JPH0275340A - Adsorbing body - Google Patents

Abstract

PURPOSE:To provide an adsorbing body having excellent adsorption capabilities which sufficiently remove harmful components and excessive components, etc., by limiting voids in an adsorbing body relative to components to be adsorbed within a specified range. CONSTITUTION:In an adsorbing body which selectively remove harmful components from humor such as blood, the ratio of volume of void in an adsorbing body which can accommodate components to be adsorbed and removed to the apparent volume of the adsorbing body, i.e., efficient voids of adsorbing body, is in the range of 0.4-0.96. Said adsorbing body is a body, wherein compounds having affinity for components to be adsorbed are immobilized on water-insoluble porous carriers or water-insoluble porous carriers themselves have affinity for components to be adsorbed. Adsorbing bodies thus prepared have excellent adsorption capabilities and can sufficiently remove harmful components and excessive components, etc.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to an adsorbent that selectively removes harmful components from body fluids such as blood.

[Problems to be solved by the conventional technology/invention] Conventionally, body fluids have been used to treat diseases in which it is assumed that substances that are directly the cause of the disease or are related to the cause of the disease, such as harmful components or excessive components, are present in the body fluids. A selective adsorbent has been developed that selectively removes harmful components and excessive components without losing much of the useful components therein.

However, the currently developed adsorbents have insufficient adsorption capacity, so it is necessary to increase the adsorber volume to obtain sufficient adsorption capacity, and the drawback is that increasing the adsorber volume increases the amount of extracorporeal circulation. There is. When treating with extracorporeal circulation, there is a limit to the amount of extracorporeal circulation that the patient can tolerate, so the adsorber volume cannot be increased too much, and it is impossible to dramatically increase the adsorption capacity, so there is no essential solution. It won't be. Furthermore, in order to increase the adsorption capacity, research has been carried out mainly on the use of ligands and spacers, but it has not yet been possible to obtain a sufficient adsorption capacity.

Therefore, the present inventors thought that in order to further increase the adsorption capacity, it is necessary to optimize the structure of the carrier or adsorbent.

As a result of intensive studies in view of the above circumstances, the present inventors discovered that the adsorption capacity can be improved by adjusting the porosity within the adsorbent to a certain range for the adsorbed and removed components, and have completed the present invention. Ta.

[Means for Solving the Problems] That is, the present invention relates to an adsorbent characterized in that the effective porosity within the adsorbent for the adsorbed and removed component is in the range of 0.4 to 0.96.

[Example] The terms used in this specification will be explained below.

The component to be adsorbed and removed refers to a component that is a harmful component or an excessive component present in a body fluid and is a target to be adsorbed and removed by an adsorbent (hereinafter also referred to as a component to be removed).

Harmful components refer to components in body fluids that directly cause or are associated with the pathogenesis, such as paraquat in balaquat poisoning and autoantibodies in autoimmune diseases.

The term "excessive component" refers to a component necessary for a living body but present in excess, such as LDL and VLDL in familial hypercholesterolemia, in a body fluid.

An adsorber is a container filled with an adsorbent and having a fluid inlet and an outlet.

Body fluid refers to blood, plasma, ascites, or other liquid components in living organisms.

The adsorption operation refers to an operation in which the component to be removed is adsorbed and removed from the body fluid by feeding the body fluid containing the component to be removed to the inlet of the adsorber and draining the fluid from the outlet.

The beginning of breakthrough is the bottom position (for example, chemical engineering handbook,
As explained in the Chemical Engineering Handbook, Revised 5th Edition, Adsorption Operation, Maruzen), in general, the concentration of the component to be removed at the outlet (hereinafter referred to as outlet concentration) is the concentration in the influent (hereinafter referred to as inlet concentration). In this specification, it refers to the time when the liquid reaches 5% to 10%.The time from when the liquid starts flowing into the adsorber until this point is called the breakthrough time. A graph showing the change in outlet concentration over time from the beginning until the outlet concentration becomes the same as the inlet concentration is called a breakthrough curve.

Adsorption capacity refers to the adsorption surplus, which is the difference between the adsorption capacity and the adsorbed amount, and/or the speed at which the target component is removed from the body fluid, and the target component is adsorbed and removed from the body fluid through an adsorption operation. It refers to overall performance, and in this specification, the length of breakthrough time is used as an index for comparison.

