Patent Application
20230173408
The present invention relates to a chromatography column, a method for priming a chromatography column and to a method for unpacking a chromatography column.
Chromatography columns provide a means for separating, purifying and isolating chemical and biological compounds. The size and type of column used in these operations typically depends upon the scale of the process in question, small glass or plastic walled columns typically being used for research purposes, while larger metal columns are employed for industrial processes. For instance, chromatography columns may be used in manufacturing processes to purify process liquids and separate substances of interest from such liquids; typical examples include large-scale preparative purification of fine chemicals and pharmaceuticals, together with biological products.
Whilst the present invention relates to both plastic walled and larger metal walled chromatography columns, it is particularly useful for manufacturing and industrial-scale chromatography columns. Industrial-scale chromatography columns typically comprise a hollow, axially vertical tubular housing including a liquid inlet at the upper end and through which the buffer and substances to be separated are dispensed to the media bed located within the cavity of the tube, and a liquid collecting system at the lower end for collecting substances and buffer. The particulate chromatographic media or bed through which the buffer fluid and/or substances to be separated and purified percolates is located between the liquid inlet and collecting system.
An adapter assembly is typically affixed to the upper end of the tubular housing and a base assembly to the lower end where it is bolted to bottom flanges. Each of these assemblies typically comprises a strong backing plate and a distributor plate which further supports a bed support; a bed support is a layer of mesh, screen, filter, sinter or other fluid-permeable media-retaining material which permits process liquid flow into and out of the chromatography bed space or cavity while retaining the bed of particulate medium. To provide adjustability and control of the bed height and bed compression, the adapter assembly is typically made in the form of a piston or sliding adapter in the column tube interior. After the column is charged with bed media, typically through a valve or nozzle, the adapter may be forced toward the bottom of the tube to compress or pressurize the media bed. Generally, the base assembly is a fixed structure which is bolted against a bottom flange of the column tube but, in some instances, may also be in the form of a movably slidable piston or adapter.
The backing plate of the base assembly generally acts as a support for the column, being itself supported on legs or some other stand arrangement, which allows clearance for outlet pipework projecting beneath the base assembly.
Liquid chromatography is a technique for separating the individual compounds that exist in a sample. In employing the technique, the sample is dissolved and carried in a liquid, called a mobile phase. The mobile phase carrying the sample is forced to migrate through a bed of particulate media, sometimes called a stationary phase. Different compounds will have differing rates of migration through the media, which effects the separation of the components in the sample. The bed of particulate media is formed under gravity, washing with a mobile phase, and/or by compression by an adapter or piston. Before a column can be used to separate compounds by liquid chromatography and before the column is filled with chromatography resin it is necessary to remove air from the bed space which has become trapped in the column otherwise the air will interfere with the separation process. The air may have been originally present in the bed space itself or have moved into the bed space from associated tubing, pumps or column components such as a bed support. Problems are often encountered in trying to remove the air from the column prior to conducting liquid chromatography. The trapped air may create air pockets in the stationary phase where chemical interaction between the sample and the media cannot take place. These air pockets thus negatively affect the separation of the different components in the sample.
U.S. Pat. No. 7,820,042 discloses a chromatography column which is specifically designed for allowing air which has been trapped within the bed space to be removed. An elongated groove is provided in an inner wall of the tubular side wall of the housing of the chromatography column.
A problem in a chromatography column according to the one described in U.S. Pat. No. 7,820,042 is that the whole column needs to be filled up with pure water during the priming process in order to get rid of the possibly trapped air. This process is both time consuming and requires a large amount of pure water.
An object of the present invention is to improve and facilitate priming of a chromatography column.
A further object of the invention is to improve the possibilities to remove all the slurry from the column during unpacking.
These objects are addressed by a chromatography column, a method for priming a chromatography column and a method for unpacking a chromatography column according to the independent claims.
According to one aspect of the invention a chromatography column is provided comprising a tubular side wall having an inner wall, an adaptor assembly and a base assembly, wherein an enclosed bed space is defined between said adaptor assembly, said base assembly and said inner wall of said tubular side wall and wherein said adaptor assembly is axially movable inside said tubular side wall in relation to said base assembly, wherein the inner wall comprises at least one groove which is positioned in a lower half of the tubular side wall of the chromatography column, via which groove air and other fluid can pass the adaptor assembly when the adaptor assembly is positioned in a priming position, in which priming position said adaptor assembly intersects with at least a part of the at least one groove.
