JPH0616843B2 - Pyrogen adsorbent - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to water effective for removing a substance that causes fever when administered into a living body by injection or the like, that is, a pyrogen (pyrodiene). It relates to an insoluble pyrogen adsorbent.

[Conventional Technology and Problems] In the production of pharmaceuticals to be administered in vivo, the technology of removing pyrogens is very important.

The pyrogens mentioned here are classified into exotoxins secreted by diphtheria and staphylococcus aureus and endotoxins which are components of the cell wall of Gram-negative bacteria such as Escherichia coli. Of these, usually
The problem is the endotoxin of the latter Gram-negative bacterium, whose identity is lipid A, which is the complex of lipids bound to polysaccharides, that is, the glycolipid that is the active center in lipopolysaccharide (LPS). It is known. It is considered that when injections containing such pyrogens are administered into the body, these pyrogens act on the heat centers of the hypothalamus and cause fever. It can lead to fever and sometimes shock death.
Therefore, in manufacturing pharmaceutical products such as injections, a fever test using rabbits is usually conducted to confirm safety,
It is mandatory to make sure that pyrogens are not included.

As a method of removing such a heat-generating substance, a method of adsorbing and removing the heat-generating substance using carbon powder or an ion exchange resin, a method of decomposing and removing the heat-generating substance using an acid or an alkali, and an ultra membrane filter are used. There is known a method of selectively removing these using them.

Further, it has been reported that an imidazole-containing compound such as histamine or a nucleic acid base is bound to a dextran-based gel carrier and applied to the adsorption and removal of a pyrogen. [Minobe et al., Journal of Chromatography, 262 p193-198 (198
3)].

However, the removal of pyrogens in the manufacture of pharmaceutical products is such that the target drug product itself is unstable, or the amount of pyrogens to be removed is very small relative to the amount of the target substance during purification. For the reason, moreover, many technical problems still remain when the above method is applied to the industrial manufacturing level, and it is desired to develop an effective and simple method capable of solving these problems. Has been done.

Under these circumstances, a novel exothermic substance adsorbent characterized by using polyamino acid spherical particles as a carrier and binding an imidazole derivative to the carrier has excellent affinity with the exothermic substance and the hardness of the carrier is high. It has been found to have advantages in the operation of adsorbing exothermic substances based on
039).

However, it is unavoidable that several steps of reaction process are required when preparing a pyrogen adsorbent, and some complexity is involved, and there is a limit to the amount of the imidazole derivative that serves as a ligand. There was a limit to the adsorption capacity.

[Problems to be Solved by the Invention] An object of the present invention is to provide a heat-generating substance adsorbent which is excellent in selective adsorption and can be applied to an industrial production level.

[Means for Solving the Problems] In view of these problems, the inventors of the present invention have conducted further diligent research, and as a result, have modified a compound having an aliphatic group and / or an aryl group at the terminal of the side chain and / or the main chain. It was found that a water-insoluble substance composed of a polyamino acid containing a group or a polyamino acid not containing any modifying group has an extremely high affinity for a heat-generating substance and can specifically adsorb and remove the heat-generating substance from a target substance. The invention was completed.

That is, the exothermic substance adsorbent of the present invention is a water-insoluble substance composed of a polyamino acid which may include only a modifying group having an aliphatic group and / or an aryl group at the end of the side chain and / or the main chain. Is characterized by.

As mentioned above, various methods for removing pyrogens have been tried, but in the process of removing pyrogens in the manufacture of pharmaceuticals, the concentration of the target substance in the sample to be treated is relatively high, but There is a problem that the amount of exothermic substances that should be used is very small, and in many cases it is not possible to adsorb only exothermic substances as you would like. Development of methods is desired. Further, in addition to the above-mentioned specificity, the method for removing a heat-generating substance as a manufacturing process of pharmaceuticals is required to be a method capable of rapid treatment.

The pyrogen adsorbent of the present invention does not use the agarose gel or the dextran-based sepharose gel used in the conventional adsorption technology, but uses a polyamino acid instead of the agarose gel or the dextran-based sepharose gel to greatly improve the affinity with the pyrogen. In addition, a special product having excellent hardness as compared with the conventional polysaccharide gel has made it possible to efficiently and promptly separate and remove the pyrogen from the final purified product of interest.

