WO2004054709A1

WO2004054709A1 - Sorbents with standard pore sizes

Info

Publication number: WO2004054709A1
Application number: PCT/EP2003/012951
Authority: WO
Inventors: Egbert Müller
Original assignee: Merck Patent Gmbh
Priority date: 2002-12-14
Filing date: 2003-11-19
Publication date: 2004-07-01
Also published as: DE10258491A1; AU2003303024A1

Abstract

The invention relates to organic separating materials with standard pore sizes for use in chromatography. In order to produce the inventive materials, a polymerisation mould is filled with monodispersable template-particles and a suitable polymerisation solution, is centrifuged, polymerised and subsequently the template-particles are eliminated preferably with hydrofluoric acid.

Description

Sorbents with a uniform pore size

The invention relates to monolithic or particulate sorbents made from organic polymers and their use in chromatography.

Monolithic shaped bodies or particulate materials made from organic or inorganic polymers are typically used as sorbents for chromatography. In order to achieve a high binding capacity of the sorbents, it is necessary to use materials with a large specific surface. For this reason, mostly porous materials are used for chromatography. Depending on the size of the pores, a distinction is made between micro, meso and macro pores. The pores can be created by chemical processes (e.g. phase separation, salt expansion) or physical processes (e.g. temperature treatment). In the case of organic polymers, the

Pores are formed by adding a porogen to the polymer solution. The statistical distribution of the size of these pores in the Sorbents is mostly of the Gauss type. The production of membranes from organic polymers is disclosed in EP 0 320 023. These polymer membranes are preferably used for the separation of macromolecular substances such as biopolymers, since macroporous materials can be used for such applications. However, mesoporous organic polymers with a narrow pore size distribution for the separation of smaller molecules cannot be produced in the necessary quality.

From the field of nano-technology it is known that porous solids can be formed by

colloidal particles are arranged in a regular package as a template, this template package is impregnated with a polymerization solution,

- is polymerized - and finally the template particles are removed from the polymer. (C.Göltner, Angew. Chem. 1999, 111, pp. 3347-3349). S. Johnson, P. Ollivier and T. Mallouk, Science, 1999, 283, pp. 963-965, use monodisperse silica gel particles with a diameter of 35 nm, which are packed into pellets under pressure, easily sintered together at 800 ° C. are and then serve as a template for copolymerization of an organic polymer. After removing the silica particles with hydrofluoric acid, porous polymer pellets are obtained.

It has now been found that certain shaped articles produced in a similar manner are outstandingly suitable as chromatographic separating materials. The materials according to the invention are produced from monodisperse silica gel particles which are centrifuged into a polymerization mold with a polymerization solution. The mixture is then polymerized and the silica gel particles are extracted. The separating material obtained in this way can be used as a monolithic sorbent or comminuted as a particulate sorbent. The separating materials according to the invention with a uniform pore size show significantly better separating properties than corresponding conventional materials with a pore size distribution of the Gauss type.

The present invention therefore relates to a process for the production of sorbents from organic polymers having a uniform pore size, with the following process steps: a) providing a polymerization form; b) filling the polymerization mold with a polymerization solution and with monodisperse template particles with a diameter between 100 nm and 300 μm; c) centrifugation of the filled polymerization mold; d) optionally pouring off the excess polymerization solution; e) polymerization; f) Removing the template particles with a washing solution which dissolves the template particles.

In a preferred embodiment, in step c) 1 to 2 hours

Centrifuged 2500 to 4000 revolutions.

In a further preferred embodiment, the treatment with hydrofluoric acid in step f) takes place at temperatures between 50 and 85 ° C.

In a further embodiment, the polymer block formed is removed from the polymerization mold after step e) and comminuted into particles.

In a preferred embodiment, a mixture of styrene and trimethyl methacrylate ester is used as the polymerization solution.

In a preferred embodiment, the sorbent obtained after step f) is dried at a temperature between 200 and 350 ° C.

