JPH0622629B2 - ▲I▼▲g▼E Adsorbents and Apparatus - Google Patents

Description

Description: FIELD OF INDUSTRIAL APPLICATION The present invention relates to an adsorbent used for efficiently adsorbing and removing IgE (immunoglobulin E) from blood, and an IgE adsorption device equipped with said adsorbent.

(PRIOR ART) Anaphylactic hypersensitivity such as bronchial asthma and nasal allergy is considered to be IgE antibody-dependent hypersensitivity.
E is a receptor on the surface of basophils and mast cells and Fc
When an antigen (allergen) enters the blood and binds to the Fab portion of IgE, an agglutination reaction progresses and chemical mediators such as histamine are released from intracellular granules, causing capillary expansion and increased secretion from mucus glands, resulting in anaphylactic hypersensitivity such as bronchial asthma and nasal allergies. Therefore, if IgE can be removed from the blood, it is expected to become an effective means of treating anaphylactic hypersensitivity such as bronchial asthma and nasal allergies.

An example of a known prior art is an IgE adsorbent comprising anti-human IgE bound to an insoluble carrier (Publication 1984-151964).

(Problems to be Solved by the Invention) However, the adsorbent to which such a protein (anti-human IgE) is bound requires complicated and time-consuming procedures to obtain anti-human IgE, and it is difficult to obtain a large amount of it. Furthermore, when a large amount of energy is loaded, such as in the case of sterilization, especially high-pressure steam sterilization, the anti-human IgE is easily absorbed.
However, there is a drawback in that the activity of gE decreases, making it difficult to put into practical use in cases where sterility must be guaranteed.

(Means for solving the problems) In view of the above-mentioned circumstances, the present inventors have conducted extensive research and have found that IgE
The present invention was made based on the discovery that an adsorbent having a hydrophobic compound bound to an insoluble carrier is effective as an adsorbent capable of efficiently adsorbing and removing IgE and which is easy to sterilize. That is, the gist of the present invention resides in an IgE adsorbent having a hydrophobic compound bound to an insoluble carrier, and in a device equipped with said adsorbent.

The hydrophobic compound used in the present invention is a compound having a solubility in physiological saline of 100 mmol/dL or less.
Among them, compounds having at least one aromatic ring give preferable results. The aromatic ring means a benzene-based aromatic ring such as benzene or naphthalene, a nitrogen-containing six-membered aromatic ring such as pyridine or quinoline, a five-membered ring or a six-membered ring containing nitrogen, oxygen, sulfur, or the like, or other heteroaromatic rings and their derivatives. Hydrophobic compounds having at least one of these aromatic rings and/or heteroaromatic rings are I.
It gives favorable results for gE adsorption.

Among hydrophobic compounds, hydrophobic amino acids can be used more safely and efficiently, and can adsorb IgE. Hydrophobic amino acids are amino acids with a solubility in physiological saline of 100 mmol/dl or less, such as lysine, valine, leucine, tyrosine, phenylalanine, isoleucine, tryptophan and its derivatives. Among these hydrophobic amino acids, tryptophan and its derivatives give good results, and can be used without distinction between d and l forms.

In addition, as a method for binding these hydrophobic amino acids to an insoluble carrier, known means such as covalent bonding, ionic bonding, and embedding methods can be used, but in terms of the elution of the bound product, the covalent bonding method is more preferred.

The activation methods used in the covalent bonding method include the cyanogen halide method, the epichlorohydrin method, the bisepoxide method,
However, the present invention is not limited to the above examples, and any method may be used as long as it can react with a reactive group having an active hydrogen, such as an amino group, a hydroxyl group, a carboxyl group, or a thiol group, of a hydrophobic compound, by substitution and/or addition reaction.

On the other hand, the insoluble carrier used in the present invention is preferably a porous carrier which is insoluble in water and has a protein exclusion limit molecular weight of 1 x ¹⁰ or more, and in addition to natural polymeric carriers such as dextran and agarose, carriers for affinity chromatography such as synthetic polymeric carriers such as acrylamide are used.

However, since the physical strength of natural polymer carriers is weak, the carriers may be destroyed during preparation of the adsorbent, or may be crushed when a viscous liquid such as blood plasma is passed through the carrier, resulting in clogging and increased pressure loss. For these reasons, cross-linked copolymers having vinyl alcohol as the main component are more preferred (JP Patent Publication 58-15924).

The shape of the carrier may be any shape, such as a sphere, flat membrane, hollow fiber, or filament, but in order to increase the adsorption surface area, a spherical insoluble carrier is preferred, with a particle size of 50 to 500 μm being preferred.

