CA1285925C - ADSORBENT FOR .beta. -MICROGLOBULIN AND IMMUNOGLOBULIN L-CHAIN - Google Patents

Abstract

Abstract of the Disclosure An adsorbent for .beta.2-microglobulin and immunoglobulin L-chain suitable for use in an extracorporeal circulation treatment, which comprises a porous water-insoluble carrier and a compound immobilized onto said carrier, said compound satisfying that the value of log P, in which P is a partition coefficient in an octanol-water system, is not less than 2.5. The adsorbent is inexpensive, and efficiently adsorb and remove .beta.2-microglobulin and immunoglobulin L-chain contained in a body fluid.

Description

~2~3592S

ADSORBENT FOR ~2-MICROGLOBULIN AND
IMMUNOGLOBULIN L-CHAIN

BACKGROUND OF THE INVENTION
The pre~ent invention relates to an adsorbent suitable for use in an extracorporeal circulation treatment, to remove ~2-microglobulin (hereinafter referred to as "32-m") and immunoglobulin L-chain thereinafter referred to as "L-chain") whic`n are contained in a body fluid.
In recent years, many of patients who have been treated by dialysis for a long piriod suffer from carpal tunnel syndrome. "Carpal tunnel syndrome" is a disease attended with symptons o~ paralysis of the median nerve due to compression of the median nerve at the part of carpal tunnel. Lately, it was found out that a ~-fibril-like amyloid protein called "AH" was deposited on the diseased part. Further, it was found out that a precursor protein ¢orresponding to the amyloid is the ~2-m which i8 contained in the patient's blood. However, there hitherto has not been found an effective treatment, especially drug treatment.
32-m~is a low molecular protein (M. W. 11,800) consisting of one hundred of amino acids. The separation method by means of membrane has been used to separate a component in blood or plasma rather selectively according to the size so ~ar. However, there are such dlsadvantages in this method as use~ul protelns except ~2-m may be removed and as the amount of removed ~2-m i9 small. Therefore a more selective and effeetive way~to remo~e a lot of ~2-m is desired.
The~meehod ~or removing ~2-m by means of adsorbent ha8 hasdly been used so far. A few examples of adsorbsnt ~o~r~purification of~B2-m are known, for insta~nce, an~immunoadsorbent in which an anti ~2-m antibody is immobilized onto its carrier, an adsorbent ~285925 based on a principle of a~finity chromatography in which concanavalin A i~ immobilized as a compound showing an affinity for ~2-m (hereinafter referred to as "ligand") onto it9 carrier, and the like. Although these adsorbents show a high selectivity ~or ~2-m, they are not practical for a treatment because of the problems such as expensive anti ~2-m antibody and ligands, for instance, concanavalin A, an unstability in preservation and a difficulty of sterilization.
On the other hand, there is a desease called amyloidosis attended with serious troubles, for example, insufficiency of organs such as heart and kidrey, disturbance of impulse conducting system, progressive dementia, cerebrovascular disease, nerve trouble and so on. Amyloidosis i9 caused by a deposition of amyloid on a blood vessel, a certain organ and so on. It is known that there are some types o~ amyloidosis, i.e. primary amyloidosis, secondary amyloidosis, familial amyloidosis, senile amyloidosis, and so on, and that the composition of proteins causing amyloidosis differs depending on the type of amyloidosis.
Primary amyloidosis is caused by a protein called "AH", and the precursor corresponding to deposited amyloid is supposed to be L-chain. However, there hltherto has not been found an e~fective treatment, especially drug treatment. L-chaih is a low molecular protein (M. W. 23,000) consisting of two hundreds of amino acids in the ~orm of monomer.
Besides the primaly amyloidosis, there are some diseases attended with an extraordinary product o~ L-chain. The typlcal disea~es are multiple myeloma, macroglobulinemia or malignant lymphoma and the highly produced L-chain in these diseases is a cloned protein called Bence Jones protein (hereinafter re~erred to as "BJP"). Usually BJP is excreted into urine. At that time, it inhibits the reabsorption of other protein~, especially albumin, and causes a myeloma kidney syndrone. Also, there are many ca~es to cause , ..~ - . . - , - .

