A preparing process for instant precipitate-free freeze-dried human fibrinogen

A technology for the preparation of human fibrinogen, applied in the field of biopharmaceuticals, can solve the problems of harsh production process, not too long storage time, protein damage, etc., to optimize the freeze-drying process conditions, shorten the total freeze-drying time, reduce protein the effect of damage

Pending Publication Date: 2017-06-06
上海洲跃生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The fibril preparation process using cold glue as raw material, because the raw material contains very little Fg, after a series of operation steps, layer by layer loss, the final yield is very limited, lacking the value of commercial production; while cold glue and components I co-precipitation is the fibroblast production technique of raw material, also there is following problem, at first, contain 8% ethanol in the precipitation, for the stability of the human coagulation factor VIII in the precipitation, be a very unfavorable factor, the storage of precipitation It must be very careful, the storage time should not be too long, and special attention must be paid to the denaturation and damage of ethanol to human blood coagulation factor VIII when feeding materials, so the existence of ethanol in raw materials has undoubtedly proposed harsh requirements for the production process of human blood coagulation factor VIII; secondly, due to The presence of a large amount of human coagulation factor VIII in the precipitate also brings hidden dangers to the production of fibrils, because fibrils are easily activated and lead to production failure. The reasons for fibril ...

Method used

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  • A preparing process for instant precipitate-free freeze-dried human fibrinogen
  • A preparing process for instant precipitate-free freeze-dried human fibrinogen

Examples

Experimental program
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Effect test

Embodiment 1

[0034] In the first step, according to the dilution ratio of 1:20, take 2kg of fresh component I and precipitate it, dilute and chop it immediately, then dissolve it in 40kg buffer 1, and stir it evenly for 3 hours; the formula of buffer 1: sodium citrate·2H 2 O640g, TRIS 108g, lysine hydrochloride 160g, sucrose 600g, glycine 600g, sodium chloride 400g, buffer 1 pH value 6.95, temperature 20°C;

[0035] In the second step, use a 30SP depth filter element in series with a 1.0μm filter element to filter the first suspension; the filtration pressure is controlled at about 0.5bar, and the filter element is washed with buffer 1 of the formula described in the first step, and the filtrate 50.1 is collected kg.

[0036] The third step is to add 2.5 kg of pre-prepared S / D mother liquor to the filtrate to make the concentration of Tween-80 in the solution to 1% and the concentration of TNBP to 0.3%. Stir slowly at 24-26°C and keep it warm for 6 hours ;

[0037] The fourth step is to cool the...

Embodiment 2

[0046] In the first step, take 10kg of fresh component I precipitate and dilute and chop it immediately. According to the dilution ratio of 1:20, put it into 200kg of buffer 1 to dissolve and stir evenly for 1.5 hours; buffer 1 formula: sodium citrate·2H 2 O 3.2kg, TRIS 540g, lysine hydrochloride 800g, sucrose 3kg, glycine 2.0kg, sodium chloride 2.0kg, buffer 1 pH value 6.95, temperature 25℃;

[0047] The second step is to filter the above suspension with a 30SP depth filter element in series with a 1.0 μm (10 inch) filter element; the filtration pressure is controlled at about 0.5 bar, the filter element is washed with about 50 kg of buffer 1 of the above formula, and 251 kg of filtrate is collected.

[0048] The third step is to add 12.5kg of pre-prepared S / D mother liquor to the filtrate to make the concentration of Tween-80 in the solution to 1% and the concentration of TNBP to 0.3%. Stir slowly at 24-26℃ and keep it warm for 6 hours. ;;

[0049] The fourth step is to cool the so...

Embodiment 3

[0058] In the first step, take 10kg of fresh component I and precipitate it, dilute and chop it immediately, according to the dilution ratio of 1:10, and then dissolve it in 100kg of buffer 1, and stir it evenly for 3 hours; the formula of buffer 1: sodium citrate·2H 2 O1.6kg, TRIS 270g, lysine hydrochloride 400g, sucrose 1.5kg, glycine 3.0kg, sodium chloride 1.0kg, buffer 1 pH value is 6.80, temperature 30℃;

[0059] The second step is to filter the above suspension with a 30SP depth filter element in series with a 1.0μm filter element; the filtration pressure is controlled at about 0.5bar, and the filter element is washed with about 50kg of buffer 1 formulated in the first step, and 152kg of filtrate is collected .

[0060] The third step is to add 7.5 kg of pre-prepared S / D mother liquor to the filtrate to make the concentration of Tween-80 in the solution to 1% and the concentration of TNBP to 0.3%. Stir slowly at 24-26°C and keep it warm for 6 hours ;;

[0061] The fourth step ...

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Abstract

The invention relates to the field of preparing processes for human fibrinogen, and particularly relates to a preparing process for instant precipitate-free freeze-dried human fibrinogen. The process includes five steps, namely the method includes a first step of component I precipitate dissolving and filtering; a second step of S/D virus inactivation; a third step of twice low-temperature ethanol precipitation rectification; a fourth step of freeze-drying; and a fifth step of virus inactivation in a dry heat manner. Through the improved fibrinogen production process, the fibrinogen which is instant, precipitate-free, opalescence-free and free of protein particles can be prepared. The process can prepare the human fibrinogen used for traditional intravenous injection, and can prepare high-concentration human fibrinogen compatible in human fibrous protein adhesives.

Description

Technical field [0001] The invention belongs to the field of biopharmaceuticals, and relates to a preparation process of blood products, in particular, a preparation process of freeze-dried human fibrinogen with no precipitation. Background technique [0002] Human fibrinogen (fibrinogen or Fg for short) is a macromolecular glycoprotein containing 2964 amino acids. It is a protein with coagulation function synthesized by the liver. It has the largest content of thirteen coagulation factors in the human body. A coagulation factor, also known as coagulation factor I. Human fibrinogen is a blood product that is frequently used clinically, and it is also an indispensable emergency medicine for hemostasis on the battlefield. However, fibrinogen has been in a clinically in short supply. Especially in recent years, human fibrin adhesive, a surgical blood product with highly effective hemostasis function, has become more and more popular among doctors and patients. As one of the compati...

Claims

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Application Information

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IPC IPC(8): C07K1/30C07K1/14
Inventor 李春洲
Owner 上海洲跃生物科技有限公司
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