In a preferred embodiment, the template particles used in step b) carry template molecules on their surface.

The present invention also relates to sorbents made from organic polymers having a uniform pore size and produced by the process according to the invention.

The present invention also relates to the use of the materials according to the invention for the chromatographic separation of at least two substances. According to the invention, sorbents made from organic polymers are sorbents which are produced by polymerizing one or more organic monomers and / or oligomers.

According to the chromatographic results obtained with the sorbents according to the invention, it is not so much a uniform size of the sorbent particles that seems to produce a good separation performance, but rather a uniform size of the pores. This can be achieved with the method according to the invention. Furthermore, the process according to the invention has the advantage that sorbents with a homogeneous pore distribution are produced which can be used both as monolithic and as particulate sorbent. The preferred thermal aftertreatment can greatly reduce the often undesirable swelling behavior of the sorbents in organic solvents.

Any form which is stable with respect to the polymerization solution and from which the polymerization block can be removed after the template particles have been washed out or preferably directly after the polymerization has taken place can be used as the polymerization form. For monolithic sorbents, the form of polymerization should match the shape of the monolith to be made. Tubular polymerization forms are therefore preferably used, the inside diameter of which typically does not exceed 1 to 2 cm, since otherwise the template particles can only be removed very slowly. Centrifuge tubes are particularly preferably used, which at both ends by means of a

Screw lid can be opened and closed. In this way, the polymer block can be pushed out of the polymerization mold after the polymerization.

Polytetrafluoroethylene is particularly preferably used as the material of the polymerization form. According to the invention, monodisperse porous or preferably non-porous silica particles, ie silica gel particles, are used as template particles. These materials can be produced in different particle sizes with good monodispersity. It is also possible to use other monodisperse particles made from inorganic oxides, such as Al ₂ O ₃ , provided that they can be produced in a suitable quality. According to the invention, these materials also fall under the term template particles. The diameter of the template particles is between 100 nm and 300 μm. Particles between 200 nm and 5 μm in diameter are preferably used. With these particle sizes, depending on the selected particle size and also depending on the polymerization solution used, sorbents with pore sizes between approx. 30 and 300 nm in diameter are obtained, which are very suitable for chromatographic separations (see Example 1).

The process according to the invention enables a wide variety of organic polymeric moldings to be produced. The moldings can be produced, for example, by radical, ionic or thermal polymerization. It can be, for example, poly (meth) acrylic acid derivatives, polystyrene derivatives, polyesters, polyamides or polyethylenes. The monomers to be used accordingly are known to the person skilled in the art in the field of organic polymers. These are, for example, monoethylenically or polyethylenically unsaturated monomers, such as vinyl monomers, vinyl aromatic and vinyl aliphatic monomers, for example styrene and substituted styrenes, vinyl acetates or vinyl propionates, acrylic monomers, such as methacrylates and other alkyl acrylates, ethoxymethyl acrylate and higher analogues and the corresponding methacyl acid esters or their corresponding amide, such as acrylamide or acrylonitrile. Further monoethylenically and polyethylenically unsaturated monomers can be found, for example, in EP 0 366 252 or US 5,858,296. The person skilled in the art is able to combine the various monomers / oligomers accordingly, optionally to choose a suitable free-radical initiator or initiator and thus to put together a polymerization solution with which the polymerization form is filled up.

The mold filled with the template particles and the polymerization solution is first centrifuged so that the particles settle and form the densest spherical packing possible. It has been found that materials with a particularly uniform pore distribution can be produced by the centrifugation step according to the invention, i.e. centrifugation leads to a much better packing of the particles than e.g. a sedimentation. Furthermore, the centrifugation step offers process engineering advantages over e.g. a pack under pressure, since the aggressive and mostly unmanageable polymerization solutions only have to be filled in and do not have to be pumped through a suitable vessel to seal the particles under pressure.

Centrifugation is preferably carried out at 2000 to 4000 revolutions for 1 to 2 hours.