The effective porosity (β) in the adsorbent for the adsorbed component to be removed is the apparent ratio of the volume (Vp) into which the adsorbed component can enter to the adsorbent volume (Vx) (β-Vp).
/ Vx), and this value is determined by the adsorbent and the component to be adsorbed and removed. The apparent adsorbent body 1M (VW) is the adsorber volume (Vt) and the void volume (Vo) which is the volume between particles.
It is calculated as the difference between

The volume (Vp) in which the component to be adsorbed and removed can enter the adsorbent is determined as the difference between the volume (Ve) in the adsorber in which the component to be adsorbed and removed can enter and the void volume (Vo).

The adsorbent of the present invention is an adsorbent in which a compound having an affinity for the component to be adsorbed and removed is immobilized on a water-insoluble porous carrier, or an adsorbent in which the water-insoluble porous carrier itself has an affinity for the component to be adsorbed and removed. The particle effective porosity for the adsorbed and removed component is in the range of 0.4 to 0.96. Effective porosity for adsorbed and removed components is 0.4
If it is less than 0.96, it becomes difficult for the component to be adsorbed and removed to enter the adsorbent and the amount of adsorption decreases, and if it exceeds 0.96, the amount of adsorption decreases as the amount of the component that has an affinity for the component to be adsorbed and removed decreases.

The water-insoluble porous carrier used in the present invention may be either organic or inorganic, but it is preferably one that has little adsorption (so-called non-specific adsorption) of body fluid components other than the target component to be removed. It is preferable that the water-insoluble porous carrier is hydrophilic rather than hydrophobic, since non-specific adsorption is less likely to occur if the carrier is hydrophilic.

Furthermore, it is advantageous if a functional group that can be used for a ligand immobilization reaction is present on the surface of the water-insoluble porous carrier. Representative examples of these functional groups include hydroxyl group, amino group, aldehyde group, carboxyl group, thiol group, silanol group, amide group, epoxy group, halogen group, succinylimide group, acid anhydride group, etc. It is not limited to these. Further, it is particularly preferable that the water-insoluble porous carrier is made of a compound having a hydroxyl group among the above-mentioned functional groups, since non-specific adsorption is small. Needless to say, water-insoluble porous carriers into which these functional groups are introduced as spacers can also be used.

Representative examples of water-insoluble porous carriers used in the present invention include:
Soft porous materials such as agarose, dextran, and polyacrylamide, inorganic porous materials such as porous glass and porous silica gel, synthetic polymers such as polymethyl methacrylate, polyvinyl alcohol, styrene, and divinylbenzene copolymers, and/or cellulose. Examples include, but are not limited to, porous polymer hard gels made from natural polymers such as .

When using the adsorbent of the present invention for extracorporeal circulation treatment, blood,
Because of the need to flow high viscosity fluids such as plasma at high speeds, it is preferred to use a rigid, water-insoluble porous carrier that has sufficient mechanical strength to avoid compaction. In other words, a hard water-insoluble porous carrier is one in which the relationship between pressure loss and flow rate when a water-insoluble porous carrier is uniformly packed into a cylindrical column and an aqueous fluid is passed through it is at least 0.3 kg/c.
It refers to something that has a linear relationship up to -.

Typical examples of compounds that have an affinity for the adsorbed and removed components used in the present invention include polyacrylic acid, polyvinyl sulfate,
Synthetic polyanionic compounds such as polyvinyl sulfonic acid, polyvinyl phosphoric acid, polystyrene sulfonic acid, polystyrene phosphate, polyglutamic acid, polyaspartic acid, polymethacrylic acid, polyphosphoric acid, styrene-maleic acid copolymer, and heparin, dextran sulfate, and chondroitin. , chondroitin sulfate, phosphomannan, chitin, chitosan, and other functional group-containing polysaccharides that have an affinity for the component to be adsorbed and removed, but are not limited to O. The number of compounds having affinity for the removed component may be one, or two or more.

The adsorbent of the present invention is an adsorbent in which a compound having an affinity for a component to be adsorbed and removed is immobilized on a water-insoluble porous carrier.

