KR20070112940A - Monolithic silica particles, preparation method thereof and stationary phase materials for liquid chromatography using the same - Google Patents

Abstract

Monolithic silica powder with improved separation efficiency, a method for preparing the monolithic silica powder and a stationary phase for liquid chromatography using the same are provided to be used as a newly shaped liquid chromatography stationary phase by improving the separation efficiency, maintaining merits of a conventional monolithic column. Monolithic silica powder has a moving-bed flow path. The powder has a particle size of 5 to 10 mum. The powder has a cavity size of 50 to 300 Å. A method for preparing monolithic silica powder comprises: a first step of preparing a silica monolith; a second step of powdering the silica monolith prepared in the first step; a third step of cleaning and sintering the powdered silica monolith of the second step to strengthen the powdered silica monolith; and a fourth step of optionally sorting particles from the strengthened silica monolith of the third step through a particle sorting process. A stationary phase for liquid chromatography is prepared by attaching a ligand to monolithic silica powder having a moving-bed flow path.

Description

Silica monolith powder with improved resolution, preparation method thereof, and stationary phase for liquid chromatography using the same {Monolithic silica particles, preparation method method and stationary phase materials for liquid chromatography using the same}

1 is a scanning electron micrograph of silica monolith powder particles according to the present invention;

2 is a nitrogen adsorption isotherm graph of silica monolith powder particles according to the present invention;

3 is a distribution diagram of pore size in silica monolit powder particles according to the present invention;

4 is a chromatogram using a column packed with silica monolit particles not subjected to particle fractionation according to an embodiment of the present invention;

5 is a chromatogram using a column packed with silica monolit particles which have been subjected to particle fractionation according to an embodiment of the present invention;

Figure 6 is a graph measuring the height of the mono monolith stationary phase and commercial stationary phase in accordance with an embodiment of the present invention; And

Figure 7 is a graph showing the mobile phase flow rate trend for the step height by group in the silica monolith stationary and commercial stationary phase according to an embodiment of the present invention.

<Description of Major Symbols in Drawing>

1: phenol

2: 2-nitroaniline

3: acetophenone

4: benzene

5: toluene

The present invention relates to silica monolit powder with improved resolution, a method for preparing the same, and a stationary phase for liquid chromatography using the same.

Generally, porous powder is mainly used as a stationary filler in liquid chromatography, and silica (SiO ₂ ), alumina (Al ₂ O ₃ ), zirconia (ZrO ₂ ) powder, and various ligands on the porous surface of the powder are used. Chemically bonded or polymer coated ones are used, and various types of block copolymer porous polymer powders are also used. Currently, the most common filler is a C ₁₈ stationary phase in which an octadecyl ligand is bound to a porous silica powder.

However, monoliths have recently been of great interest in the areas of liquid chromatography (LC) and capillary electrochromatography (CEC).

Monolit means that the stationary phase is one giant three-dimensional porous network. Monolit columns do not require frits because the stationary phase itself acts as a frit, and because of their high porosity, they require less pressure at high flow rates and faster mass transfer rates. This is high. In addition, the manufacturing cost is reduced compared to the conventional packed column. Because of these advantages, monolithic columns of 2 to 5 mm in diameter and 10 to 30 cm in length are already commercially available.

The monoliths are in the spotlight especially in the field of microcolumn chromatography and capillary electrochromatography. In microcolumn chromatography and capillary electrochromatography, frits with a very small void volume should be used. The formation and mounting of such frits is not only very difficult, but also causes many problems such as bubble formation. Is a porous net structure and at the same time interest in monolit with a frit function is increasing, and research on this is being actively conducted. However, the reproducibility of the monolithic column, etc., is not yet complete, and thus the commercialization stage of the microcolumn or capillary electrochromatography column has not been reached.

