Rabies vaccine protective agent, rabies vaccine and preparation method thereof

Abstract

The present invention relates to a rabies vaccine protective agent, a rabies vaccine and a preparation method thereof. The rabies vaccine protective agent does not contain gelatin and dextran 40, but contains human serum albumin, trehalose, maltose and an antioxidant, wherein the mass ratio of human serum albumin, trehalose, maltose and the antioxidant is (0.5-2): (0.2-1): (1-4): (0.05-0.4), and the antioxidant includes vitamins. The rabies vaccine prepared by using the rabies vaccine protective agent has good potency stability and high safety.

Description

Technical Field

[0001] The present invention relates to the field of biological products, and in particular to a rabies vaccine protective agent, a rabies vaccine and a preparation method thereof. Background Art

[0002] Rabies is a natural epidemic disease and a malignant infectious disease with a mortality rate of almost 100%. Rabies is usually transmitted to humans by infected animals through bites. The clinical manifestations include unique fear of water, fear of sound, fear of wind, fear and anxiety, pharyngeal muscle spasm, progressive paralysis, etc. According to statistics from the World Health Organization (WHO), the prevalence of rabies in all countries and regions in the world is as high as 80%. Currently, more than 30,000 to 50,000 rabies cases are reported each year. However, there is currently no effective treatment for rabies, and the mortality rate is extremely high, posing a serious threat to human life and health. Therefore, vaccination with rabies vaccine is an effective means to prevent rabies and ensure human health.

[0003] There are many types of rabies vaccines on the market; however, the potency stability of current rabies vaccines is poor. Summary of the invention

[0004] Based on this, the embodiments of the present application provide a rabies vaccine protective agent, which can improve the current problem of poor stability of rabies vaccine potency.

[0005] A rabies vaccine protective agent does not contain gelatin and dextran 40, and contains human serum albumin, trehalose, maltose and an antioxidant, wherein the mass ratio of human serum albumin, trehalose, maltose and the antioxidant is (0.5-2): (0.2-1): (1-4): (0.05-0.4), and the antioxidant includes vitamin E.

[0006] The above rabies vaccine protective agent can improve the stability of the rabies vaccine potency by combining human albumin, trehalose, maltose and antioxidants; and the above rabies vaccine protective agent does not contain gelatin and dextran 40, so that the rabies vaccine prepared by the above rabies vaccine protective agent is less likely to cause allergic reactions after vaccination, reducing the risk of potential side effects and having better safety. In addition, the freeze-dried rabies vaccine prepared by the above rabies vaccine protective agent has a very short dissolution time (30s to 35s) when used, which is conducive to clinical application.

[0007] In one embodiment, the vitamin E includes α-tocopherol and / or α-tocopherol derivatives.

[0008] In one embodiment, the α-tocopherol derivative comprises α-tocopherol acetate and / or α-tocopherol succinate.

[0009] In one embodiment, the rabies vaccine protective agent includes a solvent, and the solvent includes water. In 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 0.5g to 2g, the content of trehalose is 0.2 to 1g, the content of maltose is 1g to 4g, and the content of the antioxidant is 0.05g to 0.4g.

[0010] In one embodiment, in 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 1g to 2g, the content of trehalose is 0.5 to 1g, the content of maltose is 1g to 3g, and the content of the antioxidant is 0.1g to 0.3g.

[0011] In one embodiment, the rabies vaccine protectant does not contain antibiotics.

[0012] In one embodiment, the pH of the rabies vaccine protective agent is 7.2-7.6.

[0013] In one embodiment, the rabies vaccine protective agent is composed of human serum albumin, trehalose, maltose, the antioxidant, a pH adjuster and water. In 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 0.5g~2g, the content of trehalose is 0.2~1g, the content of maltose is 1g~4g, and the content of the antioxidant is 0.05g~0.4g.

[0014] A method for preparing a rabies vaccine comprises the following steps:

[0015] The inactivated rabies virus is mixed with the above-mentioned rabies vaccine protective agent to prepare the rabies vaccine.

[0016] In one embodiment, after the rabies virus and the rabies vaccine protective agent are mixed, the method further includes freeze-drying the mixed mixture.

[0017] In one embodiment, the rabies virus is the Pitman-Moore 1503-3M strain.

[0018] A rabies vaccine is prepared by the above-mentioned rabies vaccine preparation method.