~285g2S

amyloidosis at heart, kidney and so on as a result of a deposition of a lot of BJP in serum. Therefore, an e~fective method for removing BJP in serum is wanted.
However, there has not been found a practical method for removing 90 far, as same as that for primary amyloidosis.
The object of the present invention is to solve the mentioned problems and to provide an inexpensive adsorbent for use in an extracorporeal circulation treatment which i9 able to adsorb and remove a lot of both ~2-m and L-chain.

SUMMARY OF THE INVENTION
In accordance with the present invention, there is provided an adsorbent for ~2-m and L-chain suitable for use in an extracorporeal circulation treatment, which comprises a porous water-insoluble carrier and a compound immobilized onto said carrier, said compound satisfying that the value of log P, in whi¢h P is a partition coefficient in an octanol water system, 19 not less than ` 20 2.5.

Fig. 1 is a graph showing a relation between a flow rate and a pressure drop ~P obtained in Reference Ixample.

DETAILED DESCRIPTION OF THE INVENTION
The adsorbent of the present invention comprises a porous water-insoluble oarrier and a compound ~immobilized onto~said carrier, said compound satisfying that the value of log P is not less than 2.5.
; The logarlthm of a partition coefficient in an oo~tanol-water syst~em, i.e. log P, is a hydrophobic ;parameter of a compound. The partition coefficient P is 35~ obtained acco~rding to the ~following typical method.
First of all, a compound i 9 dissolved in an octanol (or a water) and an equal volume of a water (or an oc~tanal) is adde~d thereto. After shaking for 30 : . - . . . . .
.
.- : . . . . .

~;28592~;

minutes by Criffin flask shaker (made by Gri~fin & George Ltd.), it is centrifuged for from 1 to 2 hours at 2000 rpm. Then concentrations of the compound in both octanol and water layer can be measured by various methods such as spectroscopic method and GLC, and the value of P is obtained according to the following formula:

P - Coct/Cw Coct: a concentration of a compound in an octanol layer Cw: a concentration of a compound in a water layer Before now, many investigators have measured log P of various compounds and the found values were put in order by C. Hansch et. al. ["PARTITION COEFFICIENTS
AND THEIR USES", Chemical Revlews, 71, 525 (1971)]. As to the compounds whose found values are unknown, the calculated values using~a hydrophobic fragmental constant f (hereinafter referred to as "~f"), shown in "THE
HYDROPHOBIC FRAGMENTAL CONSTANT" (Elsevier Sci. Pub.
Com., Amsterdam, 1977) written by R. F. Rekker, can be a ~;~ 25 good guide.
A hydrophobic fragmental constant show the hydrophobicity of various fragments, whlch are determined by a statistioal management of many found values of log P,~ and the total of f of each fragment, i.e. ~f which is 30~ a constitutent of one compound almost corresponds to log In order to find out a compound whlch has an ;ef~fect on ad~o~rbing ~2-m and L-chaln, compounds havlng varlous log~P~are~immobillzed onto a carrier. As a 35~ resu~l~t,~it~ found that~compounds, whose value of log P
are~not l~ess~than 2.50, ~are effective on adsorbing 32-m and L-ch~ain and that compounds, whose value of log P are esB t~han 2.5Q,;hardly show an adsorption ability of 32-m .
.. . . ..