Subsequently, any excess polymerization solution is poured off or pipetted off, and the part of the polymerization solution remaining in the polymerization form is polymerized. The duration and temperature of the polymerization are matched to the respective monomer solution according to customary rules. It was found that it takes a little longer to complete the reaction than a corresponding polymerization in solution would require. A person skilled in the art of polymerization is able to optimize the polymerization conditions and the polymerization time accordingly. After the polymerization step has ended, the solid polymerization block formed, which consists of the inorganic template particles and the organic polymer formed around them, is removed and the material of the template particles is washed out and dissolved out. To do this, the polymer block is washed in a washing solution that

Particles dissolve, optionally with heating, stored or preferably pivoted.

Ammonia solution, solutions of amines, alkali metal solutions, such as sodium hydroxide solution, or preferably aqueous hydrofluoric acid can be used as the washing solution which dissolves the template particles.

A treatment with aqueous hydrofluoric acid to remove silica gel typically takes 2 to 10 days, depending on the size of the polymer block. This step is preferably carried out at a temperature between 50 and 85.degree.

After further washing steps to remove the aggressive washing solution and the last particles, the porous organic polymer molded body is obtained as an impression or counterpart to the polymerization mold used, which is filled with the template particles.

Alternatively, the polymerization block obtained after the polymerization can first be comminuted, for example in a mortar, before the template particles are washed out. This procedure is possible if a particulate sorbent is to be produced. By shredding the polymer block, the washing out of the template particles is accelerated. Then the fragments can e.g. be ground in a ball mill and divided by sieving.

The shaped bodies or particles are preferably dried for 1 to 7 days after treatment with hydrofluoric acid and corresponding washing at temperatures between 200 and 350 ° C., preferably at about 250 ° C. If necessary, this drying should take place under protective gas, e.g. argon, respectively. It has been found that this undesired swelling behavior of the polymeric moldings can be greatly reduced by this drying at high temperatures. For example, thermal aftertreatment (24 h at 250 ° C.) can increase the length of a 2 cm long polymer rod (diameter 1 cm) made of styrene and trimethyl methacrylate ester when treated with water / acetonitrile (50/50, v / v) by 15% 5% can be reduced.

If the resulting shaped bodies or particles already have the appropriate functionalities, they can be used directly for chromatographic separations. For example, a polymer made of polystyrene or derivatives thereof can be used directly for reversed phase separations. For this purpose, the particles are packed in appropriate chromatography columns and the shaped bodies are provided with appropriate connectors, surrounded with a jacket and integrated in a chromatographic separation column. Suitable holders and jackets are already known for inorganic monolithic sorbents (e.g. WO 98/59238 and WO 01/03797) and can be transferred to the moldings according to the invention. Especially when wrapping with plastics, e.g. PEEK or fiber reinforced

PEEK, the organic moldings according to the invention can usually be coated more effectively and more densely than the corresponding inorganic moldings, since they can form a stronger bond with the plastic coating.

Depending on the separation properties which the sorbent according to the invention is intended to have, however, further modifications may initially be necessary. If it is to be used, for example, for affinity or ion exchange chromatography, the surface must be covered with appropriate separation effectors. In some cases, suitable substances can already be added to the monomer solution and thus introduced directly into the polymer. preferably, However, according to known methods, functionalities are first introduced during the polymerization, which can then be implemented using separation effectors. Likewise, further modifications can be introduced via block or graft polymerizations on the polymer moldings. Separation effectors and

Monomers which, in addition to a polymerizable double bond, also have other functionalities, such as Oxirane rings, are known to the person skilled in the art. Examples can be found in WO 96/22316 or WO 95/10354.

In the same way, suitable functionalities of the sorbents according to the invention can be used to bind or immobilize biomolecules, such as enzymes. Macroporous moldings or particles are particularly suitable for this. Therefore, biomolecules such as Enzymes according to the invention also under the term separation effectors.