In order to obtain an adsorbent in which a compound having an affinity for the adsorbed component to be removed is immobilized, there are various known methods for introducing into a water-insoluble porous carrier a functional group that has an affinity for the adsorbed component to be removed. The method described above can be used without any special restrictions, but it is necessary to use a functional group that has an affinity for the adsorbed component to obtain an adsorbent on which a compound having an affinity for the adsorbed component is immobilized. Typical methods for introducing into the carrier are: (1) A compound containing a functional group that has an affinity for the component to be adsorbed or that can be easily converted into a functional group that has an affinity for the component to be adsorbed and removed. A method of forming an adsorbent by polymerization using a monomer or a crosslinking agent, (2) A method of fixing a compound containing a functional group that has an affinity for the adsorbed component to a water-insoluble porous carrier, (3) A method of fixing a compound containing a functional group that has an affinity for the adsorbed component to be removed. A method of immobilizing a compound having a functional group having an affinity for a component to be adsorbed and removed onto a water-insoluble porous carrier by directly reacting the compound forming a functional group having an affinity for the water-insoluble porous carrier with the water-insoluble porous carrier. can be given.

Of course, an inherently water-insoluble porous carrier such as glass, silica, alumina, etc. may be used as an adsorbent having an affinity for the component to be adsorbed and removed.

Typical examples of 7mers or crosslinking agents that contain a functional group that has an affinity for the adsorbed component to be removed or that can be easily converted into a functional group that has an affinity for the adsorbed and removed component used in method (1) Examples include, but are not limited to, acrylic acid and its esters, methacrylic acid and its esters, styrene sulfonic acid, and the like.

(Method '2J, that is, a method for immobilizing a compound containing a functional group that has an affinity for the component to be adsorbed and removed, on a water-insoluble porous carrier includes physical adsorption, ionic bonding, and covalent bonding. Any method may be used, but for the preservation and stability of the adsorbent, it is important that the functional group-containing compound that has an affinity for the component to be adsorbed and removed does not desorb. , a coexistence bonding method that allows for strong fixation is desirable.

In the case of immobilizing a compound that has an affinity for the component to be adsorbed and removed by covalent bonding, the compound that has an affinity for the component to be adsorbed and removed has a functional group that can be used for immobilization in addition to the functional group that has an affinity for the component to be adsorbed and removed. It is preferable to have

Typical examples of functional groups that can be used for immobilization include amino groups,
Examples include, but are not limited to, amide groups, carboxyl groups, acid anhydride groups, succinylimide groups, hydroxyl groups, thiol groups, aldehyde groups, halogen groups, epoxy groups, and silanol groups.

Furthermore, among compounds that have an affinity for adsorbed and removed components, typical examples of compounds containing sulfate groups include sulfate esters of hydroxyl group-containing compounds such as alcohols, sugars, and glycols. Partially sulfated alcohol ester compounds, especially saccharide sulfated ester compounds, contain both sulfate groups and functional groups necessary for immobilization, and have high biocompatibility and activity, and sulfated polysaccharides are easily hydrated. It is particularly preferred because it can be fixed on an insoluble porous carrier.

There are many compounds that have an affinity for these adsorbed and removed components, and dextran sulfate described in the Examples is one example.

Next, by the method (3), that is, by reacting the water-insoluble porous carrier with a compound that forms a functional group that has an affinity for the adsorbed and removed component, the water-insoluble porous carrier has an affinity for the adsorbed and removed component. A typical example of a method for introducing a functional group having an affinity for a component to be adsorbed and removed by immobilizing a compound having a hydroxyl group is a reaction in which a sulfate ester group is introduced into a porous carrier containing a hydroxyl group. In this case, the sulfate ester group can be directly introduced by reacting the hydroxyl group-containing water-insoluble porous carrier with a reagent such as chlorosulfonic acid or concentrated sulfuric acid.

Regarding the porous structure of the water-insoluble porous carrier, total porosity is preferable to surface porosity. The shape of the water-insoluble porous carrier can be selected from any shape such as granules, spheres, fibers, membranes, and hollow fibers. When using a granular water-insoluble porous carrier, if the particle size is less than 1 μm, the pressure loss will be large, and if it exceeds 5000 μm, the adsorption rate will be slow.
It is preferable that it is less than μl.