[0019] In one embodiment, the rabies vaccine does not contain antibiotics. DETAILED DESCRIPTION

[0020] In order to facilitate understanding of the present invention, the present invention will be described more fully below. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the disclosure of the present invention more thorough and comprehensive.

[0021] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art to which the present invention belongs. The terms used herein in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" includes any and all combinations of one or more of the related listed items.

[0022] One embodiment of the present application provides a rabies vaccine protective agent, which does not contain gelatin and dextran 40. The rabies vaccine protective agent contains human serum albumin, trehalose, maltose and an antioxidant, wherein the mass ratio of human serum albumin, trehalose, maltose and antioxidant is (0.5-2): (0.2-1): (1-4): (0.05-0.4), and the antioxidant includes vitamin E.

[0023] The stability of vaccine potency is closely related to the protective agent. The above-mentioned rabies vaccine protective agent can improve the stability of rabies vaccine potency by combining human albumin, trehalose, maltose and antioxidants. In addition, traditional rabies vaccine protective agents generally contain gelatin and dextran 40, which can easily cause allergic reactions after vaccination. The above-mentioned rabies vaccine protective agent does not contain gelatin and dextran 40, so that the rabies vaccine prepared by the above-mentioned rabies vaccine protective agent is less likely to cause allergic reactions after vaccination, reducing the potential risk of side effects and having better safety. In addition, the freeze-dried rabies vaccine prepared by the above-mentioned rabies vaccine protective agent has a very short dissolution time (30s to 35s) when used, which is conducive to clinical application.

[0024] In some embodiments, the antioxidant in the above-mentioned rabies vaccine protective agent is vitamin E. Vitamin E exists naturally in 8 different forms: four tocopherols and four tocotrienols (α-tocopherol, β-tocopherol, γ-tocopherol and δ-tocopherol; four tocotrienols: α-tocotrienol, β-tocotrienol, γ-tocotrienol and δ-tocotrienol), all of which have antioxidant properties. α-Tocopherol is the most widely distributed, most abundant and most active vitamin E in nature. In some embodiments, the vitamin E in the above-mentioned rabies vaccine protective agent includes α-tocopherol and / or α-tocopherol derivatives. Optionally, the α-tocopherol derivative is an esterified derivative of α-tocopherol. Further, the α-tocopherol derivative includes α-tocopherol acetate and / or α-tocopherol succinate. In other embodiments, the antioxidant in the above-mentioned rabies vaccine protective agent includes other substances with antioxidant effects in addition to vitamin E, such as vitamin C.

[0025] Furthermore, in the above-mentioned rabies vaccine protective agent, the mass ratio of human serum albumin, trehalose, maltose and antioxidant is (1-2): (0.5-1): (1-3): (0.1-0.3). Furthermore, in the above-mentioned rabies vaccine protective agent, the mass ratio of human serum albumin, trehalose, maltose and antioxidant is (1-1.5): (0.5-0.8): (2-3): (0.1-0.2).

[0026] In some embodiments, the above-mentioned rabies vaccine protective agent is a dry preparation, such as a dry powder preparation. In this case, the rabies vaccine protective agent needs to be mixed with water when used.

[0027] In other embodiments, the above-mentioned rabies vaccine protective agent also includes a solvent. Optionally, the solvent includes water. The rabies vaccine protective agent at this time is an aqueous solution. Further, in 100mL of the above-mentioned rabies vaccine protective agent, the content of human serum albumin is 0.5g~2g, the content of trehalose is 0.2~1g, the content of maltose is 1g~4g, and the content of antioxidant is 0.05g~0.4g. The rabies vaccine protective agent of human serum albumin, trehalose, maltose and antioxidants set according to the above-mentioned is a rabies vaccine protective agent in a use concentration state, which can be used directly at this time without dilution. It is understandable that in other embodiments, the above-mentioned rabies vaccine protective agent can also be a non-use concentration (for example, 2×, 5×, 10×, etc.) state or a concentrated state, which needs to be diluted before use.

