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Heat-resistant freeze-drying protectant for duck viral hepatitis live vaccine and its preparation method and application

A heat-resistant freeze-drying technology for duck viral hepatitis, which is applied in the direction of antiviral agents, freeze-dried delivery, and viral antigen components, can solve the problems of poor antigen activity protection ability, reduce the risk of inactivation, and reduce the cost of freeze-drying , Efficient protection of antigenic activity

Active Publication Date: 2018-08-17
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the prior art, the main components of heat-resistant freeze-drying protective agents commonly used in duck viral hepatitis live vaccines are lactose, milk, gelatin, etc., although the formula is simple, the product is stable in shape, easy to dissolve, and low in cost, but in freeze-drying and poor protection against antigenic activity during high temperature storage

Method used

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  • Heat-resistant freeze-drying protectant for duck viral hepatitis live vaccine and its preparation method and application
  • Heat-resistant freeze-drying protectant for duck viral hepatitis live vaccine and its preparation method and application
  • Heat-resistant freeze-drying protectant for duck viral hepatitis live vaccine and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 Preparation of duck hepatitis virus liquid

[0026] Take the duck hepatitis virus type 1 A66 strain for production, dilute it 100-fold with sterile normal saline, and inoculate the well-developed 9-10d SPF chicken embryo allantoic cavity, 0.2ml per embryo, seal the hole, and continue incubating at 37°C. Discard the chicken embryos that died within 24 hours, take the embryos every 4 hours, and harvest the chicken embryos that died within 24 to 72 hours in time. After cooling for 24 hours at 4°C, take out the iodine tincture disinfection air chamber, remove the eggshell with sterile surgical scissors, and harvest Diseased chicken embryo chorioallantoic membrane, embryo fluid (allantoic fluid and amniotic fluid), embryo body are mixed and homogenized at high speed to make a suspension, which is used as duck hepatitis virus fluid. Place the duck hepatitis virus liquid in a sterilized container, add penicillin and 10 mg / ml streptomycin at a final concentration of 1000...

Embodiment 2

[0027] Example 2 Preparation of heat-resistant freeze-dried protective agent

[0028] The molecular formula of pullulan in this example is C 37 H 62 O 30 , The molecular weight is 10,000. Bovine serum albumin is abbreviated as BSA.

[0029] 1. Heat-resistant freeze-dried protective agent A

[0030] The heat-resistant freeze-dried protective agent A contains the following ingredients: sucrose 70g / L, pullulan 1g / L, gelatin 6g / L, reduced glutathione 10g / L, dipotassium hydrogen phosphate 10g / L, diphosphate Potassium hydrogen 5g / L, BSA10g / L, methionine 10g / L, and the solvent is pH7.2, 20mmol / L Tris-HCl buffer.

[0031] The preparation method of heat-resistant freeze-dried protective agent A includes the following steps:

[0032] (1) Prepare a solution containing 140g / L sucrose, 12g / L gelatin and 2g / L pullulan with pH7.2, 20mmol / L Tris-HCl buffer as solvent, and perform high-pressure steam quenching at 110°C Bacteria 20min;

[0033] (2) Use pH7.2, 20mmol / L Tris-HCl buffer as the solvent to ...

Embodiment 3

[0040] In order to better set the freeze-drying curve, heat-resistant freeze-drying protectant B (prepared in Example 2) was mixed with physiological saline and duck hepatitis virus solution (prepared in Example 1) at a volume ratio of 1:1, and the collapse temperature was measured. , Eutectic point and common melting point.

[0041] Table 1 Collapse temperature, eutectic point and eutectic point of heat-resistant freeze-dried protective agent B

[0042] Numbering

[0043]

[0044] It can be seen from Table 1 that the collapse temperature of the heat-resistant freeze-dried protective agent B is -34~-35℃; the eutectic point temperature is -23.8~-24℃; the eutectic point temperature is -24.6~-24.8℃, please refer to this The conditions set the best freeze-drying process.

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Abstract

The invention provides a heat-resisting freeze-drying protecting agent for duck viral hepatitis (DVH) live vaccine and a preparation method and application thereof, and belongs to the field of veterinary use biological product manufacturing. The heat-resisting freeze-drying protecting agent includes 70-100 g / L of saccharose, 1-10 g / L of Pullulan, 6-10 g / L of gelatin, 10-20 g / L of reduced glutathione, 10-20 g / L of dipotassium phosphate, 5-10 g / L of monopotassium phosphate, 10-30 g / L of bovine serum albumin (BSA) and 10-20 g / L of methionine. The invention further discloses the preparation method for protection of the heat-resisting freeze-drying protecting agent and the live vaccine including the heat-resisting freeze-drying protecting agent. The protecting agent can effectively protect antigenic activity in the freeze-drying and high-temperature storage processes and is safe to ducks, the low-temperature cold-chain problem of the vaccine in the storage and transportation processes is effectively solved, and the low-temperature cold-chain problem in remote area villages is solved particularly; meanwhile, the storage and transportation cost of the vaccine is lowered.

Description

Technical field [0001] The invention belongs to the technical field of veterinary biological product manufacturing, and in particular relates to a heat-resistant freeze-dried protective agent for duck virus hepatitis live vaccine, and a preparation method and application thereof. Background technique [0002] Duck viral hepatitis (duck viral hepatitis, DVH) is an acute, severe, and highly lethal hepatic hemorrhagic inflammation in ducklings caused by duck hepatitis virus (DHV; belonging to picornavirus). infectious disease. The clinical manifestations are neurological symptoms such as spasms, convulsions and reflexes, and liver swelling and bleeding are characteristic lesions. The mortality rate of ducklings after infection can reach 90%-95%, which has become one of the main diseases that harm the duck industry and will cause great economic losses in the duck industry. [0003] Duck hepatitis virus (DHV) has 4 independent serotypes, type 1, type 2, type 3 and new type. Fan Shuca...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K9/19A61K47/42A61K47/36A61K47/26A61K47/18A61K47/02A61K39/12A61P31/14
Inventor 赵艳红吕芳卢宇彭苗苗邓碧华张金秋候继波
Owner JIANGSU ACAD OF AGRI SCI
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