JPH0557015A - Adsorbent and sterilizing method therefor - Google Patents

Abstract

PURPOSE:To provide a blood purification adsorbent having a sulfated polysaccharide on the surface thereof for adsorbing and removing a harmful ingredient from blood, and a sterilizing method of high safety therefor, ensuring freedom from a drop in adsorption and elimination capability due to high pressure steam sterilization. CONSTITUTION:A blood purification adsorbent having a sulfated polysaccharide on the surface thereof is sterilized under high pressure steam in water solution containing 0.001 to 10% of pyrosulfurous acid and/or a salt thereof and having a pH value controlled within the range of 5 to 9.5.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an adsorbent for blood purification having on its surface a sulfated polysaccharide for adsorbing and removing harmful components in blood, and a sterilizing method thereof.

[0002]

2. Description of the Related Art For the purpose of removing harmful components in blood, a substance having an affinity for the harmful components, that is, an adsorbent in which a ligand is immobilized on a water-insoluble carrier by a covalent bond, is clinically utilized. Attempts have been made.

Sterilization is indispensable for clinical use of these adsorbents. As the sterilization method, high-pressure steam sterilization is preferable from the viewpoints of handleability and safety. However, high pressure steam sterilization of these adsorbents causes deactivation and desorption of ligands,
As a result, the removal performance of the adsorbent is greatly reduced, and there is a problem in safety in clinical practice. Particularly, in the case of sulfated polysaccharides such as dextran sulfate, the covalent bond between the sulfated polysaccharide and the water-insoluble carrier is cleaved by heat during high-pressure steam sterilization, the sulfated polysaccharide molecule itself is decomposed, and occurs in the process of its decomposition. It is considered that the sulfuric acid compound further accelerates the self-decomposition.

By the way, high pressure steam sterilization of an adsorbent having a sulfated polysaccharide on its surface is carried out by citric acid, phosphoric acid, acetic acid, boric acid,
There has been an attempt to reduce the decomposition of sulfated polysaccharides by carrying out in a buffer solution of at least one of tartaric acid, carbonic acid, maleic acid and glycine at pH 5 to 9 (JP-A-61-100261). ). However, although the above-mentioned method can prevent self-decomposition due to pH decrease due to the generated sulfuric acid compound, it does not prevent decomposition due to heat itself, and there is a considerable decrease in performance and desorption of ligand, which is a serious problem. there were.

[0005]

DISCLOSURE OF THE INVENTION As a result of intensive studies conducted by the present inventors to find a better sterilization method, it was found that pyrosulfite or / And its salt 0.00
We have found an epoch-making fact that the removal performance of adsorbents can be suppressed from decreasing by carrying out in an aqueous solution containing 1 to 10%, and we invent an adsorbent that is free from deterioration of removal performance and excellent in safety. Came to.

[0006]

That is, the gist of the present invention is to provide pyrosulfite or / and its salt in an amount of 0.001-10.
%, And high-pressure steam sterilization in an aqueous solution whose pH is adjusted to a range of 5 to 9.5, and a method for sterilizing an adsorbent for blood purification having a sulfated polysaccharide on its surface, and this sterilization In the adsorbent for blood purification obtained by the method.

According to the results of various studies by the present inventors,
Pyrosulfurous acid and / or its salt have an effect of preventing both the cleavage of the covalent bond between the sulfated polysaccharide and the water-insoluble carrier due to heat during high-pressure steam sterilization and the decomposition of the sulfated polysaccharide molecule itself. it is conceivable that.

To explain the present invention in more detail, it is desirable that the concentration of pyrosulfite or / and its salt is 0.001 to 10%. More preferably, the range is 0.01 to 2%. At this time, depending on the alkaline substance,
A value of 9.5 is effective for preventing self-decomposition at low pH, and an excellent sterilization method can be achieved only by combining these two.

The pH of 5 to 9.5 means that the pH of the aqueous solution immediately after sterilization is 5 to 9.5. The pH before sterilization is preferably 5 to 9.5, but is not necessarily limited to the case where the adsorbent is immersed in an aqueous solution and then rapidly sterilized. However, the pH before sterilization is preferably 5 or higher because the sulfated polysaccharide is rapidly decomposed at pH 4.5 or lower.

