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Purificatino of Insulin Glargine

  • Categories:Industry News
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  • Time of issue:2014-01-03 00:00
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Purificatino of Insulin Glargine

Information

nsulin glargine is a type of human insulin analogues which has low solubility in neutral PH solution. It dissolves completely in acid (PH 4) injecta. After injecting into subcutaneous tissue, micro-sediments created by neutralization of acid solution could release small amount of insulin glargine constantly, therefore, it produce expectable, long-lasting, stable, and blood concentration with no peak value.

Chemical name: 21A-Gly-30B a-L-Arg-30Bb-L-Arg-human insulin.

Molecular formula: C267H404N72O78S6.

Molecular weight: 6063.

Purification method:

Zymotic fluid (prosomal)

Remove cells by centrifugal force

Supernatant fluid

Phosphoric acid PH 3.8

CQK30S

Loading amount < 50g/L

Eluate by ammonium acetate

Crystallize

Add 0.5% of Phenol into eluant, 4% of AnCl2 and NaOH adjust PH to 4.08, stir slowly for 15min, transfer to 2-8℃environment, stay overnight, centrifugal collecting and precipitating

Enzymatic conversion

GH C8 reverse phase

GH C8 (10µm, 120Å) column, 10% acetonitrile-0.25M acetic acid equilibrium liquid, enzymatic conversion liquid and water with 1:5 portion dilution then put the sample, gradient elution by acetonitrile, get product with 94.5% purity; dilute the product from previous step, then use GH C8 (the same as before), acetronitrile-0.2M sodium acetate (PH 5.0) equilibrium liquid, get 98.5% purity product; dilute the product from previous step by water with 1:3 portion, use GH C8 (the same as before), ethanol-0.05M acetic acid equilibrium liquid, gradient elution by ethoanol, get 99% pure insulin glargine finished product

Crystallize

Put the sample from previous step into citric acid, ZnCl2 and adjust PH to get crystallized sediments, wash the sediment with cold water

Insulin glargine finished product (purity >99%)

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