Vaccine adjuvant as well as vaccine composition and vaccine preparation both containing vaccine adjuvant

Abstract

The invention discloses a vaccine adjuvant as well as a vaccine composition and a vaccine preparation both containing the vaccine adjuvant. The vaccine adjuvant comprises chitooligosaccharides which is prepared from the following method: chitosan is subjected to hydrolysis using a biological enzyme method, degradation using a chemical method, or schizolysis using a physical method to produce chitosan oligosaccharide with the polymerization degree of 2-10; alternatively, chitin is directly subjected to degradation using an enzymic method or a chemical method to obtain chitooligosaccharides with the polymerization degree of 2-10; the acetylation degree of chitooligosaccharides is higher than 90%, and the molecular weight range is 400-2000 Da. The vaccine adjuvant provided by the invention can be used as the adjuvant of attenuated vaccine, inactivated vaccine, DNA vaccine, protein vaccine and polypeptide vaccine.

Description

technical field

[0001] The invention belongs to the technical field of medicine, and relates to a vaccine adjuvant, a vaccine composition and a vaccine preparation containing the vaccine adjuvant. Background technique

[0002] Adjuvant is a kind of substance that can enhance the immune effect of antigen when applied in advance of antigen or at the same time with antigen. Its main function is to enhance the specific immune response of the body's immune system to antigen. Since the British doctor Jenner vaccinated against smallpox more than 200 years ago and achieved success in human trials, humans can prevent and control certain infectious diseases through vaccination. In 1925, French immunologist and veterinary scientist Gaston Ramon discovered that adding tapioca starch, agar, pyogenic bacteria and other substances unrelated to antigens to diphtheria and tetanus vaccines could specifically enhance the body’s resistance to diphtheria and tetanus toxins force. In the subseq...

Examples

Embodiment 1

[0035] Example 1: Preparation and physicochemical properties of chitooligosaccharides

[0036] 1. Preparation of chitooligosaccharides:

[0037] Add deionized water into the batching tank, dissolve chitosan in 1% acetic acid solution, stir and heat up to 38±2.0°C. Add 5% (to the substrate) enzyme preparation, stir, and keep the temperature at 38°C. After the viscosity of the enzymolysis solution drops to 60%, start ultrafiltration (cut molecular weight at 6000). The part smaller than the cut-off molecular weight permeates the membrane, and the part larger than the cut-off molecular weight returns to the reactor to continue to degrade. The ultrafiltration permeate is passed through a nanomembrane filter (cutting molecular weight 300), and the permeated part is water, acetic acid and small molecule sugar (the permeated part can be recycled for preparing chitosan solution), and the non-permeated part is Concentrate into chitosan oligosaccharide solution and spray dry to obtai...

Embodiment 2

[0045] Embodiment 2: Chitosan oligosaccharide is to the adjuvant activity determination of porcine PRRS inactivated vaccine (1)

[0046] The chitosan oligosaccharide in Example 1 was used as a vaccine adjuvant, and the inactivated porcine PRRSV vaccine (PRRSV, 1.6 mg / ml) was used as an antigen, and the two were used in combination to immunize mice by intramuscular injection, and the antibody titer was determined. The specific method is as follows:

[0047] Experimental animals: BALB / c mice, 6-8 weeks old, 6 mice / group, female, purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd., license number SCXK (Beijing) 2012-0001.

[0048] Drug concentration: by chitosan oligosaccharide (COSNAC) prepared in the above-mentioned embodiment 1: 4mg / ml; Porcine PRRSV inactivated vaccine (PRRSV): 1.6mg / ml; ISA206 (Seppic company, oil adjuvant)

[0049] Control solvent: physiological saline (Saline).

[0050] Drug concentration after mixing: chitosan oligosaccharide...

Embodiment 3

[0066] Embodiment 3: The adjuvant activity assay (two) of chitosan oligosaccharide to porcine PRRS inactivated vaccine

[0067] The chitosan oligosaccharide in Example 1 was used as a vaccine adjuvant, and the porcine blue ear disease inactivated vaccine (PRRSV, 1.6 mg / ml) was used as an antigen, and the two were used in conjunction with intramuscular injection to immunize mice, and the antibody subtype and subtype were determined. kind. Concrete experiment is with embodiment 2.