[0028] Optionally, in 100mL of the above-mentioned rabies vaccine protective agent, the content of human serum albumin is 0.5g, 0.8g, 1g, 1.2g, 1.5g, 1.8g or 2g. Optionally, in 100mL of the above-mentioned rabies vaccine protective agent, the content of trehalose is 0.2g, 0.4g, 0.5g, 0.7g, 0.8g or 1g. Optionally, in 100mL of the above-mentioned rabies vaccine protective agent, the content of maltose is 1g, 1.5g, 2g, 2.5g, 3g, 3.5g, 4g, 4.5g or 5g. Optionally, in 100mL of the above-mentioned rabies vaccine protective agent, the content of antioxidant is 0.05g, 0.08g, 0.1g, 0.15g, 0.2g, 0.25g, 0.3g, 0.35g or 0.4g.

[0029] Furthermore, in 100 mL of the above-mentioned rabies vaccine protective agent, the content of human albumin is 1g to 2g, the content of trehalose is 0.5 to 1g, the content of maltose is 1g to 3g, and the content of antioxidant is 0.1g to 0.3g. Further, in 100 mL of the above-mentioned rabies vaccine protective agent, the content of human albumin is 1g to 1.5g, the content of trehalose is 0.5 to 0.8g, the content of maltose is 2g to 3g, and the content of antioxidant is 0.1g to 0.2g.

[0030] In some embodiments, in 100mL of the above-mentioned rabies vaccine protective agent, the content of human albumin is 0.5g~2g, the content of trehalose is 0.2~1g, the content of maltose is 1g~4g, and the content of vitamin E is 0.05g~0.4g. Further, in 100mL of the above-mentioned rabies vaccine protective agent, the content of human albumin is 1g~2g, the content of trehalose is 0.5~1g, the content of maltose is 1g~3g, and the content of vitamin E is 0.1g~0.3g. Further, in 100mL of the above-mentioned rabies vaccine protective agent, the content of human albumin is 1g~1.5g, the content of trehalose is 0.5~0.8g, the content of maltose is 2g~3g, and the content of vitamin E is 0.1g~0.2g.

[0031] In some embodiments, the pH of the above-mentioned rabies vaccine protective agent is 7.2-7.6. According to such a setting, the stability of the viral antigen can be better. In an optional specific example, the pH of the above-mentioned rabies vaccine protective agent is 7.2, 7.3, 7.4, 7.5 or 7.6. Further, the pH of the above-mentioned rabies vaccine protective agent is 7.3-7.5.

[0032] In some embodiments, the above-mentioned rabies vaccine protectant does not contain antibiotics.

[0033] In some embodiments, the above-mentioned rabies vaccine protective agent is composed of human serum albumin, trehalose, maltose, antioxidants, pH regulators and water. In 100mL of rabies vaccine protective agent, the content of human serum albumin is 0.5g~2g, the content of trehalose is 0.2~1g, the content of maltose is 1g~4g, the content of antioxidants is 0.05g~0.4g, and the pH of the above-mentioned rabies vaccine protective agent is 7.2~7.6. Optionally, the pH regulator includes disodium hydrogen phosphate. It is understandable that in other embodiments, the pH regulator is not limited to disodium hydrogen phosphate, and other substances for adjusting the pH of the solution can also be used. Of course, the antioxidant is as described above. Furthermore, the above-mentioned rabies vaccine protective agent is composed of human serum albumin, trehalose, maltose, antioxidants, pH adjusters and water. In 100mL of rabies vaccine protective agent, the content of human serum albumin is 1g~2g, the content of trehalose is 0.5~1g, the content of maltose is 1g~3g, the content of antioxidant is 0.1g~0.3g, and the pH of the above-mentioned rabies vaccine protective agent is 7.2~7.6. Furthermore, the above-mentioned rabies vaccine protective agent is composed of human serum albumin, trehalose, maltose, antioxidants, pH adjusters and water. In 100mL of rabies vaccine protective agent, the content of human serum albumin is 1g~1.5g, the content of trehalose is 0.5~0.8g, the content of maltose is 2g~3g, the content of antioxidant is 0.1g~0.2g, and the pH of the above-mentioned rabies vaccine protective agent is 7.2~7.6.

[0034] In addition, one embodiment of the present application also provides a use of a rabies vaccine protective agent according to any of the above embodiments in the preparation of a rabies vaccine.

[0035] In addition, one embodiment of the present application also provides a method for preparing a rabies vaccine, which comprises the following steps: mixing the inactivated rabies virus with the rabies vaccine protective agent of any of the above embodiments to prepare the rabies vaccine.