The process according to the invention can also be used to produce sorbents with special separating properties. As with "molecular imprinting", template molecules can be bound to the surface of the, in this case porous or non-porous, template particles.

The gaps or pores are then filled with monomer solution and polymerized. During the polymerization, cavities form that enclose the template molecules. Then the template particles and the template molecules bound to them are washed out. In contrast to the known methods of molecular imprinting, the method according to the invention has the advantage that the template molecules assume a defined aligned position due to the binding to the template particles. In this way, more defined cavities are created, which can enter into clearer and stronger interactions during the chromatographic separation. Another great advantage of the process according to the invention is that after the polymerization, all template molecules can be washed out. In conventional processes, part of the template molecules are completely surrounded by the polymer after the polymerization and cannot be removed from the polymer at all or only very slowly. Since template molecules can still be released from the polymer even when the polymers are later used for chromatographic separation and falsify the analysis, the use of such materials is mostly limited to purification or qualitative analysis. A trace analysis is hardly possible.

Polymers produced according to the invention do not show this bleeding. The reason for this is the preferred covalent binding of the template molecules to the template particles. In this way, they are never completely surrounded and held in place by the polymerized molded body, but are completely removed by washing out the template particles together with them.

In addition, to produce an imprint polymer according to the prior art, a large amount of cleaned template is necessary, which can only be recovered with difficulty. In the process according to the invention, however, the template is first bound to the template particles. Excess template molecules can be washed off and collected. Only in a second step is the monomer solution added and polymerized. The polymer moldings produced by the process according to the invention using a template can, as already explained in the description of the production process, be used both as moldings or can also be comminuted into particles for certain applications.

The method according to the invention thus offers the possibility of producing organic polymer moldings with a defined, narrow pore size manufacture. The pores can be defined by the template particles themselves or by a modification of the particles with template molecules. The method according to the invention is simple and reliable. The centrifugation step ensures that a dense and homogeneous packing of the template particles is generated.

The materials according to the invention enable more effective chromatographic separations in comparison to conventional sorbents with a Gaussian-like distribution of the pore size. In addition, the surface of the template particles and / or the surface of the shaped body according to the invention can be modified, so that there are a multitude of possibilities for ideally adapting the shaped bodies to the particular separation problem.

Even without further explanations, it is assumed that a person skilled in the art can use the above description in the broadest scope. The preferred embodiments and examples are therefore only to be regarded as descriptive, in no way as in any way limiting in any way.

The complete disclosure of all of the applications, patents and publications listed above and below, and of the corresponding application DE 102 58 491, filed on December 14, 2002, are incorporated by reference into this application. Examples

1. Production of moldings with different sized pores

Approximately 1 g of monodisperse silica gel particles are suspended in about 2 ml of a styrene / divinylbenzene polymerization solution (1: 1, v: v) with the addition of 1% (m: m) azoisobutyronitrile. The solution is evacuated and filled into a plastic sleeve made of PTFE (polytetrafluoroethylene). It is polymerized at 70 ° C for 20 hours. The polymerization block is then removed and shaken in aqueous hydrofluoric acid for 144 hours at room temperature. Then it is carefully washed with water.

The completeness of the acid treatment was checked by Si elemental analysis. All of the moldings produced were free of silica gel. The following table 1 lists the morphological properties of the moldings produced using different template particles.