According to another aspect of the invention a method for priming a chromatography column according to the invention is provided. Said method comprises the steps of lowering the adaptor assembly to a priming position within the tubular side wall such that the adaptor assembly intersects with at least a part of the at least one groove of the inner wall such that air and other fluid can pass the adaptor assembly via said at least one groove; providing a priming liquid to fill up the enclosed bed space with priming liquid; removing air by providing more priming liquid to the enclosed bed space such that air can pass the adaptor assembly via the at least one groove and/or by lowering the adaptor assembly axially within the tubular side wall such that air can be pressed out via the at least one groove.
According to another aspect of the invention a method for unpacking a chromatography column according to the invention is provided, wherein said method comprises the steps of: resuspending a packed bed of chromatography media provided in said chromatography column by providing a fluid to said bed space; removing resuspended chromatography media via a chromatography media outlet of the chromatography column; lowering the adaptor assembly to a position within the tubular side wall such that the adaptor assembly intersects/fluidically couples with at least a part of the at least one groove; flowing a fluid through at least one priming fluid passage which is provided passing through the adaptor assembly, whereby said fluid will be flowed out through a first end opening of each of the at least one priming fluid passage and via said groove into the bed space for resuspending any remaining chromatography media in the bed space; removing resuspended chromatography media via the chromatography media outlet.
Hereby a chromatography column is provided in which trapped air can be removed by lowering the adaptor to a priming position in a lower half of the column, where the groove is positioned in the inner wall. When the adaptor is positioned in the priming position which is in line with the groove, i.e. the adaptor is aligned to intersect with at least a part of the groove, a priming liquid is provided into a comparatively small compartment provided between the adaptor and the base assembly. Priming liquid is provided until any possible air has passed the adaptor assembly via the at least one groove. Alternatively, when the bed space between the adaptor assembly and the base assembly has been filled with priming liquid, the adaptor can be pushed a small distance downwards thereby forcing some of the priming liquid together with any possible air to pass the adaptor assembly via the at least one groove. Thanks to the comparatively low position of the at least one groove in the chromatography column wall, close to the base assembly, a very small amount of priming liquid is needed compared to previous methods where the whole column had to be filled up for performing the priming. This both saves time and costs. A further advantage with a groove positioned in a lower half of the column is that the groove can be used for flowing a fluid from outlets which are provided around the circumference of the adaptor assembly. Such a flow of a fluid can then pass the adaptor assembly via the at least one groove and hereby flow into the bed space. This can be used during unpacking of the chromatography column. Any possibly remaining chromatography material can be flushed out by this flow. This is much more useful when the groove is provided close to the base assembly compared to prior art where the groove was provided in an upper part of the column.
In one embodiment of the invention the at least one groove is provided within a distance from the base assembly which is less than 20 cm or less than 10 cm.
In one embodiment of the invention the at least one groove is a continuous groove around the circumference of the inner wall.
In one embodiment of the invention there are a number of separate grooves distributed around the circumference of the inner wall.
In one embodiment of the invention the at least one groove is provided as a continuous spiral groove provided around the circumference of the inner wall, starting less than 20 cm from the base assembly and ending less than 15 cm from the base assembly.
In some embodiments of the invention the adaptor assembly comprises at least one priming fluid passage which is passing through the adaptor assembly, wherein each priming fluid passage has a first end opening which is provided at a position around the circumference of the adaptor assembly, whereby if there are more than one priming fluid passage provided, the first end openings for each of the priming fluid passages are distributed around the circumference of the adaptor assembly, whereby said at least one priming fluid passage is configured for transferring a fluid between an outside of the chromatography column and a circumference of the adaptor assembly.
In some embodiments of the invention said at least one first end opening of each of said at least one priming fluid passage is provided between two scarpers of the adaptor assembly which scrapers are sealing against the inner wall of the tubular side wall of the chromatography column.
In some embodiments the method for priming the chromatography column comprises a further step d) which is performed after step c), wherein said step d) comprises raising the adaptor assembly axially within the tubular side wall away from the base assembly such that the adaptor assembly is removed from the at least one groove whereby no more air or other fluid can pass the adaptor assembly.
In some embodiments of the method for unpacking the chromatography column the step of flowing a fluid through the at least one priming fluid passage further comprises simultaneously moving the adaptor assembly axially within the tubular side wall such that the adaptor assembly passes over a distance where the groove is provided, wherein the groove is provided as a spiral groove, whereby the flow of fluid out through the at least one first end openings is transferred out via the spiral groove into the bed space from different directions depending on the position of the adaptor assembly.
