Patent Application
20060118471
The invention relates to liquid phase chromatography.
Liquid phase chromatography is conventionally carried out in a column containing a chromatographic mixture comprising, for example, a gel constituted by microparticles in suspension in a buffer solution contained in a cylindrical tube which is supplied with liquid to be chromatographed, said liquid then being recovered at the column outlet.
That proven technique suffers from a certain number of disadvantages. Firstly, the chromatography columns are bulky, expensive installations. Because of their capacity, filling them involves handling large quantities of chromatographic mixture, which has to be stored both before and after use.
Attempts have been made to find alternatives to such columns. Reference may in particular be made to published French patent application FR-A-2 645 965, which proposes a modular chromatography device constituted by a juxtaposition of chromatography modules into each of which the liquid to be chromatographed is introduced under pressure.
The technical solution proposed by that document is not entirely satisfactory, however. The chromatography modules described are relatively complex to produce. Further, the structure of the device poses problems regarding supply and distribution of the liquid to be chromatographed between the juxtaposed modules. Further, the complex path followed by the liquid to be chromatographed limits the throughput of the device.
The aim of the invention is to overcome the disadvantages cited above by proposing a chromatography module which can facilitate assembly and dismantling of the chromatography column while guaranteeing high quality chromatography.
To this end, the invention proposes a chromatography module comprising:
said module being characterized in that the ends of the container are open and have complementary shapes, and in that the filtering membrane is fixed to the lower end of the container.
Such a module, which is easy to handle, allows direct storage of the chromatographic mixture. Because it fits directly on an identical module, its structure means that a chromatography column can be produced wherein the chromatographic mixture, which is monoblock, enables chromatography to be carried out in a single step.
The invention also provides a chromatography column comprising a stack of said modules fitted together.
Further aims and advantages of the invention will become apparent from the description below made with reference to the accompanying drawings in which:
Said module 1 comprises a container 2, shown alone in
The container 2, preferably produced from a plastic material such as polypropylene, has an upper end 4 and a lower end 5, both open, each formed by a circular free edge, an upper edge 6 and lower edge 7 respectively.
Between its ends 4, 5, the container 2 defines a chamber 8 with volume V1, in which a chromatographic mixture 9 (also termed the stationary phase) is received comprising, for example, a chromatographic gel constituted by microparticles (such as microspheres) in suspension in a buffer solution, for example in a ratio of 1:1 (i.e. one kilogram of microparticles per 1 litre of buffer solution).
A filter membrane 10 the diameter of which corresponds to the internal diameter of the wall 3 is fixed to the lower end 5 of the container 2. Said membrane 10, produced from polyethylene, for example, has a porous structure and guarantees retention of the chromatographic mixture 9 in the container 2.
As can be seen in
In the chamber 8, the countersink 12 has a secondary volume V2 determined as a function of the compressibility of the chromatographic mixture 9 and the desired final pressure thereof, as will become apparent below.
The ends 4, 5 of the container thus have complementary shapes, so that it is possible to stack several modules 1 by fitting them one into another by inserting the lower end 5 of a first module 1 into the upper end 4 of a second module 1 until the lower edge 7 of the first abuts against the shoulder 11 of the second.
We shall discuss the advantages of such a disposition below.
As can be seen in
Once the mixture 9 has been introduced into the container 2, it is sealed by means of two films 16, 17 formed from plastics material, one of which, 16, covers the upper end 4 and the other of which, 17, covers the lower end 5.
Further, a rubber O-ring 18 is placed at the lower end 5 of the container 2. Although not clearly visible in the figures, said ring 18 may be fixed in an annular groove provided in the lower edge 7 of the container 2; the O-ring extends slightly out of the groove to allow it to be compressed when stacking the module 1.
Regarding the dimensions, the module 1 has a thickness—i.e. the distance separating its edges 6, 7—in the range 50 mm [millimeters] to 100 mm, while its diameter (i.e. the external diameter of the wall) is in the range 100 mm to 2000 mm. The example illustrated shows a relatively “square” module 1, but it may also be more flattened, whereupon its diameter will be much larger than its thickness.
Module 1 is constituted as follows.
Starting from the container 2, the O-ring 18 is attached along with the membrane 10 at its lower end 5. The chromatographic mixture is then introduced under pressure so that the mixture 9 is level with the upper edge 6 without overflowing, however. The container 2 is then sealed using films 16, 17 disposed on the upper 4 and lower 5 edges. The pressure of the chromatographic mixture 9 in the module 1 is P1.
A chromatography column 19 is shown in
Either side of the modules 1, column 19 comprises two closing plates, namely:
When stacking the chromatographic modules 1, the lower end 5 of an upper module 1 is introduced into the upper end 4 of a lower module 1, after removing films 16, 17. It is easy to understand that said introduction tends to compress the chromatographic mixture 9 of the lower module 1 the secondary volume V2 of which, previously occupied by the mixture 9, is, in the end, occupied by the fitted portion of the upper module 1. Because of the nature of the chromatographic mixture 9 (we saw above that it is a gel), such compression can only occur by forced fitting.
For this reason, to keep the stack together, connecting rods 15 (made in the shape of threaded rods) are introduced into the openings, bolts (not shown) then being screwed onto the ends of the connecting rods 15 to keep them fitted together.
Thus, the pressure in the chromatographic mixture 9 increases in proportion to the reduction in the volume occupied by the mixture 9. More precisely, if P2 is the pressure of the chromatographic mixture 9 in a stacked module 1, and if the thickness of the membrane 10 is ignored, the pressure P2 satisfies the following relationship:
P2/P1=(V1−V2)V1
It will be understood that adjusting the height of the countersink 12 can proportionately modify the final pressure of the chromatographic mixture 9 to adjust the flow rate in the column 19.
The liquid to be chromatographed, introduced via the upper plate 20, passes through all of the modules 1 in the stack in succession before being evacuated via the lower plate 23. The openings in modules 1 at their ends means that flow of the mobile phase is laminar in the stationary phase 9, while the membranes 10, which maintain the stationary phase in place in each separate module do not oppose passage of the mobile phase. Thus, the conditions inside the column are as if the stationary phase 9 were a monoblock.
Clearly, the chromatographic mixture 9 will be selected as a function of the type of chromatography which is to be carried out (ion exchange chromatography, gel filtration chromatography, hydrophobic chromatography, affinity chromatography, silica gel chromatography, etc), for example for protein filtration, purification or reagent preparation.
| Number | Date | Country | |
|---|---|---|---|
| Parent | PCT/FR04/02837 | Nov 2004 | US |
| Child | 11266493 | Nov 2005 | US |