[0036] In some embodiments, the above preparation method includes step S110 and step S120. Specifically:

[0037] Step S110: preparing inactivated rabies virus.

[0038] In some implementations, the step of preparing the inactivated rabies virus includes step S111, step S112, step S113 and step S114. Specifically:

[0039] Step S111: inoculating rabies virus into human diploid cells.

[0040] In some embodiments, the rabies virus is the Pitman-Moore 1503-3M strain. It is understood that in other embodiments, the rabies virus is not limited to the above, and can also be other strains, such as the CTN-1 strain.

[0041] In some embodiments, before inoculating the rabies virus into the human diploid cells, the method further includes a step of thawing the human diploid cells.

[0042] In some embodiments, the human diploid cell is MRC-5 or KBM17. It is understood that in other embodiments, the human diploid cell is not limited thereto, and may also be other cells.

[0043] Step S112: Cultivate human diploid cells inoculated with rabies virus to obtain virus liquid.

[0044] In some embodiments, the step of culturing human diploid cells inoculated with rabies virus includes: pre-culturing human diploid cells inoculated with rabies virus; when the virus is amplified to a virus titer of not less than 6.01 gPFU / mL, continuously perfusing and harvesting the virus liquid for 8 to 10 days to obtain the virus liquid. In some embodiments, the next step of purification is performed after all the virus liquid is collected. In other embodiments, the subsequent purification and inactivation steps can be performed while the virus liquid is collected, that is, the purification and inactivation steps are performed after a certain amount of the virus liquid is collected instead of after all the collection is completed.

[0045] In some embodiments, antibiotics are not added when culturing human diploid cells.

[0046] Step S113: purify the virus solution to obtain a purified virus solution.

[0047] In some embodiments, the step of purifying the virus liquid includes: using the harvested virus liquid to clarify and filter with a 0.65 μm filter membrane, concentrate with a 300 kDa ultrafiltration, and then purify with ultracentrifugation and column chromatography molecular sieve to obtain a purified virus liquid. It is understood that in other embodiments, the method of purifying the virus liquid is not limited to the above, and can also be other methods.

[0048] Step S114: inactivate the virus purification solution to obtain inactivated rabies virus.

[0049] In some embodiments, beta-propiolactone (BPL) is used to inactivate the virus. Optionally, the step of inactivating the purified virus solution includes: adding a beta-propiolactone solution with a final concentration of 1:4000 to 1:6000 to the purified virus solution, inactivating it in an environment of 2 to 8°C for 20 to 24 hours, and hydrolyzing it at 37°C for 2 hours. The inactivated purified virus solution has an antigen content of no less than 8 IU / dose (determined by ELSIA method), thereby obtaining the inactivated rabies virus.

[0050] Step S120: Mix the inactivated rabies virus with the rabies vaccine protective agent of any of the above embodiments to prepare a rabies vaccine.

[0051] In some embodiments, the antigen content of the inactivated rabies virus in the rabies vaccine is 8 IU / dose or more, for example 8 IU / dose or 10 IU / dose. At this time, the amount of inactivated rabies virus added can be adjusted according to the specifications of the rabies vaccine to be prepared.

[0052] In some embodiments, the rabies vaccine is an aqueous solution. In this case, the inactivated rabies virus is mixed with the rabies vaccine protective agent of any of the above embodiments to prepare the rabies vaccine.

[0053] In other embodiments, the rabies vaccine is a freeze-dried preparation. In this case, after the rabies virus is mixed with the rabies vaccine protective agent of any of the above embodiments, the process further includes freeze-drying the mixed mixture. Optionally, the freeze-drying step includes: (1) pre-freezing: setting the temperature to quickly drop to -45°C to -50°C within 1 to 2 hours, and pre-freezing at -45 for 4 to 6 hours. Vacuum control: (2) pre-vacuuming: pre-vacuuming at -45°C to -50°C to 0.2 mbar; (3) primary drying: adjusting the temperature to -30°C-35°C, maintaining the vacuum degree at 0.15 mbar, and keeping warm for 14 to 16 hours; adjusting the temperature to -25°C-28°C, maintaining the vacuum degree at 0.15 mbar, and keeping warm for 10 to 12 hours; (4) analytical drying: quickly heating to 10 to 15°C within 1 hour, with a vacuum degree of 0.1 mbar, and maintaining for 1 hour; then heating to 30°C, with a vacuum degree of 0.1 mbar, and keeping warm for 4 to 6 hours.

