JPH04129567A - Regeneration liquid and regeneration method for adsorbent - Google Patents

Abstract

PURPOSE:To obtain the regeneration liquid, regeneration method and ex vivo circulation system suitable for the regeneration of the adsorbent of an in vivo high molecular substances by using a buffer solution containing one kind of a compound selected among a group of salt with high chaotropisity, urea and glycol. CONSTITUTION:Salt with high chaotropisity 3mol or below is preferably used for a buffer solution 1l, urea 8mol or below for the buffer solution 1l, and glycol 50wt.% or below for the buffer solution 1l to obtain a regeneration liquid. A phosphoric acid buffer solution is used for the buffer solution, for example. For the method to regenerate a used adsorbent with the regeneration liquid, the adsorbent is floated and stirred in the regeneration liquid, the adsorbed in vivo high molecular substances is moved to the regeneration liquid, then the regeneration liquid is removed by filtering. In an ex vivo circulation system incorporated with a column of the adsorbent, a passage is switched, and the regeneration liquid can be fed to the column of the adsorbent.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention relates to a regenerating liquid used for regenerating a biopolymer adsorbent;
The present invention relates to a regeneration method and an extracorporeal circulation system.

The regeneration solution and regeneration method of the present invention can be applied to biopolymers such as the following, disease-causing factors such as rheumatoid factors, anti-DNA antibodies, and anti-acetylcholine antibodies, or pharmaceutical intermediates and drugs.
Alternatively, it is particularly suitable for regenerating an adsorbent for the purpose of adsorbing pyrodiene and the like.

[Prior art and its problems]

Currently, various adsorbents and extracorporeal circulation systems have been developed to remove factors causing diseases such as autoimmune diseases. However, in actual clinical practice, the problem is that the adsorbent itself has a small adsorption capacity and cannot sufficiently remove disease-causing factors. Furthermore, since the adsorbent itself is a disposable product, there is also the problem of increasing the financial burden on patients.

In order to solve this problem, attempts have first been made to increase the adsorption capacity of the adsorbent itself.

However, since there is a limit to the amount of extracorporeal circulation in a patient, the adsorption capacity cannot be increased significantly, so this did not fundamentally solve the problem.

Therefore, attempts have been made to regenerate the adsorbent itself by washing it with a salt solution or buffer solution. However, since the adsorption mechanism between the adsorbent and the adsorbed substance is often based on cooperative effects such as hydrophobic bonds, electrostatic bonds, and hydrogen bonds, it is not possible to completely regenerate the adsorbent using a buffer solution alone. could not.

Among them, only L D L (Low Density
Lipoprotein) and VLDL (νery L
An extracorporeal circulation system has been put into practical use that adsorbs and removes (Density Lipoprotein) with immobilized dextran sulfate and regenerates the immobilized dextran sulfate by washing it with saline. However, this does not mean that LDL or V
The extremely high electrostatic interaction between LDL and dextran sulfate is destroyed with saline, and adsorbed LDL and VLDL are removed. Therefore, substances adsorbed to the adsorbent through hydrophobic bonds and hydrogen bonds, such as disease-causing factors, could hardly be removed by saline.

[Problems to be solved by the present invention] The present invention provides a regenerating solution suitable for regenerating a biopolymer adsorbent;
The purpose is to provide a regeneration method and an extracorporeal circulation system.

Specifically, the following biopolymers, disease-causing factors contained in patient's body fluids such as blood and plasma, such as rheumatoid factors, anti-DNA antibodies and anti-acetylcholine antibodies, pharmaceutical intermediates and drugs, or pyrogens. It is an object of the present invention to provide a regeneration liquid, a regeneration method, and an extracorporeal circulation system that are particularly suitable for regenerating adsorbents for the purpose of adsorption, such as .

[Configuration of the Present Invention] The regenerating solution of the present invention is a regenerating solution consisting of a buffer solution containing at least one compound selected from the group consisting of salts with high chaotropicity, urea, or glycols.

The regenerating solution of the present invention must contain at least one of salts with high chaotropicity, urea, or glycols. And it is more preferable that it contains any two or more of these, and it is most preferable that it contains all three of these.

In the regenerating solution of the present invention, in the case of a salt with high chaotropicity, the amount is 3 m with respect to buffer solution 11.

! Below, in the case of urea, the ratio is 8 m to buffer solution 11.

! or less, and in the case of glycols, it is preferably 50 or less with respect to the buffer if.

Salts with high chaotropicity refer to salts of monovalent anions with a large ionic radius, specifically 5NC-1-,
It is a salt having anion such as Cl0a-, NO2-1, Br-, etc. In the present invention, among these salts, alkali metal salts, alkaline earth metal salts, ammonium salts, amine salts, quaternary ammonium salts, etc. are preferred. Particularly preferred are salts, Na salts, K salts, Ca salts, and ammonium salts.

Glycols include ethylene glycol, propylene glycol, 1,3-propanediol, glycerin, butylene glycol, 1.3-butanediol, 1,4
-butanediol, l.

