JP3317749B2 - Inorganic porous column - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an inorganic porous column. The inorganic porous column of the present invention supports an enzyme such as glucose isomerase, a catalyst such as platinum or palladium, or a functional group such as an octadecyl group, and is suitable for chromatography columns such as liquid chromatography and gas chromatography. Can be used for Also,
It can also be used for syringes and blood infusion catheters.

[0002]

2. Description of the Related Art Chromatography columns of this type are known to be composed of an organic polymer such as a styrene-divinylbenzene copolymer or a column filled with an inorganic filler such as silica gel in a cylinder. .

A column made of an organic material has disadvantages such as low pressure resistance due to low strength, swelling / shrinking by a solvent, and inability to heat sterilize. Therefore,
Inorganic materials without such difficulties, especially silica gel,
It is widely used.

[0004]

However, even with the conventional inorganic filler, it may be because the shape is bead-like.
It was found to have disadvantages different from those of organic columns.

[0005] That is, a column constituted by packing an inorganic filler such as silica gel in a cylinder first has a large resistance to the flow of a fluid and therefore a large pressure loss. As a result, the flow rate per unit time decreases, and the analysis requires a long time to be used as chromatography. Similarly, when used for instruments such as syringes due to large pressure loss,
The flow rate of the sample solution becomes extremely slow.

[0006] Further, since the flow rate changes depending on the packing state, the analysis value greatly varies between the production lots of the column.
In addition, when trying to use it for blood infusion catheters and syringes,
The beads may be spilled.

The inventors of the present invention have studied and found that an inorganic column which is not a packed column but which is of an integral type and whose pore diameter and pore distribution are appropriately controlled is a disadvantage of the conventional inorganic packed column. Did not occur.

The present invention has been made based on such findings. It is an object of the present invention to solve the problems of conventional inorganic columns, and to provide an inorganic porous column with small pressure loss and easy handling. Another object is to provide an inorganic porous column having a large contact area per unit volume with a sample fluid.

[0009]

The means is substantially made of glass or glass ceramic and has a pore diameter of 500 nm.
The above has a through-hole continuous in a three-dimensional network and pores having a pore diameter of 5 to 100 nm formed on the inner wall surface of the through-hole, and the total volume of the pores is 10 m ³ / t or less, The volume ratio of the through-holes to the whole is 20 to 90%, preferably 50 to 80%, and the volume ratio of the pores in all the pores is 10 to 90%.
% Or more, desirably 50% or more , and is an integral type porous column. In this means, it is desirable that the glass or glass ceramic is mainly composed of silica SiO ₂ .

In the inorganic porous column of the present invention, for example, after a silicon alkoxide is hydrolyzed and polymerized in an acidic solution of an organic polymer to gel a reaction solution, an organic polymer in the formed porous gel is formed. It can be produced by removing the molecules and then firing.

If the step of immersing the solidified porous gel in an aqueous ammonia solution is added between the hydrolysis step and the calcination step, the pore diameter and the pore volume ratio can be easily controlled. Further, a mixture of formamide and a polyhydric alcohol may be used instead of the organic polymer.

Further, the inorganic porous column of the present invention comprises:
Important component requirements are the pore diameter of the through-holes and the pores formed on the inner wall surface, the total volume of the pores, the volume ratio of the through-holes to the entire column, and the volume ratio of the pores to the total volume of the pores and the through-holes. And The three-dimensional structure of the column varies depending on the composition of the reaction system, the temperature and pH value, the molecular weight of the organic polymer, and other various conditions that affect the reactivity of the silicon alkoxide. Therefore, it is difficult to uniformly describe a method of controlling the three-dimensional structure, but if the conditions described above are the same, an object having substantially the same pore diameter and the like can be provided with good reproducibility.

The removal of the organic polymer from the porous gel formed as an intermediate can be done to some extent by washing the gel before drying, but after the washing process, up to the temperature at which the organic polymer decomposes or burns. It is more effective to completely remove the gel by heating it for a sufficiently long time.

The organic polymer used in the production of the inorganic porous column of the present invention is a water-soluble organic polymer which can theoretically be formed into an aqueous solution having an appropriate concentration, and is formed by hydrolysis of silicon alkoxide. Any one can be used as long as it can be uniformly dissolved in the reaction system containing alcohol, and specifically, a sodium or potassium salt of polystyrene sulfonic acid, which is a polymer metal salt, or a polymer acid, which is dissociated to form a polyanion. Polyacrylic acid, polyallylamine and polyethyleneimine which are polymer bases and generate polycations in aqueous solution, or polyethylene oxide which is a neutral polymer and has an ether bond in the main chain, and has a cyclic amide in the side chain Polyvinylpyrrolidone and the like are preferred.

