JPS6267451A - Packing agent for chromatography - Google Patents

Abstract

PURPOSE:To obtain a packing agent which is excellent in the endurance and reproducibility with a large mechanical strength, by spraying a material solution in which a calcium compound and a phosphor compound are mixed with water or an organic solvent compatible with water into a flame or a heated zone. CONSTITUTION:Water or an organic solvent compatible with water are used as solvent separately or a mixed solvent of water and the organic solvent compatible with water is used as solvent. A soluble calcium and phosphor compound are added to individual solvents and mixed to prepare a material solution. Then, the solutions obtained by such a preparation are sprayed into a flame by a widely-accepted method or into a heated zone heated up to a temperature range in which the material solution is thermally decomposed. Hydroxyapatite spherical particles with the size of several mum - several ten mum formed by this thermal decomposition by spraying are used as column packing agent without an additional treatment intact or after classified to uniform the size.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention provides hyroxyapatite having novel properties,
This invention relates to a chromatography packing material consisting of ca5 (PO4)30H1. More specifically, the present invention provides a chromatographic packing material comprising microspherical hydroxyapatite particles. The chromatography packing material according to the present invention is made of microspherical hydroxya, e-tite, which has high mechanical strength, excellent durability, and reproducibility, and is particularly suitable as a packing material for column chromatography. It is extremely useful for industrial applications.

Recently, high-performance liquid chromatography and electrophoresis equipment are often used for the separation and production of chemical and biochemical substances. Among these, high-performance liquid chromatography uses columns filled with substances that have adsorption capacity. It is widely used because it is easy to handle, as it only requires separating and passing the target material through. Until now, silica and alumina have been the mainstream filler materials used in such applications. However, there is a limit to the separation of biopolymers with large molecular weights using existing high-performance liquid chromatography using packing materials such as ion exchange type and molecular sieve type. Separation and purification of molecular weight proteins is considered impossible. On the other hand, recently, biotechnology's 4(v)
Along with the exhibition, there is a strong desire to develop technological means to break through these barriers of separation limits.

By the way, recently, columns packed with hydroxya♀tite have become commercially available, and it has become possible to separate and purify biopolymers with a size of 10 to 5 Daltons or more, which was previously impossible. Separation of proteins, nucleic acids, m-element, etc.
It is attracting attention as a solution to the industrialization of biochip, which requires purification.

However, the conventional hydroxy/a-tite fillers have low mechanical strength, are easily divided into particles, have poor reproducibility of filling characteristics and chromatographic characteristics, are poor in durability, and have a short lifespan. It is excellent for pot purification of biopolymers due to various drawbacks such as being expensive.Despite having excellent properties, it has not been widely used, and there is an urgent need to resolve these drawbacks. has been done.

That is, the conventional hydroquina g-tite was produced using the Teiselius method (A, Ti5elius, S, Hjerius method).
tenand I'j, Levin, ArCh+E
1i0Chem, EliOphys, 65.132-
155 (1956)), Ca/P It is difficult to control the ratio and the phosphoric acid content cannot be removed sufficiently, so it has the disadvantage of being weak against strong alkalis and organic solvents.Also, the shape of the filler particles is not easy to break, such as plate-like crystals or It is composed of an aggregate of microcrystals and has low mechanical strength, so it has the disadvantage of being easily pulverized during filling or measurement operations under high pressure. Furthermore, since the particles are composed of aggregates of plate-like or columnar microcrystals, they have poor fluidity during filling, and as mentioned above, their mechanical strength is low.
It has been extremely difficult to apply high pressure to densely pack the filler particles and make the gaps between the filler particles uniform. In addition, there are many practical difficulties, such as the need to be extremely careful in handling the packed column so as not to change its filling state.

