Method for stablizing protein medicine and its application in microball preparation

A protein drug and stabilization technology, which is applied in the preparation of microspheres, improves the stability of protein drugs, and can solve problems such as hidden dangers of formulation stability and decreased stability.

Inactive Publication Date: 2005-02-09
FUDAN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After contacting with organic solvents, a part of the soluble protein is transformed into a dimer, and the stability of this existing form is greatly lower than that of the protein in the natural conformation, which lays a hidden danger for the stability of the preparation

Method used

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  • Method for stablizing protein medicine and its application in microball preparation
  • Method for stablizing protein medicine and its application in microball preparation
  • Method for stablizing protein medicine and its application in microball preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] growth hormone stability test : Prepare a phosphate buffer solution (PBS, 10mM, pH8.0) containing various stabilizers and 50mg / ml recombinant human growth hormone (rhGH), mix the solution with 0.2ml of dichloromethane, stir vigorously or ultrasonically for 1min, Immediately transfer the water phase and repeatedly extract the oil phase with PBS (pH8.0) for 8 to 10 times, combine the extracts and dilute to 10ml. The solution was centrifuged at 4° C. and 3000 rpm for 30 min, and the supernatant was injected into a high-performance liquid chromatography system for determination.

[0026] The concentration of rhGH in the samples was determined by size exclusion chromatography (SEC). Chromatography system: Agilent1100series; Chromatographic column: TSK-Gel G2000SWXL (equipped with TSK-Guardcolumn SWXL pre-column); Column temperature: 25°C; Mobile phase: 3% isopropanol / 97% phosphate buffer solution (0.063M, pH7.0) ; Flow rate: 0.6ml / min; Detection wavelength: 214nm; Injecti...

Embodiment 2

[0047] Effect of Poloxamer Concentration on Growth Hormone Stability : In the PBS (10mM, pH8.0) solution that contains 50mg / ml rhGH, add the poloxamer 407 of different concentrations, this solution is mixed with dichloromethane as internal water phase, all the other operations and assay methods are with embodiment 1 . Table 2 is the effect of non-ionic surfactants on the stability of rhGH.

[0048] Table 2

[0049] High speed stirring Ultrasonic mixing

[0050] concentration

[0051] Stabilizer Soluble Protein Monomer Soluble Protein Monomer

[0052] mg / ml

[0053] % (a) % % %

[0054] control (b) 64.1 30.1 64.2 23.6

[0055] 2 57.9 34.2 63.1 33.6

[0056] Poloxamer 407 20 87.2 73.0 56.6 48.6

[0057] 50 97.5 90.0 97.0 89.1

[0058] 20 74.9 59.3 86.1 63.7

[0059] Tween 20 50 41.3 38.1 - (c) -

[006...

Embodiment 3

[0063] Preparation of Recombinant Human Growth Hormone Microspheres 1 : Dissolve 0.75 grhGH in 8 ml of PBS solution (10 mM, pH 8.0) containing 50 mg / ml poloxamer 407 and 45 mg / ml sucrose. This solution was gradually added dropwise to 20ml of dichloromethane solution containing 5g PLGA (75:25, MW 15kD). ), stirred at 1800rpm for 1min, poured the solution into 2L of distilled water and continued to stir at a lower speed for 4h to completely volatilize the organic solvent to obtain PLGA microspheres loaded with somatotropin. The obtained microspheres were collected by suction filtration on a suction filter, washed three times with distilled water and 0.2% poloxamer 188 solution, and dried in vacuum for 24 hours to obtain microsphere powder.

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Abstract

The present invention belongs to the field of medicine preparation, and relates to one method of raising the stability of protein medicine and its application in preparing microballoon. Non-ionic surfactant polyoxypropylene ppolyoxyethylene copolymer is added into water phase to raise the stability of protein medicine during preparing microballoon. The method of the present invention can raise obviously the tolerance of protein medicine to organic solvent, avoid the aggregation and deactivation of protein medicine during preparing microballoon, and further raise the stability and validity of protein medicine preparation and make it possible to prepare even more long-acting protein medicine preparations.

Description

technical field [0001] The invention belongs to the field of pharmaceutical preparations, and relates to a method for improving the stability of protein drugs and the application of the method in the preparation of microspheres. Background technique [0002] In recent years, with the rapid development of life-related disciplines, a large number of protein and peptide drugs have emerged. Compared with traditional small-molecule organic drugs, protein and peptide drugs have poor physical and chemical stability, and are prone to aggregation, oxidation, and deamidation reactions, resulting in loss of biological activity. Among them, the impact of aggregation on the activity of protein drugs is more prominent. [0003] Protein drugs all have a certain three-dimensional spatial conformation, which is an important condition to ensure that they bind to receptors and exert therapeutic effects. However, protein structures are easily disrupted during formulation preparation, leading t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K9/14A61K38/00A61K47/32
Inventor 魏刚陆丽芳陆伟跃
Owner FUDAN UNIV
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