When a mixture of formamide and a polyhydric alcohol is used in place of the organic polymer, glycerin is most suitable as the polyhydric alcohol. As silicon alkoxide, tetramethoxysilane, tetraethoxysilane,
Further, those obtained by appropriately polymerizing these to increase the oxide content can be mentioned.

Incidentally, the inorganic porous column of the present invention exerts the operation and effect described later as long as the three-dimensional structure satisfies the above conditions. Therefore, the manufacturing method is not limited.

[0017]

A fluid such as a sample solution enters from one end of the column, passes through a continuous through-hole formed in a three-dimensional network, and exits from the other end. During the passage, there is no obstacle such as beads in a conventional packed column, and the through-hole has a diameter of 500 nm or more, so that the fluid receives little resistance. Therefore, the pressure loss is also small.

If the diameter of the through-holes is the same, the higher the volume ratio of the through-holes to the whole column, the smaller the pressure loss, which is preferable. However, if the volume ratio exceeds 90%, the mechanical strength is significantly impaired. As a result, it becomes difficult to constitute a column for chromatography or the like by itself. On the other hand, if the volume ratio is less than 20%, the pressure loss is larger than that of the packed column. A preferred range of volume fraction for chromatography is 50-80%.

Since the pores are formed on the inner wall of the through hole, the specific surface area is high. Therefore, for example, when a functional group such as an octadecyl group is fixed to the pores by chemical modification, or an enzyme such as glucose isomerase, or a catalyst such as platinum or palladium is supported on the pores, these fluids pass through in the process of passing the fluid. Reacts efficiently with molecules. Moreover, since the functional groups are fixed in the pores, the functional groups will not flow even if the flow of the fluid is fast.

However, the volume ratio occupied by the pores in all the pores is 1
0% or more is required. If it is less than 10%, the functional groups can hardly be fixed even if the volume ratio of the through holes is increased to 90%. On the other hand, if the total volume of the pores exceeds 10 m ³ / t, the mechanical strength is significantly impaired, and it becomes difficult to constitute a single column for chromatography or the like. The range of the pore volume ratio and the total volume suitable for chromatography is 50% or more with respect to all pores, and the total volume of pores is 1 m ³ / t.
It is.

Incidentally, even if the volume ratio of the through-holes and the total volume of the pores with respect to the entire column are higher than the above-mentioned preferable upper limits, within the scope of the present invention, the outer periphery is covered with a cylindrical body or the like. It can be applied to chromatography by reinforcement.

[0022]

【Example】

Example 1 First, sodium polystyrene sulfonate (product number 24305-1 manufactured by Aldrich), which is a polymer metal salt, was added to 1
It was dissolved in 5.51 g of a normal nitric acid aqueous solution to obtain a 20% by weight solution. 5 ml of methanol was added thereto to make a homogeneous solution, and then 5 ml of tetramethoxysilane was added dropwise over about 1 minute to carry out a hydrolysis reaction. After stirring for several minutes, the resulting clear solution was transferred to a closed vessel and kept in a constant temperature bath at 40 ° C., whereupon it solidified after about 20 hours.

The solidified sample was further aged for several days,
It was immersed in a 1N aqueous ammonia solution at room temperature for 7 days while renewing the solution every day, dried at 60 ° C., and then heated to 500 ° C. at a heating rate of 100 ° C./h. The decomposition products of the high-molecular metal salt are washed with distilled water.
Heat treatment was performed at 0 ° C. for 2 hours. After this, the sample was 150 m long.
m and machined into a column shape with an inner diameter of 4.6 mm. Thus, a column made of porous silica glass was obtained.

It was confirmed by an electron microscope and a mercury intrusion measurement that through holes of about 3 μm were present in the column of the obtained porous silica glass in a three-dimensionally meshed structure. Then, it was confirmed by nitrogen adsorption measurement that many pores of about 10 nm were present on the inner wall of the through hole. The pore volume and pore diameter of the sample dried at 60 ° C. were almost the same as those of the porous silica glass after the heat treatment.

Example 2 First, a 25% by weight aqueous solution of polyacrylic acid which is a high molecular acid (manufactured by Aldrich, product number 19205-8; molecular weight 9)
Was diluted with distilled water to give a 7.4% aqueous solution, and then concentrated nitric acid was added to make the solution 1N nitric acid. Polyacrylic acid
To this solution consisting of 4 g and 5.51 g of 1N nitric acid was added 7 ml of tetraethoxysilane under stirring to effect a hydrolysis reaction. A few minutes later, the obtained clear solution was transferred to a closed container and kept in a thermostat at 60 ° C., and solidified after about 2 hours.