Examples of the modifying group having an aliphatic group used in the exothermic substance adsorbent of the present invention include an alkyl group represented by a methyl group and a cyclohexyl group, an allyl group such as a vinyl group, an ether group such as a methoxy group, and a carboxyl group. Examples include ester groups represented by ethoxycarbonyl groups, acyl groups such as acetyl groups and cyclohexylcarbonyl groups, alkylamino groups such as methylamino groups and aminoethylamino groups, carbamoyl groups, and mesyl groups.

The modifying group having an aryl group is an aryl group such as a phenyl group, an aralkyl group such as a benzyl group, an ether group such as a phenoxy group or a benzyloxy group, an ester group typified by a benzyloxycarbonyl group, and a benzoyl group. And an amino group bonded to an aromatic compound via an aliphatic spacer such as a benzylamino group, and a toluenesulfonyl group.

The polyamino acid used in the pyrogen adsorbent of the present invention includes an inherently hydrophobic amino acid polymer composed of neutral amino acids such as alanine, phenylalanine, valine and leucine.

In addition to this, a hydrophilic polyamino acid hydrophobized (hydrophilic amino acid polymer having a hydrophobic group introduced) can be used. Examples of such hydrophilic polyamino acid derivatives include those obtained by subjecting acidic polyamino acids such as polyglutamic acid and polyaspartic acid to hydrophobic esterification, for example, alkyl esters of these amino acids, benzyl esters,
Dicyclohexane methyl ester, tetrahydropyran methanol ester and the like can be mentioned, and also basic amino acids such as lysine and ornithine protected with a hydrophobic group, for example, carbobenzoxylated products of such amino acids and carboethoxy. Compounds can be mentioned.

The polyamino acid used in the present invention may be either a polymer composed of a single amino acid or a copolymer composed of a mixture of amino acids.

In addition, examples thereof include those obtained by acylating the terminal amino group of the polyamino acid, such as acetylated products and benzoylated products of polyamino acids, and those reacted with an isocyanate compound, such as carbamide compounds of polyamino acids. You can also

Further, a copolymer of polyamino acid or a polyamino acid derivative with a urethane prepolymer, an epoxy resin or the like can also be used. As a polyamino acid derivative not suitable for the heat-generating substance adsorbent of the present invention, polyglutamic acid whose side chain is amidated with ethylenediamine And a polyglutamic acid derivative obtained by further modifying the above with glutaric acid.

The activity of adsorbing a heat-generating substance can be increased by further introducing a basic functional group into the polyamino acid derivative or the polyamino acid derivative, which is the heat-generating substance adsorbent of the present invention.
Therefore, a polyamino acid derivative hydrophobized by esterification, for example, polyglutamic acid methyl ester, can be hydrophobized by originally aminating the ester portion of lysine, ornithine, etc., which has an amino group in its side chain. Derivatives such as poly (Nε-carbobenzoxylidine) are
By removing the hydrophobic group by a conventionally known method, the ability to adsorb the heat-generating substance can be further increased.

The basic functional group mentioned here is particularly preferably an amino group, and particularly preferably an aliphatic amino group.

That is, an alkylamine represented by ethylami, an aliphatic amine such as ethylenediamine, diethylenediamine, and benzylamine, or an amine bonded to an aromatic compound through a spacer is used. Most of the basic functional groups used in the present invention have pKa6.5.
It has the above acid ionization index values. When these basic functional groups are introduced into the polyamino acid or the polyamino acid derivative, they can be introduced directly or via a spacer. It is also possible to prepare the adsorbent of the present invention, which has excellent affinity with a heat-generating substance, by introducing it through a spacer. Such a spacer needs to be selected according to the polyamino acid or polyamino acid derivative constituting the adsorbent of the present invention. For example, a polyamino acid derivative including a glutamic acid derivative or an aspartic acid derivative has 2 carbon atoms.
Polyvalent amines of ~ 6 are preferred.

There is no restriction on the number of basic functional groups introduced here.