[0068] Intramuscular injection of animals, 2 immunizations within 4 weeks of the first immunization, 3 immunizations within 4 weeks of the 2nd immunization, and determination of mouse serum antibody subtypes and subtypes after 2 weeks after the 3rd immunization.

[0069] ELISA method to detect serum-specific antibody subtypes and subtypes The experimental method is as follows:

[0070] The N protein antigen of PRRSV was diluted to 5 μg / ml with antigen coating solution, added to a 96-well plate (C...

technical field

[0001] The invention belongs to the technical field of medicine, and relates to a vaccine adjuvant, a vaccine composition and a vaccine preparation containing the vaccine adjuvant. Background technique

[0002] Adjuvant is a kind of substance that can enhance the immune effect of antigen when applied in advance of antigen or at the same time with antigen. Its main function is to enhance the specific immune response of the body's immune system to antigen. Since the British doctor Jenner vaccinated against smallpox more than 200 years ago and achieved success in human trials, humans can prevent and control certain infectious diseases through vaccination. In 1925, French immunologist and veterinary scientist Gaston Ramon discovered that adding tapioca starch, agar, pyogenic bacteria and other substances unrelated to antigens to diphtheria and tetanus vaccines could specifically enhance the body’s resistance to diphtheria and tetanus toxins force. In the subseq...

Examples

Embodiment 1

[0035] Example 1: Preparation and physicochemical properties of chitooligosaccharides

[0036] 1. Preparation of chitooligosaccharides:

[0037] Add deionized water into the batching tank, dissolve chitosan in 1% acetic acid solution, stir and heat up to 38±2.0°C. Add 5% (to the substrate) enzyme preparation, stir, and keep the temperature at 38°C. After the viscosity of the enzymolysis solution drops to 60%, start ultrafiltration (cut molecular weight at 6000). The part smaller than the cut-off molecular weight permeates the membrane, and the part larger than the cut-off molecular weight returns to the reactor to continue to degrade. The ultrafiltration permeate is passed through a nanomembrane filter (cutting molecular weight 300), and the permeated part is water, acetic acid and small molecule sugar (the permeated part can be recycled for preparing chitosan solution), and the non-permeated part is Concentrate into chitosan oligosaccharide solution and spray dry to obtai...

Embodiment 2

[0045] Embodiment 2: Chitosan oligosaccharide is to the adjuvant activity determination of porcine PRRS inactivated vaccine (1)

[0046] The chitosan oligosaccharide in Example 1 was used as a vaccine adjuvant, and the inactivated porcine PRRSV vaccine (PRRSV, 1.6 mg / ml) was used as an antigen, and the two were used in combination to immunize mice by intramuscular injection, and the antibody titer was determined. The specific method is as follows:

[0047] Experimental animals: BALB / c mice, 6-8 weeks old, 6 mice / group, female, purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd., license number SCXK (Beijing) 2012-0001.

[0048] Drug concentration: by chitosan oligosaccharide (COSNAC) prepared in the above-mentioned embodiment 1: 4mg / ml; Porcine PRRSV inactivated vaccine (PRRSV): 1.6mg / ml; ISA206 (Seppic company, oil adjuvant)

[0049] Control solvent: physiological saline (Saline).

[0050] Drug concentration after mixing: chitosan oligosaccharide...

Embodiment 3

[0066] Embodiment 3: The adjuvant activity assay (two) of chitosan oligosaccharide to porcine PRRS inactivated vaccine

[0067] The chitosan oligosaccharide in Example 1 was used as a vaccine adjuvant, and the porcine blue ear disease inactivated vaccine (PRRSV, 1.6 mg / ml) was used as an antigen, and the two were used in conjunction with intramuscular injection to immunize mice, and the antibody subtype and subtype were determined. kind. Concrete experiment is with embodiment 2.

[0068] Intramuscular injection of animals, 2 immunizations within 4 weeks of the first immunization, 3 immunizations within 4 weeks of the 2nd immunization, and determination of mouse serum antibody subtypes and subtypes after 2 weeks after the 3rd immunization.

[0069] ELISA method to detect serum-specific antibody subtypes and subtypes The experimental method is as follows:

[0070] The N protein antigen of PRRSV was diluted to 5 μg / ml with antigen coating solution, added to a 96-well plate (C...

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