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Curdlan sulfate/6-O-quaternized chitosan nanoparticles and application thereof in mucosal vaccines

A technology of quaternized chitosan and mucosal vaccines, which is applied to medical preparations containing active ingredients, antibody medical ingredients, drug combinations, etc., can solve the problems of difficult passage of macromolecular protein antigens and weak enhancement effects, and achieve Improve the level of cellular immunity and humoral immunity, stimulate proliferation and differentiation, and promote the effect of activation

Inactive Publication Date: 2018-01-26
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, previous experiments found that when CS alone was used as a nasal mucosal vaccine adjuvant, it had a weak effect on enhancing the humoral immunity of mice. Protein antigens are difficult to pass through the dense mucous layer

Method used

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  • Curdlan sulfate/6-O-quaternized chitosan nanoparticles and application thereof in mucosal vaccines
  • Curdlan sulfate/6-O-quaternized chitosan nanoparticles and application thereof in mucosal vaccines
  • Curdlan sulfate/6-O-quaternized chitosan nanoparticles and application thereof in mucosal vaccines

Examples

Experimental program
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preparation example Construction

[0051] Concrete, the preparation method of described nanoparticle, comprises the following steps:

[0052] 1) Put sulfated codran polysaccharide (CS) in deionized water, dissolve, filter and dilute to obtain solution A;

[0053] 2) Place 6-O-quaternized chitosan (O-HTCC) in deionized water, dissolve, filter, and dilute to obtain solution B;

[0054] 3) Slowly add solution B to solution A under stirring, and continue to stir for a period of time to obtain sulfated codranan / 6-O-quaternized chitosan nanoparticle solution;

[0055] Preferably, the filtration in step 1) and step 2) is treated with a filter membrane with a diameter of 0.22 μm;

[0056] Preferably, the molecular weight of sulfated candella polysaccharide is 103kD, the molecular weight of 6-O-quaternized chitosan is 100kD~200kD (most preferably 100kD), and the degree of quaternization is not less than 30%;

[0057] Preferably, the concentration ratio of the solution A to the solution B is 1:4-6 (most preferably 1:5)...

Embodiment 1

[0077] The preparation of embodiment 1 OVA / CS / O-HTCC nanoparticles

[0078] The preparation process is as figure 1 As shown, 10 mg of sulfated candlan was dissolved in 10 mL of deionized water to prepare a 1 mg / mL sulfated candlan mother liquor. After filtering with a filter membrane with a diameter of 0.22 μm, take 100 μL of the mother liquor of sulfated ketanan and dilute it with 900 μL of deionized water to obtain a 100 μg / mL sulfated ketanan solution; take 10 mg of 6-O-quaternized chitosan and dissolve it in In 10 mL of deionized water, a 1 mg / mL 6-O-quaternized chitosan mother solution was prepared. After filtering with a filter membrane with a diameter of 0.22 μm, 500 μL of the 1 mg / mL 6-O-quaternized chitosan mother solution was diluted with 500 μL of deionized water to obtain a 500 μg / mL 6-O-quaternized chitosan solution.

[0079]Take 500 mL of 100 μg / mL sulfated kurtlan solution and add 500 mL of 500 μg / mL 6-O-quaternized chitosan prepared in the previous stage drop...

Embodiment 2

[0083] Example 2 Detection of uptake capacity of FITC-labeled OVA / CS / O-HTCC nanoparticles

[0084] To investigate the ability of OVA / CS / O-HTCC nanoparticles to be taken up by epithelial cells, the Calu-3 cell line with nasal epithelial cell characteristics was selected as the experimental model, and the specific implementation steps were as follows:

[0085] 1 mL of 1 mg / mL FITC solution was added dropwise to 1 mL of OVA solution (10 mg / mL), stirred overnight at 4°C, and 7 mg of ammonium chloride was added to terminate the reaction. The reaction solution was dialyzed in the dark for 48 hours using a dialysis bag with a molecular weight cut-off of 1 kD, and then freeze-dried to obtain FITC-OVA.

[0086] Take Calu-3 cells in the logarithmic growth phase, and use 1×10 5 Inoculate into a culture dish at a density of one / dish, culture in a cell incubator for 4 days, discard the medium, add FITC-OVA, a mixture of FITC-OVA and CS, a mixture of FITC-OVA and O-HTCC or FITC -OVA / CS / O-...

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Abstract

Nanoparticles are prepared from curdlan sulfate and 6-O-quaternized chitosan by an ion gelation method for the first time; the method is simple and convenient, green, safe and pollution-free, and through controlling experimental steps and parameters, the prepared curdlan sulfate / 6-O-quaternized chitosan nanoparticles have uniform size and good stability. Evaluation of the membrane penetrating ability, the immune regulation ability and the mucosal adjuvant effect of the curdlan sulfate / 6-O-quaternized chitosan nanoparticles shows that the positively charged characteristic of the nanoparticles make the nanoparticles more easily act on effective parts through epithelium mucosae, so as to play a role of immune enhancers and help an antigen reach the effective parts to play relatively good immune adjuvant effect; the nanoparticles have great industrial application value.

Description

technical field [0001] The invention belongs to the field of pharmacy, and in particular relates to a sulfated codran polysaccharide / 6-O-quaternized chitosan nanoparticle and its application in mucosal vaccines. Background technique [0002] Vaccination is the most effective measure to prevent and control the spread of infectious diseases. However, the adjuvants approved by FDA and CFDA for human vaccines are still only traditional aluminum salts. Aluminum adjuvant has the advantages of simple composition and low cost, but it also has the disadvantages of easy deposition at the injection site, and can only induce Th2 type immune response in the body and basically has no effect on Th1 type immune response. Therefore, the development of new human adjuvants has increasingly become a research hotspot. [0003] As a vaccine adjuvant, polysaccharides have the advantages of low toxicity, safety, and the ability to induce immune responses in the body. A series of polysaccharides e...

Claims

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Application Information

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IPC IPC(8): A61K39/39A61K39/00A61P11/02A61P11/00A61P31/00
Inventor 王凤山张舒路璐李平利程艳娜
Owner SHANDONG UNIV
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