The polyamino acid or polyamino acid derivative used in the adsorbent for the heat-generating substance of the present invention is not particularly limited, but the raw material is easily available, that is, the synthesis is easy, and the chemical modification after particle preparation is easy. In consideration, it is preferable to use, for example, polyglutamic acid ester or polyaspartic acid ester, a heteropolymer partially containing other hydrophobic amino acids, which partially includes glutamic acid ester or aspartic acid ester, hydrophobized polylysine or hydrophobized polyornithine, or hydrophobized polylysine or hydrophobic. A heteropolymer or the like which partially contains a modified polyornithine and is composed of another hydrophobic amino acid is used. For example, in the preparation of the adsorbent of the present invention composed of a glutamic acid derivative and / or an aspartic acid derivative, the target polyamino acid derivative is prepared by using methyl ester, ethyl ester, propyl ester, benzyl ester or the like of these amino acids as raw materials. can do.

The degree of polymerization of the polyamino acid or the polyamino acid derivative is not particularly limited, but 100 or more is preferable from the viewpoint of strength, and 100 to 1000 is generally selected.

The shape of the heat generating substance adsorbent of the present invention is not particularly limited, and may be, for example, any of spherical particles, fibers, films, powders and the like.

In particular, the spherical particles, which are one form of the heat-generating substance adsorbent in the present invention, use polyamino acid (synthetic polyamino acid), which is a completely different material from the conventional spherical particles of the polysaccharide system. The inventors of the present application have previously filed a patent application for such a basic spherical particle of polyamino acid and a method for producing the same (JP-A-62-1728).

In order to prepare the polyamino acid spherical particles which are the heat-generating substance adsorbents used in the present invention, the originally hydrophobic polyamino acid and the hydrophobized polyamino acid derivative are used as raw materials to obtain these spherical particles. You can

That is, a solution of a hydrophobic polyamino acid or a hydrophobized polyamino acid derivative dissolved in an organic solvent is added to an aqueous medium (a medium in which the solution is insoluble or only slightly soluble), and suspended to remove the organic solvent. This forms spherical particles, which may be further modified if desired.

Further, as long as the spherical particles as described above, particularly the form of the hydrophobic polyamino acid derivative is adopted during the preparation of the dispersion,
After preparation of the particles, any spherical particles of the hydrophobic polyamino acid derivative and the hydrophilic polyamino acid derivative (including the amphipathic polyamino acid derivative) can be finally obtained by means such as elimination of the protecting group. In selecting the prepared hydrophobic or hydrophilic polyamino acid derivative particles, the properties of the target substance from which the heat-generating substance is to be removed have a great influence. That is, in general, if the target substance to be purified is hydrophilic, both hydrophilic and hydrophobic polyamino acid derivative particles can be used without particular restrictions, but the target substance is hydrophobic. If so, hydrophilic polyamino acid derivative particles are preferably adopted.

Among the pyrogen adsorbents of the present invention, the organic solvent used in the method for preparing the polyamino acid spherical particles is well soluble in hydrophobic polyamino acids or polyamino acid derivatives and insoluble in water, and has a boiling point higher than that of the aqueous solvent. The lower one is the best. Preferred organic solvents are chloroform, dichloromethane, dichloroethane and halogenated hydrocarbons similar thereto, benzene, and mixed solvents thereof. For such a reaction, a solution obtained by polymerizing a hydrophobic polyamino acid or a polyamic acid derivative from an amino acid or an amino acid derivative in an organic solvent can be used as it is.

The particle size of the spherical particles can be easily controlled by the viscosity of the organic solvent-aqueous medium system and the stirring speed. In general, the higher the concentration of polyamino acid or polyamino acid derivative in the organic solvent, and the lower the viscosity of the aqueous medium system, the larger the particle size of the spherical particles obtained. Particles are obtained. Further, the control becomes easier by adding a viscosity modifier such as partially acetylated polyvinyl alcohol or gelatin.

