A method for packing Source packing material into a column and its application.

Abstract

This invention provides a method for packing Source packing material into a column and its application. The method includes the following steps: displacing the packing material with a solution, then loading the packing material into a container and connecting it to a chromatography column; removing air bubbles from the tubing and chromatography column, drawing the packing material into the chromatography column, allowing it to stand, and then pressurizing and lowering the column head to compress the column height to the target height, thus completing the packing; the pressure applied is below 3 bar and not less than 1 bar. The packing method provided by this invention can achieve the packing of Source packing material without high pressure, allows for a large column diameter, low packing pressure, and low packing cost, and the resulting chromatography column product meets the application requirements.

Description

Technical Field

[0001] This invention belongs to the field of biopharmaceuticals, specifically relating to a column packing method for source packing materials and its application, and more particularly to a column packing method for source packing materials that does not require high pressure and its application. Background Technology

[0002] Source 30Q is a single-pore anion exchange packing material with a particle size of 30μm. In production, it is typically packed under high pressure using stainless steel chromatography columns such as Fineline. The column diameter is generally less than 400mm, and the column body is made of stainless steel. It needs to withstand pressures of over 5 bar, and the packing process itself is also carried out at pressures above 5 bar. This places high demands on the packing equipment. Many manufacturers lack the necessary equipment to produce this type of column, and consumers are even less likely to perform on-site packing after purchasing the packing material. This contributes to the high cost of this type of column.

[0003] CN106552443B discloses a method for packing a reversed-phase chromatography column, comprising the following steps: 1) loading a slurry of reversed-phase packing material into the reversed-phase chromatography column; 2) removing air bubbles from the column head of the reversed-phase chromatography column; and 3) compacting the reversed-phase packing material inside the reversed-phase chromatography column. This method for packing a reversed-phase chromatography column simplifies the operation and improves the packing effect by removing air bubbles from the column head. It also reduces the need for repeated packing, saving operation time, manpower, and resources.

[0004] CN112121466B discloses a chromatography column apparatus and a chromatography column packing method. The chromatography column apparatus includes a chromatography column tube, a column head, and a base plate. The column head is sealed inside the chromatography column tube and can move up and down along the axial direction of the chromatography column tube. The column head has a first upper channel connecting the upper and lower sides of the column head. The base plate is sealed and covered at the bottom end of the chromatography column tube. The base plate has a first lower channel and a second lower channel connecting the upper and lower sides of the base plate respectively. The base plate also has a third lower channel connecting the lower side of the base plate and the second lower channel. The second lower channel is configured to have a first state where it is connected to the upper side of the base plate but disconnected from the third lower channel, and a second state where it is connected to the third lower channel but disconnected from the upper side of the base plate. A clearance structure is formed on the inner wall of the chromatography column tube, which is configured to connect the upper and lower sides of the column head when the column head moves to the clearance structure. This invention can achieve positive pressure column packing, overcoming various defects caused by negative pressure column packing in the prior art.

[0005] CN114217001A discloses an apparatus for packing a liquid chromatography column and its method of use. The apparatus includes a computer control unit, a liquid supply unit, a pressure unit, and a waste liquid recovery unit. The liquid supply unit includes a switching valve and at least one displacement tank. The pressure unit includes a pressure pump, a homogenizing tank, and a liquid chromatography column. The apparatus provided by this invention automates the entire column packing process by precisely controlling the displacement liquid flow rate, column pressure, column packing time, and the rates of pressure increase and decrease, greatly ensuring column packing quality and inter-column consistency. The apparatus also allows for free switching of the displacement liquid during the packing process, providing flexibility for column packing method development. Using the apparatus provided by this invention, the packed liquid chromatography column can exhibit good repeatability and high separation efficiency.

[0006] However, due to the small particle size of Source 30Q packing material, the aforementioned packing devices and methods cannot effectively pack Source 30Q packing material, resulting in column efficiency and symmetry factors that fail to meet requirements. Therefore, providing a packing method for Source 30Q packing material that has low packing requirements and good packing results has become an urgent problem to be solved. Summary of the Invention

[0007] To address the shortcomings of existing technologies, the present invention aims to provide a method for packing source packing materials and its applications, particularly a method for packing source packing materials without high pressure and its applications. The packing method provided by this invention achieves source packing without high pressure, allows for large column diameters, low packing pressure, and low packing costs, and yields chromatographic column products that meet application requirements.

