Classifications

• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
  • G01N30/02—Column chromatography
  • G01N30/50—Conditioning of the sorbent material or stationary liquid
  • G01N30/56—Packing methods or coating methods
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
  • G01N30/02—Column chromatography
  • G01N30/50—Conditioning of the sorbent material or stationary liquid
  • G01N30/56—Packing methods or coating methods
  • G01N2030/562—Packing methods or coating methods packing
  • G01N2030/565—Packing methods or coating methods packing slurry packing
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
  • G01N30/02—Column chromatography
  • G01N30/60—Construction of the column
  • G01N30/6004—Construction of the column end pieces

Definitions

• the present invention relates to a column for liguid chromatography, which includes a column tube intended to be filled with a separation medium, and inlet and outlet means.
• the invention also relates to a method of filling such a column tube with separation medium.
• a chromatographic column In order for a chromatographic column to be able to separate biological substances effectively, it is usual to pack fine particles of separation material as tightly and as uniformly as possible in the column tube. Filling of the column, or column packing as it is often referred to, is normally effected by closing one end of the column with an outlet means which includes a filter element, and pumping a liquid suspension of the particles under pressure into the other end of the column. Whereas the pumped liquid is able to pass through the filter element essentially unobstructed, the particles are retained by the filter element, so as to build up a particle bed along the length of the tube.
• the particles are pressed out towards the wall of the tube and the particle bed obtains a stable compaction state with the particles well distributed, this state being maintained during the whole of the filling process.
• the stable restraining force in the particle bed is partially lost, resulting in expansion of the particle bed. Consequently, when the column tube is once again placed under pressure, disturbing heterogeneities or irregularities are liable to occur in the particle bed, such as the formation of channels and dead volumes. Numerous attempts have been made to solve this problem.
• EP-A-0040633 suggests that the inlet element is provided with a spring construction which will maintain the column bed under pressure and therewith improve bed stability.
• a similar construction based on the effect produced by a pressure plunger is described in DE-A- 3021366 and US-A-4 , 350, 595.
• DE-A-3000475 describes a column having a flexible wall which stabilizes packing of the particles by radial compression of the column packing.
• a column-filling device including a pressure plunger which functions to compress the bed during the actual filling process but which is then removed is described in US-A- 4,549,584.
• an object of the present invention is to provide a column having an inlet part which, while being structurally simple, enables the original compactness/density of the particle bed to be maintained, so as to avoid completely those problems that are associated with a drop in pressure in the column bed and expansion of the column bed subsequent to filling the column.
• Another object of the invention is to provide a method for filling a column in accordance with this concept.
• Fig. 2 is a view corresponding to the view of Fig. 1, but shows the column inlet element in a locked position after completion of a filling operation;
• Fig. 3 is a cross-sectional view taken on the line A-A in Fig. 1;
• Fig. 4 is a partial longitudinal-sectioned view of another embodiment of the separation column and shows the column in a filling position
• Fig. 5 is a view corresponding to the view of Fig. 4, but with the inlet part in a locked position after filling of the column has been completed.
• the separation column illustrated in the drawings is comprised of a column tube 1, for instance made of glass, PEEK (a polyether-ether-ketone sold by ICI) or like material, and an inlet means in the form of a top element 2, made for instance of PEEK.
• the inlet means can be moved axially along the tube 1 and locked thereto, as will be described in the following.
• the bottom part of the tube 1 includes an outlet passage 3 which opens into a connecting recess or aperture 4 which is screw-threaded for instance (not shown) .
• a bottom filter 5 is sealingly mounted on top of the outlet passage 3.
• the top element 2 Similar to the bottom part of the tube, the top element 2 also has a connecting recess or aperture 6 (for instance screw-threaded) which includes an inlet passage 7 which opens into a central recess 8 in the bottom part of the top element 2.
• the inner diameter of the recess 8 corresponds to the outer diameter of the column tube 1, so that the top element 2 can be moved vertically, i.e. axially, in relation to the tube 1.
• the recess 8 is provided inwardly of the sealing groove 10 with a circular recessed-part 11 which is adapted to accommodate a top filter 12, made for instance of sintered PEEK.
• the recessed-part 11 has a depth which is slightly smaller than the thickness of the filter 12. Furthermore, the diameter of the recessed-part 11 in relation to the filter is such as to enable the filter to be fixated in any desired position in the recessed-part by virtue of a force-fit, and so that the filter can only be moved vertically in the recessed-part by applying to the filter a force which is greater than the force to which the filter is subjected during filling of the column.
• radial slots or channels 13 Arranged around the periphery of the recessed-part 11 are a number, in the illustrated case six, radial slots or channels 13 which connect the bottom of the recessed-part 11 with the space below the sealing groove 10, and therewith also with the column tube interior 14.
• the configuration of these channels can be varied within wide limits, for instance with regard to both number, peripheral extension and radial depth.
• the top element 2 When filling the column tube 1 with packing material, the top element 2 will have the position shown in Fig. 1, in which it rests against the uppermost part of the tube 1 via the sealing ring 9.
• the filter 12 is placed in the recessed-part 11 so as to leave a gap 15 between the bottom of said recessed-part and the upper side of the filter, and also/ so as to form a gap between the upper edge of the tube and the underside of the filter. This enables the space 15 above the filter 11 to communicate with the column tube interior 14 through the channels 13 in the side wall of the recessed-part 11.