Furthermore, porous spherical particles can be easily prepared. That is, an organic solvent solution of a hydrophobic polyamino acid or a hydrophobized polyamino acid derivative is incompatible with an organic solvent in which the polyamino acid or polyamino acid derivative that is incompatible with the polyamino acid or polyamino acid derivative is dissolved. Additives that are soluble and have a boiling point higher than those of the organic solvent and the aqueous solvent are added, and the same reaction as described above is performed. Such additives include decalin,
Tetralin, toluene, xylene, ethylbenzene, diethylbenzene, anisole, hexanol, octanol, dibutyl ether, cyclohexane, paraffins, phthalates such as dibutyl phthalate, higher fatty acid esters such as methyl dodecanoate, oleic acid, etc. And higher saturated fatty acids. By adjusting the type and amount of such additives, the pore size corresponding to the molecular weight of the water-soluble polysaccharide is 10 ² to 10 ⁷ , and any porosity with a porosity of 10 to 99%. Spherical particles can be obtained. Particularly preferred particles of the present invention are those having a pore size of 10 ³ to 10 ⁷ . In addition, such a spherical particle can be arbitrarily controlled in size by being prepared according to the method described above. Generally, the particle size of the spherical particles obtained here is in the range of 1 to 300 μm, but it is also possible to granulate in the range of 0.1 to 1 mm by adjusting the viscosity of the reaction solution or the stirring speed. The size of the particles is appropriately selected according to the application, but for example, 5 is used when the column chromatography method is used.
It is preferably in the range of up to 300 μm.

The characteristic of such spherical particles is that they are harder than the conventionally used polysaccharide-based spherical particles. When the infrared absorption spectrum of the spherical particles was measured and analyzed, the β structure was found to exist at least partially, and it is considered that the existence of the β structure further contributes to the hardening of the particles.

The fibrous exothermic substance adsorbent is easily prepared by a conventional method. That is, fibers can be obtained by preparing a solution of polyamino acid or polyamino acid derivative in an organic solvent and spinning the solution in an organic solvent that is incompatible with the polyamino acid or polyamino acid derivative. The thickness of the fiber is appropriately selected depending on the diameter of the spinning nozzle used, but is usually 30 to
Fibers with a diameter of 60 μm are used.

The film-like exothermic substance adsorbent can be easily prepared by a conventional method. Prepare an organic solvent solution of polyamino acid or polyamino acid derivative, cast on a glass plate,
It can be obtained by removing the organic solvent. The thickness of the film is appropriately selected at the time of casting.

The powder of polyamino acid or polyamino acid derivative can be easily prepared by heterogeneous polymerization. That is, a polymerization reaction is carried out using an organic solvent that is compatible with the amino acid N-carboxy anhydride, which is a polymerizable monomer, and is incompatible with the polyamino acid produced, and the polyamino acid or polyamino acid derivative that precipitates is separated. By doing so, powder can be obtained.

It is also possible to surface-treat and use a polyamino acid or a polyamino acid derivative on a porous carrier such as silica or alumina.

For example, a porous carrier surface-treated with the polyamino acid or the polyamino acid derivative can be easily obtained by dispersing the porous carrier in an organic solvent solution of the polyamino acid or the polyamino acid derivative and then separating and drying. The porous carrier used is not particularly limited,
An inorganic carrier such as silica or alumina having a large specific surface area is advantageously used.

Furthermore, it is also possible to use those obtained by coating the surface of fibers made of other materials such as cellulose, polyester, polyacrylonitrile, and polyolefin with polyamino acid or polyamino acid derivative. The fiber used is not particularly limited, but a fiber having a large specific surface area is advantageously used.

The characteristics of such a porous carrier or fiber surface-treated with a polyamino acid or a polyamino acid derivative are that the hard point and the specific surface area are larger than the spherical particles of the polysaccharide type which have been conventionally used like the spherical particles. The point is that the pyrogen can be efficiently and rapidly separated and removed from the target final purified product. In addition, the amount of polymer used is smaller than that of spherical particles, which is economically advantageous.

The operation for removing the heat-generating substance using the heat-generating substance adsorbent can be carried out by any of a chromatographic method using a column and a batch treatment method.

In the case of the column method, the pyrogen adsorbent of the present invention is packed in a column, washed with an appropriate buffer, and then the desired solution containing the pyrogen is passed through to collect the flow-through fraction. Thus, the target substance from which the heat-generating substance has been removed can be obtained.

