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Thrombin preparation method

A technology of thrombin and porcine thrombin, applied in biochemical equipment and methods, enzymes, peptidases, etc., can solve problems such as high impurity content in products, restrictions on clinical application of thrombin products, and potential safety hazards

Active Publication Date: 2020-02-25
长春雷允上药业有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Further, no matter whether the thrombin product is prepared from animal blood or human blood, the existing preparation methods will have the following problems: (1) the titer of the thrombin product is unstable; (2) the concentration of impurities and miscellaneous proteins in the product (3) There is a risk of introducing various viruses, which will limit the clinical application of thrombin products and cause safety hazards
The patent application does not disclose the types of reagents and nanomembranes used for virus inactivation, and the product obtained by the method has a relatively high impurity content, and the potency of the product needs to be further improved

Method used

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Effect test

Embodiment 1

[0070] The preparation of embodiment 1 porcine thrombin freeze-dried product

[0071] 1) Collect pig blood and prepare anticoagulant pig plasma, inhale the anticoagulant pig plasma into the high-level storage tank, flow into the tubular centrifuge through the lower discharge valve of the tank, and turn on the tubular centrifuge for centrifugation. Specifically, add anticoagulant to fresh pig blood according to the ratio of anticoagulant to fresh pig blood volume ratio of 1:9, stir gently, let it stand at 10°C, and within 5 to 8 hours Centrifuge at 19000r / min, the centrifugation temperature is 10°C, and the centrifugation time is 15 minutes, and the upper plasma is collected. Described anticoagulant is the trisodium citrate aqueous solution of 38 grams / liter. And step by step through the 70 mesh, 100 mesh, 200 mesh filter to remove impurities;

[0072] 2) Add the pre-treated sorbent to the porcine plasma that has been filtered according to the mass ratio of 1000:2 between the...

Embodiment 2

[0091] Example 2: Effects of different virus inactivation and removal methods on the potency of porcine thrombin freeze-dried products

[0092] Among the various preparation methods in this example, the virus inactivation and removal methods are respectively S / D method combined with membrane filtration method (for specific methods, see ZL201410591661.6 Chinese invention patent) and S / D method combined with dry heat method (for specific methods Referring to Example 1 of the present application, the remaining conditions used in the preparation method of the porcine thrombin freeze-dried product are the same as in Example 1. Specifically, the dry heat method controls the moisture content of the freeze-dried product within 2% by weight. The dry heat method first raises the temperature to 105°C, and then continues to control the temperature at 100±1°C for 45 minutes of dry heat treatment. The dry heat method uses water Spray type. Three batches of products were produced by various...

Embodiment 3

[0096] Example 3: Effects of different dry heat methods on the virus inactivation of porcine thrombin freeze-dried products

[0097] In the preparation method of this embodiment, the following dry heat methods are respectively used for virus inactivation:

[0098] (1) Water spray dry heat method: first raise the temperature to 104°C, and then continue to control the temperature at 100±1°C for 30 minutes of dry heat treatment. The dry heat method adopts the water spray dry heat method to produce three Batch, batch number is B. All the other preparation conditions of the porcine thrombin freeze-dried product are the same as in Example 1.

[0099] (2) The dry heat method is adopted in a dry heat box, and the temperature is directly controlled at 100±1°C for 30 minutes of dry heat treatment, and three batches are produced, and the batch number is C. All the other preparation conditions of the porcine thrombin freeze-dried product are the same as in Example 1.

[0100] Table 2 C...

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Abstract

The invention provides a thrombin preparation method which includes the steps: freeze-drying extracting solution containing thrombin and / or increasing the temperature of a freeze-dried product containing thrombin to reach 101-107 DEG C; performing water spraying dry-heat treatment at the temperature of 100+ / 3 DEG C. The water content of the freeze-dried product containing thrombin by weight is nothigher than 3% and preferably not higher than 2.5% and more preferably not higher than 2%. According to the preparation method, the impurity content of thrombin products is further reduced, the valence of thrombin products is remarkably improved, viruses are effectively removed, and the preparation method is quite suitable for large-scale industrial production of the thrombin products.

Description

technical field [0001] The invention belongs to the field of biochemical pharmacy, relates to a preparation method of thrombin, in particular to a preparation method of thrombin capable of increasing potency, removing viruses and reducing impurities, and suitable for large-scale industrial production. Background technique [0002] Thrombin is a serine protease widely present in human and animal blood, and it is the main effector protease in the blood coagulation cascade, showing procoagulant and anticoagulant properties. When circulating coagulation factors come into contact with tissue factor in exposed extravascular tissue, thrombin accumulates on the tissue. Thrombin plays a central role in the initiation and development of thrombotic diseases by activating platelets to catalyze the conversion of fibrinogen into fibrin and promoting the stability of blood clots. Because thrombin can directly act on fibrinogen in the blood, prompting it to transform into fibrin, accelerat...

Claims

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Application Information

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IPC IPC(8): C12N9/74
CPCC12N9/6429C12Y304/21005
Inventor 孙丽
Owner 长春雷允上药业有限公司
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