[0008] To achieve this objective, the present invention adopts the following technical solution:

[0009] On one hand, the present invention provides a column packing method for Source packing material, the column packing method comprising the following steps:

[0010] The packing material is replaced with a solution, then loaded into a container and connected to the chromatography column. Air bubbles are removed from the tubing and the chromatography column, the packing material is drawn into the chromatography column, and allowed to stand. Then, pressure is applied and the column head is lowered to compress the column height to the target height, thus completing the column packing.

[0011] The pressurized pressure is less than 3 bar and not less than 1 bar, such as 1 bar, 1.5 bar, 2 bar, 2.5 bar or 2.9 bar, but not limited to the values ​​listed above. Other unlisted values ​​within the above range are also applicable.

[0012] The above method, by adopting a method of first allowing the column to stand and then pressurizing, can effectively complete the packing of the source packing material without the use of high-pressure column pressing. The chromatography column has a large diameter, low packing cost, and the column efficiency and symmetry factor of the obtained chromatography column can meet the application requirements.

[0013] Preferably, the Source filler includes Source 30S or Source 30Q.

[0014] Preferably, the solution is an aqueous solution of ethanol.

[0015] Preferably, the volume fraction of the ethanol-water solution is 15-25%, such as 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25%, but not limited to the values ​​listed above. Other unlisted values ​​within the above range are also applicable.

[0016] Preferably, the settling time is at least 24 hours, such as 24 hours, 25 hours, 26 hours, 27 hours, 28 hours, 29 hours or 30 hours, but is not limited to the values ​​listed above. Other unlisted values ​​within the above range are also applicable.

[0017] Preferably, the chromatography column is an plexiglass column.

[0018] The column loading method of the present invention eliminates the need for high-pressure loading, thus eliminating the need for stainless steel columns and significantly reducing costs.

[0019] Preferably, the diameter of the chromatography column is not less than 800 mm, such as 800 mm, 850 mm, 900 mm, 950 mm or 1000 mm, but not limited to the values ​​listed above. Other unlisted values ​​within the above range are also applicable.

[0020] On the other hand, the present invention also provides the application of the column packing method described above in the preparation of Source chromatographic columns.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] This invention provides a method for packing source packing material into a column. By adopting a method of first allowing it to stand and then pressurizing it, the packing of source packing material into a column can be effectively completed without the use of high-pressure column pressing. The resulting column has a large diameter, low packing cost, and the column efficiency and symmetry factor of the obtained column can meet the requirements of use. Detailed Implementation

[0023] To further illustrate the technical means and effects of the present invention, the following describes the technical solution of the present invention in conjunction with preferred embodiments of the present invention. However, the present invention is not limited to the scope of the embodiments.

[0024] The following example uses a column loading workstation for column loading.

[0025] Example 1

[0026] This embodiment provides a column packing method for Source packing material, the specific steps of which are as follows:

[0027] The Source 30Q packing was replaced with a 20% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0028] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0029] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0030] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0031] Let the homogenate in the chromatography column stand for 24 hours.

[0032] After 24 hours, the column head position of the chromatography column is lowered by 2.5 bar to compress the column height as much as possible to the target column height range (15-18 cm), thus obtaining the target chromatography column.

[0033] Example 2

[0034] This embodiment provides a column packing method for Source packing material, the specific steps of which are as follows:

[0035] The Source 30Q packing was replaced with a 20% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0036] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0037] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0038] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0039] Let the homogenate in the chromatography column stand for 28 hours.

[0040] After 28 hours, the column head position of the chromatography column is lowered by 2 bar to compress the column height as much as possible to the target column height range (15-18 cm), thus obtaining the target chromatography column.

[0041] Example 3

[0042] This embodiment provides a column packing method for Source packing material, the specific steps of which are as follows:

[0043] The Source 30Q packing was replaced with a 10% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0044] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0045] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0046] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0047] Let the homogenate in the chromatography column stand for 28 hours.