When the pore volume of the water-insoluble porous carrier is small and the component to be adsorbed cannot enter the particles, the effective porosity becomes 0, but since it does not enter the particles, the amount of adsorption becomes 0. On the other hand, if we hypothetically increase the pore volume of the water-insoluble porous carrier and assume that there is no water-insoluble porous carrier in the particles, the adsorbed and removed components will freely enter the particles, and in this case, the effective porosity will be 1. but,
Since there is no water-insoluble porous carrier to immobilize the ligand, the amount of ligand becomes 0 and the amount of adsorption becomes 0.

In order to increase the adsorption capacity, it is necessary to set the effective porosity to an appropriate value, but at this time, it is necessary to consider not only the adsorption capacity but also the adsorption rate.

As a guideline when using an adsorber, generally, use of the adsorber during adsorption operation should be stopped when breakthrough begins, and immediately replaced with a new adsorber. Therefore, as an index of adsorption capacity, it is considered that the later the point of breakthrough when adsorption operation is performed under the same conditions, that is, the longer the breakthrough time, the better the adsorption capacity, and this index is used for determination.

The present invention will be explained in more detail below using an example in which the component to be adsorbed and removed is a riboprotein and the water-insoluble porous carrier is granular, but the present invention is not limited to this example.

Example 1 Porous cellulose gel CKA-22 (trade name, manufactured by Chisso ■, polyethylene glycol exclusion limit molecular weight 1
500,000 to 3 million, particle size 45 to 105) 2 per 60 m1
After adding 32 ml of N-NaOH and 60 ml of water and stirring at 40°C for 30 minutes, epichlorohydrin fig was added.
Stir for hours. The gel was washed with water and filtered to obtain a cellulose gel into which epoxy groups were introduced.

The resulting cellulose gel with epoxy groups introduced 55m
1, 0.47 g of dextran sulfate (molecular weight: 5000, sulfur content: 15% by weight) was added, water was further added to make 97.4 ml, the pH was adjusted to 9, and the mixture was allowed to stand at 45° C. for 18 hours. The gel was washed and filtered with water, 55 ml of water, monoethanolamine o, and osg were added, and the unreacted epoxy groups were sealed by standing at 45°C for 3 hours, followed by washing and filtration with water to obtain a cellulose gel with fixed dextran sulfate. I got it.

A glass cylindrical column (inner diameter IO-■, length approximately IQ, 50 m, hereinafter referred to as blade ram) equipped with filters at both ends was uniformly filled with cellulose gel on which dextran sulfate was fixed as an adsorbent.

To measure the apparent adsorbent volume (V'x), use the nonionic surfactant Tween 20 (trade name, manufactured by Kao Atlas ■).
A particle size of 1μ that does not enter the pores of the adsorbent in a 0.2% solution of
A solution in which polystyrene latex (manufactured by Sigma) was dissolved (hereinafter referred to as solution 1) was used. After replacing the inside of the column with the surfactant solution in advance, solution 1 was flowed at a flow rate of 0.5 ml/sin using a bersal pump, and the change in column outlet concentration over time was measured using a υV meter (manufactured by Tosoh Corporation, wavelength 250).
nm), and the time-dependent change in the value, which was made dimensionless by dividing the outlet concentration by the inlet concentration, was measured. As shown in Figure 1,
Integrate the curve section from 0 to 1 (until it becomes equal to the inlet concentration) of the obtained dimensionless value to determine the volume between the gels used, that is, the void volume (Vo),
The apparent adsorbent volume (VX) was determined by subtracting from the column internal volume, that is, the adsorber volume (Vt).

In Figure 1, the shaded area is the volume (Vo) between the gels used.
shows. The ratio of the volume between the gels used to the internal volume of the column (Vo/Vt) was 0.384.

To measure the effective porosity (β) in the adsorbent for the component to be adsorbed and removed (riboprotein), NaCf is added to the plasma so that the NaCl concentration is approximately IH so that the riboprotein does not adsorb to the adsorbent.
A solution (hereinafter referred to as solution 2) was used.