Packing materials for high performance liquid chromatography need to have extremely small particle sizes. In other words, the diffusion coefficient of a substance in a liquid, K, is significantly smaller than that in a gas, so in order to reduce the effects of non-equilibrium partitioning in liquid chromatography, the mass transfer distance in the mobile phase must be sufficiently Therefore, in order to speed up the analysis of liquid chromatography, a packing material with an extremely small particle size is required compared to that of gas chromatography.Recently, various packing materials are being used. 3-5
Particles with particle diameters on the order of μm are also coming into practical use.

Particle miniaturization has been achieved to some extent with regard to HIF CI gysiapatite, but as mentioned above, conventional hydroxya♀tite particles have low mechanical strength and have defects that make them easy to pulverize. Furthermore, since the microvessels are composed of a monomer of plate-shaped or cast-shaped microcrystals, when used as a filler, it was inevitable that the particles would have irregular shapes and sag.

In addition to reducing the particle size of the filler, making the shape of the filler particles spherical has the practical effect of making the filling operation easier.
Not only is this an advantage, but it also reduces the variation in the gaps between packed particles, which is one of the causes of peak broadening in chromatograms. This is extremely important from the standpoint of improving characteristics. However, regarding the spheronization of hydroxya and e-tight fillers using conventional techniques, only spherical molded particles with a size of about 100 μm, which are technically easy to granulate, have been produced.
Several μm x suitable for high performance liquid chromatography
Extremely minute spherical hydroxya of several μm in size.
Regarding the actual situation of Q-tight, although its realization is strongly desired, it has not been realized to date due to its technical difficulties.

The present inventors previously proposed a novel method for producing hyroxyapatite, in which water or a water-compatible organic solvent was used alone as a solvent, or a water-compatible organic solvent and water were used. Using a mixed solvent with as a solvent, add and mix a soluble calcium compound and a phosphorus compound to the used solvent to prepare a raw material solution, and spray the raw material solution into a flame or heating zone. We provide a method for producing microspherical hydroxyapatite by causing a thermal decomposition reaction, and by selecting the forming conditions at that time, it can be produced directly without any forming operation. It has been discovered that it is possible to produce hydroquina Q-tite particles having a spherical shape of submicron to several 10 microns (Japanese Patent Application No. 59-265270, Japanese Patent Application No. 60-3).
(See No. 8655). Further, through subsequent research, the present inventors found that the spherical particles thus obtained were not only the most desirable shape for particles of chromatography fillers, but also had extremely excellent chromatographic properties. It was discovered that it has certain characteristics. In other words, the hydroxyapatite particles with this novel property not only have the advantage of being able to directly produce spherical particles of several μm and 1 ft without any molding operation, which was impossible with the conventional technology, but also have the advantage that Oa Because the manufacturing method allows easy control of the /P ratio, the produced hydroxya Q-tite spherical particles have excellent chemical stability, extremely high mechanical strength, and excellent durability, making them suitable as fillers. It has been found that it has many excellent properties. The present invention has been made based on this knowledge.

The present invention will be explained in detail below.

In the present invention, the hydroxya. It is used as a.

Since the hydroxyapatite mentioned above has a spherical particle shape, it has good fluidity during filling, and does not cause the bridging phenomenon that tends to occur when filling angular particles, and can be filled using normal operating procedures. A uniformly packed column can be easily produced by simply carrying out the following steps. Furthermore, when filling well-shaped spherical particles with uniform particle diameters, the filling operation is not only easier, but also the interparticles formed by uniformly packed hydroxya Q-tite particles are Since the gaps are also extremely uniform, the vortex causes a difference in the linear velocity of the solute flow in the direction of the column, eliminating variations in the gaps between the packed particles, which is one of the causes of reduced column separation performance. It has the advantage of being able to

Minimization of particle size and uniformity of particle size have been achieved to some extent with hydroxya/Q-tight packing materials produced by conventional techniques, and also with respect to column packing, the gap between packed particles has been reduced. Although careful efforts have been made to reduce variation,
Hydroxia Q-tite fillers produced by conventional techniques have various particle shapes such as plate-like or columnar microcrystal aggregates, and have low mechanical strength.
It was not possible to completely eliminate variations in interparticle gaps by applying sufficient pressure to achieve uniform filling.