2-bentanediol, 1,3-bentanediol, 1
．． 4-bentanediol, 1.5-bentanediol,
Polyethylene glycol, polypropylene glycol, etc. can be used. Among these, ethylene glycol, propylene glycol, polyethylene glycol,
Alternatively, polypropylene glycol is preferred. Particularly preferred are ethylene glycol and propylene glycol.

A normal buffer solution can be used.

For example, phosphate buffer, acetic acid-sodium acetate buffer, Menzel's buffer, Michaelis buffer, Sørensen's buffer, Clark-Laps buffer, pinequilvaine buffer, Britton-Robinson buffer, etc. are used.

A method for regenerating a used adsorbent using the regenerating liquid of the present invention is to suspend the adsorbent in the regenerating liquid, stir it, transfer the adsorbed biopolymer to the regenerating liquid, and then filtrate, etc. A method of removing the regenerating liquid is possible.

Alternatively, the adsorbent after use may be packed in a column, and the regeneration liquid may be passed through the column.

Furthermore, in an extracorporeal circulation system incorporating an adsorbent column, the flow path may be switched to allow the regenerated liquid to flow through the adsorbent column. In this extracorporeal circulation system, as shown in FIG. 2, it is preferable to use two or more adsorbent columns.

When one column processes the patient's body fluid and the adsorbent reaches equilibrium adsorption, the column can be switched and the other column processes the patient's body fluid, and the column that has reached adsorption equilibrium can be regenerated by passing the regenerating liquid. This is because the patient's body fluids can be processed continuously.

〔Example〕

Hereinafter, the present invention will be specifically explained with reference to Examples.

A reference flame-crosslinked agarose (trade name: Sepharose CL-4B (manufactured by Pharmacia)) was repeatedly washed with distilled water, and the water was thoroughly squeezed out by suction filtration.

Dimethyl sulfoxide (DM
SO) 50d and stirred at room temperature for 2 hours. After removing DMSO in a moisture-free system, newly dehydrated DMS
030d was added and stirred at room temperature for 1 hour. Similarly below, 3
0mj! (L30mj! (3 hours) operation was repeated for 12.5 hours, and finally DMSO20- was added. At this time, the amount of water in DMSO in the system was analyzed by Karl Fischer method, and it was found to be about 10 ppm.

In this system, hexamethylene diisocyanate [・(HMDI
) was added to 10 d of dehydrated DMSO, and the mixture was stirred at 100°C for 2 hours to carry out the reaction. After removing the reaction solution, freshly charged MSO25d and stirred at room temperature for 1.
Washing was performed by stirring for 5 hours.

After that, 25me (1.5 hours), 20ml1 (1
, 5 hours), 2O- (1,5 hours), 20 mfl (
11.5 hours), 20m1! (2.5 hours).

Titration analysis of isocyanate groups in the final 20 washes was performed, but no isocyanate groups were detected.

Thereafter, 30-liters of water was added to stop the reaction. The reaction product was suction filtered, stirred in 20 mji of DMSO at room temperature overnight, filtered again, and washed repeatedly with large amounts of water to obtain an adsorbent. The shape of the adsorbent is bead-like, and the particle size is 20
-100 μm, average particle size was 60 μm. After suction filtration, this adsorbent was immersed in distilled water for injection.

The adsorption test and regeneration test were conducted using an adsorbent from which water had been squeezed out by suction filtration.

Disaster ■Soil 0.5 m (dry weight 0.1 g) of the above adsorbent was packed into a mini-column with a diameter of 2 columns and a length of 15 cm to adsorb various disease-causing factors and biopolymers, and then the regeneration solution was added. It was circulated, and then the same disease-causing factors were adsorbed, and changes in the adsorption rate were measured.

Human IgG, anti-acetylcholine receptor, rheumatoid factor, anti-DNA antibody, and β2-microglobulin were investigated as disease-causing factors, and pyrodiene was investigated as a biopolymer.

In addition, as a regenerating solution, phosphate buffer 11 and Na5CN
mo I! Dissolved in the proportion of 1 of the same buffer! 8 mof of urea dissolved in the same buffer solution, 100 mof
A solution containing 50% by weight of ethylene glycol was used. A solution of Na5CN, phosphoric acid, and ethylene glycol was used.

The results are shown in Table 1.

From this result, it was found that the adsorbent could be effectively regenerated by using the above-mentioned regenerating liquid.

As a regenerating solution, a solution containing Na5CN and urea dissolved in a phosphate buffer was used, and the adsorption and desorption of IgG was repeated using the same adsorbent as in Example 1. Changes in adsorption performance were investigated.

Although the adsorption and underwear were repeated 11 times, almost no change was observed in the adsorption performance of the adsorbent.

EXPERIMENTAL The adsorbent used in Example 1 was packed into a 200 ml O column and used to treat a patient with myasthenia gravis.

The anti-acetylcholine receptor antibody titer in the patient's plasma was 120 pmof/m before treatment.

