JP2568846B2 - Myoglobin adsorbent - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to a myoglobin adsorbent that selectively adsorbs myoglobin, which is considered to be closely related to certain types of acute renal failure.

Myonephropatic metabolic syndr, a complication of so-called crush syndrome, an acute arterial occlusion, in which myoglobin in the muscle flows out due to extensive muscle crush and causes renal damage
Many diseases and injuries, such as ome (MNMS), lightning, diabetic coma, and sunstroke, have been shown to increase myoglobin in the blood and cause kidney damage, resulting in renal failure. Therefore, a technique for selectively removing myoglobin from blood to prevent renal damage has been desired.

Existing technologies Existing technologies that can be used for the above purposes include (1) plasma exchange and (2) plasma filtration.

Plasmapheresis is a method in which plasma, which is a humoral component, is separated from blood containing myoglobin by filtration and centrifugation techniques, the plasma containing myoglobin is discarded, and frozen fresh plasma and albumin solution are replenished. is there. However, in this method, together with myoglobin, useful components in blood such as albumin, immunoglobulin, blood coagulation protein,
Complement components, hormones, etc. are discarded at the same time.Frozen fresh plasma or albumin solution as a replenisher is derived from living organisms, so it is easy for infections to occur, and it is difficult to obtain raw plasma. There are various problems such as a problem and a problem of high price. Plasma filtration is a method of filtering and discarding all molecules smaller than myoglobin from blood and replenishing the electrolyte. This method has less material to discard than plasma exchange. However, since the pore size distribution of the blood filtration membrane is not constant, it is inevitable that many proteins larger than myoglobin will be lost and hypoproteinemia will occur, and useful blood components of molecules smaller than myoglobin will be There was a drawback that it was thrown away.

(Problems to be Solved by the Invention) As described above, removal of useful components in blood is unavoidable with the conventional technology, and it has been impossible to selectively remove myoglobin in blood.

An object of the present invention is to provide a technique for selectively removing myoglobin in blood, and an adsorbent capable of selectively adsorbing and removing myoglobin, which rarely removes useful substances such as albumin and immunoglobulin. To provide.

(Means for Solving the Problems) The present inventors have conducted intensive studies in accordance with the above-mentioned object, and
It has been found that myoglobin can be adsorbed at a surprisingly high rate by using a water-insoluble porous body made of a hydrophobic material, and only when the degree of hydrophobicity of the material is 20 degrees or more in contact angle, The inventors have found that high adsorption ability can be exhibited, and have completed the present invention.

That is, the gist of the present invention resides in a myoglobin adsorbent, which is a water-insoluble porous material having a contact angle of 20 degrees or more, and the material surface of which is a myoglobin adsorption surface.

The substance to be adsorbed in the present invention is myoglobin. To be more specific, it is a monomer having a molecular weight of about 1.7 × 10 ⁴ and, like hemoglobin, a protein that reversibly binds oxygen.

The “contact angle” referred to in the present invention is a contact angle of air bubbles on a solid surface in water, and is referred to as WCHamilton, J. Colloid.
Interface Sci., 40, 219-222 (1972) and JDAndra
de, J. Polym. Sci. Polym. Symp., 66, 313-336 (1979) refers to a measured contact angle. As a sample, a molded product such as a sheet or a film was prepared. The contact angle was measured at a temperature of 25 ° C., measured 10 times or more, and the average value was used as the value of the contact angle of the material.

The material used for the water-insoluble porous body of the present invention has a contact angle of 20.
Must be at least degrees. If the contact angle is smaller than this, the surface of the water-insoluble porous body is too hydrophilic, and the adsorbing ability of myoglobin is extremely reduced, and the adsorbent for blood myoglobin is poor in practical ability. Further, if the contact angle is too large, that is, if the degree of hydrophobicity is too high, the adsorption capacity of myoglobin is high, but non-selective adsorption tends to increase slightly, the preferred contact angle range is
20 degrees to 110 degrees, a more preferred range is 25 degrees to 100 degrees
Degrees, more preferably in the range of 30 to 90 degrees.

The water-insoluble porous body having a contact angle of 20 degrees or more referred to in the present invention is a porous body made of a material that is solid in water and has a contact angle of 20 degrees or more measured by the above-described method. In other words, any inorganic or organic compound may be used as long as it is a porous body made of a material that is biased toward hydrophobicity.However, the amount of eluted substances with respect to warm water is small, and the control of the pores of the porous body can be performed more easily and precisely. High molecular compounds are preferred. Such examples include polypropylene, polystyrene, polymethacrylate esters, polyacrylate esters,
Examples thereof include polymers and copolymers of vinyl compounds such as polyacrylic acid and polyvinyl alcohol, polyamide compounds such as nylon 6, 66, and polyester compounds such as polyethylene terephthalate. The material of the porous body used in the present invention may have a contact angle of 20 degrees or more, and is not limited to the above examples.