2. Production of polymer particles

50 g of Monospher ^® 250 (diameter 0.25 μm) particles are suspended in a polymerization solution consisting of 50 ml trimethylolpropane trimethacrylate, 50 ml styrene and 0.4 g azoisobutyronitrile and filled into three centrifuge tubes made of PTFE (inner diameter 26 mm) and settled to let. The supernatant solution is suctioned off and the centrifuge tubes are centrifuged for 1 hour at 3000 rpm. They are then stored in the oven at 70 ° C. for 20 hours for the polymerization. Allow to cool, push the polymer blocks out of the centrifuge tubes and crush them (eg with a hammer). Removing the template

Particles take place over 10 days in HF at 70 ° C. Then it is shaken twice with deionized water and twice with acetone and dried in vacuo at 70 ° C. A silicon analysis showed <0.3%. The fragments obtained are ground in a ball mill with porcelain balls with an average diameter of 5 cm (duration approx. 20 hours) and then dried at 250 ° C. for 5 days. Then they are pre-sorted on a 250 μm sieve and added for particle size distribution and sieving. 5

3. Production of molded polymer articles

5 g of Monospher ^® 250 (diameter 0.25 μm) particles are suspended in a polymerization solution consisting of 5 ml trimethylolpropane trimethacrylate, 5 ml ⁰ styrene and 0.04 g azoisobutyronitrile and filled into three centrifuge tubes made of PTFE (inner diameter 1 cm) and allowed to settle. The supernatant solution is suctioned off and the centrifuge tubes are centrifuged for 1 hour at 3000 rpm. They are then stored in the oven for 18 hours at 70 ° C for polymerization. Allow to cool 5 and push the polymer blocks out of the centrifuge tubes. The template particles are extracted in HF at 70 ° C for 10 days. The mixture is then shaken twice with deionized water and twice with acetone, dried in vacuo at 70 ° C. and then dried at 250 ° C. for 5 days. ° A silicon analysis showed <0.3%. 4. Comparison with conventional sorbents

The following sorbents were examined: a) LiChrospher ^® WP 3005 μm RP 18 particles (in chromatography column with length: 125 mm and diameter: 4 mm) b) Chromolith ^® Performance RP 18 (length: 100 mm, diameter: 4.6 mm) c ) according to the invention particles, prepared according to example 2, mean particle size 25 microns, packed in a Superformance ^® cartridge with bed height 1.5 cm (diameter: 1 cm).

Elution conditions: isocratic

Eluent: acetonitrile / water (40/60, v / v) with the addition of 0.1% trifluoroacetic acid.

The theoretical floor height for human insulin was determined by measuring the peak widths: a) LiChrospher ^® WP 300 3500 theoretical floors per mb) Chromolith ^® RP 18 8040 theoretical floors per mc) particles according to the invention 8100 theoretical floors per m

It can be seen from this that a very high number of trays can be achieved when using the particles according to the invention. The theoretical plate number of the LiChrospher ^® WP 300 material should be 10 μm (twice the particle size), which corresponds to a plate number of 100,000 for toluene. With the 25 μm material produced according to the invention, the number of trays would have to be 5 times worse. Instead, it is almost three times as good.

Claims

Expectations

1. A process for producing sorbents from organic polymers with a uniform pore size, comprising the following process steps: a) providing a polymerization mold; b) filling the polymerization mold with a polymerization solution and with monodisperse template particles with a diameter between 100 nm and 300 μm; c) centrifugation of the filled polymerization mold; ° d) optional pouring of excess polymerization solution; e) polymerization; f) Removing the template particles with a washing solution which dissolves the template particles.

5 2. The method according to claim 1, characterized in that in step c) is centrifuged for 1 to 2 hours at 2500 to 4000 revolutions.

3. The method according to any one of claims 1 or 2, characterized in that the treatment with hydrofluoric acid in step f) takes place at a temperature between 50 and 85 ° C.

4. The method according to any one of claims 1 to 3, characterized in that after step e) the resulting polymerization block is removed from the polymerization mold and comminuted. 5

5. The method according to any one of claims 1 to 4, characterized in that the sorbent obtained after step f) is dried at a temperature between 200 and 350 ° C.

0

6. The method according to any one of claims 1 to 5, characterized in that the template particles used in step b) carry template molecules on their surface.

7. Separating materials made of organic polymers produced by a process according to one of claims 1 to 6.

8. Use of the separation materials according to claim 7 for the chromatographic separation of at least two substances.