Treatment was carried out by passing the patient's plasma through the above column for 2 hours at a flow rate of Q p = 20 ax. The antibody titer after treatment decreased to 24 pmoE/all.

Next, a regeneration solution prepared by dissolving NaSCN in a phosphate buffer was passed through the column, and the antibody titer of the desorbed anti-acetylcholine receptor was measured. The results are shown in Table 2. As a result, it was found that 95% of the adsorbed anti-acetylcholine receptorase was recovered.

The adsorbent used in Senniwa Medical Soil Example 1 was packed into a 50 d column,
It was incorporated into the circulation system shown in FIG. 1, and adsorption and desorption of anti-acetylcholine receptors were repeatedly performed. Anti-acetylcholine receptor antibody titer in pooled plasma was 240.
The amount of plasma was 2000 pmoffi.
ml. A solution of Na5CN dissolved in a buffer solution was used as the regeneration solution.

Two columns were used, and adsorption and desorption were repeated alternately.

As a result, it was found that anti-acetylcholine receptorase could be removed 100%. The results are shown in Table 2.

Using the extracorporeal circulation system shown in Figure 2, anti-acetylcholine receptor antibodies, which are the disease-causing factor of myasthenia gravis, were adsorbed and removed. 2 columns with a capacity of 50ml
I used wood. The same regenerating liquid as in Example 4 was used.

The time required for the 50-column to reach adsorption equilibrium from the anti-acetylcholine receptor antibody titer was measured in advance, and 25 minutes of that time was taken as the column switching time. Plasma flow rate Qp=
Treatment was performed with 20 ml for 150 minutes (3 cycles). As a result, it was found that the same amount of anti-acetylcholine receptor antibody could be adsorbed and removed as that which could be adsorbed and removed by a 300 ml column.

In the same manner as in Example 1, various disease-causing factors and biopolymers were adsorbed onto the adsorbent, and then saline solution was passed through the soil, the concentration of disease-causing factors, etc. on the skin was measured, and the cleaning effect was evaluated. It was measured.

Human IgG, anti-acetylcholine receptor, rheumatoid factor, anti-DNA antibody, and β2-microglobulin were investigated as disease-causing factors, and pyrodiene was investigated as a biopolymer.

The results are shown in Table 1.

From this result, it was found that the adsorbent was hardly regenerated in saline solution.

Comparison 1 The adsorbent used in Example 1 was packed into a 200-0 column and incorporated into the circulation system shown in FIG. 2 to repeatedly adsorb and desorb anti-acetylcholine receptor antibodies.

The anti-acetylcholine receptor antibody titer in pooled plasma was adjusted to 240 pmol, and the amount of plasma was 2.
000 mfl.

One column was used. As a result, only 50% of anti-acetylcholine receptor antibodies could be removed. The results are shown in Table 2.

Table [For lfF of the present invention] Adsorbents usually form strong hydrogen bonds and hydrophobic bonds with biopolymers such as disease-causing or causative factors. On the other hand, among the components contained in the regenerating solution of the present invention, high chaotropic salts and glycols both have the function of increasing the water solubility of biopolymers and weakening hydrophobic bonds. Urea also has the function of breaking hydrogen bonds with the adsorbent.

Therefore, when the regeneration liquid of the present invention is brought into contact with this adsorbent,
The water solubility of biopolymers adsorbed on the adsorbent increases due to the action of high chaotropicity salts in the regenerating solution, and the biopolymers are removed from the adsorbent. The adsorbent is thus regenerated.

[Effects of the present invention]

The regenerating solution and regenerating method of the present invention are applicable to adsorbents for the purpose of adsorbing the following biopolymers, such as rheumatoid factor and anti-D
It is particularly suitable for regenerating adsorbents for the purpose of adsorbing disease-causing factors such as NA antibodies and anti-acetylcholine antibodies contained in body fluids such as patient's blood and plasma, pharmaceutical intermediates and drugs, or pyrogienes.

Furthermore, it is most preferable that the above-mentioned adsorbent is formed by bonding a polymer compound having a hydroxyl group, such as agarose, with an isocyanate compound having two or more incyanate groups.

Using the extracorporeal circulation system of the present invention, it is possible to treat a patient while washing multiple columns alternately.

Therefore, even in long-term treatment, only a small amount of adsorbent is required.

[Brief explanation of drawings]

FIG. 1 is a schematic diagram of the circulation system used in Example 4. FIG. 2 is a schematic diagram of the extracorporeal circulation system used in Example 5.

Claims (3)

[Claims] (1) A regenerating solution for an adsorbent for biopolymers, which comprises a buffer containing at least one compound selected from the group consisting of salts with high chaotropicity, urea, or glycols. (2) A method for regenerating an adsorbent for biopolymers, which comprises bringing it into contact with the regeneration liquid according to claim 1. (3) In the extracorporeal circulation system incorporating the adsorption column packed with the above-mentioned adsorbent, the flow path is switched.
An extracorporeal circulation system capable of regenerating the adsorption column by flowing the regenerating liquid described in Section 1 through the adsorption column.