There are two types of monoliths for microcolumn or capillary electrochromatography columns: inorganic polymers and organic polymers. In general, a monomer mixture, a polymerization catalyst, and a solvent which does not dissolve the resulting polymer are mixed to form a solution. After polymerization, the polymerization is carried out by raising the temperature, and finally, by washing off the solvent and the unreacted monomer. Inorganic polymer monoliths are predominantly silica type (Bartle et al . , J. Chromatogr . A , 2000 , 892, 279-290; Tang et al . , J. High Resolut . Chromatogr . , 2000 , 23, 73), organic The polymer type is a styrene-divinylbenzene copolymer (Gusev et al . , J. Chromatogr . A , 1999 , 855, 273-290), a methacrylate copolymer (Peters et al . , Anal. Chem. , 1988 , 70, 2296; Coufal et al. , J. Chromatogr . A. , 2002 , 946, 99-106), acrylamide-based copolymers (Fujimoto et al . , J. Chromatogr . A. , 1995 , 716 107, Hoegger et al., J. Chromatogr.A . , 2001 , 914, 211-222).

The monolith can be made variously through a modification reaction to attach the ligand again after the first preparation. The ligands include C ₁₈ ligands (Minakuchi et al . , Anal. Chem . , 1996 , 68, 3498-3501; Minakuchi et al . , J. Chromatogr. A. , 1997 , 762, 135-146; Ishizuka et al . , J. Chromatogr . ., 2002, 960, 85-96), a polar ligand or ligands for ion exchange (such as Suzuki, J. Chromatogr. A., 2000, 873, 247-256), ligand chiral separation (Chen, etc., Anal. Chem. , 2001 , 73, 3348-3357; Chen et al . , J. Chromatogr.A . , 2002 , 942, 83-91). One unusual monolit is one in which a silica capillary is filled with a powdery stationary phase, followed by sintering the entire stationary phase with a cyclic heating line (Adam et al . , J. Chromatogr . A. , 2000 , 887, 327-337).

However, since monolith tends to shrink during manufacturing, it is not possible to make a column directly in a stainless steel tube, but to make a monolith first and then shrink the tube by putting a monolith in a teflon tube that shrinks by heating. There are limitations to be prepared (Majors, LC- GC , 2000 , 18, 586-598). In addition, a monolith column ends its life immediately when a part of the column (mainly the inlet) is blocked or broken.

Furthermore, since a series of processes such as washing of unreacted material after monolith production, ligand addition reaction, endcapping reaction, and additional washing process are inefficient, a lot of time and effort are consumed.

Therefore, the inventors of the present invention have superior resolution than the liquid chromatography stationary phase using silica, and during the research to solve the problem of the conventional monolit column, the monolit is prepared through a high polymerization reaction and then pulverized into fine powder. After the washing process and the ignition process, the high performance liquid chromatography prepared by attaching the ligand was confirmed that the resolution can be improved and the monolith column problem can be solved and the present invention has been completed.

It is an object of the present invention to provide silica monolit powder with improved resolution.

Another object of the present invention to provide a method for producing the monolit powder.

Still another object of the present invention is to provide a stationary phase for liquid chromatography using the silica monolit powder.

In order to achieve the above object, the present invention provides a silica monolit powder.

The present invention also provides a method for producing the silica monolith powder.

The present invention also provides a stationary phase for liquid chromatography using the silica monolith powder.

Hereinafter, the present invention will be described in detail.

The present invention provides silica monolit powder.

Silica monolith powders according to the present invention have the characteristics of typical three-dimensional monolithic structures with coarse mobile phase flow passages. Thus, for example, the general monolit column has advantages such as low pressure and high mass transfer speed even at high flow rates.

The particle size of the silica monolith powder according to the present invention is preferably 5 to 10 μm, and the size of the pupil of the silica monolith powder is preferably 50 to 300 mm 3. It is based on a commercial liquid chromatography stationary phase that is conventionally used. The commercial liquid chromatography stationary phase generally has a particle size of 3, 5, and 10 μm, and a pupil size of 60 to 300 mm 3, each of which has various sizes. Can be controlled by a factor.

