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Preparation method for intravenous injection human immune globulin (PH4)

A human immunoglobulin, intravenous injection technology, applied in the direction of serum immunoglobulin, can solve the problems of low product purity, protein denaturation damage, protein co-precipitation, etc., to reduce the probability of protein denaturation damage, The effect of less process and short process

Inactive Publication Date: 2016-01-20
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

[0009] Since most low-temperature ethanol methods require multiple ethanol precipitations, it will inevitably cause denaturation and damage to some proteins, and it is also easy to cause protein co-precipitation, so the purity of the product is not very high, and often contains more impurity proteins such as IgA, IgM and Polymers, etc.; although some are also combined with column chromatography for purification, two-step column chromatography is used, which makes the process complicated and the yield decreases

Method used

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  • Preparation method for intravenous injection human immune globulin (PH4)
  • Preparation method for intravenous injection human immune globulin (PH4)

Examples

Experimental program
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Effect test

Embodiment 1

[0036] 1. Suspension of component II+III precipitate: Weigh 1 kg of component II+III precipitate and put it into 8 kg of dissolution buffer (20mmol / L acetic acid-sodium acetate buffer, pH4.10-4.20, 0~1℃) , stirred evenly for 3 hours to fully suspend the precipitate;

[0037] 2. Caprylic acid extraction: Mix the above suspension with 9kg extract (20mmol / L sodium acetate, 40mmol / L caprylic acid, PH5.90-6.00, 5~6℃), and stir for 5 hours to fully extract the target protein from the precipitate , to obtain a homogeneous suspension;

[0038] 3. Pressure filtration: Press filter the above suspension at 2~3°C, and then wash the precipitate, collect a total of 18.6kg of filtrate, add 0.19kg of glycine, stir well to dissolve;

[0039]4. Anion exchange column chromatography: put the CaptoQ column on the column at a linear velocity of 90-100cm / h. The column is pre-balanced with an equilibrium buffer (20mmol / L acetic acid-sodium acetate solution, pH5.20-5.30), and the collected flow Thro...

Embodiment 2

[0045] 1， Component II+III precipitation suspension: Weigh 1kg of component II+III precipitation, put it into 10kg dissolution buffer (10mmol / L acetic acid-sodium acetate buffer, pH4.40-4.50, 4~5℃), and stir evenly 2 hours to fully suspend the precipitate;

[0046] 2 , octanoic acid extraction: mix the above suspension with 12kg of extract (30mmol / L acetic acid-sodium acetate buffer, 50mmol / L octanoic acid, pH5.50-5.60, 9~10°C), stir for 3 hours to make the target protein Fully extract from the precipitate to obtain a homogeneous suspension;

[0047] 3 , Press filtration: press filter the above suspension at 5-7°C, and wash the precipitate, collect a total of 24.7kg of filtrate, add 0.50kg of glycine, stir fully to dissolve;

[0048] 4. Anion exchange column chromatography: use 60-70cm / h upper column flow rate on DEAESepharoseFF column, pre-equilibrate with equilibrium buffer (10mmol / L acetic acid-sodium acetate solution, pH5.50-5.60), collect the flow-through solution; a...

Embodiment 3

[0052] 1 , Component II+III precipitate suspension: Weigh 1kg of component II+III precipitate, put into 12kg dissolution buffer (30mmol / L acetic acid-sodium acetate buffer, pH4.70-4.80, 2~3℃), evenly Stir for 1 hour to fully suspend the precipitate;

[0053] 2 , Octanoic acid extraction: Mix the above suspension with 10.4kg of extract (10mmol / L acetic acid-sodium acetate buffer, 60mmol / L octanoic acid, pH5.20-5.30, 5~6°C), stir for 4 hours, and make the target The protein is fully extracted from the precipitate to obtain a homogeneous suspension;

[0054] 3 , Pressure filtration: Press filter the above suspension at 3~4°C, and then wash the precipitate, collect a total of 24.1kg of filtrate, add 0.73kg of glycine, stir well to dissolve;

[0055] 4 , Anion exchange column chromatography: put the QSepharoseFF column on the column at a linear velocity of 50-60cm / h, the column is pre-balanced with an equilibrium buffer, the equilibrium solution is 15mmol / L acetic acid-sodium ...

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Abstract

The invention discloses a method for preparing intravenous injection human immune globulin (PH4) from plasma fractions II+III. The method comprises the following steps of 1 fraction II+II precipitate suspending; 2 octanoic acid solution extracting; 3 filter pressing; 4 anion exchange column chromatography performing; 5 ultra-filtering, concentrating and regulating; 6 sterilizing and filtering; 7 low-PH incubating; 8 virus removing with nano films and filtering; 9 sterilizing, filtering and subpackaging. According to the method, a low-PH suspended buffer solution is selected and used to enable fraction II+II precipitates to be fully suspended, an octanoic acid extracting solution is mixed with the suspension liquid to enable IgG to be selectively dissolved out, a crude IgG solution is obtained, and purification on IVIG is completed through a step of anionic column chromatography; the method is short in technological process, few in procedure and low in energy consumption, the product is high in yield and stable in quality, the key technical indexes such as IgA, ACA, PKA and polymers are all superior to those of a traditional technology, and the yield can reach up to about 2200 bottles per ton of plasma.

Description

technical field [0001] The invention belongs to the field of biopharmaceuticals and relates to a preparation process of blood products, in particular to a method for preparing intravenous human immunoglobulin (PH4) from components II+III. Background technique [0002] Human immunoglobulin (immunoglobulin) is a protein with antibody activity, mainly present in plasma, but also in other body fluids, tissues and some secretions. Immunoglobulins can be divided into five categories: IgG, IgA, IgM, IgD, and IgE. The main component of human serum immunoglobulins is IgG, accounting for 70-75% of the total immunoglobulins. [0003] Human serum immunoglobulin IgG is the most persistent and important antibody in the primary immune response and exists only in monomeric form. Most antibacterial, antiviral and antiviral antibodies belong to IgG, which play a leading role in anti-infection, can promote the phagocytosis of mononuclear macrophages, neutralize the toxicity of bacterial toxin...

Claims

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Application Information

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IPC IPC(8): C07K16/06C07K1/36C07K1/34C07K1/18
Inventor 李春洲
Owner 上海洲跃生物科技有限公司