Further, in the batch method, the exothermic substance adsorbent of the present invention is added to the exothermic substance-containing solution and stirred, and then the adsorbent alone is separated and removed from the liquid to obtain a target substance containing no exothermic substance.

Compared with conventional agarose and dextran-based particles, the adsorbent of the present invention shows almost no swelling property and can be subjected to column chromatography, etc. in an extremely short time, and is industrially used as a high-speed separating agent for pyrogens. Ideal when used for.

The exothermic substance adsorbent of the present invention has an excellent affinity with the exothermic substance, and in addition to this, the adsorbent is extremely hard as compared with the particles using the conventionally used agarose or dextran type substance as a material. Since it is rich in properties and stable, the degree of swelling is extremely small, and it is possible to carry out the operation of removing the heat-generating substance in a short time.

Further, the exothermic substance adsorbent can be prepared very easily without passing through a special reaction route. Further, since the exothermic substance adsorbent of the present invention has a low affinity for the target substance, it is possible to extremely selectively remove only the exothermic substance without lowering the recovery rate of the target substance.

As described above, the factors of the features of the pyrogen adsorbent of the present invention having the extremely optimum function as the pyrogen adsorbent are the effect of the structure of the polyamino acid or the polyamino acid derivative on the adsorption mechanism of the pyrogen, and the polyamino acid. Alternatively, it can be considered as a synergistic effect of the affinity between the basic functional group introduced in the polyamino acid derivative, especially the weakly basic amino group, and the heat generating substance. Further, it is also noteworthy that the types of the basic functional groups can be appropriately selected when the basic functional groups are introduced, and the introduction rate of these can be easily controlled.

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

[Example] Preparation of pyrogen adsorbent Example 1 10 g of methyl poly-L-glutamate, 10 m of decalin
1 is dissolved in 400 ml of dichloroethane, and this is heated at 50 ° C.
Was added dropwise to 2000 ml of an aqueous solution of 1.5 w / v% partially acetylated polyvinyl alcohol. Upon vigorous stirring at the same temperature for 24 hours, dichloroethane was evaporated, and spherical particles of poly-L-glutamic acid methyl ester containing decalin were obtained.

This was washed with acetone by a Soxhlet extraction method to remove decalin, and then suspended in water to obtain spherical particles having a particle diameter of 64 to 105 μm through a JIS standard compliant sieve. The maximum pore diameter of the spherical particles was 2 × 10 ⁴ in terms of the molecular weight of the water-soluble polysaccharide, and the porosity was 55%. The particles thus obtained are referred to as adsorbent A (PG-OMe).

Example 2 5 g of the adsorbent A obtained in Example 1 was added to 50 m of methanol.
1 and 50 ml of ethylenediamine were suspended in the mixed solution and gently stirred at 65 ° C. for 48 hours. The particles were collected by filtration and washed with methanol and water to obtain amino group-introduced particles. The amount of amino groups introduced in this case was 3.2 meq / g (1
It was 3.2 meq / g particle). The particles thus obtained are referred to as adsorbent B (PG-NH ₂ ).

Comparative Example 1 4 g of the adsorbent B obtained in Example 2 was suspended in 100 ml of tetrahydrofuran, 0.14 g of glutaric anhydride was added, and the mixture was gently stirred at 40 ° C. for 24 hours. The particles were collected by filtration and washed with methanol and water to obtain particles having carboxyl groups introduced. By the titration method,
It was confirmed that 98% or more of the amino groups in the particles were carboxylated. The particles thus obtained are adsorbent C (P
G-COOH).

Comparative Example 2 3.5 g of the adsorbent C obtained in Comparative Example 1 and 3 g of histamine dihydrochloride were suspended in 50 ml of water, and triethylamine 4.
5 ml, water-soluble carbodiimide (for example, 1-cyclohexyl-3- (2-morpholinoethyl) carbodiimide-p-toluenesulfonate, etc.) 0.56 g were added,
The mixture was gently stirred at room temperature for 2 days. The formed particles were collected by filtration and washed with water, alcohol and ether in this order. The histamine content in the produced particles was 0.2 meq / g by the amino acid analysis method. The particles thus obtained are referred to as adsorbent D (PG-His).