To adjust the pH of the aqueous solution to 5 to 9.5, alkali metal or alkaline earth metal carbonates, hydroxides, or hydrogen carbonate compounds, such as sodium carbonate, sodium hydrogen carbonate, sodium hydroxide, This can be done by adding a basic substance such as potassium hydroxide into the aqueous solution. The optimum concentration of the basic substance at this time varies depending on the concentration of pyrosulfurous acid or / and its salt and cannot be specified. However, usually 0.001 to 5% can be preferably used, and if a particularly preferable range is mentioned, For example, it is 0.005 to 1.5%.

According to the research conducted by the present inventors, sodium carbonate and sodium hydrogencarbonate were the best among these, considering the handling property. The most preferable examples are 0.01-0.2% sodium pyrosulfite and 0.005-0.2% sodium carbonate.

The adsorbent for blood purification having a sulfated polysaccharide on its surface as used in the present invention is an adsorbent in which a sulfated polysaccharide is insolubilized in a water-insoluble carrier through a covalent bond. As the shape of the water-insoluble carrier, any of spherical shape, granular shape, thread shape, hollow fiber shape, flat membrane shape and the like can be effectively used. Among these, spherical or granular ones are most preferable because they can have a large adsorption area per unit volume.

The spherical or granular average particle size is 10 to 2,5.
It is easy to use the one of 00 μm, but it is 25-1,000 μm
The range of is preferable. The water-insoluble carrier is preferably porous from the viewpoint that a large adsorption area for harmful components can be obtained and a practical adsorption capacity can be obtained. The exclusion limit molecular weight of porosity depends on the molecular weight of the harmful component to be adsorbed, but is preferably 1 × 10 ⁴ or more and 1 × 10 ⁸ or less. To further limit, 1 × 10 ⁵ or more,
It is more preferably 5 × 10 ⁷ or less.

Specific examples of the water-insoluble carrier include, for example, one of soft gels made of agarose, dextran, polyacrylamide, etc., methyl methacrylate, polyvinyl alcohol, styrene, divinylbenzene, vinyl ether, maleic anhydride, polyamide, etc. Alternatively, a synthetic polymer having a plurality of constituents, and / or a hard gel made of a porous polymer obtained from a natural polymer such as cellulose as a raw material. Further, a polymer material having a hydroxy group such as hydroxyethyl methacrylate and hydroxyethyl acrylate, a monomer having a basic nitrogen-containing functional group such as vinylamine and dimethylaminoethyl (meth) acrylate, and a basic nitrogen-containing functional group are included. A copolymer with a polymerizable monomer, a polymer material having a negatively charged functional group such as a sulfonic acid group or a carboxylic acid group, a segmented polyurethane, a block copolymer such as a segmented polyester, or a polyethylene oxide chain. It may have a surface layer such as a graft copolymer such as a copolymer of the monomer having it and another polymerizable monomer.

Of these, hard gels are preferably used from the viewpoint of the flowability of body fluids. Still more preferably, a synthetic polymer hard gel made of polyvinyl alcohol or the like is preferable for practical use because an active group can be relatively easily obtained on the gel surface. The active group on the surface of the water-insoluble carrier that is covalently bonded to the sulfated polysaccharide may be any group that can be substituted or / and addition-reacted with a reactive group having an active hydrogen such as a hydroxyl group or a carboxyl group of the sulfated polysaccharide.

Known examples of methods for obtaining an active group on a water-insoluble carrier include a cyanogen halide method, an epichlorohydrin method, a bisepoxide method, and a bromoacetyl bromide method. Specific examples thereof include an amino group, a carboxyl group, a hydroxyl group, a thiol group, an acid anhydride group, a succinylimide group, a chlorine group, an aldehyde group, an amide group and an epoxy group.

Of these, an epoxy group derived from the epichlorohydrin method or the like is particularly preferable because of its stability during heat sterilization. The sulfated polysaccharide covalently bound through the above active group is a polysaccharide having a sulfate group in its molecule, heparin, dextran sulfate, chondroitin sulfate, chondroitin polysulfate, heparan sulfate,
Keratan sulfate, heparitin sulfate, xylan sulfate, caronine sulfate, cellulose sulfate, chitin sulfate, chitosan sulfate,
Examples thereof include pectin sulfate, inulin sulfate, alginate sulfate, glycogen sulfate, polylactose sulfate, carrageenin sulfate, starch sulfate, polyglucose sulfate, ramillarin sulfate, galactan sulfate, levan sulfate, and mepesulfate. Of these, heparin and dextran sulfate are the most preferable examples from the viewpoint of clinical practicality.