The pores to be formed are determined by the materials used in polymer polymerization, and in order to achieve effective pore formation, the unreacted materials and materials remaining inside and outside the silica monolith structure must be effectively washed.

The present invention also provides a method for producing the silica monolit powder.

The method for producing silica monolit powder according to the present invention,

Preparing a silica monolit (step 1);

Powdering the silica monolith prepared in step 1 (step 2);

Washing and heating the silica monolith powdered in step 2 to strengthen it (step 3); And

It comprises a step (step 4) to selectively separate the particles through the particle fractionation process in the silica monolit reinforced in step 3.

Step 1 is to prepare silica monolit.

The silica monolith may be synthesized through a known method or commercially available, and may be synthesized using copolymerization.

In order to prepare silica monolith powder using the copolymerization, a reaction mixture consisting of tetramethylorthosilicate (hereinafter referred to as TMOS), polyethylene glycol (PEG) and urea is first made. At this time, the mixing ratio of the reaction mixture is 60 to 80% by weight TMOS, 15 to 25% by weight PEG, 0 to 25% by weight of urea is preferred.

The reaction proceeds by adding and stirring an aqueous acetic acid solution to the reaction mixture. At this time, the concentration of the acetic acid aqueous solution is preferably 0.002 ~ 0.1 N, it is preferable that the reaction mixture is 20 to 45% by weight, the acetic acid aqueous solution is mixed at 55 to 80% by weight.

The reaction temperature of the silica monolit according to the present invention is preferably 40 to 80 ° C., and the temperature is raised to 100 to 150 ° C. to heat the monolith lump. The solidified monolit mass is then removed from the container. As the polymerization proceeds, the monolit shrinkage occurs, so that the monolit mass can be easily taken out of the container.

Next, step 2 is a step of powdering the silica monolit prepared in step 1.

In the powdering of the silica monolith according to the present invention, a conventional method may be used, and for example, a ball mill or agate mortar and pestle may be used to gently grind the fine powder.

Next, step 3 is a step of strengthening by washing and heating the silica monolith powdered in step 2.

In this step, the monolith powder is washed with a sufficient amount of water while stirring to solidify the structure of the monolith powder, followed by washing with methanol and drying. It is preferable that the temperature at the time of drying is 100-150 ^degreeC . The dried monolith powder is sintered at 250-350 ^° C. for 12-48 hours to strengthen the structure of the monolith.

Next, step 4 is a step of selectively separating the particles through the particle fractionation process in the silica monolit reinforced in step 3.

In this step, the particles are selectively fractionated using a particle fractionation apparatus, wherein the size of the fractionated particles is preferably 5 to 10 µm. If the fractionation process is not performed, the average size of the particles is large and the size distribution is wide, so that the column cannot be efficiently filled when the column is filled, and the separation efficiency of the column is reduced.

The silica monolith powder prepared by the above method has superior separation efficiency than the conventional liquid chromatography stationary phase and can solve the problem of washing and ligand adhesion, which is a problem of the conventional monolith column, and thus can be used as a new type of liquid chromatography stationary phase. .

The present invention also provides a stationary phase for liquid chromatography using the silica monolith powder.

The stationary phase may be prepared by attaching a ligand to the obtained silica monolit powder, and the ligand attachment method may be a conventional method. For example, by attaching a C ₁₈ ligand using chlorodimethyloctadecylsilane and deactivating the silanol groups remaining through end-capping using chlorotrimethylsilane. Stationary phases can be prepared.

The column may be prepared by filling the still phase produced by the above method in a stainless steel tube coated with glass on the inner wall. The filling method may be carried out in a conventional manner, for example, a slurry filling method may be used to apply vibration for 10 minutes at 14,000 psi, 2 minutes at 10,000 psi, and 30 minutes at 8,000 psi.

Hereinafter, the present invention will be described in more detail with reference to Examples.

However, the following examples are merely to illustrate the present invention, the contents of the present invention is not limited to the following examples.