[0054] The preparation method of the rabies vaccine is simple and can be used for large-scale production of rabies vaccines.

[0055] In addition, an embodiment of the present application further provides a rabies vaccine, wherein the rabies vaccine protective agent of any of the above-mentioned embodiments is used in the preparation process of the rabies vaccine.

[0056] In some embodiments, the rabies vaccine comprises, in addition to the rabies vaccine protective agent of any of the above embodiments, other pharmaceutically acceptable excipients. Pharmaceutically acceptable excipients are compatible with other ingredients of the pharmaceutical preparation and suitable for contact with tissues or organs of the recipient (e.g., human or animal). There is no or little toxicity, irritation, allergic reaction, immunogenicity or other complications when used.

[0057] In some embodiments, the dosage form of the rabies vaccine is powder or liquid. It is to be understood that the dosage form of the rabies vaccine is not limited to the above, and may also be other dosage forms. Of course, for different dosage forms, pharmaceutically acceptable excipients are added accordingly during the preparation process.

[0058] In some embodiments, the rabies vaccine described above does not contain antibiotics.

[0059] The above-mentioned rabies vaccine uses the rabies vaccine protective agent of any of the above-mentioned embodiments as a protective agent during its preparation process. It has been verified that it has good potency stability and safety, and the above-mentioned rabies vaccine in a freeze-dried state can also be quickly dissolved in water, which is convenient for clinical application. Specific embodiments

[0061] The following is a detailed description in conjunction with specific examples. The following examples do not include other components except for inevitable impurities unless otherwise specified. The reagents and instruments used in the examples are conventionally selected in the art unless otherwise specified. The experimental methods for which specific conditions are not specified in the examples are implemented according to conventional conditions, such as the conditions described in the literature, books, or methods recommended by the manufacturer. The vitamin E used in the following examples is alpha-tocopherol acetate.

[0062] Example 1

[0063] 1.1. Preparation of rabies vaccine semi-finished products

[0064] (1) The revived human diploid cells are digested and passaged, and then cultured and amplified in a bioreactor, and then rabies virus is inoculated on the human diploid cells; the virus is amplified until the virus titer is not less than 6.0lgPFU / mL, and the virus liquid is harvested by continuous perfusion for 10 days, and then the virus liquid is further purified to improve the purity. The purification process is as follows: the virus harvest liquid is first clarified and filtered using a 0.65μm filter membrane, then concentrated by 300kDa ultrafiltration, and then purified by ultracentrifugation and column chromatography molecular sieve purification to obtain a purified virus liquid.

[0065] (2) Adding a BPL solution with a final concentration of 1:4000 to the virus purification solution obtained in step (1), inactivating the solution at 4°C for 24 hours, and hydrolyzing the solution at 37°C for 2 hours to obtain an inactivated virus purification solution.

[0066] (3) Mixing the purified virus solution obtained in step (2) with the vaccine protective agent in an amount of about 8 IU / dose to obtain a semi-finished vaccine. The rabies vaccine protective agent is composed of: 0.5% (w / v) human albumin, 0.2% (w / v) trehalose, 4% (w / v) maltose, 0.05% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4. "w / v" in this article represents the mass volume percentage content, for example, 0.5% (w / v) human albumin means that 100 mL of rabies vaccine protective agent contains 0.5g of human albumin.

[0067] 1.2. Freeze-drying of rabies vaccine semi-finished products

[0068] The semi-finished vaccine product obtained in step 1.1 (3) is freeze-dried by the following freeze-drying method:

[0069] (1) Cleaning and sterilization of freeze dryer: Clean the inner cavity of the freeze dryer once with water for injection, then sterilize it with pure steam at 121°C for 30 min, and dry and cool it to room temperature.

[0070] (2) Packaging and loading: Pack the prepared vaccine semi-finished product into neutral borosilicate vials, 1.0 ml / vial, and half-fill the vials with stoppers. Set the freeze dryer plate layer to 5°C and place the vials on the freeze dryer plate layer. After loading, close the box door and continue to maintain for 1 hour.

[0071] (3) Prefreezing: Set to quickly cool to -45°C within 1 hour, and prefreeze at -45°C for 5 hours. Vacuum control 0 bar;

[0072] (4) Pre-vacuuming: Turn on the vacuum pump and pre-vacuum to 0.2 mbar at -45°C.