The molecular weight of these sulfated polysaccharides may be any molecular weight, but if it is dared to say, a molecular weight of 5,000 or more is particularly preferable from the viewpoint of performance as a ligand.

The above-mentioned adsorbent for blood purification having a sulfated polysaccharide on the surface, which is filled in an aqueous solution containing 0.001 to 10% of pyrosulfurous acid and / or its salt, has a low density or / And extremely low density lipoprotein, sulfatide adhesion protein, activated complement component, amyloid protein A, immune complex, autoantibody such as anti-DNA antibody and rheumatoid factor, and / or immune B cell producing the autoantibody, immunity Globulin L chain, blood coagulation factor VIII, blood coagulation factor IX
Factor, β ₂ microglobulin and the like. Among them, the use of low density and / or very low density lipoprotein as an adsorbent is highly clinically useful.

[0020]

By sterilizing the present invention in an aqueous solution containing 0.001 to 10% of pyrosulfurous acid and / or a salt thereof, it is possible to prevent the deterioration of the performance which has occurred during the sterilization operation up to now, and the ligand It is possible to obtain an adsorbent of excellent quality with little desorption of sulfated polysaccharides.

[0021]

EXAMPLES The present invention will be described below with reference to specific examples, but the present invention is not limited thereto.

Example 1. Using triallyl isocyanurate as a cross-linking agent, suspension polymerization of vinyl acetate in a phosphate buffer aqueous solution was performed to obtain a polyvinyl alcohol copolymer.
The obtained polyvinyl alcohol copolymer was used as a water-insoluble carrier. This water-insoluble carrier has an average particle size of 100.
μm, vinyl alcohol unit per unit weight (qO
H) was 6.0 meq / g, and the exclusion limit molecular weight was 5 × 10 ⁶ .

After transesterification and thorough washing with water, epichlorohydrin was reacted in a mixed solution of dimethyl sulfoxide and an aqueous sodium hydroxide solution to obtain an epoxy activated carrier. This epoxy-activated carrier has a molecular weight of 500,000 and a sulfur content of 1 via an epoxy group.
A dextran sulfate-immobilized adsorbent was obtained by covalently bonding 8.2 wt% of dextran sulfate. The amount of dextran sulfate fixed on this adsorbent was 5.
It was 9 mg.

10 ml of this adsorbent was taken and subjected to high-pressure steam sterilization at 121 ° C. for 30 minutes in 100 ml of an aqueous solution containing 0.02% of sodium pyrosulfite and 0.016% of sodium carbonate. At this time, pH before and after sterilization and 1 ml of adsorbent
The fixed amount of dextran sulfate and the adsorption / removal performance of cholesterol were measured. The cholesterol adsorption performance was measured using human plasma with a cholesterol concentration of 410 mg / d1. The results are shown in Table 1.

[0025]

[Table 1]

Example 2. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1 was used as an aqueous solution 1 containing sodium pyrosulfite 0.06% and sodium carbonate 0.03%.
High-pressure steam sterilization was carried out for 30 minutes at 121 ° C. in 00 ml, and the adsorption performance was measured as in Example 1. The results are shown in Table 1.

Example 3. Example 1 10 ml of the obtained dextran sulfate-fixed adsorbent was added to an aqueous solution 1 containing 0.05% sodium pyrosulfite and 0.06% sodium carbonate.
High-pressure steam sterilization was carried out for 30 minutes at 121 ° C. in 00 ml, and the adsorption performance was measured as in Example 1. The results are shown in Table 1.

Example 4. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1 was used as an aqueous solution 1 containing sodium pyrosulfite 0.24% and sodium carbonate 0.09%.
High-pressure steam sterilization was carried out for 30 minutes at 121 ° C. in 00 ml, and the adsorption performance was measured as in Example 1. The results are shown in Table 1.

Example 5. 10 ml of an aqueous solution containing 0.12% of sodium pyrosulfite and 0.06% of sodium carbonate was added to 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1.
Perform high-pressure steam sterilization in 0 ml at 121 ° C for 30 minutes,
The adsorption performance was measured in the same manner as in Example 1. The results are shown in Table 1.