~8S925 and L-chain.
In a ca~e of alkyl amine being immobilized, for example, an ad~orption ability of ~2-m and L-chain is extraordinarily increa~ed by using n-octyl amine (log P =
2.90) instead of u~ing n-hexyl amine (log P - 2.06).
These re~ults indicate that an adsorption of 32-m and L-chain on an ad~orbent of the present invention is caused by the hydrophobic interaction between atomic groups, which are introduced into a carrier through immobilizing compounds who~e value of log P are not les~ than 2.50J and ~2-m and L-chain, and that compound~ who~e value of log P are less than 2.50 have no ad~orption ability on an account of it~ too small hydrophobicity.
In the present invention, compounds immobilized onto a porou~ water-in~oluble carrier, ~atisfying that the value of log P are not less than 2.50 can be u~ed without no particular limitation. However, in a case of immobllizlng a oompound onto a carrier by chemical bond, a part o~ the compound is frequently eliminated. When the elimination group greatly contributes to the hydrophobicity o~ the c~ompound, i.e. when the ~f of the atomic group immobilized onto a carrier by elimlnation is smaller than 2.50, the compound is not ~uited for using at the present lnvention ln vlew of the ob~ect of the present invention. The typlcal example is that benzoic acid isopentylester (~f - 4.15) is immobilized onto a aarrier, having a hydroxy group, by an ester interchange. In this case, the atomic group immobilized 30 ~ onto a carrie;r ls~C6H5C0- and ~f of this group i9 under Thérefore, whether a compound is sufficient as a compound used in the~present inventlon or not may be determlned depending on whether the value of log P, when an elimination part o~ the group i9 substituted by 35;~ hydr~ogen,~i~ les~ than 2.50 or not.
Among~the compounds whose~value of log P i~ not Less than 2.50, compound~ having functional groups which are available for i~mmobilization onto a carrier, e.g.

~ . . . i - ~ . ' . - ~' . . . .

unsaturated hydrocarbons, alcohols, amines, thiols, carbonic acids and its derivatives, halides, aldehydes, hydrazide~, isocyanates, compound~ having an oxirane ring ~uch as glycidyl ethers, halogenated silanes and so on 5 are preferable. The typical examples of these compounds are amines such as n-heptylamine, n-octylamine, decylamine, dodecylamine, hexadecylamine, octadecylamine, 2-aminooctene, naphtylamine, phenyl-n-propylamine and diphenylmethylamine, alcohols such as n-heptylalcohol, n-10 octylalcohol, dodecylalcohol, hexadecylalcohol, 1-octene-3-ol, naphtol, diphenylmethanol and 4-phenyl-2-butanol, glycidylethers thereof, carbonic acids such a~ n-octanoic acid, nonanoic acid, 2-nonenoic acid, decanoic acid, dodecanoic acid, stearic acid, arachidonic acid, oleic 15 acid, diphenylacetic acid and phenylpropionic acid, carbonic acid derivatives thereof such as acid halides, ester and amide, halides such as octylchloride, octylbromide, decylchloride and dodecylchloride, thiols ~uch as octanethiol and dodecanethiol, halogenated 20 silanes such as n-octyltrichlorosilane and octadecyltrichlorosilane, and aldehydes such as n-octylaldehyde, n-caprinaldehyde and dodecylaldehyde.
Moreover, among the above-mentioned compounds whose hydrogen in their hydrocarbon moiety is substituted 25 by substituents having hetero atoms such as hoalogen, nitrogen, oxygen and sulfur, other alkyl groups and 90 on, the compounds whose log P are not less than 2.50 can be used. And the compounds shown on the tables of from page 555 to page 613 in the above mentioned reference, 30 "PARTITION COEFFICIENTS AND THEIR USES", satisfying that the value of log P are not less than 2.50 can also be used. However, the present invention is not limited thereto.
These compound may be used alone, in 35 combination thereof~or even together with a compound having a value of log P of less than 2.5.
The typical examples of the water-insoluble carriers in the present invention are inorganic carrlers : :
: :

. - ~ . . .
.
.