As is well known, there are three factors that reduce the separation ability of column chromatography: (1) diffusion, (2) nonequilibrium, and (3) vortex flow.

All of these have the effect of increasing the band broadening of the solute and therefore increasing the height of the theoretical plate. (1) is the diffusion of solute in the axial direction of the column, but pumps that can stably pump liquid at high pressure are now in practical use, so chromatographic separation can be performed at a high flow rate where solute diffusion can be ignored. By doing so, it is possible to actually eliminate the effect of (1). (2) is an effect that occurs when equilibrium is not necessarily established in each part of the column, and is more pronounced as the particle size of the packing material becomes larger. In addition, (3) (A vortex flow occurs because the packing material is positioned so as to block the flow, which causes a difference in the linear velocity of the solute in the column axis direction. This phenomenon becomes more pronounced as the particle size increases. , and if the particle shape is irregular or the packing method is not uniform, the effect will be greater.As a corollary of the well-known theory of Section 1H, by decreasing the height of the Lirin stage, It can be seen that in order to improve the separation ability of the column, it is important to use a packing material with particles as fine as possible, and it is also important to pack the particles evenly and in a uniform manner with as much uniformity as possible.

From this, it is clear that the microspherical hydroquina g-tite used in the present invention satisfies the conventional customization of fillers desired.

In addition, the microspherical Hydroquina Q-Tite manufactured by the above-mentioned method has extremely high mechanical strength, with the tightly packed particles not breaking even when a filling pressure of 500 kg/T2 is applied, for example. This has an important practical advantage in that the packing operation and the handling of the filled column are extremely easy. In order to improve column chromatography, the particle size of the packing material can be reduced to ff1l, as described in Section 7 above, but on the other hand, the column chromatography can be improved by packing fine particles. The particles 11''5 isi within the tube also become wasteful, thus resulting in a dropout that significantly reduces the flow rate.

In reality, to solve this problem, a method is used to increase the flow rate by applying high pressure to the column, but in this case, a considerable amount of royal force is applied to the packing material. .

Therefore, if the mechanical strength (W) of the filler particles used is small, the packed particles will be destroyed during packing or measurement and become fine, which will cause clogging of the column. This causes a shortened lifespan.The conventional hydroxya gutite consists of plate-like or deep coalescence of microcrystals.
Compared to the disadvantage that it is easily broken, the filler of the present invention is made of particles with extremely high mechanical strength, so this point can be solved in four points. .

Furthermore, whether or not it is possible to reliably apply light to a column with a packing material consisting of particles as small as a few μm to 10 μm is a matter of practical practicality, as a characteristic that the packing material should have. This is an extremely important matter in IT. However, the hydroquina/Q-tite filler produced by conventional wave techniques has poor fluidity and is difficult to fill uniformly because its particle shapes are composed of various shapes such as plate shapes and aggregates of micro-folds. was not easy and required special ingenuity.

Therefore, uniform charging of large columns with larger diameters is further complicated by the low mechanical strength of the particles and the problems of technicians trying to suppress variations in the interparticle spacing. However, the ease of filling operation was not as expected.

However, since the microspherical hydroxya-Q-tite used in the present invention has a well-ordered spherical shape, it has good fluidity during filling and has high mechanical strength, so it can be easily reproducible. High filling operation 2 can be performed.Moreover,
Not only for packing analytical columns with small column diameters, but also for preparative separation columns with large column diameters, the packing operation can be easily performed without worrying about destroying the packed particles or making the packing unrestricted. There is an advantage in being able to do this.