Example 3 10 g of poly-L-leucine and 10 m of diethylbenzene
1, and 500 ml of bendene were added, and the mixture was stirred at room temperature for several hours for swelling, and then heated to 70 ° C. to obtain a viscous transparent solution. This was dropped into 2000 ml of an aqueous solution of 2.5 w / v% partially acetylated polyvinyl alcohol kept at 60 ° C., and vigorously stirred for 24 hours while gradually lowering the temperature to 40 ° C., benzene was evaporated and polybenzene containing diethylbenzene was contained. Spherical particles of leucine were obtained. This is washed with Soxhlet extraction method using acetone to remove diethylbenzene, and then 10-2 by a JIS standard sieving screen.
Spherical particles with a particle size of 5 μm were obtained. The particles thus obtained are called adsorbent E (PLeu).

Example 4 10 g of poly (Nε-carbobenzoxy-L-lysine) was dissolved in 10 ml of diethylbenzene and 400 ml of chloroform, and the solution was added dropwise to 2000 ml of an aqueous solution of 1.5 w / v% partially acetylated polyvinyl alcohol kept at 40 ° C. did. Upon vigorous stirring at the same temperature for 24 hours, chloroform was evaporated and spherical particles of poly (Nε-carbobenzoxy-L-lysine) containing diethylbenzene were obtained.
After this was washed with acetone by a Soxhlet extraction method to remove diethylbenzene, spherical particles having a particle size of 64 to 105 μm were obtained through a JIS standard compliant sieve.

To 10 g of the particles, 30 ml of 25% hydrogen bromide / acetic acid and 300 ml of ether were added and suspended at room temperature for 1 hour to obtain poly-L-lysine particles in which carbobenzoxy groups were partially removed. The particles thus obtained are used as adsorbent F
(PLys).

Example 5 1.0 g of L-leucine N-carboxyanhydride and 0.2 g of L-isoleucine N-carboxyanhydride were dissolved in 15 ml of acetonitrile and heated to 40 ° C. When 4 μl of triethylamine was added, a powdery polymer started to precipitate.

After 6 hours, the mixture was cooled to room temperature, the precipitated powdery polymer was separated by filtration and dried.

0.9 g of poly (L-leucyl-L-isoleucine) was obtained.

The particles thus obtained are referred to as adsorbent G (PLeu-Ile).

Example 6 A 10 wt% dichloroethane solution of methyl poly-L-glutamate was heated to 40 to 50 ° C. and defoamed under reduced pressure.

The above spinning dope was added to a spinning syringe and discharged into kerosene at a constant rate. Fiber was obtained by drawing out poly-L-methyl glutamate coagulated at the tip of the nozzle of the syringe into a coagulation bath and winding it with a bobbin rotating at a constant speed.

The fibers thus obtained are adsorbent H (fibrous PG-OME)
Called.

Example 7 1.4 g of methyl L-glutamate N-carboxyanhydride
Was dissolved in 10 ml of ethyl acetate and heated to 40 ° C. When 5 μl of triethylamine was added, methyl poly-L-glutamate began to precipitate. Cool to room temperature after 4 hours,
The precipitated methyl poly-L-glutamate was filtered off and dried.

1 g of methyl poly-L-glutamate was obtained. The particles thus obtained are referred to as adsorbent I (powder PG-OME).

Example 8 1 g of poly-L-glutamate methyl was added to dichloroethane 1
Dissolve in 00 ml, porous silica (particle size 0.5 ~ 11μ
m) 1 g was added, and the mixture was gently stirred at room temperature for 2 hours. It was degassed for 5 minutes under reduced pressure, and the inside of the porous silica was replaced with the polymer solution.

After repeating the above operation twice, the porous silica was filtered off, washed with methanol and then dried.

1 g of porous silica surface-treated with methyl poly-L-glutamate was obtained.

The particles thus obtained are adsorbent J (surface-treated PG-OM
E).