The adapter assembly 120 is a cylindrical disc type structure that operates by moving axially inside the centre of the tubular side wall 110 in the direction of arrow A. The disc circumference is such that it fits almost flush with the face of inner wall 112. The bed space 130 is designed to contain a bed of particulate media suspended in a liquid carrier. The adapter assembly 120 is used to settle the bed of particulate media, which is initially in the form of a slurry, by pushing down on it; the adapter assembly 120 may subsequently be used to compress and pack the particulate media to achieve a packed bed suitable for chromatographic separation. The volume of the media bed decreases as the adapter 120 is pushed down the column 100 (i.e. in the direction of arrow A in
In order to fully confine the carrier liquid and the particulate media it must be ensured that the solution cannot pass through the adapter assembly 120. A sealing assembly 160 provided at the circumference of the adapter assembly 120 is used to form a dynamic seal between the adapter assembly 120 and the inner wall 112. One example of a sealing assembly 160 is better seen in
With reference to both
The adaptor assembly 120 comprises one or more priming fluid passages 171. One such priming fluid passage 171 can be seen in
According to the invention the inner wall 112 comprises at least one groove 180 via which groove 180 air and other fluid can pass the adaptor assembly 120 when the adaptor assembly is positioned in a priming position, wherein said priming position is a position where the adaptor assembly is provided in line with the groove 180 for allowing a passage of fluid. The at least one groove 180 is formed in the inner wall 112 such that when the lower sealing arrangement 160b is aligned to intersect with the at least one groove 180 there is formed at least one fluid passage between a volume of the chromatography column 100 provided below the adaptor assembly 120, i.e. the enclosed bed space 130, and the first end openings 172 of the fluid passages 171 in the adaptor assembly 120. The at least one groove is provided within a distance from the base assembly 125 such that the amount of priming liquid needed for priming the column can be limited. By providing the at least one groove 180 in a lower half of the column only a part of the column needs to be filled up with priming liquid during priming. The at least one groove 180 can be positioned in a lower half of the tubular side wall 110 of the chromatography column 100 where lower refers to a direction toward the base assembly 125, i.e. that the at least one groove 180 is provided closer to the base assembly 125 than to a top part 110a of the tubular side wall 110. In one embodiment the at least one groove 180 is positioned in a lower third or in a lower quarter of the tubular side wall. In some embodiments the at least one groove is positioned close to the base assembly 125 however giving place for a lower part of the adaptor to pass the at least one groove 180. In some embodiments of the invention the at least one groove 180 is positioned within a distance from the base assembly 125 which is less than 20 cm (e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 cm), and preferably within a few centimetres (e.g. about 3-5, about 3-10 cm). The at least one groove 180 may be substantially circumferential. Said at least one groove 180 may have an axial dimension thereto. For example, said at least one grove 180 may extend in the tubular side wall over an axial distance range. For example, such an axial distance range may be from about 2, 3, 4, or 5 to about 20 cm, about 2, 3, 4, or 5 to about 10 cm, about 2, 3, 4, or 5 to about 15 cm, about 10 to about 20 cm, about 10 to about 15 cm, about 15 to 20 about cm, etc. of the distance from the base assembly 125 within the chromatography column 100. The distance between the at least one groove 180 and the base assembly 125 can be measured between a bottom part 181 of the at least one groove 180 and a bottom bed support 121b of the base assembly 125 and the distance can be less than 20 cm or in another embodiment of the invention less than 10 cm or even less than 5 cm. The lower the position of the at least one groove 180 in the column is the less amount of priming liquid is needed for the priming and the less time will be required for the priming. Furthermore, during unpacking of the column a media bed can be effectively flushed out by providing a flow via the at least one groove and a low position of the groove may be more effective. When the adaptor assembly 120 is positioned in a low position, i.e. in the priming position which is in line with the at least one groove 180, the fluid flow provided via the fluid passages 171 can be effectively used for flushing out remaining parts of a media bed. The fluid flow will hereby be provided from a short distance and a high flow can be provided which will be effective for flushing out the media. With previous systems there have been problems getting rid of all chromatography media during unpacking. Furthermore, when more than one fluid passages 171 are provided in the adaptor assembly 120 the fluid flow can be increased in order to unpack and flush out the media bed even more effectively. However, for unpacking of some types of chromatography media the position of the at least one groove may suitably by a bit higher for being most effective. For some embodiments 10 cm or even 15 cm or 20 cm may be a suitable position of the groove. In
A method for priming a chromatography column as described above is also provided according to the invention. The method steps are shown in the flow chart of
S1: Lowering the adaptor assembly 120 to a priming position within the tubular side wall 110, which priming position is a position where the adaptor assembly 120 is in line with the at least one groove 180 of the inner wall 112 such that air and other fluid can pass the adaptor assembly 120 via said at least one groove 180.
