Preparation method for Sincalide

A mass percent, ammonia water technology, applied in the field of polypeptide drug preparation, can solve the problems of difficult removal of reagents, unsatisfactory yield, unstable product performance, etc., to reduce pressure, simplify operation steps, and ensure stability.

Active Publication Date: 2018-05-22
GL BIOCHEM SHANGHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It mainly solves the technical problems that the reagents in the existing enzymatic preparation are difficult to remove and the product performance in the chemical method is unstable and the yield is not ideal

Method used

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  • Preparation method for Sincalide
  • Preparation method for Sincalide
  • Preparation method for Sincalide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] 1. Desalting of crude samples

[0027] (1) Fully dissolve 5 g of the crude peptide obtained by solid-phase synthesis with 500 ml of ammonia water with a concentration of 1% by mass, and set aside;

[0028] (2) First use a glass column with a diameter of 2cm. Fill the upper layer with 300g of polystyrene matrix, and the lower layer with 100g of C18 silica gel matrix, soak the entire matrix with methanol, and set aside;

[0029] (3) Connect the glass column to the high-pressure pump, and adjust the flow rate of the high-pressure pump to 5ml / min;

[0030] (4) The mass percentage concentration of the proportioned acetic acid aqueous solution is 0.15%, and the volume is 500ml;

[0031] (5) First extract all the methanol solution from the matrix, pour the crude product dissolved in 1% ammonia solution into the glass column, adjust the flow rate of the high-pressure pump to 5ml / min until the crude product liquid penetrates the flat top matrix, Add 500ml of acetic acid water...

Embodiment 2

[0039] 1. Desalting of crude samples

[0040] (1) Fully dissolve 5 g of the crude peptide obtained by solid-phase synthesis in 500 ml of ammonia water with a concentration of 1.2% by mass, and set aside;

[0041] (2) First use a glass column with a diameter of 2cm. Fill the upper layer with 350g of polystyrene substrate, and the lower layer with 150g of C18 silica gel substrate, soak the entire substrate with methanol, and set aside;

[0042] (3) Connect the glass column to the high-pressure pump, and adjust the flow rate of the high-pressure pump to 6.5ml / min;

[0043] (4) The mass percent concentration of the proportioned acetic acid aqueous solution is 0.18%, and the volume is 500ml;

[0044] (5) First extract all the methanol solution from the matrix, pour the crude product dissolved in 1% ammonia solution into the glass column, adjust the flow rate of the high-pressure pump to 6.5ml / min until the crude product liquid penetrates the flat top matrix , add 500ml acetic ac...

Embodiment 3

[0052] 1. Desalting of crude samples

[0053] (1) Fully dissolve 10 g of the crude peptide obtained by solid-phase synthesis with 800 ml of ammonia water with a concentration of 1.7% by mass, and set aside;

[0054] (2) First use a stainless steel column with a diameter of 3cm. Fill the upper layer with 450g of polystyrene matrix, and the lower layer with 200g of C18 silica gel matrix, soak the entire matrix with methanol, and set aside;

[0055] (3) Connect the glass column to the high-pressure pump, and adjust the flow rate of the high-pressure pump to 10.5ml / min;

[0056] (4) The mass percent concentration of the proportioned acetic acid aqueous solution is 0.2%, and the volume is 800ml;

[0057] (5) First extract all the methanol solution from the matrix, pour the crude product dissolved in 1.5% ammonia solution into the glass column, adjust the flow rate of the high-pressure pump to 10.5ml / min until the crude product liquid penetrates the flat top matrix , add 800ml of...

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Abstract

The invention relates to a preparation method for Sincalide. The invention is mainly to solve the technical problems of difficult removal of a reagent in a conventional enzyme method and unstable product performances and unsatisfactory product yield in a chemical method for preparation of the Sincalide. The method comprises the following steps: 1) allowing crude peptide obtained by solid-phase synthesis to be completely dissolved in ammonia water, and carrying out ultrafiltration through an ultrafiltration column; 2) allowing the ultrafiltration column to be connected with a high-pressure single pump, selecting an acetic acid aqueous solution as an eluting agent so as to obtain a TFA-salt-removed Sincalide crude product intermediate liquid; 3) adjusting a pH value of the TFA-salt-removed Sincalide crude product intermediate liquid to 7.25 with an NH4HCO3 solution; 4) purifying the TFA-salt-removed Sincalide crude product intermediate liquid by using a reverse phase silica gel column ofoctadecylsilane chemically bonded silica gel as a solid phase and using an A phase of an acetic acid aqueous solution with a concentration of 0.2% and a B phase of an acetonitrile solution as liquidphases so as to obtain a highly-pure Sincalide liquid with an acetate form; and 5) subjecting the obtained highly-pure Sincalide liquid with the acetate form to concentration and freeze-drying so as to finally obtain solid Sincalide with a content of more than 98.5%.

Description

technical field [0001] The invention relates to the field of preparation methods of polypeptide drugs, in particular to a preparation method of sincalide synthesized in solid phase. Background technique [0002] Sincalide is a synthetic cholecystolin octapeptide, its structural sequence is: H-Asp-Tyr(SO 3 H)-Met-Gly-Trp-Met-Asp-PHe-NH 2 , Mainly used for promoting cholecystokinin contraction, cholecystokinin diagnostic reagent, has all the biological activities of cholecystokinin. Studies have shown that sincalide can reduce pulmonary interstitial edema and leukocyte infiltration, inhibit the production of pro-inflammatory cytokines, improve lung tissue structure damage, improve hemodynamic disorders, delay the decline in mean arterial pressure and pulmonary artery pressure, and improve liver and kidney. Microcirculatory blood flow, restore heart rate, reduce pulmonary hypertension, improve tidal volume, etc. Clinically, it can be used to treat chronic pancreatitis, endot...

Claims

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Application Information

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IPC IPC(8): C07K14/595C07K1/20C07K1/34C07K1/04C07K1/36
CPCC07K14/595
Inventor 闫凤徐红岩
Owner GL BIOCHEM SHANGHAI
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