After replacing the inside of the column with IM NaCl solution in advance, solution 2 was passed through a peristaltic bomb, and the effluent was collected using a fraction collector. The fractionated liquid was treated with Cholesterol C Test Fuco (trade name, manufactured by Wako ■).
Measured based on cholesterol concentration. Below, riboprotein concentration was similarly expressed on the basis of cholesterol concentration. We measured the change over time in the dimensionless value obtained by dividing the outlet concentration by the population concentration. As shown in Figure 2, by integrating the area surrounded by the obtained time-course curve and the previous elution curve with latex, we can find that the volume in which riboproteins can enter in the adsorbent body is calculated as follows:
In other words, the volume (
Vp) was calculated. In FIG. 2, the shaded area indicates the volume (Vp) in which riboproteins can fit inside the adsorbent. The apparent value (Vp) obtained can be calculated by dividing the apparent adsorbent volume (Vx) to determine the effective porosity (β) in the adsorbent for riboproteins.
I asked for

Next, after replacing the inside of the column with physiological saline in advance, the cholesterol concentration was adjusted to 184 mg using a beristal bomb.
/dl of plasma was flowed at a flow rate of 2.0 ml/sin, the effluent was collected using a fraction collector, and the cholesterol concentration was measured. The time-dependent change in the value made dimensionless by dividing the outlet concentration by the inlet concentration was measured. The amount of cholesterol adsorbed in the column was determined by integrating the area surrounded by the obtained time course curve and the latex elution curve shown in FIG. Here, the amount of cholesterol that has entered the adsorbent is conveniently referred to as the amount of cholesterol adsorbed.

Separately, by drying an unused adsorbent and measuring its weight, the amount of cholesterol adsorbed per the actual weight of the adsorbent was determined to be 127 mg/g-adsorbent.

The particle diameter of the adsorbent was measured by photographing the adsorbent on which dextran sulfate was immobilized, and was defined as the average particle diameter.

Table 1 shows the carrier group used, the average particle diameter of the adsorbent, the effective porosity (β) for riboprotein, the cholesterol concentration, and the amount of cholesterol adsorbed.

Examples 2 and 3 Porous cellulose gel CKA-32 (trade name, manufactured by Chisso ■, polyethylene glycol exclusion limit molecular weight 1
000,000 to 2,000,000, particle size 45 to 105 μm) (Example 2
), CKA-3-C (trade name, manufactured by Chisso ■, exclusion limit molecular weight of polyethylene glycol 2 million to 5 million, particle size 210 to 315 μm) (Example 3) were taken, and the same method as in Example 1 was taken. Dextran sulfate was fixed under certain conditions and operations.

The effective porosity (β) for the adsorbed component (riboprotein) is
It was determined by performing the same measurements as in Example 1 using Solution 1 and Solution 2. Each value is 0.488 (Example 2)
and 0.958 (Example 3).

Furthermore, a column was filled with each adsorbent on which dextran sulfate was immobilized, and the same operation as in Example 1 was performed to measure the breakthrough curve and determine the total adsorption amount of cholesterol. The amount of cholesterol adsorbed per actual weight of the adsorbent was determined by dividing by the dry weight of the adsorbent, which was measured separately. The amount of cholesterol adsorbed was 79.1 mg/g-adsorbent (Example 2) and 75 mg/g, respectively.
．． 9-g/g-adsorbent (Example 3).

The respective values are shown in Table 1 together with Example 1.

As shown in Table 1 in Examples 1, 2 and 3, the inlet concentrations are different, so the adsorption amounts cannot be directly compared.

In liquid phase adsorption, Freundlich is generally
h) Type adsorption and crosstalk is established. CKA-3 (trade name, manufactured by Chisso ■, polyethylene glycol exclusion limit molecular weight 2
An adsorbent on which dextran sulfate was immobilized was prepared in the same manner as in Example 1 using dextran sulfate as a carrier, and the amount of cholesterol adsorbed was measured using the following Freundlich type adsorption isotherm. : Cholesterol adsorption amount - coefficient x cholesterol concentration 0.424 (Cholesterol adsorption amount E-g/g-adsorbent)
, cholesterol concentration [■g/dll) was obtained. By using this, comparisons can be made at the same cholesterol concentration.