• a slurry comprising the desired particles of separation medium is pumped into the inlet passage 7 in per se conventional manner through a flexible tube connected to the connection 6. Because of the presence of the channels 13, the slurry will pass along the side of the filter 11 and down into the column tube 1 and will continue to the outlet passage 3 in the tube bottom. The particles are retained at the bottom of the tube by the bottom filter 5, while liquid will pass essentially unimpeded through the filter.
• the sealing ring 9 prevents liquid/slurry from leaking along the contacting surfaces between the top element 2 and the column tube l. Eventually, the interior 14 of the tube will have been filled with homogeneously packed particles (not shown) .
• this may be seen as the pumping ceasing, by the particles clogging the channels 13 and stopping the delivery of slurry.
• a suitable indicator which will indicate when filling of the tube is complete, for instance a suitably positioned optical sensor which detects when the particles begin to pack at the connection 6, or a pressure sensor which detects the change in pressure that occurs when the channels 13 become blocked or clogged, etc.
• the top element 2 When the column tube is full and pumping has been stopped, the top element 2 is pressed down over the column tube 1 to the position shown in Fig. 2, either manually or mechanically, and is firmly locked in this position.
• the top filter 12 is compressed slightly between the top element 2 and the upper edge of the tube, so that the filter abutment surfaces will seal effectively and close the earlier open communication of the channels 13 with the column tube interior.
• the sealing ring 9 will have been compressed in the groove 10 at the same time, so as to ensure effective sealing against leakage when the packed column is used.
• the filling arrangement is then disconnected and the packed separation column is ready for use.
• the particle bed will have the same degree of compaction as that during filling of the column tube, in other words that the stability of the particle bed achieved while filling the column tube will be retained and that this stability can be maintained even at high liquid pressures, such as in HPLC.
• the top element 2 can be locked in the position shown in Fig. 2 in a number of different ways.
• the top element 2 may be constructed for screw-engagement with the upper part of the column tube 1, in which case the top element is brought to its sealing position by screwing said element down along the column tube.
• the top element may be provided with an internal (e.g. circular) recess which coacts with a stop shoulder (e.g. also circular) on the outside of the column tube, or vice versa.
• a stop shoulder e.g. also circular
• the use of cotter-pins or like devices is a further alternative.
• top element 2 In order to ensure that the top element 2 will be held in its sealing position (Fig. 2) in use, at least when using the stop shoulder (shoulders) alternative, it may be convenient to place a locking sleeve or the like over the top element such as to fix the element against the column tube, by preventing radial expansion. It is, of course, also conceivable for the top element itself to lack a locking function and to lock the element in its sealing position totally with the aid of an additional element instead, such as a sleeve-like element of the aforesaid kind or a similar sleeve-like element which can be placed over the top element and which includes means for locking coaction with the separation column.
• the material in the top filter 12 will need to have at least a certain degree of rigidity, in order to achieve the intended function. If the filter material used is not sufficiently rigid in this regard, appropriate supportive means may be used, for instance the filter may be placed in a cassette or like supportive device.
• small spacer elements in the form of pegs or the like, for instance, may be provided on the filter surface that abuts the column tube. These pegs, or like projections, will preferably be configured to pierce the filter and/or be deformed to a sufficient extent as the filter is pressed down onto the column tube when locking the top element (Fig. 2) . Such spacer elements may replace totally the frictional engagement of the filter with the side-wall of the recessed-part 11.
• the sealing ring 9 will be pressed against the partition wall 11a between the groove 10 and the channels 13 as the top element 2 is pressed down into its locked position, so as to deform the partition wall and press said wall against the filter 12, at least to a substantial extent. This will reduce or even essentially completely eliminate the pocket or dead volume that is formed between the channel wall and the filter in the Fig. 2 illustration.
• the inner wall 11a of the recessed-part 11 may be extended right down to the column tube 1 in the filling position, and then be deformed when the top element 2 is brought to its locked state.
• the bottom part of the wall may optionally extend radially inwards beneath the filter to some extent.
• the partition wall 11a may extend down into a corresponding groove in the upper side of the column tube 1 and therewith effectively prevent access to the sealing groove.
• the filter 12 when the filter 12 is in a locked state the filter will lie against the abutment surfaces of both the top element 2 and the column tube 1. As will be understood, it may be sufficient for the top filter 12 to seal solely against one of these abutment surfaces. If desired, means may be provided for holding the filter 12 spaced from each of these surfaces, for instance spacer elements in the bottom of the recess- part 11, so as to ensure the provision of a liquid distributing space above the filter 12. Similarly, the aforesaid spacer elements on the column tube abutment surface need not necessarily be configured for piercing of the filter 12, provided that an effective seal can be achieved against the bottom-part of the recessed-part 11.
• FIGs. 4 and 5 illustrate a slightly modified embodiment of the invention.
• This embodiment includes a column tube 16 which is provided with an end-piece 17 having a larger inner diameter than that of the column tube.
• a top element 18 can be screwed onto the end piece 17 and has a connection 19 which is joined to an inlet passage 20.
• the inlet passage opens into a recessed-part 21 in the bottom of the top element 18 and is adapted to accommodate a top filter 22 provided with a peripheral sealing ring 23 (for instance pressed therein) .
• FIG. 4 shows the top element 18 in the filling state of the column, wherein the top filter 22 (similar to the filter 12 in the Fig. 2 embodiment) is in frictional engagement with the side-wall of the recessed-part 21 at a distance from the bottom of said recessed-part such that the channels 24 connect the space above the filter 22 with the space beneath the filter, and therewith with the interior of the column tube 16.
• the bypass channels are delimited between the top filter and recesses in the top element.
• the channels may be formed instead by peripheral recesses in the filter.
• different combinations of these alternatives are also conceivable.