Example 6. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1 was treated with an aqueous solution 1 containing sodium pyrosulfite (1.00%) and sodium carbonate (0.50%).
High-pressure steam sterilization was carried out for 30 minutes at 121 ° C. in 00 ml, and the adsorption performance was measured as in Example 1. The results are shown in Table 1.

Example 7. A cellulose gel having an average particle size of 80 μm and an exclusion limit molecular weight of 5,000,000 (manufactured by Chisso Corporation) was used as a water-insoluble carrier, and a dextran sulfate-fixed adsorbent was obtained in the same manner as in Example 1.

The amount of dextran sulfate fixed on this adsorbent was 0.62 mg per 1 ml of the adsorbent. Then, 10 ml of the dextran sulfate-fixed adsorbent was subjected to high-pressure steam sterilization at 121 ° C. for 30 minutes in 100 ml of an aqueous solution containing sodium pyrosulfite 0.12% and sodium carbonate 0.06%, and the adsorption performance was the same as in Example 1. Was measured. The results are shown in Table 1.

Comparative Example 1. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1 was used as an aqueous solution 1 containing 0.24% of sodium pyrosulfite and 0.06% of sodium carbonate.
High-pressure steam sterilization was carried out for 30 minutes at 121 ° C. in 00 ml, and the adsorption performance was measured in the same manner as in Example 1. The results are shown in Table 1.

Comparative Example 2. Using 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1, potassium dihydrogen phosphate 0.27%, disodium hydrogen phosphate 0.67%
High-pressure steam sterilization was carried out at 121 ° C. for 30 minutes in 100 ml of an aqueous solution containing, and the adsorption performance was measured as in Example 1. The results are shown in Table 1.

Comparative Example 3. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 1 was autoclaved at 121 ° C. for 30 minutes in 100 ml of an aqueous solution containing 0.65% potassium dihydrogen phosphate and 0.29% disodium hydrogen phosphate. The adsorption performance was measured in the same manner as in Example 1. The results are shown in Table 1.

Comparative Example 4. 10 ml of the dextran sulfate-fixed adsorbent obtained in Example 7 was autoclaved at 121 ° C. for 30 minutes in 100 ml of an aqueous solution containing 0.27% potassium dihydrogen phosphate and 0.67% disodium hydrogen phosphate. The adsorption performance was measured in the same manner as in Example 1. The results are shown in Table 1.

Sodium pyrosulfite is present and pH
In the range of 5 to 9.5, there was almost no change in dextran sulfate fixed amount and cholesterol adsorption / removal performance before and after sterilization. On the other hand, in the example in which sodium pyrosulfite was not present, both the fixed amount of dextran sulfate and the adsorption / removal performance of cholesterol were decreased before and after sterilization even when the pH was in the range of 5 to 9.5.
Furthermore, even if sodium pyrosulfite is present, the pH is 5-9.
When it was not within the range of 5, the dextran sulfate fixed amount and the cholesterol adsorption / removal performance were also decreased before and after sterilization.

Claims (8)

[Claims] 1. Pyrosulfurous acid or / and its salt is 0.0
A method for sterilizing an adsorbent for blood purification having a sulfated polysaccharide on the surface, which comprises performing high-pressure steam sterilization in an aqueous solution containing 01 to 10% and having a pH adjusted to a range of 5 to 9.5. 2. The method for sterilizing an adsorbent according to claim 1, wherein the aqueous solution contains 0.001 to 5% of at least one alkaline substance. 3. The method for sterilizing an adsorbent according to claim 2, wherein the alkaline substance is at least one compound selected from alkali metals and alkaline earth metals. 4. The method for sterilizing an adsorbent according to claim 2, wherein the alkaline substance is at least one of sodium carbonate, sodium hydrogen carbonate, sodium hydroxide and potassium hydroxide. 5. The amount of pyrosulfurous acid and / or its salt added to 0.0
High pressure steam sterilization was performed in an aqueous solution containing 01 to 10% and having a pH adjusted to a range of 5 to 9.5,
An adsorbent for blood purification having a sulfated polysaccharide on the surface. 6. The adsorbent according to claim 5, wherein the aqueous solution contains 0.001 to 5% of at least one alkaline substance. 7. The adsorbent according to claim 6, wherein the alkaline substance is at least one compound selected from the group consisting of alkali metals and alkaline earth metals. 8. The adsorbent according to claim 6, wherein the alkaline substance is at least one of sodium carbonate, sodium hydrogen carbonate, sodium hydroxide and potassium hydroxide.