~;~8592~i such as glas~beads and silicagel, organic carriers made - of synthetic polymers such as cross linked polyvinylalcohol, cross linked polyacrylate, aross linked polyacrylamide, cro~s linked polystyrene and polysaccharides ~uch as crystal cellulose, cross linked cellulose, cross linked agaro~e and cross linked dextran, and complex carrier obtained by combining them, e.g.
organic-organic carrier, organic-inorganic carrier and so on. Especia11y, hydrophilic carriers are preferable since non-specific adsorption is rather small and the adsorption selectivity for ~2-m and L-chain is good.
The term "hydrophilic carriers" means the carriers which contact with water at an angle of not more than 60 when compounds constltuting the carriers are made plane. Typical examples of these carriers are made of cellulose, polyvinylalcohol, saponi~icated compound of ethylene-vinyl acetate vinyl copolymer, polyaarylamide, polyacrylic acid, polymethacrylic acid, polymethyl methacrylate, acrylic acid grafted polyethylene, acrylamide grafted polyethylene, glass and 90 on. In particular, the porous cellulose gels are preferred for the reasons that (1) they are hard to be broken into pieces or fine powder by operation such as agitation beoause of a relatively high mechanical strength and toughness, and also even if a body fluid is passed through~ a column packed therewith at a high flow rate, oonsolidation and choking do not ocour and, thererore, it 19 pQssible to pass a body fluld at a high flow rate, and further the pore structure is hard to suffer change upon high pressure steam sterilization, (2) the gels are hydrophilic because Or being made of cellulose and plenty of~hydroxyl group utilizable for bonding the sulfated palysaccharide, and also nonspecific adsorption is a lL~ttle,~(3) an adsorption capacity comparable to that of 35~the soft gel4 i8 obtained, because the strength i9 maintained relatively~high even if the porosity volume i~
incr;easej and (4)~the safety i9 high as compared with the ynthetic polymer gel~. However, the present invention - ~ ~