The solidified sample was aged for several hours, washed several times with distilled water and ethanol, and immersed in a 0.1 N aqueous ammonia solution at room temperature for 7 days while renewing the solution every day. Dried at 60 ° C. Then, it was machined into a column shape having a length of 150 mm and an inner diameter of 4.6 mm.

It was confirmed by an electron microscope and a mercury intrusion measurement that through-holes having a uniform size of about 3 μm were present in the dried sample in a three-dimensionally meshed structure. Then, it was confirmed by nitrogen adsorption measurement that many pores of about 10 nm were present on the inner wall of the through hole.

When ethanol was added to the above reaction solution to a maximum of 5 ml and solidified, the diameter of the through-hole of the obtained porous material became small, and it could be continuously controlled to a minimum of about 0.5 μm. Also, by changing the amount of the 1N nitric acid aqueous solution to be used from a minimum of 3.3 g to a maximum of 11 g, the diameter of the through-hole of the resulting porous body could be controlled in a range of a maximum of about 10 μm to a minimum of about 0.2 μm. Furthermore, the through-hole diameter could be controlled similarly by changing the concentration of polyacrylic acid and the reaction temperature.

These dried samples were heated to 800 ° C. at a rate of temperature increase of 100 ° C./h and maintained at this temperature for 2 hours. A column made of porous silica glass having almost the same structure as the dried samples was obtained. was gotten.

Example 3 First, a mixture of formamide and glycerin at a molar ratio of 3: 2 is prepared. Then, tetramethoxysilane, formamide in glycerin, and a 1 N nitric acid (HNO ₃ ) aqueous solution are mixed at a molar ratio of 1: 3: 1.5,
The mixture was stirred to homogenize to obtain a sol-gel reaction solution. 10 ml of the reaction solution is placed in a cylindrical closed container having a length of 200 mm and an inner diameter of 6 mm, and the container is placed in a thermostat, and the temperature of the sol-gel reaction solution is maintained at 60 ° C. for 2 hours to carry out hydrolysis / condensation polymerization. And solidified to obtain a column-shaped sample.

The solidified column-shaped sample was immersed in a 1N aqueous nitric acid solution at room temperature for 1 day, and then in a 0.1N aqueous ammonia solution at room temperature for 7 days while renewing the solution every day, Next, after drying at 60 ° C. for 10 hours, it was heated to 700 ° C. at a rate of 100 ° C./hour and heat-treated at 700 ° C. for 2 hours. Thus, a column made of porous silica glass was obtained.

It was confirmed by an electron microscope and a mercury intrusion measurement that through-holes of about 3 μm were present in the column of the obtained porous silica glass in a structure entangled in a three-dimensional network. Then, it was confirmed by nitrogen adsorption measurement that many pores of about 8 nm were present on the inner wall of the through hole.

-Experimental Example 1- In this experiment, the same raw materials as those used for manufacturing the porous column of Example 1 were used, and the amount of tetramethoxysilane dropped was varied as shown in Table 1. A porous column was manufactured, and the relationship between the microstructure of the column and its performance was evaluated.

[0034]

[Table 1] The performance evaluation was performed by measuring the pressure loss when a fluid was passed through the column.

That is, first, a porous column is assembled into a liquid chromatography apparatus, and a sample solution is flowed through the column using a pump that automatically sets a pressure necessary for obtaining a constant flow rate, and an outlet flow rate is 1 ml / min. The inlet pressure required to satisfy the condition was determined. Since the outlet side is open to the atmosphere, this inlet pressure directly corresponds to the pressure loss. As a sample solution, a 9: 1 (volume ratio) mixture of normal hexane and isopropyl alcohol was used.

Then, ten columns were manufactured under the same manufacturing conditions, and the sample solution was flowed, and the maximum value and the minimum value of the pressure loss were recorded. The measurement results are shown in Table 2 together with the microstructure of the column.

[0037]

[Table 2] As seen from Table 2, column N belonging to the scope of the present invention
o. In Nos. 2 to 4, the pressure loss was small and the variation was small.

On the other hand, column No. which does not belong to the scope of the present invention. In Nos. 1 and 5, since the volume ratio of the through-holes was lower than 20%, the pressure loss was so large that the pressure loss could not be measured.

Experimental Example 2 In this experiment, the same raw materials as those used for producing the porous column of Example 2 were used, and the amount of tetraethoxysilane dropped was varied as shown in Table 3. A porous column was manufactured, and the relationship between the microstructure of the column and its performance was evaluated.