[0048] After 28 hours, the column head position of the chromatography column was lowered to 2.9 bar, and the column height was compressed as much as possible to the target column height range (15-18 cm), thus obtaining the target chromatography column.

[0049] Example 4

[0050] This embodiment provides a column packing method for Source packing material, the specific steps of which are as follows:

[0051] The Source 30Q packing was replaced with a 20% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0052] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0053] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0054] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0055] Let the homogenate in the chromatography column stand for 12 hours.

[0056] After 12 hours, the column head position of the chromatography column is lowered by 2.5 bar to compress the column height as much as possible to the target column height range (15-18 cm), thus obtaining the target chromatography column.

[0057] Example 5

[0058] This embodiment provides a column packing method for Source packing material, the specific steps of which are as follows:

[0059] The Source 30Q packing was replaced with a 20% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0060] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0061] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0062] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0063] Let the homogenate in the chromatography column stand for 24 hours.

[0064] After 24 hours, the column head position of the chromatography column is lowered by 1 bar to compress the column height as much as possible to the target column height range (15-18 cm), thus obtaining the target chromatography column.

[0065] Comparative Example 1

[0066] This comparative example provides a column packing method for Source packing material, the specific steps of which are as follows:

[0067] The Source 30Q packing was replaced with a 20% ethanol aqueous solution, then transferred to a packing mixing tank, and samples were taken to test the packing concentration.

[0068] An 800mm acrylic chromatography column is connected to a packing mixing tank via a pressure-resistant pipe. Then, the air bubbles in the chromatography column and pipeline are removed through the chromatography system.

[0069] Enter the column packing information in the column packing workstation, open the bottom valve of the packing homogenizer and the packing valve of the chromatography column, and draw the well-mixed packing into the chromatography column.

[0070] After the gel aspiration is complete, close the packing valve of the chromatography column and recover the packing material in the pipeline.

[0071] Then, lower the column head position by 2.5 bar to compress the column height as much as possible to the target column height range (15-18 cm), thus obtaining the target column.

[0072] The column height, column efficiency, and symmetry factor of the chromatography columns provided in Examples 1-5 and Comparative Example 1 were tested and compared with the recommended parameters (column height requirement 15-18 cm, column efficiency requirement ≥2000, symmetry factor requirement 0.8-2.0).

[0073] The results are as follows:

[0074]

[0075]

[0076] The data above shows that the method provided by the present invention can achieve effective column packing under low pressure, saving costs. Comparing Examples 1-5 and Comparative Example 1, it can be found that the present invention can achieve effective column packing by adopting a static method and controlling relevant parameters. All relevant parameters can meet the requirements, and the symmetry factor can be significantly reduced, reducing tailing. Other methods are difficult to achieve the target column packing effect.

[0077] The applicant declares that this invention illustrates the column packing method of the Source packing material and its application through the above embodiments, but this invention is not limited to the above embodiments, that is, it does not mean that this invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvements to this invention, equivalent substitutions of the raw materials of the product of this invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of this invention.

[0078] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

[0079] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable way without contradiction. In order to avoid unnecessary repetition, the present invention will not describe the various possible combinations separately.

Claims

1. A method of packing a column with Source packing, characterized by, The column packing method comprises the following steps: The packing material is replaced by a solution, then the packing material is packed into a tank and connected with a chromatographic column; the pipeline and the chromatographic column are drained of air bubbles, the packing material is sucked into the chromatographic column, and then the chromatographic column is left to stand for at least 24 hours, and then the chromatographic column is compressed to the target column height under pressure, so that the column packing is completed. The pressure of the compression is lower than 2.9 bar and not lower than 1.5 bar.

2. The column packing method according to claim 1, characterized by, The Source packing material comprises Source 30S or Source 30Q.

3. The column packing method according to claim 1, characterized by, The solution is an ethanol aqueous solution.

4. The column packing method according to claim 3, characterized by, The volume fraction of the ethanol aqueous solution is 15-25%.

5. The column packing method according to claim 1, wherein, The chromatographic column is made of organic glass.

6. The column packing method according to claim 1, wherein, The diameter of the chromatographic column is not less than 800 mm.

7. Application of the column packing method according to any one of claims 1-6 in the preparation of a Source chromatographic column.

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