Furthermore, the hydroxy/anotite used in the present invention is not only a large spherical particle with a mechanically strong shape as described above, but also the Ca/p ratio can be easily controlled. Therefore, it has excellent chemical stability and is organic)
It is resistant to chemicals and strong alkalis, and will not turn into powder after one use. In this way, since it uses hydroxyapatite, which is extremely stable both physically and chemically,
The reproducibility of filling is also high, so retention time, retention capacity, etc.
It is possible to provide a column with high reproducibility of chromatographic characteristics and high fire resistance that can withstand repeated use. This makes it possible to provide a hydroxyapatite column with a long life, and has the effect of greatly reducing the cost of using the column, which is of extremely great practical significance.

Next, microspherical hydroxya♀ used in the present invention
The manufacturing method of tight will be explained in detail.

First, as a starting material, water or a water-compatible solvent is used as a solvent, or a mixed solvent of water and an organic solvent that is compatible with water is used as a solvent, and each of these solvents is used as a starting material. A soluble calcium compound and a phosphorus compound are added and mixed, and the whole is dissolved to form a raw material solution.At this time, the order of mixing 1 raw material for making solution 1 is not necessarily in the above order. They are not specific and only need to be mixed together in an order that does not cause precipitation.Also, with regard to the above raw material solution, depending on the combination of calcium compound, phosphorus compound and solvent, uniform The solution may be difficult to prepare, or the prepared solution may become cloudy over time; In addition, when using an organic compound as a phosphorus compound, adding aqueous ammonia to the solution can suppress the formation of by-product acids and free 111t CaO, and improve the formation of roxy/Qtite. It has the effect of increasing or decreasing the production rate.If necessary, acid or aqueous anthonia can be added to the solution.The time for mixing will vary depending on the raw materials selected, but the total Stir thoroughly until the raw materials are dissolved and a solution with a uniform composition is obtained.

Next, the solution prepared in this way is sprayed into a flame such as a gas burner by a known spraying method using a pressurized nozzle or a rotating disk, or is sprayed into a flame such as a gas furnace, electric furnace, high temperature gas, etc. The raw material solution is sprayed into a heating zone that has been preheated to a desired temperature in the temperature range where thermal decomposition occurs. When a flammable organic solvent that is compatible with water or a mixed solvent of a flammable organic solvent that is compatible with water and water is used as a solvent, the tail may be caused by combustion of the flammable organic solvent itself. The heat of combustion can also be used as a heat source for the pyrolysis reaction. In addition, when used to promote evaporation of a mixed solvent of water and an organic solvent that is compatible with water without burning the organic solvent, or when using water alone as a solvent, Requires a source of heat, such as a gas furnace as described above. Alternatively, both the combustion heat of the organic solvent and a heating source such as a gas furnace can be used in combination. Note that even when an organic solvent that is compatible with water is used alone as a solvent, depending on the combination of raw material compounds, the reaction system may be mixed with water due to residual moisture or crystal water in the raw material compounds. In some cases, the system uses a mixed solvent with an iTI medium, but in any of these cases, as long as the raw material solution can be prepared without precipitation, it can be used as a solvent in this method. I can do it.

Calcium and phosphorus compounds used in this method include Ca(NO3)2.4 H2O, C
a(OH3Coo)2. + H2O, Ca0J2 ・
2H20, CaBr2-2H20, C! aHPO4-2
H20, N FL4 H2PO4, NH4H2PO3,
NH4H2PO2, H3P0. , H3PO3, H3P0
2, (CH30)3F, (C2H50)3F.

((CH3)2CHO)3F, (OH3(CnH2)3
0:13F, (C6H5)3F.

Examples include (C2H3O)2POH1(06H5)3PO. Are there still methanol, ethanol, and isozorono as organic solvents? It is preferable to use alcohols such as mer, or acetone, but there is no particular limitation to these, and any combination of raw materials that can be mixed to produce a uniform solution that does not cause precipitation may be used. . In addition, for this purpose, the acid added as necessary is as follows:
Nitric acid and salinity are suitable.