[0040]

[Table 3] The performance evaluation was performed by measuring the pressure loss when a fluid was passed through the column in the same manner as in Experimental Example 1. The measurement results are shown in Table 4 together with the microstructure of the column.

[0041]

[Table 4] As shown in Table 4, column N belonging to the scope of the present invention
o. In Nos. 7 to 9, the pressure loss was small and the variation was small.

On the other hand, column No. which does not belong to the scope of the present invention. 6, since the volume ratio of the through-hole is lower than 20%,
The pressure loss was so large that it could not be measured. Also,
Column No. In No. 10, although the volume ratio of the through-hole was barely within the range of the present invention, the diameter of the through-hole was so small that the pressure loss was too large to be measured.

Experimental Example 3 In this experiment, the same raw materials as those used for manufacturing the porous column of Example 3 were used, and the mixing ratio of the raw materials and the reaction temperature were varied as shown in Table 5. Thus, a porous column was manufactured, and the relationship between the microstructure of the column and its performance was evaluated.

[0044]

[Table 5] The performance evaluation was performed by measuring the pressure loss when a fluid was passed through the column in the same manner as in Experimental Example 1. The measurement results are shown in Table 6 together with the microstructure of the column.

[0045]

[Table 6] As shown in Table 6, column N belonging to the scope of the present invention
o. Nos. 12 to 14 had a small pressure loss and a small variation.

On the other hand, column No. which does not belong to the scope of the present invention. In Nos. 11 and 15, although the volume ratio of the through-hole was within the range of the present invention, the diameter of the through-hole was too small, so that the pressure loss was too large to be measured.

-Comparative Experiment 1- This experiment was conducted using a conventional packed inorganic column for comparison with Experimental Examples 1 to 3.

That is, a diameter of 10 μm and a density of 1.5 g /
silica gel beads length 150mm of cm ^3, filled with void ratio of 40% to the tubular body having an inner diameter of 4.6 mm, which assembled to chromatography, column, measured like the pressure loss as in the above Experimental Examples and flow went.

As a result, the pressure loss is 70 kg / c at maximum.
m ² , a minimum of 30 kg / cm ² , and there was a considerable difference depending on the column.

Experimental Example 4 and Comparative Experiment 2 In this experiment, a functional group was actually immobilized on a porous column.
The analytical performance is evaluated. First, a mixed solution of nitrobenzene and toluene was added to a normal mobile phase, and this was used as a sample solution.

The column no. 13 was chemically modified with an octadecyl group, and then assembled into a liquid chromatography apparatus. Then, the sample solution was passed through liquid chromatography. Thereafter, the column manufactured in Comparative Experiment 1 was also chemically modified with an octadecyl group, and then assembled in a liquid chromatography apparatus, and a sample solution was flowed.

With both columns, the peak of nitrobenzene and the peak of toluene could be similarly separated. The pressure loss when using No. 13 was 1/10 or less of the pressure loss when using the comparative column.

[0053]

The inorganic porous column of the present invention has the following great effects. (1) Since the column is integrated, there is no problem that the flow rate changes depending on the packing state, and the variation between lots is small.

(2) The pressure loss is small because of the through-holes whose pore diameter and volume ratio are appropriately controlled. Therefore, if the inlet pressure is the same, the flow rate per unit time increases, and analysis can be performed in a shorter time than in the past. Moreover, since all the functional groups fixed to the pores react with the fluid, the efficiency of functional group consumption is high. Therefore, analysis can be performed with a short column.

(3) Easy handling when used for blood infusion catheters and syringes. (4) Since the shape and size of the fluid flow path are highly uniform,
The distribution of the analyte between the solution and the inner surface of the column does not vary from place to place.

Continuation of the front page (56) References JP-A-4-187237 (JP, A) JP-A-54-9969 (JP, A) JP-A-2-291963 (JP, A) JP-A-2-290552 (JP, A) , A) Special table Hei 4-500726 (JP, A) (58) Field surveyed (Int. Cl. ⁷ , DB name) G01N 30/60

Claims (3)

(57) [Claims] 1. A through hole substantially made of glass or glass ceramic having a hole diameter of 500 nm or more and continuous in a three-dimensional network, and a hole diameter of 5 to 5 formed on the inner wall surface of the through hole.
100 nm pores, and the total volume of the pores is 10 m ³ /
t, the volume ratio of the through-holes to the whole is 20 to 90%, and the volume ratio of the pores in all the pores is 10 to 90%.
% Or more , and is an integral type . 2. The inorganic porous column according to claim 1, wherein the glass or glass ceramic is mainly composed of silica SiO ₂ . 3. The inorganic porous column according to claim 1, which is used alone for chromatography.