< Example  1> silica Monolit  Powder manufacturing

4.0 ml of TMOS, 880 mg of PEG, and 900 mg of urea were dispersed in 10.0 ml of 0.01 N acetic acid aqueous solution and then stirred at 0 ° C. for 40 minutes. Thereafter, the mixed solution was placed in a glass container and copolymerized at 40 ° C. for 24 hours. The monolith produced after the reaction was maintained at 120 ° C. and dried for 3 hours to complete the porous pupil structure of the monolith. The dried monolit was finely powdered, and then polyethylene glycol was removed using water and methanol and dried. The dried monolit powder was ignited at 300 ° C. for 24 hours to form a rigid silica structure. The formed silica monolit powder was selectively fractionated only particles of 5 to 10 μm using a particle sorting apparatus (Sonic Sifter Separator).

 <Analysis>

Referring to the particle shape and size of the silica monolith powder obtained by the method with reference to Figure 1 , the powder is composed of small-scale monolith particles having the characteristics of a typical three-dimensional monolith structure with a thick mobile phase flow passage, It can be seen that each particle consists of a set of elliptical basic units about 2 microns long and 3 microns long.

As shown in FIG . 3 prepared with reference to FIG. 2 , it was confirmed that the average pore size of the silica monolith particles was about 100 μs.

< Experimental Example  1> Silica according to particle fractionation Monolit Stationary phase  Chromatography Resolution  Measure

In order to determine the resolution of the silica monolith stationary phase according to the particle fractionation process, the following experiment was performed.

660 mg of chlorodimethyloctadecylsilane and 600 μl of pyridine were added to the silica monolit powder prepared in Example 1 and the silica monolit powder, which was not subjected to the particle fractionation process, to the control, and 20.0 ml of xylene After dispersion, the reaction was carried out at 110 ° C. for 24 hours. After the reaction, the mixture was washed with toluene, tetrahydrofuran, methanol, methanol / water (50:50 by volume) and finally methanol solution, and dried to attach C ₁₈ ligand. 800 μl of chlorodimethyloctadecylsilane and 600 μl of pyridine were added to the dried particles, and then dispersed in 20.0 ml of xylene, followed by a reaction process at 110 ° C. for 24 hours. It was washed under the conditions to prepare a stationary monolith powder.

The monolith powder was dispersed in a methanol solution, and packed into a column for 10 minutes at 14,000 psi, 2 minutes at 10,000 psi, and 30 minutes at a pressure of 8,000 psi to prepare a microcolumn. The prepared microcolumn is 10 μl of solvent mixed with 80/20% (v / v) of methanol and water as a moving solvent in a test sample consisting of phenol, 2-nitroaniline, acetophenone, benzene, and toluene as analyte solute. The separation efficiency was measured by moving at the flow rate of minutes.

The results are shown in FIGS . 4 and 5 .

4 is a chromatogram of a column prepared from a stationary phase made of a monolit powder, which has not been subjected to a particle size screening process, and FIG. 5 is a chromatogram of a column prepared from a stationary phase made of a monolit powder, which has been subjected to a particle size screening process. As shown in FIG . 4 , the retention time between peaks of phenol (1), 2-nitroaniline (2) and acetophenone (3) when the monolith powder without the particle size selection process is used as the stationary phase Although the difference between the two is small, the effective separation was not achieved . However, as shown in FIG . 5 , when the monolit powder, which had undergone the particle size selection process, was used as the stationary phase, the five test samples, phenol (1) and 2- It was confirmed that nitroaniline (2), acetophenone (3), benzene (4) and toluene (5) were satisfactorily separated.

< Experimental Example  2> Hem height (Height Equivalent to Theoretical Plate, HETP ) Measure

In order to determine the superiority of the separation efficiency of the monolith powder according to the present invention, a column made of the monolith powder according to Example 1 and a column made of Altima (Alltima C ₁₈ , particle size: 5 μm), which is a commercial stationary phase, were prepared. The following experiment was performed.