The powder produced by the spray pyrolysis reaction by the above method can be made into hollow spherical micropowder, tightly packed microspherical powder, or by selecting the four types and combinations of raw material compounds and the spray pyrolysis conditions. It can be changed in various ways, such as spherical powder being broken into powder of various shapes. Among these, well-shaped spherical particles can be suitably used as the chromatography filler of the present invention, regardless of whether they are hollow or not.

In addition, when the powder produced by the above-mentioned spray pyrolysis reaction is further exposed to various atmospheres such as air, air with water vapor added, and inert ``A'' air, hydroquina/Q-tite stably exists without decomposition. By heat-treating at a desired temperature within a possible temperature range, it is possible to improve the degree of crystallinity and sintering and further increase the mechanical strength.

If necessary, as described above, the microspherical hydroxya and Q-tite produced can be classified by various known methods in order to make them uniform throughout the particle size.

Hereinafter, the present invention will be specifically explained with reference to Examples. However, the present invention is not limited to the following examples.

Implementation row 1 ■) Manufacturing ability of microspherical hyroxyapatite
(Nf)3)z ・4H20 and H3P0. is dissolved in methanol to give a total solute concentration of 2 mo/! A solution was prepared so that the amount of HNO39-5me was 1 liter/liter, and HNO39-5me was added to 1 liter of this solution to prepare a homogeneous solution. This solution was then applied to a two-fluid atomizing nozzle at a rate of 3 rrrl per minute (atomizing air pressure = 0.7 kg/l,
x2) into the flame of a gas burner to cause a thermal decomposition reaction, and the generated powder was collected with a cyclone. As a result of X@ diffraction analysis of this powder, it was found to be hyrroxy, e-tight, and observation using an electron microscope (SKM) revealed that it was a powder containing spherical particles of approximately 1 to 137 μm. The powder was classified using an air classifier, and only well-shaped spherical hirroxia ξtite with a particle size of 5 to 15 μm was eliminated.

2) Pressure test during filling The hirroxy/ξtite particles obtained in 1) were dispersed in pure water and slurry filled using a stainless steel column for high performance liquid chromatography with an inner diameter of 4 mm and a length of 250 plates. Filling tests were conducted by varying the pressurizing pressure from 100 to 500':g/CTl12 according to the method. pressurized test)
The packed column was disassembled and the packed hydroxyanonotite was observed using SEM, but no deformation or destruction of the particles was observed even after applying a high pressure of 9/cTL2 to 500. Therefore, we conducted a packing test again using the hydroxyl 7ξ tight particles recovered by disassembling the first column using the same procedure as described above, and were able to perform the same packing as the first time. No changes such as deformation or destruction were observed in the hydroxya.ξtite particles inspected by SEM.

3) Column chromatography The hydroxya♀tite spherical particles produced by the method described in
The inner diameter is 4 in order and the length is 150 mm by slurry filling method.
Pack into an xtm stainless steel column.

(Filling pressure = 170 kg/Crn2).

Using this column, egg white lysozyme was measured by a linear gradient method (gradient = 3.49 x 10-'' M/ml!) using a soda buffer solution at pH 6 and 8. Liquid flow rate from 0.5 to 2.0
ml/min (3 when converted to a column with an inner diameter of 1 cIrL)
．． The constant pressure of 1 jill when varied from 13 to 12.5 ml/min) is 19 kg/crrt2 (0.5 ml/min).
ml! / min) to 81 kQ/CIn”
(In the case of 2.0 ml/min), it was possible to perform stable measurements under each condition, and no troubles such as column clogging or breakage occurred. This shows that the packing material of the present invention is suitable for chromatography at a high pressure of about 80:9/CIrL2. Furthermore, when the packing material of the present invention is used for a protein solution with low viscosity such as zozyme, the flow rate converted to 1 column inner diameter is 10 q'l.
This shows that this column has superior characteristics compared to conventional columns, which allow solutions to flow at a high flow rate of less than 1/min.

