate injection or insulin and dexamethasone sodium phosphate compound preparation (batch G) was started, at a dose of 10 μg/dose; immunotherapy was performed once a week for a total of 9 times. The results of the blood glucose changes in mice are shown in FIG. 16.

The results show that: early intervention of the T1D treatment regimen with the insulin and dexamethasone sodium phosphate compound formulation (lot G) can delay the glycemic progression in diabetic mice.

Example 10: delay control of late-stage T1D blood sugar by insulin and dexamethasone sodium phosphate compound preparation

STZ formulation concentration was 10mg/mL. The injection quantity of the mice is 50mg/kg. I.e. 20g mice were injected at a volume of 100 μl. STZ was injected intraperitoneally, five times for five consecutive days. After five continuous STZ injections were successfully molded and the blood sugar of the mice was in a disease state (day 17), the administration of the commercial insulin injection combined with the commercial dexamethasone sodium phosphate injection or the administration of the insulin and dexamethasone sodium phosphate compound preparation (batch G) was started, and the dosage was 10 mug/dose; immunotherapy is performed once a week. The results of the blood glucose changes in mice are shown in FIG. 17.

The results show that: the commercial insulin in combination with dexamethasone, insulin and dexamethasone sodium phosphate formulation (batch G) was able to lower blood glucose in diabetic mice after treatment of diabetic mice.

Finally, it should be noted that the above description is only for illustrating the technical solution of the present invention, and not for limiting the scope of the present invention, and that the simple modification and equivalent substitution of the technical solution of the present invention can be made by those skilled in the art without departing from the spirit and scope of the technical solution of the present invention.

Claims

1. A stable pharmaceutical composition comprising both insulin and dexamethasone sodium phosphate, characterized in that: the pharmaceutical composition consists of insulin, dexamethasone sodium phosphate, zinc, buffer solution, phenols and surfactant;

the pharmaceutical composition comprises 10-50 mug/mL insulin, 10-50 mug/mL dexamethasone sodium phosphate, 0.01-0.5 mug/mL zinc, 5-50mmol/L buffer solution, 5-50 mug/mL phenols and 0-100 mug/mL surfactant; the weight ratio of the insulin to the dexamethasone sodium phosphate is 5:1-1:5, a step of; the buffer comprises one or more of phosphate, acetate, citrate, arginine, N-glycylglycine and Tris buffer.

2. The pharmaceutical composition according to claim 1, wherein: the zinc is added in the form of zinc salts including one or more of zinc chloride, zinc bromide, zinc iodide, zinc fluoride, zinc sulfate and zinc acetate.

3. The pharmaceutical composition according to claim 1, wherein: the phenols include one or more of cresol, chlorocresol, and phenol.

4. The pharmaceutical composition according to claim 1, wherein: the surfactant comprises one or more of Tween 20, tween 40, tween 80, poloxamer 188, polyoxyethylene 35 castor oil, polyoxyethylene 40 hydrogenated castor oil, 15-hydroxystearic acid polyethylene glycol ester, polyethylene glycol 300, polyethylene glycol 400 and polyethylene glycol 600.

5. Use of a pharmaceutical composition according to any one of claims 1-4 for the preparation of a compound injection.

6. An HPLC method for simultaneous detection of insulin and dexamethasone sodium phosphate in a pharmaceutical composition according to any one of claims 1-4, characterized in that: the HPLC method uses a 250mM 4.6mM 5 μm column of Sepax Bio-C8 with 200mM Na ₂ SO ₄ : acetonitrile=74: 26, run at column temperature 45 ℃ for 50min using a flow rate of 0.8 mL/min; the Na is ₂ SO ₄ The pH of (C) was 2.3.