Among the examples, polymers and copolymers of vinyl compounds are more preferably used in view of ease of polymerization and ease of pore control. Examples of such are styrene,
Polymers of styrene-based compounds such as p-methylstin and p-ethylstyrene, polymers of (meth) acrylate-based compounds such as methyl (meth) acrylate, ethyl (meth) acrylate, propyl (meth) acrylate, and the above Copolymers of the compound with a vinyl compound such as divinylbenzene, methacrylonitrile, vinylpyrrolidone, ethylene glycol, and methyl acrylate can be exemplified.

Among them, a polymer of a (meth) acrylate compound such as methyl (meth) acrylate, ethyl (meth) acrylate, and propyl (meth) acrylate, and a copolymer with the above vinyl compound are more preferably used.

Furthermore, the polymer of the (meth) acrylate compound and the copolymer with the vinyl compound containing 70% by weight or more of methyl methacrylate is a material used for the water-insoluble porous material of the present invention. Gives more favorable results.

The shape of the water-insoluble porous body is spherical, granular, thread-like,
Any of a hollow fiber shape, a flat membrane shape and the like can be effectively used, but a spherical shape or a granular shape is particularly preferably used from the surface of the body fluid circulation during body fluid circulation. Spherical or granular average particle size is 10-2500
Although a micrometer is easy to use, a range of 25 μm to 300 μm is preferably used.

The water-insoluble porous body needs to be a porous body from the viewpoint that a large adsorption surface area of myoglobin can be obtained and a practical adsorption ability can be obtained. Exclusion limit molecular weight of the porous body from it (protein) molecular weight of myoglobin is 1.7 × 10 ^4, it is necessary that more than this value, 2 × 10 ⁴ 2
A range of × 10 ⁷ is preferred. More preferred is 2 × 10 ⁵ to 2 × 10 ⁶ . The pore distribution of the porous body can be obtained from a mercury intrusion curve obtained by a mercury intrusion method (for example, Catalyst Engineering Course-4, Catalyst Measurement Method, edited by the Catalysis Society of Japan, Jinjinshokan, pp. 69-73). It is preferable that there are many pores having a diameter of 50 ° or more, and it is more preferable that the pores be collected in a range of 50 ° to 2000 °. More preferably, it is in the range from 60 ° to 1000 °, most preferably in the range from 65 ° to 400 °.

Hereinafter, a method for producing a water-insoluble porous body will be described, but the present invention is not limited to this example.

The method for producing the water-insoluble porous material of the present invention is a generally known method such as bulk polymerization, solution polymerization, suspension polymerization, and emulsion polymerization. And the solvent for dissolving the monomer, and the method for producing each polymer. For example, in a styrene-divinylbenzene copolymer, styrene, ethylbenzene, divinylbenzene and toluene, octanol and AIBN
By stirring in the presence of (azobisisobutyronitrile), porous particles having a sphere diameter of about 50 to 1000 μm can be produced. Further, particles having various particle diameters and pore diameters can be produced by radical polymerization in a suspension polymerization system.

The water-insoluble porous body of the present invention can be subjected to a surface treatment for suppressing platelet adhesion in order to improve affinity with blood. This can be achieved, for example, by providing a coat layer of a platelet low-adhesive material on the surface of the water-insoluble porous body that comes into contact with blood cells.

Platelet low-adhesive materials include high molecular weight materials having a hydroxyl group such as hydroxyethyl methacrylate and hydroxyethyl acrylate, and monomers having a basic nitrogen-containing functional group such as vinylamine and dimethylaminoethyl (meth) acrylate. Block copolymers such as copolymers with polymerizable monomers having no nitrogen-containing functional groups, polymer materials having negatively charged functional groups such as sulfonic acid groups and carboxylic acid groups, segmented polyurethanes and segmented polyesters Examples thereof include a graft copolymer such as a copolymer of a monomer having a polyethylene oxide chain and another polymerizable monomer.

The myoglobin adsorbent of the present invention is generally used by being filled and held in a container having a body fluid outlet. In the accompanying drawings, reference numeral 1 denotes an example of an adsorption device filled with the myoglobin adsorbent of the present invention, and a packing 4 in which a filter 3 is stretched inside one end opening of a cylinder 2.
A cap 6 having a bodily fluid outlet 5 is screwed into the cap 2, and a cap 8 having a bodily fluid outlet 7 is screwed into the other end opening of the cylinder 2 through a packing 4 'having a filter 3' inside. The container is formed by fitting, and the adsorbent is filled and held in the gap between the filters 3 and 3 'to form the adsorbent layer 9.

The adsorbent layer 9 may be filled with the myoglobin adsorbent of the present invention alone, or may be mixed or laminated with another adsorbent. When the volume of the adsorbent layer 9 is used for extracorporeal circulation,
About 50-500 ml is appropriate. When the apparatus of the present invention is used in extracorporeal circulation, there are roughly the following two methods. One is to separate blood taken out from the body into a plasma component and a blood cell component using a centrifuge or a membrane-type plasma separator, pass the plasma component through the device, purify the blood, Another method is to return blood to the body together with the components, the other is to pass blood taken from the body directly to the device,
It is a method of purification.