S2: Providing a priming liquid to fill up the enclosed bed space 130 with priming liquid. The priming liquid could be water, for example purified water, or a buffer. The priming liquid is provided through the inlet/outlet 225 in the bottom of the column or through a bottom mobile phase (not seen in these figures).
S3: Removing air by providing more priming liquid to the enclosed bed space 130 such that air can pass the adaptor assembly 120 via the at least one groove 180 and further through the one or more priming fluid passages 171 and/or by lowering the adaptor assembly 120 axially within the tubular side wall 110 such that air can be pressed out via the at least one groove 180 and further through the one or more priming fluid passages 171.
Possibly the method further comprises the step:
S4: Raising the adaptor assembly 120 axially within the tubular side wall 110 away from the base assembly 125 such that the adaptor assembly 120 is removed from the at least one groove 180 whereby no more air or other fluid can pass the adaptor assembly 120. Hereby a sealed bed space 130 free from air is achieved. During this step the flow rate of priming fluid into the enclosed bed space 130 is kept higher than a corresponding rate of the adaptor movement, i.e. a fluid flow rate measured in cm/h of the column height (i.e. depending on the column cross section area) is kept higher than the adaptor movement measured in cm/hour along the column tube height. Hereby a sealed bed space free from air is ensured.
S5: After the priming of the column a step of introducing slurry into the column is usually provided for packing a media bed from the slurry in the column. Slurry is introduced via the chromatography media inlet/outlet 225.
An advantage with a spiral groove 180′″ is that a process for unpacking chromatography media from the chromatography column can be improved. This unpacking process is further described below. Furthermore, priming may be more effective with a spiral groove thanks to a possibility to provide a higher flow over a smaller part of an outer periphery of the lower sealing arrangement 160b since fluid flow is only passing the lower sealing arrangement 160b where the spiral groove intersects with the lower sealing arrangement 160b which will be different for different positions of the adaptor assembly 120.
According to the invention a method for unpacking a chromatography column is also provided. The method steps of this method are illustrated in the flow chart of
A1: Resuspending a packed bed of chromatography media provided in said chromatography column 100; 100′ by providing a fluid to said bed space 130. This fluid is provided from a mobile phase bottom which is not seen in these views of the chromatography column, but which is provided in a bottom of the column.
A2: Removing resuspended chromatography media via a chromatography media outlet of the chromatography column 100; 100′.
A3: Lowering the adaptor assembly 120 to a position within the tubular side wall 110 such that the adaptor assembly 120 is in line with the at least one groove 180; 180′; 180″; 180′″. This could be the priming position mentioned above where the lower sealing assembly 160b intersects with at least a part of the at least one groove 180; 180′; 180′″ (depending on if the groove is a spiral groove or not).
A4: Flowing a fluid through at least one priming fluid passage 171 which is provided passing through the adaptor assembly 120, whereby said fluid will be flowed out through a first end opening 172 of each of the at least one priming fluid passage 171 and via said at least one groove 180; 180′; 180″; 180′″ into the bed space 130 for resuspending any remaining chromatography media in the bed space.
A5: Removing resuspended chromatography media via the chromatography media outlet.
The step A4 of flowing a fluid through the at least one priming fluid passage 171, may comprise the simultaneously movement of the adaptor assembly 120 axially within the tubular side wall 110. This is advantageous in embodiments where the groove is a spiral groove 180′″. Hereby the adaptor assembly 120 is moved over a distance of the tubular side wall 110 where the groove 180′″ is provided, whereby the flow of fluid out through the at least one first end opening 172 of the at least one priming fluid passage 171 will be transferred out via the spiral groove 180′″ into the bed space 130 from different directions depending on the position of the adaptor assembly 120. This is illustrated in
A combination of providing at least one groove 180, 180′, 180″, 180′″ in the inner wall 112 in a low position close to the base assembly 125 and providing at least one and suitably more than one priming fluid passages 171 with first end openings 172 distributed around the circumference of the adaptor assembly 120 is especially advantageous for both effective priming and effective unpacking of the chromatography column.
Various alternative embodiments may also be provided. For example, various standard conventional columns may be modified to provide a priming groove in an upper portion thereof (e.g. in an upper half). Such a groove may be formed as a spiral, and/or with multiple channel (e.g. arcuate, linear, etc.) portions, or the like. Such a modification could thus be used to provide a better chromatographic performance aspect (e.g. packed bed).
| Number | Date | Country | Kind |
|---|---|---|---|
| 2009557.6 | Jun 2020 | GB | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/EP2021/065962 | 6/14/2021 | WO |