Adsorption removal of heat-generating substances Example 9 Various adsorbents prepared in Examples 1 to 4 and Comparative Examples 1 and 2, and commercially available adsorbents (Company A and Company B) 0.1 to 0.1, respectively.
0.2 g (dry weight) of pyrogen-free 10 mM phosphate buffer pH 7.5 (containing 1.0 M NaCl), 1
0 mM phosphate buffer pH 7.5 (0.07M NaC
It was sequentially washed with 1). These adsorbents were brought into contact with 1 to 2 ml of various bacterial toxoid solutions containing pyrogens, and the amount of pyrogens remaining in the eluate or supernatant was measured.

The amount of the heat-generating substance remaining in the eluent or the supernatant was subtracted from the amount of the heat-generating substance initially contained, and the value was divided by the amount of the heat-generating substance initially contained and multiplied by 100 to express the adsorption rate. The results are shown in Table 1.

As a result, it was revealed that the adsorbent A itself has a considerable ability to adsorb a heat-generating substance, and the weakly basic amino group is introduced into the side chain via a spacer to further increase the adsorbability in the adsorbent B.

On the other hand, the adsorbent C having a carboxyl group introduced into the side chain shows almost no adsorbability of a heat generating substance, and it is considered that the carboxyl group suppresses the heat generating substance adsorption ability.

Imidazole derivative (histidine) on amino acid spherical particles
Although the adsorbent D having immobilized thereon has the same exothermic substance adsorbing ability as that of the adsorbent A, it does not exceed the adsorbent B. In addition, when an adsorbent of Company A having chitosan as an adsorbent body and an adsorbent of Company B in which sepharose is used as a carrier and imidazole is bound via a spacer, some effects can be expected in the present measurement system, but it is expected. No good adsorption capacity was observed.

Further, the adsorbent E having a particle body of polyleucine, which is a hydrophobic polyamino acid, or the adsorbent F having an amino group on its side chain has adsorbing activity of a heat-generating substance. Supporting results were obtained.

Example 10 0.05 g (dry weight) of adsorbent A prepared in Example 1
Then, 0.05 g (dry weight) of the adsorbent E prepared in Example 3 was mixed and treated in the same manner as in Example 9 to measure the amount of the remaining exothermic substance.

The adsorption rate was determined by the same method as in Example 9. As a result, a value of 81% was obtained as the adsorption rate of the exothermic substance.

This value was almost the same as the result of evaluating Adsorbent A and Adsorbent E under the above conditions.

Example 11 0.1 g (dry weight) of each of the various adsorbents prepared in Examples 5 to 8 was added to a pyrogen-free 10 mM phosphate buffer solution pH.
The cells were sequentially washed with 7.5 (containing 1.0 M NaCl) and 10 mM phosphate buffer (pH 7.5) (0.07 M NaCl). These adsorbents have various bacteria-derived pyrogens 200-
250 ng dissolved phosphate buffer (pH 7.2) 1-
The eluent or the supernatant was contacted with 2 ml and the amount of the remaining pyrogen was measured.

The adsorption rate was determined by the same method as in Example 9. The results are shown in Table 2.

Excellent adsorbing activity of exothermic substances was observed in all of the various adsorbents.

In addition, the detection of the exothermic substance was performed by a toxinometer (made by Wako Pure Chemical Industries, Ltd.).

[Advantages of the Invention] The exothermic substance adsorbent of the present invention has an effect of extremely selectively removing only the exothermic substance without lowering the recovery rate of the target substance.

Further, its preparation method is also very easy, and it is an excellent adsorbent applicable to industrial production of pharmaceuticals.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Hirotaka Ihara 854-2 Takahira, Shimizu-cho, Kumamoto-shi, Kumamoto Prefecture (72) Inventor Toshinori Tsunoda 2136-58 Ki-mizu, Shimizu-cho, Kumamoto-shi, Kumamoto Prefecture (72) Inventor Katsutoshi Aihara 970-8 Iwano, Ueki-machi, Kumamoto-gun, Kumamoto Examiner Koji Amamiya

Claims (1)

[Claims] 1. A water-insoluble exothermic substance adsorbent composed of a polyamino acid which may contain only a modifying group having an aliphatic group and / or an aryl group at the end of a side chain and / or a main chain.