A method for preparing cytokine and bone protein microspheres by thin film evaporation

A technology of cytokines and morphogenic proteins, which is applied in the field of biomedicine, can solve the problems of low bioavailability, high protein loss, and difficult measurement of microglobulin encapsulation rate, and achieve simple and stable preparation methods, uniform volume, and improved The effect of bioavailability

Inactive Publication Date: 2018-02-27
SHENZHEN POLYTECHNIC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the disadvantage of its application to bone tissue is that the protein is easily diluted and fused with tissue, making its bioavailability very low
A suitable carrier is needed to release BMP slowly. Chinese patent CN101816634B discloses a microsphere prepared by spraying technology, but there is still a lot of protein loss in the package due to spraying, and the protein encapsulation rate of the microspheres produced by spraying is not easy. Determination (there is no measurement about encapsulation rate and release rate in its embodiment), the controllable release degree of microspheres is unstable, etc.

Method used

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  • A method for preparing cytokine and bone protein microspheres by thin film evaporation
  • A method for preparing cytokine and bone protein microspheres by thin film evaporation
  • A method for preparing cytokine and bone protein microspheres by thin film evaporation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] [Example 1] Optimization of membrane composition

[0043] Orthogonal test design without considering the interaction was used to determine the optimal formulation of lipid microsphere membrane materials, and the best composition of three components including lecithin, cholesterol and octadecylamine for injection in lipid microsphere membrane materials was selected. proportional formula. Its preparation method is to adopt the optimized thin film evaporation method, and use an ultra-high pressure homogenizer for uniform crushing, and then conduct particle size analysis, encapsulation efficiency, stability and other indicators related to lipid microspheres, and set the parameters according to each individual indicator. There are 5 grades from poor to excellent, according to the grades (1-5, 5 points), and finally the best formula is obtained according to the analysis of variance.

[0044] (1) The selection results of the best membrane materials are shown in Table 1

[00...

Embodiment 2

[0050] [embodiment 2] the selection of solvent

[0051] The effects of four different solvents on the prepared lipid microspheres were compared, and the results are shown in Table 2:

[0052] Table 2: Comparison of the effects of four different solvents on the prepared lipid microspheres (0 month)

[0053]

[0054] Note: The smaller the oxidation index value, the smaller the degree of oxidation of phospholipids

[0055] ★★ The smaller the diameter value, the particle size tends to be normally distributed, and the lipid microsphere particles are more uniform

[0056] See Table 3 for the influence of four different solvents on the particle size stability of the prepared lipid microspheres:

[0057] Table 3: Effects of four different solvents on the particle size stability of the prepared lipid microspheres

[0058]

[0059] The solvent of the membrane material is an important factor affecting the morphology and encapsulation efficiency of lipid microspheres during the ...

Embodiment 3

[0061] [Example 3] Preparation of Lipid Microspheres

[0062] The steps of preparing lipid microspheres by thin film evaporation method are as follows: ①Film formation: membrane material ingredients: lecithin for injection, cholesterol and octadecylamine (or vitamin E) are weighed according to the ingredient ratio of 3-7:1-3:1, Then dissolve it in dichloromethane or trichloromethane with a final concentration of 0.5-1.5%, put the liquid in an eggplant-shaped bottle of a rotary evaporator, use a water bath at 60°C as the evaporation temperature, rotate to evaporate the solvent, and use nitrogen gas in the final stage. Dry the solvent to make the liposomes form a film in an eggplant-shaped bottle. ②Hydration: Add about 50ml of the aqueous phase solution of cytokine recombinant human IFNβ1a (final concentration: 100ng / ml) and bone morphogenetic protein BMP-2 (final concentration: 80ng / ml) (the concentration of the membrane material in the solution is 1% ), add a few small glass ...

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Abstract

The invention discloses a preparation method of a cell-carried factor and a bone protein microsphere. The preparation method comprises the steps of: proportioning of raw materials of a film material: proportioning lecithin for injection, cholesterol, octadecyl amine or vitamin E; film formation by using microspheres: mixing the proportioned raw materials of the film material in an organic solvent, adding in an eggplant-shaped reaction bottle of a rotary evaporator, and forming a thin film under the conditions of constant temperature, vacuum degree and rotational speed; preparation of protein solution for wrapping: adding a cell factor protein and a bone morphogenetic protein in a buffer solution to obtain a protein solution; hydration: adding the protein solution to hydrate the thin film; high-pressure homogenizing: carrying out high-pressure homogenizing on rough microspheres which are coated with the protein to obtain a liposome solution; freeze-drying: adding a freeze-drying protective agent, precooling the liposome solution in a refrigerator, and then carrying out vacuum drying to prepare a freeze-dried product of the microspheres; and then hydratation: adding water for injection in the freeze-dried product of the cell-carried microspheres, and carrying out shock hydratation to obtain a milk white solution.

Description

technical field [0001] The invention belongs to the field of biomedicine, and more precisely, the invention relates to a method for preparing lipid microspheres loaded with cytokines and bone morphogenetic proteins by thin film evaporation. technical background [0002] Drug-loaded lipid microspheres are developed with the development of biomaterials and material technology equipment, combined with pharmacy and clinical medicine, and have been developed and applied in the past two decades. Lipid microspheres can embed drug powder or solution in particles with a diameter of nanometers. This particle can have a cell-like structure, enter the human body and be mainly swallowed by the reticuloendothelial system to activate the body's autoimmune function and change The distribution of the encapsulated drug in the body makes the drug mainly accumulate in tissues and organs such as liver, spleen, lung and bone marrow, thereby improving the therapeutic index of the drug, reducing th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K9/16A61K9/127A61K38/19A61K38/18A61P19/08A61L27/54A61L27/50
Inventor 张英王妍张登央张丽君张水华
Owner SHENZHEN POLYTECHNIC
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