~2~5 is not limited thereto. The above-mentioned carriers may b~e used either alone or in combination thereof volantarily.
The mo~t essential property for a water-insoluble carrier of the pre~ent invention is to containa lot of pore~ having a preferable size, i.e. to have a porous structure. ~2-m and L-chain, both of which are objects to be removed by the adsorbent of the present invention, are proteins having molecular weights 11,800 and 23,000 respectively as mentioned above. And it is preferable for an effective adsorption of these proteins that ~2-m and L-chain can enter the pores of adsorbent at a certain high probability, but on the other hand, that other proteins can hardly enter the pores. There are many methods for measuring pore size. Though mercury porosimetry is most widely used now, sometimes it cannot be applied to the porous water-insoluble carrier of the present invention. In such case, it i9 proper to use the molecular weight of the excluslon limit as a measure of the pore size.
The term "the molecular weight of the exclusion limit" means, for instance, as described in the literature such as "Jikken Kosoku Ekitai Chromatography (Experimental High Performance Liquid Chromatography)"
written by Hiroyuki Hatano and Toshihiko Hanai, pùblished by Kabu~hiki Kaisha Kagaku Do~in, the minimum molecùlar weight of the molecule which cannot permeate into a pore, i.e. which i~ excluded, in a gel permeation chromàtography.
The moleoular weight of the exclusion limit is generally well~studied about globular protein, dextranj polyethylene~-glycol and 80 on and, in case o~ the carrier of the present inventi~on, it is proper to use the value obtained by globular protein.
3~5~ As-~the r~esult of an investigation using oarriers having various molecular welghts of the exclusion limit, it is shown that the preferable range of pore size indicated~by molecular weight of the exclusion ~..2~S925 limit i~ ~rom 1 x 104 to 6 x 105. That is, it is ~ound that a carrier having a molecular weight of the exclusion limit of less than 1 x 104 can hardly adsorb the ~2-m and L-chain and i~ not suited for practical use and that a carrier having a molecular weight of exclusion limit of over 6 x 105 adsorbs a large amount of proteins other than ~2-m and L-chain, mainly albumin, and is not suited for practical u~e in the point of selectivity.
Therefore, the moleculr weight of the exclusion limit of the carrier employed in the present invention is preferably from 1 x 104 to 6 x 10 5, more preferably from 2 x 104 to 3 x 105.
With respect to the porous structure of a carrier, a structure uniformly having pores at any part of the carrier is more preferable than a structure having pores only on the surface from the viewpoint of adsorption capacity per unit volume. And it is preferred that the porosity of a ¢arrier is not less than 20 ~ and the specific surface area is not less than 3 m2/g.
The 9hape of a carrier can be optionally selected from such shapes as particle, fiber and hollow fiber.
In the present invention, it is preferable that there are some functional group9 which are available for immobilizing a ligand onto the surface of the carrier.
Typical examples of the above functional groups are hydroxyl group, amino group, aldehyde group, carboxyl group, thiol group, silanol group, amide group, epoxy group, halogen group, succinimide group, acid anhydride 30~ group and the like.
Both a~soft carrier and a hard carrier may be employed~in the~ pre9ent invention. However, when the carrier is used as an adsorbent for an extracorporeal c~irc~ulation treatment, it i9 important that the carrier 35~;~ has a sufficlent~mechanical strength 90 as to prevent consolidation when a body fluid is passed through a column packed with the adsorbent.
Therefore~, it is preferable that a carrier :: . : ~ .: , ; , ' . -~2~35925 employed in the present invention is a hard carrier. The term "hard carrier" means, as shown in the Reference Example herein below, that the relation between a pressure drop ~P and a flow rate determined by passing an aqueous fluid through a cylindrical column uniformly packed with the gel, such as particle gel, keeps a linear relationship until the pressure drop is increased to 0.3 kg/cm .
The adsorbent of the present invention is obtained by immobilizing a compound, whose value of log P
is not less than 2.50, on a porous water-insoluble carrier and there are many known methods available for immobilization without any particular limitation.
However, it i9 important in view of the safety to minimize the elimination and elution of ligands during a sterilization or a treatment because the adsorbent of the present invention is employed for an extracorporeal circulation treatment. Therefore, it is most preferable that a compound is immobilized onto a carrier by a covalent bond.
There are many ways of using the adsorbent of the present invention in a treatment. For the most simple example, the adsorbent of the present invention can be used as follows: i.e. partient's blood is introduced outside of hiq body so as to be put into a blood bag and then mixed with the adsorbent of the present invention to remove ~2-m and L-chain, followed by removing the adsorbent through filter. Consequently, the blood treated in this way is returned back to the patient .
himself. Though this method does not need an intricate apparatus, there are some defects such as a small amount of a treated blood at one time, a lot of time for treatment and a complicated operation.
For another method, a column packed with the 3~5~ adsorbent of the present invention is incorporated into an ex~tracorporeal circulation clrouit, and then removal o~ 32-m and L-chain by adsorption is taken by on-line sy~stem. There are two tre~atment methods: i.e. one is ~. : ,:

.

. , . . .. , . .. - . . . .

i~85925 that whole blood i5 directly perfused and another is that only plasma separated from the blood is passed through the colu~n.
The adsorbent of the present invention can be used in any of above method~ and, as mentioned above, on-line system is most preferable.
Present invention is more specifically described and explained by the following Re~erence Example, Examples and Comparative Examples. It is to be understood that the present invention is not limited to the Reference Example, Examples and Compar~tive Examples and various changes and modifications can be made without departins Lrom the scope and spirit of the present invention.
eference Example A relation between a flow rate and a pressure drop ~P was determined by passing water by means of a peristaitic pump through cylindrical glass columns (inner diametar: 9mm, column length: 150 mm) equipped at both ends with filters having a pore size of 15 ~m, in which an agarose gel (Biogel A5m made by Biorado Co., particle size: 50 to 100 mesh) a vinyl polymer gel (Toyopearl HW-65 made by Toyo Soda Manufacturing Co., Ltd., particle size: 50 to 100 ~m, and a cellulose gel (Cellulofine GC-~; 700m made by Chisso Corporation, particle size: 45 to 105 ~m) were packed respectively. The results are shown ~0 in Fig. 1.
As shown in Fig. 1, an increase o~ a flow rate ~::: ::
i9 nearly proportional to that of a pressure drop in case of Toyopearl HW-65 and Cellulofine GC-700m, whereas in case of Biogel A-5m, consolidation occurrs and a flow rate does not increase even if a pressure drop increases. In the present invention, the term "hard gel"
means a gel having the relation between a pressure drop ~P and a ~low rate keeps a liner relationship until the pressure drop is increased to 0.3 kg/cm2, as in the former case of Toyopearl HW-5 and Cellulofine GC-700m.