Implementation 1] 2 The hydroxyapatite particles produced in 1) of Example 1 were heat-treated at 800° C. for 2 hours in the air. As a result of analyzing the obtained particles, the X-ray diffraction intensity of hydroxyano-tight increased with crystal growth, but no decomposition was observed, and no destruction of the shape of the spherical particles was observed by SEM observation. Next, the heat-treated hydroxya.

Using this column, egg white lysozyme and cytochrome C
The measurements were carried out in random order, and the total number of cycles was measured for egg white lysozyme, the total head circumference for cytochrome C, and the repeated use test of the blade tip was performed to determine whether the column was clogged or not and the degree of reproducibility of retention time. I looked into it. All measurements were performed using a linear gradient method using -6,8 sodium phosphate buffer (gradient = 3.49 x 10 M/m
e), the flow rate was 0.5 ml/min. The pressure at the time of measurement was 20-Z3 Kf/lri. Until the end of the test, no clogging of the column occurred, and an extremely good ROMAT column was obtained. Regarding reproducibility, the variation in retention time of egg white lysozyme in this measurement was investigated, and the minimum and maximum values throughout the measurement were 22.05 minutes and 22.37 minutes, respectively. This measurement is not intended to be a rigorous test of the retention time repeatability or retention time reproducibility that can be obtained through statistical processing of the measurement data, but it is usually described in the liquid chromatography instruction manual. It is within the 2-coupled range, which is the stated standard for evaluating reproducibility, and it can be seen that the column used for this measurement has high reproducibility. After repeated tests, the column was disassembled and SEM observation of hydroxya, e-tight spherical particles was performed.
No abnormalities such as particle deformation or destruction were observed.

Example 3 C
a(NO3)2 ・4H20 and H3P0. is dissolved in 2-methoxyethanol to give a total solute concentration of 1.0 m
The solution was prepared to be at/t. Using this solution, spray pyrolysis was carried out in the same manner as the spray pyrolysis method used in Example 1 1). The product was mixed with A/2 tight lumpy powder. This product is classified using an air classifier, and
Only 6 μm spherical hydroxya gutite was taken out.

The spherical hydroxyapatite particles obtained by this classification were packed into a stainless steel column (inner diameter 4M, length 100ff) with 4QQ Ka 7cm2 by a slurry filling method, and an evaluation test as a filler was conducted in the same manner as in Example 2. Ta. The hydroxyaq-tite filler obtained in this example gave good results in terms of mechanical strength, durability, reproducibility, etc., as in Example 2.

Example 4 ca(cH3cOo)2 in 450ml of 6N HNO3
・H2PO4 (content 85.6%) 8 in a solution of H2O220.25f and 300rnl of methanol
A homogeneous solution was prepared by mixing with a solution in which 5.85f was dissolved. This solution was subjected to spray pyrolysis in the same manner as used in 1) of Example 1 to obtain spherical hydroxya Qtite with a size of approximately 1-25 μm. This product was classified with an air classifier, hydroxyapatite particles with a particle size of 5 to 15 μm were taken out, and after heat treatment at 600°C for 2 hours, column filling and liquid chromatography evaluation test were carried out in the same manner as in Example 3. I did it.

The shape of the hydroxya IQtite particles obtained in this example was not as well spherical as the particles obtained in Example 3, and the spherical particles were somewhat distorted and had irregularities. However, the same results as in Example 3 were obtained regarding the chromatographic properties.

Example 5 C
Dissolve a(NO3)2.4H20 and (C2H50)3P in ethanol to make the total solute concentration 1 mot/L.
A solution was prepared, and 34 at of ammonia water (concentration 28%) was added to 500 m of this solution and mixed to obtain a uniform solution. This solution was then used to carry out spray pyrolysis in the same manner as used in Example 1, 1) to obtain a product consisting of spherical hydroxya-tite with a size of approximately 1-8 μm. . 80% of this product
After heat treatment at 0°C for 1 hour, 500 K97
By applying a filling pressure of cm2, the inner diameter is 4rtrm and the length is 50.
It was packed into an R stainless steel column.