[0073] (5) Primary drying: Adjust the plate temperature to -35°C, maintain the vacuum degree at 0.15 mbar, and keep warm for 16 hours; Adjust the plate temperature to -25°C, maintain the vacuum degree at 0.15 mbar, and keep warm for 12 hours.

[0074] (6) Desorption and drying: Adjust the plate temperature, quickly raise the temperature of the partition to 15°C within 1 hour, and maintain the vacuum degree to 0.1 mbar for 1 hour; then raise the temperature of the partition to 30°C, and maintain the vacuum degree to 0.1 mbar for 4 hours. After the freeze-drying is completed, lower the plate vacuum to press the bottle stopper, take it out of the box, and seal it with a cap to obtain the rabies vaccine.

[0075] 1.3. Quality evaluation of rabies vaccines

[0076] (1) Vaccine finished product testing: refer to the standard of "Freeze-dried human diploid cell rabies vaccine" in the "Chinese Pharmacopoeia". The inspection items include appearance, moisture, dissolution time, potency, abnormal toxicity inspection, etc. The inspection results are shown in Table 1.

[0077] (2) 37°C accelerated stability test: The vaccine was placed in an environment with a temperature of 37°C and a humidity of 60-70%. Samples were taken at 28 days, 60 days, and 90 days to test moisture, potency, etc. The test results are shown in Tables 2 and 3.

[0078] Example 2

[0079] The preparation method of the rabies vaccine of this embodiment is substantially the same as that of Example 1, except that the rabies vaccine protective agent used in this embodiment is different from that in Example 1. The composition of the rabies vaccine protective agent used in this embodiment is: 1% (w / v) human serum albumin, 0.5% (w / v) trehalose, 3% (w / v) maltose, 0.1% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0080] Example 3

[0081] The preparation method of the rabies vaccine of this embodiment is substantially the same as that of Example 1, except that the rabies vaccine protective agent used in this embodiment is different from that in Example 1. The composition of the rabies vaccine protective agent used in this embodiment is: 1.5% (w / v) human serum albumin, 0.8% (w / v) trehalose, 2% (w / v) maltose, 0.2% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0082] Example 4

[0083] The preparation method of the rabies vaccine of this embodiment is substantially the same as that of Example 1, except that the rabies vaccine protective agent used in this embodiment is different from that in Example 1. The composition of the rabies vaccine protective agent used in this embodiment is: 2% (w / v) human serum albumin, 1% (w / v) trehalose, 1% (w / v) maltose, 0.4% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0084] Example 5

[0085] The preparation method of the rabies vaccine of this embodiment is substantially the same as that of Example 1, except that the rabies vaccine protective agent used in this embodiment is different from that in Example 1. The composition of the rabies vaccine protective agent used in this embodiment is: 0.5% (w / v) human serum albumin, 1% (w / v) trehalose, 4% (w / v) maltose, 0.2% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0086] Control group 1

[0087] The preparation method of the rabies vaccine of this control group is roughly the same as that of Example 1, except that the rabies vaccine protective agent used in this example is different from that in Example 1. The composition of the rabies vaccine protective agent used in this control group is: 0.5% (w / v) human serum albumin, 0.2% (w / v) trehalose, 0.5% (w / v) maltose, 0.2% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0088] Control group 2

[0089] The preparation method of the rabies vaccine of this control group is roughly the same as that of Example 1, except that the rabies vaccine protective agent used in this example is different from that in Example 1. The composition of the rabies vaccine protective agent used in this control group is: 1% (w / v) human serum albumin, 1% (w / v) trehalose, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0090] Control group 3

[0091] The preparation method of the rabies vaccine of this control group is roughly the same as that of Example 1, except that the rabies vaccine protective agent used in this example is different from that in Example 1. The composition of the rabies vaccine protective agent used in this control group is: 1% (w / v) human serum albumin, 2% (w / v) maltose, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0092] Control group 4

[0093] The preparation method of the rabies vaccine of this control group is roughly the same as that of Example 1, except that the rabies vaccine protective agent used in this control group is different from that in Example 1. The composition of the rabies vaccine protective agent used in this control group is: 0.5% (w / v) human serum albumin, 1.5% (w / v) trehalose, 4% (w / v) maltose, 0.05% (w / v) vitamin E, disodium hydrogen phosphate and water, wherein 18% (w / v) disodium hydrogen phosphate is used to adjust the pH to 7.4.