* Trade-mark ~28592S

Example 1 After water was added to 170 mQ of a porous cellulose hard gel Cellulofine GC-200m (made by Chisso Corporation, exclusion limit of globular proteins:
120,000) to make the whole volume 340 mQ, 90 mQ of 2M
aqueous solution of sodium hydroxyde is added thereto and the temperature of reaction mixture was adjusted to 40C. Then there was added 31 mQ of epichlorohydrin and the reaction mixture was reacted with stirring for 2 hours at 40 C. After the reaction, the resultant was thoroughly washed with water to give an epoxy-activated gel.
To 10 mQ of the obtained epoxy-activated gel was added 200 mg of n-octylamine (log P - 2.90). After 6 days reaction by allowing it to stand in a 50 % by volume aqueous solution of ethanol at 45C, the resultant was throughly washed with a 50 % by volume aqueous solution of ethanol, ethanol, again a 50 % by volume aqueous solution and then water to give an n-octylamine-immobilized gel.
To 0.5 m~ of the obtained n-octylamine-immobilized gel was added 2 m~ of serum of a patient who is under dialysi~ treatment, the serum containing ~2-m in a concentration of 65 ~g/mQ and the mixture was incubated for 2 hours at 37C. The concentrations of 3 -m and : albumin in the supernatant were measured to obtain the amount of adsorbed 32-m, ratio of adsorbed 32-m based on:32-m added first and amount of adsorbed albumin.
The results are shown ln Table 1.
30::~
Example 2 : The procedures in Example 1 were repeated except that 3;15~:mg of dodecylamine (~f - 5.10) was used lnstead of n-oe~tylamlne to glve a dodecylamlne-`35~ immobilized~gel:.:
~ ;-The~adsorption of the~obtained dodecylamine-immobilized gel: was~examined by means of the same : procedure:s as:in Example 1.

.. ,~ , . .. .. ..

-, ~2~3592S

The result is ~hown in Table l.

Example 3 Tne procedure~ in Example 1 were repeated except that 170 mQ of a porous cellulose hard gel Cellulofine GC-700m (made by Chisso Corporation, exclu~ion limit of globular proteins: 400,000) was used as a carrier instead of Cellulofine GC-200m to give an n-octylamine-immobilized gel.
The adsorption of the obtained n-octylamine-immobilized gel was examined by means of the same procedures as in Example 1.
The re~ult is shown in Table 1.

Example 4 The procedure~ in Example 3 were repeated except 315 mg of that dodecylamine was used in~tead of n-octylamine to give a dodecylamine-lmmobilized gel.
The adsorptlon of the obtained dodecylamine-immobiliæed gel was examined by means of the ~ame procedures as in Examplle 1.
The results is shown in Table 1.

Example 5 The procedures in Example 3 were repeated except that 410 mg of cetylamine (~f - 7.22) was used instead of n-ootylamine and that ethanol was u~ed as a .:
solvent in immobilizatlon instead o~ a 50 S by volume aqueous ~olution of ethanol to give a cetylamine-immobilized gel.
The adsorption of the obtained cetylamine-immobilized gel was examined by mean~ of the same procedures in Example l.
The result is shown in Table 1.
; Example 6 The procedures in Example 2 were repeated except that 170 m~ of a porous cellulo~e hard gel ., ,, ~ . ~ . . . . - - . ~ , . - .