The adsorption device of the present invention may have a structure as shown in the drawing, but is not limited to this structure and may have any structure as long as it has an inlet and outlet for fluid, contains an adsorbent inside, and can hold it with a filter or the like. In the drawing, 1 is the adsorption device, and IgE adsorbent 9 is held by a cylindrical part 2 and filters 3 and 3'. 4 and 4'
The symbol indicates a packing, and the symbols 6 and 8 indicate caps.
The gas enters the adsorption device 1 , is subjected to adsorption treatment by the adsorbent 9 , and is discharged from the outlet 7 .

The IgE adsorbent and adsorption device of the present invention extract IgE from blood.
It can efficiently and safely remove IgE
The present invention is expected to be an effective means of treating anaphylactic hypersensitivity disorders such as bronchial asthma and nasal allergy, which are believed to be caused by the above-mentioned substances. The present invention will be specifically described below with reference to examples.

(Example) Vinyl acetate 1000g, triallyl isocyanurate 4
14g, ethyl acetate 1000g, heptane 1000g,
Polyvinyl acetate (degree of polymerization 500) 70 g, and 2, 2
A homogeneous mixture of 36 g of 1-azobisisobutyronitrile and 4 g of water containing 1% by weight of polyvinyl alcohol, 0.05% by weight of sodium dihydrogen phosphate dihydrate, and 1.5% by weight of disodium hydrogen phosphate dodecahydrate was placed in a flask, thoroughly stirred, and then heated and stirred at 65°C for 18 hours and then at 75°C for 5 hours to carry out suspension polymerization, thereby obtaining a granular copolymer. The mixture was filtered, washed with water, and then extracted with acetone, 46.5 g of NaOH, and 20 g of methanol.
The copolymer was subjected to an ester exchange reaction at 40° C. for 18 hours in a solution consisting of the above. The average particle size of the resulting particles was 15.
The particle size was 0.0 μm and the hydroxyl group density was 7.5 meq/g.

The hydroxyl group density can be determined by reacting the carrier with acetic anhydride in a pyridine solvent and measuring the amount of acetic anhydride consumed by reaction with the hydroxyl groups. When 1 g of the dry carrier reacts with 1 mmol of acetic anhydride, the hydroxyl group density is 1 meq/g.

Next, 150 g of the transesterified gel was thoroughly washed with water and dried. The gel was suspended in a solution of 1800 ml of dimethyl sulfoxide and 1200 ml of epichlorohydrin, and 150 ml of 30% aqueous NaOH solution was added.
The mixture was reacted at 0.5 °C for 5 hours with stirring. After the reaction was completed, the mixture was filtered through a glass filter and washed with 3 ml of dimethyl sulfoxide and then with 20 ml of water to obtain an epoxy group-bonded gel. The amount of epoxy group bonded was 0.11 mmol per ml.

To prepare an adsorbent by binding tryptophan as a ligand using the epoxy group-bound gel, tryptophan was dissolved in a carbonate buffer solution of pH 9.8 at 0.05 mol/L.
The epoxy bonding gel 20 is dissolved to a concentration of 100/1000.
Add 300 ml of tryptophan solution to each 50 ml of
After reacting and binding for 16 hours at 0°C, the adsorbent was repeatedly washed with a large amount of water, then washed with physiological saline, and drained to obtain an adsorbent bound with tryptophan. The amount of tryptophan bound was determined by the 280 nm absorption of the amount of tryptophan remaining in the reaction solution, and was found to be 0.058 mmol per 1 ml of gel.

0.5 g and 0.25 g of the above adsorbent were placed in test tubes, and 1 ml of human plasma was added to each tube. The tubes were then shaken in a constant temperature bath at 37° C. for 30 minutes, followed by centrifugation (×1000 g) to separate the plasma from the adsorbent. The plasma was then extracted and the Ig in the plasma was measured.
The IgE concentration was determined by radioimmunoassay, and the remaining rate relative to the original plasma before adsorption was calculated. This test was performed five times, and the results are shown in the table (No. 1 to No. 5). As is clear from the table, the use of the adsorbent of the present invention makes it possible to efficiently adsorb and remove IgE from plasma.

[Brief description of the drawings]

The attached drawing is a cross-sectional view showing an example of the device of the present invention. 1, adsorption device, 2, cylinder, 3, 3', filter, 4,
4', packing, 5, inlet, 6, cap, 7, outlet, 8, cap, 9, adsorbent

Claims (4)

[Claims] 1. An IgE adsorbent comprising an insoluble carrier to which a hydrophobic compound is bound. 2. The IgE adsorbent according to claim 1, wherein the hydrophobic compound is a hydrophobic amino acid. 3. The IgE adsorbent according to claim 2, wherein the hydrophobic amino acid is tryptophan or a derivative thereof. 4. An IgE adsorption device, comprising an adsorbent comprising an insoluble carrier and a hydrophobic compound bound thereto, which is mounted in a vessel having an inlet and an outlet for fluid.