0.001 ml / min, 0.002 ml / min, 0.003 ml / min, 0.005 ml / min, 0.01 ml / min, 0.02 ml / min, and 0.03 ml under 80/20% (v / v%) mixed solvent of methanol and water For various flow rates of / min, a test sample consisting of phenol, 2-nitroaniline, acetophenone, benzene, and toluene was isolated to measure the height of the stage. The height of the column is a measure of the separation efficiency as the length of the column division unit required for the solute, which is a sample molecule, to reach an equilibrium between the mobile phase and the stationary phase. The lower the height, the higher the separation efficiency.

The measured results are shown in FIGS . 6 and 7 .

As shown in FIG . 6 , it can be seen that the stationary phase made of the monolit powder of the present invention has a higher separation efficiency than that of the commercial stationary phase because the stationary phase is lower.

FIG. 7 is a graph comparing curves for optimizing a trend between stage height versus mobile phase flow rate for each stationary phase and commercial stationary phase using silica monolith of the present invention. As shown in FIG . 7 , not only the stationary phase using the silica monolit of the present invention showed excellent results in terms of the step height than the commercial stationary phase, but also the tendency of the step height increased slowly at high flow rates, and thus the commercial mass monolith column It can be seen that it also has the advantage characteristics. This is a result of the presence of the coarse mobile phase flow passages contained in the monolit. It was also confirmed that the coarse mobile phase flow passages contained in the monolit resulted in lower column pressures compared to columns made of commercial stationary phases.

Therefore, the silica monolit powder of the present invention can be used as a new liquid chromatography stationary phase because it has a considerably superior effect in comparison with existing commercial stationary phases in terms of manufacturing convenience, separation efficiency and column load pressure.

As described above, the silica monolith powder according to the present invention has superior separation efficiency than the conventional liquid chromatography stationary phase, retains the advantages of the conventional monolithic column, and is a problem of washing and ligand of the monolithic column. It can be used as a new type of liquid chromatography stationary phase because it can solve the problem of adhesion.

Claims (14)

Silica monolit powder with mobile phase flow passage. The method of claim 1, wherein the particle size of the silica monolith powder, characterized in that 5 ~ 10 ㎛. The silica monolith powder according to claim 1, wherein the pore size of the powder is 50 to 300 mm 3. Preparing a silica monolit (step 1); Powdering the silica monolith prepared in step 1 (step 2); Washing and sintering the silica monolith powdered in step 2 to strengthen it (step 3); And A method for producing silica monolith powder having a mobile phase flow passage comprising the step (step 4) of selectively separating particles through a particle fractionation process in the silica monolit reinforced in step 3. The method of claim 4, wherein the silica monolith of step 1 is prepared using copolymerization of TMSO, PEG and urea. The method of claim 5, wherein the amount of the reaction mixture used in the copolymerization is 60 to 80% by weight of tetramethyl ortho silicate, 15 to 25% by weight polyethylene glycol and 0 to 25% by weight of urea silica monolite powder Manufacturing method. The method for preparing silica monolith powder according to claim 5, wherein the concentration of acetic acid aqueous solution added to the reaction mixture used in the copolymerization is 0.002 to 0.1 N. The method for preparing silica monolith powder according to claim 5, wherein in the copolymerization, the reaction mixture is 20 to 45 wt% and the acetic acid aqueous solution is mixed at 55 to 80 wt%. The method for producing silica monolith powder according to claim 5, wherein the reaction temperature in the copolymerization is 40 to 80 ° C. 5. The method of claim 4, wherein the powdering of Step 2 comprises a ball mill or agate mortar and pestle. 5. The method of claim 4, wherein in the washing step 3, water and methanol are used sequentially. The method of claim 4, wherein the sintering temperature of step 3 is 250 ~ 350 ^° C. The method for preparing silica monolith powder according to claim 4, wherein the size of the particles to be fractionated in the particle fractionation process of step 4 is 5 to 10 µm. A stationary phase for liquid chromatography prepared by attaching a ligand to the silica monolit powder of claim 1.

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