~2~35925 Cellulofine GCL-300m (made by Chisso Corporation, exclu~ion limit of globular protein~: 90,000) was used as a carrier instead of Cellulofine GC-200m to give a dodecylamine-immobilized gel.
The ad~orption of the obtained dodecylamine-immobilized gel was excamined by means of the qame procedureq as in Example 1.
The result i_ shown in Table 1.

Example 7 The procedures in Example 1 were repeated except that 1 T mQ of a porouq cellulo_e hard gel Cellulofine GC-lOOm (made by Chisso Corporation, exclusion limit of globular protein~: 60,000) was used as a carrier instead of Cellulofine GC-200m to give an n-octylamine-immobilized gel.
The adsorption of the obtained n-octylamine-immobilized gel wa9 examined by mean9 of the same procedure~ as in Example 1.
' The result i 9 shown in Table 1.

Example 8 The prooedures ln Example 1 were repeated exoept that 170 mQ of a porous cellulose hard gel Cellulofine GCL-9Om (made by Chi~qso Corporation, excluqion limit of globular proteins: 35,000) waq used as ; a~carrler instead of Cellulofine GC-200m to give an n-octylamine-immobilized gel.
M ~ ~ ~ The ad~orption of the obtained n-octylamine-30~ lmmobilized~gel was~-~examined by mean_ o~ the same p~rocedures~as in~Example 1.
The~reqult is qhown in Table 1.

Example 9 35~ The~procedures in; Example 1 were repeated e~xoept~that 1~7~0~mQ~or~a porous vlnyl polymer hard gel Toyo~pearl HW-SO~o~oarse ~made;by Toyo soda Manufacturing Co.~ td.,'~exolusion limit of~globular proteins: 80,000) .... .. .. . . . . .... .. ..

~2~35925 was used as a carrier instead of Cellulofine GC-200m to give an n-octylamine-immbolized gel.
The adsorption of the obtained n-octylamine-immobilized gel was examined by means of the same procedures a~ in Example 1.
The result is shown in Table 1.

Example 10 The procedures in Example 1 were repeated except that 170 mQ o~ a porous cellulose hard gel Cellulofine GCL-300m was used as a carrier instead of Cellulo~ine GC-200m and that 200 mg of dodecylamine was used instead of n-octylamine to give a dodecylamine-immobilized gel.
To 0.5 mQ of the obtained dodecylamine-immobilized gel was added 3 mQ of plasma of a IgA myeloma patient, the plasma containing Bence Jones protein which i9 the immunoglobulin L-chain in a concentration of 200 ~g/mQ, and the mixture was incubated for 2 hours at 3~C. The concentrations of 8JP and albumin in the supernatant were measured to obtain the amount of adsorbed BJP, ratio of adsorbed BJP based on BJP added first and amount o~ adsorbed albumin.
The~results are shown in Table 2.

Example 11 The procedures in Example 10 were repeated except that 410 mg o~ cetylamine was used instead of dodecylamine to give a oetylamine-immobilized gel.
30~ The ad~sorption of the obtained oetylamine-immobilized ge~l wa9~examined by means of the same pro~cedure~s~as~in~Example 10.
The~result is ~shown in Table 2.

35~ Example 12 The~procedures in Example 10 were repeated exo~ept~that~170~mQ~of a~porous oellulose hard gel ellu1ofi~nè~OC-700~m~was used as a oarrler instead of ~Z~3592~;.

Cellulofine &CL-300m and that 220 mg of n-octylamine was used instead of dodecylamine to give an n-octylamine-immobilized gel.
The adsorption of the obtained n-octylamine-immobilized gel was examined by means of the sameprocedures as in Example 10.
The re~ult is shown in Table 2.

Example 13 The procedures in Example 12 were repeated except that 315 mg of dodecylamine was used instead of n-octylamine to give a dodecylamine-immobilized gel.
The adsorption of the obtained dodecylamine-immobilized gel was examined by means of the same procedures as in Example 10.
The result is shown in Table 2.