When comparing breakthrough times, since some of the adsorbents used in the experiments of Examples 1, 2 and 3 have different particle sizes, the effect of particle size is included when considering the adsorption rate. Therefore, we used the equation for adsorption using a packed column in Sokoki (Kei Tominaga, Adsorption, Kyoritsu Shuppan) to determine the diffusion coefficient, etc., and made it possible to compare on the basis of the same particle size. At this time, it was confirmed by observation using a scanning electron microscope that the particles were uniform from near the surface to inside.

In order to compare breakthrough times based on the same particle size, the relationship between the effective porosity (β) in the adsorbent, the adsorbed amount of cholesterol, and the diffusion coefficient in the adsorbent was divided into sections as shown in Figures 4 and 5, respectively. In the case of the same particle size of 72.5μ, when plasma with a cholesterol concentration of 150mg/di is poured into an adsorbent with an inner diameter of 1cm and a length of locm at a flow rate of 1.65ml/sin, When the breakthrough curve was determined, as shown in FIG. 6, the breakthrough time was upwardly convex with respect to the effective porosity. From this result, considering the breakthrough time which is an index of adsorption performance, the effective porosity within the adsorbent for the adsorbed and removed components of an adsorbent having excellent adsorption performance is in the range of 0.4 to 0.96. More preferably, the effective porosity is 0. B~0.7.

The value of the effective porosity within the adsorbent at which the breakthrough time was maximized was larger than the value of the effective porosity within the adsorbent at which the amount of adsorption was maximized. This is thought to be because the greater the effective porosity for the adsorbed component to be removed, the fewer obstacles there are to the diffusion of the adsorbed component within the particles, and the larger the diffusion coefficient becomes. This shows that optimizing the carrier by focusing only on the amount of adsorption is insufficient to judge the adsorption performance when considering actual use.

In the method for measuring the effective porosity within an adsorbent for riboproteins as explained in Example 1, depending on the adsorbed component and the adsorbent, the adsorbent and the adsorbed component are covered within a range where there is no denaturation or change in shape. Appropriate conditions may be selected by changing the conditions of the solution such as pH and ionic strength, or by adding urea or the like so that the components to be adsorbed and removed are not adsorbed.

By using this method, the harmful components to be adsorbed and removed have a molecular weight distribution, and even in diseases where the exact molecular weight cannot be determined, it is possible to determine a single value for the adsorbed component.

In the examples, an adsorbent having a ligand immobilized on a cellulose-based carrier was used, but an agarose-based carrier or other carrier may be used. Furthermore, as described above, the adsorbent may be an adsorbent in which the carrier itself has an affinity for the component to be adsorbed and no ligand is immobilized thereon.

As a method for measuring the effective porosity of an adsorbent, the method of flowing a solution through a column packed with an adsorbent was explained in the example.
The concentration may be determined by placing an adsorbent and a solution containing a substance that does not fit into the adsorbent such as latex or a component to be adsorbed in one container, and leaving the solution to reach equilibrium, based on the degree of decrease in concentration.

[Margins below] Table 1 [Effects of the Invention] The adsorbent of the present invention, in which the effective porosity within the adsorbent for the adsorbed and removed components is set in the range of 0.4 to 0.96, has excellent adsorption ability. It has the effect of sufficiently removing harmful components and excessive components.

[Brief explanation of the drawing]

Figures 1 and 2 are graphs showing the relationship between outlet concentration/inlet concentration and liquid volume to explain the method of calculating the effective porosity within the adsorbent, which is the main factor of the present invention. 4 is a graph showing the relationship between the outlet concentration/inlet concentration and the liquid volume to explain the method for quantifying the amount of cholesterol adsorbed by the adsorbent of the present invention. FIG. FIG. 5 is a graph showing the relationship between the effective porosity within the adsorbent and the diffusion coefficient within the adsorbent, and FIG. 6 is a graph showing the relationship between the effective porosity within the adsorbent and the breakthrough time. This is a graph showing. 1st - 2nd Mouth 30 Work 0 α2 0.4 0.6 0.8 1.
0 Effective porosity inside the adsorbent (β) [-] Fig. 5 0 0.2 0.4 0.6 α8! ,0
Effective porosity inside the adsorbent (β)

Claims (1)

[Claims] 1 The effective porosity within the adsorbent relative to the adsorbed and removed component is 0.
An adsorbent characterized in that the adsorbent is in the range of 4 to 0.96.