What Is Recombinant Insulin?

This product is a protein consisting of 51 amino acid residues produced by recombinant DNA technology. The host cell is E. coli. Calculated based on dry products, each 1mg containing recombinant human insulin shall not be less than 27.5 units. Residual host DNA and host cell protein in recombinant human insulin are potential specific impurities related to the production process and must be strictly controlled during the production process, and their limits should meet relevant regulations.

Recombinant human insulin

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This product is

Phonetic name: Chongzu Ren Yidaosu

English name: Recombinant Human Insulin

Page number: 2000 edition two -497

C257H383N65O77S6 5807.69

This product is white or off-white crystalline powder.

The product is almost insoluble in water, ethanol and ether, and in dilute hydrochloric acid and dilute sodium hydroxide solution.

(1) In the chromatogram recorded under the titer determination item, the retention time of the main peak of the test substance should be the same as that of the peak of the recombinant human insulin reference substance.

Stay the same.

(2) Take an appropriate amount of this product, use 0.1% trifluoroacetic acid solution to make a solution containing 10mg per 1ml, take 20l, and add 0.2mol / L trimethylolamino

20 l of methane-hydrochloric acid buffer (pH 7.3), 20 l of 0.1% V8 enzyme solution and 140 l of water, mix well, place in a 37 ° C water bath for 2 hours, add 3 l of phosphoric acid as a test solution; take another recombinant human insulin control Appropriate amount of product, prepared in the same way, as a reference solution. According to the method under the titer measurement method, a gradient of 0.2 mol / L sulfate buffer (pH 2.3) -acetonitrile (90:10) as mobile phase A and acetonitrile-water (50:50) as mobile phase B was performed. Elution.

Time Mobile Phase A Mobile Phase B

0 points 90% 10%

5 minutes 80% 20%

45 minutes 40% 60%

50 minutes 40% 60%

Take 25l each of the reference solution and the test solution, and inject them into the liquid chromatograph respectively to record the chromatogram. The peptide map of the test product should be consistent with the peptide map of the reference.

Relevant substances: take an appropriate amount of this product and use 0.01mol / L hydrochloric acid solution to make a solution containing 3.5mg per 1ml as the test solution. According to the method under the titer determination method, a gradient of 0.2 mol / L sulfate buffer (pH 2.3) -acetonitrile (82:18) as mobile phase A and acetonitrile-water (50:50) as mobile phase B was performed. Elution.

Time mobile phase A and work phase B

0 points 78% 22%

36 points 78% 22%

61 points 33% 67%

67 points 33% 67%

The flow comparison example is adjusted so that the retention time of the main peak of recombinant human insulin is about 25 minutes, and the system suitability test should meet the requirements under the titer determination item. Take 20l of the test solution and inject it into the liquid chromatograph, record the chromatogram, and calculate based on the area normalization method. The total related substances shall not exceed 3.0%. Take an appropriate amount of the polymer protein, and make a solution containing 1mg per 1ml with a dissolving solution (300mg of tetrasodium ethylenediamine tetraacetate, 250ml of water to dissolve, 3ml of 0.1mol / L hydrochloric acid solution, and 500ml of water). As a test solution; take an appropriate amount, and dilute with diluted solution (600 mg of ethylenediamine tetraacetic acid tetrasodium, 900 ml of water to dissolve, adjust the pH to 8.5 with 1 mol / L hydrochloric acid solution, and add water to 1000 ml). A solution containing 0.01 mg was used as a control solution. According to

According to high performance liquid chromatography (Appendix VD).

Chromatographic conditions and system suitability test Use octadecylsilane-bonded silica gel as filler (5-10 m); 0.2mol / L sulfate buffer solution (take 28.4g of anhydrous sodium sulfate, dissolve in water, add 2.7ml of phosphoric acid 800ml of water

Hypoglycemic drugs.

Shaded, sealed, and stored below -15 ° C.

(1) Recombinant human insulin injection (2) Protamine recombinant human insulin injection