Example 14 The procedures in Example 12 were repeated eXcept that 410 mg of cetylamine was u9ed instead of n-octylamine to give a cetyIamine-immobilized gel.
The adsorption of the obtained cetylamine-mmobilized gel~was examined by means o~ the same prooedure~ as in Example 10.
2~5 The result is~3hown in Table 2.

; Example 15 The procedures in Example 10 were repeated exoept that 170~mQ of a porous oellulose hard gel Cellulofine GC-200m wa~ u9ed a9~a carrier instead of C~el;lulofine GCL-300m to give a dodecylamine-immobilized The~adsorption of the obtalned dodecylamine-immobl1l~zed~gel wa~s examined by means o~ the same 35 ~procedures~as in~Example 10.
The~result 1s shown~in Table 2.

Example 16 :~ :~ . - , . , . - . , . : - .

The procedures in Example 15 were repeated except that 410 mg of cetylamine was used instead of dodecylamine to give a cetylamine-immobilized gel.
The adsorption of the obtained cetylamine-immobilized gel wa~ examined by means o~ the sameprocedures as in Example 10.
The result is shown in Table 2.

Comparative Example 1 The procedures in Example 3 were repeated except that 172 mg of n-hexylamine (log P - 2.06) was u~ed instead of n-octylamine to give an n-hexylamine-immobilized gel.
The adsorption of the obtained n-hexylamine-immobilized gel was examined by means of the same procedures as in Example 1.
The result is shown in Table 1.
.
Comparative Example 2 : ~ 20 The pro¢edures in Example 3 were repeated except that 125 mg of n-butylamlne (log P - 0.97) was used instead of:n-o¢tylamlne and that water was used as a solvent in immobilization instead of a 50 % by volume aqueous so1ution of ethanol to give an n-butylamine-immobilized g~e1.
The adsorption of the obtained n-butylamine-immobilized gel wa9 examlned by mean9 of the same procedures as in Example 1.
The re3u1t 19 shown in Table 1.

Comparative Example 3 The;p:rocedures :ln ExampIe 10 were repeated exc~ept that 170~mQ of a porous cellulose hard gel . `:`Celluloflne~GC-7QOm~wa9~u9ed as a car~rler lnstead of 35,~ :C~ellùlofine~:GCL-300m and~that 500 ~Q of 70 % by welght a~que~ous:so:1utio'n~of ethyla~mlne (log P - -0.13) was used in:s~t,ead of~d:odecylamine to give an ethylamine-lmmobillzed ~2~3~92 The adsorbent of the obtained ethylamine-immobilized gel was examined by means of the same procedures as in Example 10.
The result i9 shown in Table 2.

Comparative Example 4 The procedure~ in Comparative Example 3 were repeated except that 125 mg of n-butylamine was used instead of ethylamine to give an n-butylamine-immobilized gel.
The adsorbent of the obtained n-butylamine-immobilized gel was examined by means of the same procedures as in Example 10.
The result i9 shown in Table 2.

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~28S9æ~i As ~hown in Tableq 1 and 2, the adsorbent of the present invention e~fectively adsorbs both ~2-m and L-chain, while albumin is hardly adsorbed.
In addition to the ingredients used in the Examples, other ingredients can be used in the Examples a~ set forth in the specification to obtain substantially the same re~ults.

, . : , .: ,. .

Claims (4)

1. An adsorbent for .beta.2-microglobulin and immunoglobulin L-chain suitable for use in an extracorporeal circulation treatment, which comprises a porous water-insoluble carrier and a compound immobilized onto said carrier, said compound satisfying that the value of log P, in which P is a partition coefficient in an octanol-water system, is not less than 2.5.

2. The adsorbent of Claim 1, wherein said porous water-insoluble carrier has a molecular weight of the exclusion limit measured with a globular protein from 1 x 104 to 6 x 105.

3. The adsorbent of Claim 1, wherein said porous water-insoluble carrier is a hydrophilic carrier.

4. The adsorbent of Claim 1, wherein said porous water-insoluble carrier is a hard carrier.