Using this column, linear gradient method (gradient = 3.49 x 10-2M/rrtl)
Cytochrome C was measured using a pH 6,8 sodium phosphate buffer at a liquid flow rate of 0.5 me/min. A chromatogram obtained during this measurement is shown in FIG. Cytochrome C contains at least two types of iron atoms, one in an oxidized state and one in a reduced state, so the two clearly separated peaks in Figure 1 are due to these two types. This is presumed to be due to the molecular species (the peak on the left is due to oxidized molecules, and the peak on the right is due to reduced molecules), and it can be seen that the column of this example has a high degree of separation.

Example 6 C was added so that the atomic ratio of cal/ram and phosphorus was 1.67.
A solution was prepared by dissolving a(NO3)2.4H2o and H3PO3 in methanol so that the total solute concentration was 0.5rnO1/l. This solution was then subjected to spray pyrolysis in the same manner as used in Example 1, 1) to obtain a product consisting of spherical hydroxyanoqtite with a size of approximately 1-9 μm. This product was heat treated at 600° C. for 2 hours and then packed into a 5Qwpr column in the same manner as in Example 5. Cytochrome C was measured using this column in the same manner as in Example 5, and showed the same high degree of separation as in Example 5.

Example 7 Calcium removal/C so that the atomic ratio of
Dissolve a(NO3)2.4H20 and (CH30)3F in methanol to prepare a solution so that the total solute concentration is 1m0t/l, and add aqueous ammonia (concentration 28 parts) to 1t of this solution. 68 pieces were added and mixed to form a homogeneous solution. Then, using this solution, 1 of Example 1
) Spray pyrolysis was carried out using the same method used in
．． A product consisting of spherical hydroxya 2-tite with a size of 5-5 μm was obtained. After classifying this product using an air classifier to remove particles of 1 μm or less, 3) of Example 1
500KQ using the same slurry filling method as used in
Applying a filling pressure of 7 cm2, the inner diameter is 4 cm and the length is 30 cm.
It was packed into a rItM stainless steel column.

Using this column, lysozyme was measured under the same conditions as in Example 5.
Example 2 of tight filler, its mechanical strength, reproducibility etc.
Similarly, it was found that it had fairly good chromatographic properties.

Example 8 A solution with a total solute concentration of 0.5 mob/l was prepared in the same manner as in Example 7, except that (C2H50)2POH was used as the phosphorus compound and ethanol was used as the organic solvent. Perform spray pyrolysis to approximately 0.5-3
A product consisting of spherical hydroxoa Q-tites with a size of μm was obtained. Classification, filling, and measurement of this product were performed in the same manner as in Example 7. As a result, good chromatographies similar to Example 7 were obtained.

[Brief explanation of drawings]

FIG. 1 is a diagram showing a chromatogram of cytochrome C obtained using the packing material for chromatography of the present invention obtained in one example (Example 6) of the present invention.

Claims (1)

[Claims] 1. Water or a water-compatible organic solvent is used alone as a solvent, or a mixed solvent of a water-compatible organic solvent and water is used as a solvent; A raw material solution is prepared by adding and mixing a calcium compound and a phosphorus compound that are soluble in a solvent, and the raw material solution is sprayed into a flame or a heating zone to form microspheres produced by a pyrolysis reaction. A chromatography packing material made of hydroxyapatite. 2. Use water or a water-compatible organic solvent alone as a solvent, or use a mixed solvent of a water-compatible organic solvent and water as a solvent, and remove calcium soluble in the used solvent. A compound and a phosphorus compound are added and mixed to prepare a raw material solution, and the raw material solution is sprayed into a flame or a heating zone to further heat-treat the microspherical hydroxyapatite produced by a thermal decomposition reaction. A packing material for chromatography consisting of hydroxyapatite particles obtained by