Further, regarding the passage speed of plasma or plasma, since the adsorption efficiency of the adsorbent is very high, the particle size of the adsorbent can be coarsened and the degree of filling can be lowered, so that the shape of the adsorbent layer can be controlled. Regardless, a high transit speed can be provided. Therefore, a large amount of body fluid processing can be performed.

As a method of passing bodily fluids, the fluid may be passed continuously or intermittently according to clinical necessity or the state of the equipment in the facility.

(Effects of the Invention) As described above, the myoglobin adsorbent of the present invention selectively adsorbs and removes myoglobin in a body fluid at a high rate,
The adsorption device using the adsorbent is very compact, simple and complete.

INDUSTRIAL APPLICABILITY The present invention is applicable to general uses for purifying and regenerating body fluids such as blood and plasma, and is effective for safe and reliable treatment of diseases associated with an increase in blood myoglobin concentration.

In addition, the adsorbent of the present invention can be used not only as an affinity adsorbent for separation and purification of myoglobin, but also used as a therapeutic device after being filled in an apparatus.

(Examples) Hereinafter, embodiments of the present invention will be described in more detail with reference to Examples.

Example 1 As a water-insoluble porous body, a methyl methacrylate-divinylbenzene copolymer (80: 20% by weight, Ambersep, manufactured by Organo Corporation) was used. The contact angle of the air bubbles in water of the material used for the water-insoluble porous body was 65 ± 3 degrees.

Using the water-insoluble porous material as a myoglobin adsorbent, the following adsorption experiment was performed.

Myoglobin (Kappel, human myoglobin positive control) per 1 ml of water-insoluble porous material
6 ml of a 120 μg / ml (dissolved in heparinized human plasma) solution was added, and the mixture was incubated with shaking at 37 ° C. for 1 hour. Thereafter, the water-insoluble porous body was sedimented, and the myoglobin concentration in the supernatant was measured.

As a result of the adsorption experiment, the myoglobin concentration after adsorption was 9.4 μg
/ ml (8% before adsorption).

Example 2 An experiment was conducted in the same manner as in Example 1 except that a vinyl alcohol / triallyl isocyanurate copolymer was used as the water-insoluble porous body.

The contact angle of air bubbles in the vinyl alcohol / triallyl isocyanurate copolymer in water was 25 ± 4 degrees.

The vinyl alcohol / triallyl isocyanurate copolymer was obtained as follows. That is, 100 g of vinyl acetate,
64.3 g of triallyl isocyanurate and water in which 2,2′-azobisisobutyronitrile and polyvinyl alcohol, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate are dissolved are placed in a flask, and sufficiently stirred. 75
The suspension polymerization was carried out at 5 ° C. for 5 hours to obtain a granular polymer having a diameter of 350 to 500 μm.

As a result of the adsorption experiment, the myoglobin concentration before adsorption was 120 μg
was 60 μg / ml after adsorption (50 μg before adsorption).
%).

Comparative Example 1 An experiment was performed in the same manner as in Example 1 except that agarose gel (Sepharose 4B, Pharmacia Japan) was used as the water-insoluble porous body.

The contact angle of air bubbles in agarose water is 11 ± 6.
Degree. As a result of the adsorption experiment, the myoglobin concentration before adsorption was 120 μg / ml, whereas after adsorption 97 μg / ml
(81% before adsorption) did not drop much.

Clinically, it is said that renal failure does not occur if the blood myoglobin concentration of 100 μg / ml is reduced to half or less. With the adsorption capacity of this comparative example, the blood myoglobin concentration cannot be reduced to half or less unless one or more adsorbents are used, which is not practical.

Example 3 Except that a styrene / divinylbenzene copolymer (HP-10, Mitsubishi Kasei Kogyo) was used as the water-insoluble porous material,
An experiment was performed in the same manner as in Example 1.

The contact angle of air bubbles in the styrene-divinylbenzene copolymer in water was 76 ± 5 degrees.

As a result of the adsorption experiment, the myoglobin concentration before adsorption was 120 μg
8.3 μg / ml after adsorption (7 μg / ml before adsorption).
%).

[Brief description of the drawings]

The figure is a schematic cross-sectional view showing one example of an embodiment of an adsorption device using the myoglobin adsorbent of the present invention. 1 ... adsorption device, 2 ... cylinder, 3 ', 3 ... filter, 4,4' ... packing, 5 ... body fluid inlet, 6 ...
... cap, 7 ... body fluid outlet, 8 ... cap, 9 ...
… Adsorbent layer

Claims (1)

(57) [Claims] 1. A myoglobin adsorbent, which is a water-insoluble porous material having a contact angle of 20 degrees or more, wherein the surface of the material is used as a myoglobin adsorption surface.