[0094] Table 1 Test results of human diploid cell rabies vaccine

[0095]

[0096] Table 2 Water content results of human diploid cell rabies vaccine at 37℃ accelerated stability test

[0097]

[0098] Table 3 The titer results of human diploid cell rabies vaccine at 37℃ accelerated stability test

[0099]

[0100] It can be seen from Tables 1 to 3 above that the quality standards of the human diploid cell rabies vaccine of Examples 1 to 5 meet the quality standards of the "freeze-dried human diploid cell rabies vaccine" in the 2020 edition of the Chinese Pharmacopoeia, and the safety of the abnormal toxicity test meets the requirements of the Pharmacopoeia. In addition, the dissolution time, moisture, and thermal stability test results of the human diploid cell rabies vaccine of Examples 1 to 54 under the same conditions are significantly better than the existing (control group) vaccine preparation method, and have good market application prospects.

[0101] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0102] The above-described embodiments only express several implementation methods of the present invention, which is convenient for understanding the technical solutions of the present invention in detail, but cannot be understood as limiting the scope of protection of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention. It should be understood that the technical solutions obtained by those skilled in the art through logical analysis, reasoning or limited experiments on the basis of the technical solutions provided by the present invention are all within the protection scope of the claims attached to the present invention. Therefore, the protection scope of the patent of the present invention shall be based on the contents of the attached claims, and the description can be used to interpret the contents of the claims.

Claims

1. A rabies vaccine protective agent, characterized in that: The rabies vaccine protective agent does not contain gelatin and dextran 40, and contains human serum albumin, trehalose, maltose and an antioxidant, wherein the mass ratio of human serum albumin, trehalose, maltose and the antioxidant is (0.5-2): (0.2-1): (1-4): (0.05-0.4), and the antioxidant includes vitamin E.

2. The rabies vaccine protective agent according to claim 1, characterized in that: The vitamin E includes α-tocopherol and / or α-tocopherol derivatives; the α-tocopherol derivatives include α-tocopherol acetate and / or α-tocopherol succinate.

3. The rabies vaccine protective agent according to claim 2, characterized in that: The rabies vaccine protective agent includes a solvent, and the solvent includes water. In 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 0.5g to 2g, the content of trehalose is 0.2 to 1g, the content of maltose is 1g to 4g, and the content of the antioxidant is 0.05g to 0.4g.

4. The rabies vaccine protective agent according to claim 3, characterized in that: In 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 1 g to 2 g, the content of trehalose is 0.5 to 1 g, the content of maltose is 1 g to 3 g, and the content of the antioxidant is 0.1 g to 0.3 g.

5. The rabies vaccine protective agent according to claim 3, characterized in that: The rabies vaccine protective agent does not contain antibiotics.

6. The rabies vaccine protective agent according to any one of claims 1 to 5, characterized in that The pH of the rabies vaccine protective agent is 7.2-7.

6.

7. The rabies vaccine protective agent according to claim 1 or 2, characterized in that: The rabies vaccine protective agent consists of human serum albumin, trehalose, maltose, the antioxidant, a pH regulator and water. In 100 mL of the rabies vaccine protective agent, the content of human serum albumin is 0.5g to 2g, the content of trehalose is 0.2 to 1g, the content of maltose is 1g to 4g, and the content of the antioxidant is 0.05g to 0.4g.

8. A method for preparing a rabies vaccine, characterized in that: The following steps are involved: The inactivated rabies virus is mixed with the rabies vaccine protective agent according to any one of claims 1 to 7 to prepare a rabies vaccine.

9. The preparation method according to claim 8, characterized in that: After the rabies virus and the rabies vaccine protective agent are mixed, the method further comprises the step of freeze-drying the mixed mixture.

10. The preparation method according to claim 8 or 9, characterized in that: The rabies virus is the Pitman-Moore 1503-3M strain.

11. A rabies vaccine, characterized in that: The rabies vaccine is prepared by the rabies vaccine preparation method according to any one of claims 8 to 10.

12. The rabies vaccine according to claim 11, characterized in that The rabies vaccine does not contain antibiotics.

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