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• Physics & Mathematics (AREA)
• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Analytical Chemistry (AREA)
• Biochemistry (AREA)
• General Health & Medical Sciences (AREA)
• General Physics & Mathematics (AREA)
• Immunology (AREA)
• Pathology (AREA)
• Treatment Of Liquids With Adsorbents In General (AREA)
• Filtration Of Liquid (AREA)

Priority Applications (5)

Applications Claiming Priority (2)

Publications (1)

Family

ID=20389712

Family Applications (1)

Country Status (7)

Families Citing this family (26)

Citations (1)

Family Cites Families (18)

• 1993
  • 1993-04-26 SE SE9301395A patent/SE501195C2/sv not_active IP Right Cessation
• 1994
  • 1994-04-25 WO PCT/SE1994/000362 patent/WO1994025134A1/en active IP Right Grant
  • 1994-04-25 JP JP6524155A patent/JPH08509551A/ja active Pending
  • 1994-04-25 AT AT94914661T patent/ATE188391T1/de active
  • 1994-04-25 US US08/537,819 patent/US5714074A/en not_active Expired - Lifetime
  • 1994-04-25 EP EP94914661A patent/EP0696223B1/en not_active Expired - Lifetime
  • 1994-04-25 DE DE69422512T patent/DE69422512D1/de not_active Expired - Lifetime

Patent Citations (1)

Non-Patent Citations (2)

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