MULTIMODAL ADSORPTION MEDIUM WITH MULTIMODAL LIGANDS, METHOD FOR THE PREPARATION AND USE THEREOF

Abstract

The present invention relates to a multimodal adsorption medium, in particular a multimodal chromatography medium, a method for its production, as well as use of the adsorption medium according to the invention or an adsorption medium produced according to the invention for the purification of biomolecules.

Claims

1. A multimodal adsorption medium, comprising a polymeric carrier material C to which multimodal ligands of the following structure -G-(CO.sub.2H)n are covalently bonded via an X(CO) group, ##STR00029## where X denotes NR, O or S and R denotes alkyl, alkenyl, aryl, heteroaryl or hydrogen, G denotes a group selected from the group composed of a branched or unbranched C.sub.2-20 alkyl group, a substituted or unsubstituted C.sub.3-10 cycloalkyl group, a branched or unbranched C.sub.2-20 alkenyl group, a substituted or unsubstituted C.sub.6-20 aryl group, and a substituted or unsubstituted C.sub.4-20 heteroaryl group, wherein n is a whole number that is 1 or higher.

2. The multimodal adsorption medium as claimed in claim 1, wherein the X(CO) group is NH(CO),

3. The multimodal adsorption medium as claimed in claim 1, wherein the polymeric carrier material C comprises at least one material selected from the group composed of natural or synthetic fibers, (polymer) membranes, porous, polymeric monolithic molded bodies, polymer gels, films, nonwovens and wovens.

4. The multimodal adsorption medium as claimed in claim 1, wherein the multimodal ligands are bound to the surface of the carrier material C via polymeric spacer elements.

5. The multimodal adsorption medium as claimed in claim 4, wherein the polymeric spacer elements are polyamines with at least one primary amino group, which as an X(CO) bond forms an amide bond with the multimodal ligands,

6. The multimodal adsorption medium of claim 1, wherein the G group is a branched or unbranched C.sub.4-20 alkyl group or a branched or unbranched C.sub.3-20 alkenyl group.

7. The multimodal adsorption medium of claim 1, wherein the multimodal ligands have the following structure: ##STR00030## wherein G is a substituted or unsubstituted C.sub.2-3 alkyl group, a substituted or unsubstituted C.sub.3-10 cycloalkyl group, a substituted or unsubstituted C.sub.2-3 alkenyl group, a substituted or unsubstituted C.sub.6 aryl group or a substituted or unsubstituted five-membered or six-membered heteroaromatic group, wherein the substituents are selected from the group composed of a branched or unbranched C.sub.1-10 alkyl group, a branched or unbranched C.sub.2-10 alkenyl group, a C.sub.6-20 aryl group, and a C.sub.4-20 heteroaryl group.

8. The multimodal adsorption medium as claimed in claim 7, wherein G denotes a branched or unbranched C.sub.3-10 alkenyl group.

9. The multimodal adsorption medium of claim 1, wherein the multimodal ligands have one of the following structures: ##STR00031## wherein R is selected respectively from the group composed of hydrogen, F, Cl, Br, I, OH, NH.sub.2, SH, CO.sub.2H, a branched or unbranched C.sub.1-10 alkyl group, a branched or unbranched C.sub.2-10 alkenyl group, a C.sub.6-20 aryl group, and a C.sub.4-20 heteroaryl group, wherein A is a C.sub.6 aryl group or a five-membered or six-membered heteroaromatic group, and wherein m is a whole number from 1 to 3.

10. A method for producing an adsorption medium as claimed in claim 1, comprising the following steps: (a) providing a polymeric carrier material C, wherein the carrier material C has at least one XH group that is reactive with carboxylic acid derivatives while forming a covalent bond X(CO), where X denotes NR, O or S and R denotes alkyl, alkenyl, aryl, heteroaryl or hydrogen; and (b) reacting the at least one XH group of the polymeric carrier material C with a carboxylic acid derivative as a precursor of a multimodal ligand such that the covalent bond X(CO) is formed via which the multimodal ligand is bonded to the carrier material C.

11. The method as claimed in claim 10, wherein the covalent bond X(CO) is a secondary amide bond that is formed by reacting a carboxylic anhydride as a precursor of the ligand and amine groups of the carrier material C, and wherein the multimodal ligand has at least one free carboxylic acid group.

12. Use of the multimodal adsorption medium of claim 1 or an adsorption medium produced by the method as claimed in claim 10 for the purification of biomolecules.

13. Use as claimed in claim 12, wherein the biomolecules are proteins, peptides, amino acids, nucleic acids, viruses, virus-like particles and/or endotoxins,

14. Use as claimed in claim 13, wherein the proteins are antibodies.

15. The multimodal adsorption medium of claim 1, wherein the G group denotes a group selected from the group composed of a branched or unbranched C.sub.2-20 alkyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one aromatic substituent, a substituted or unsubstituted C.sub.3-10 cycloalkyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one aromatic substituent, a branched or unbranched C.sub.2-20 alkenyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one aromatic substituent, a substituted or unsubstituted C.sub.6-20 aryl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and a substituted or unsubstituted C.sub.4-20 oheteroaryl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens,

16. The multimodal adsorption medium of claim 6, wherein the G group denotes a group selected from the group composed of a branched or unbranched C.sub.4-20 alkyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one aromatic substituent, and a branched or unbranched C.sub.3-20 alkenyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one aromatic substituent.

17. The multimodal adsorption medium of claim 7, wherein G is a substituted or unsubstituted C.sub.2-3 alkyl group, a substituted or unsubstituted C.sub.3-10 cycloalkyl group, a substituted or unsubstituted C.sub.2-3 alkenyl group, a substituted or unsubstituted C.sub.6 aryl group or a substituted or unsubstituted five-membered or six-membered heteroaromatic group, wherein the substituents are selected from the group composed of a branched or unbranched C.sub.1-10 alkyl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one hydroxyl, carbonyl, carboxyl, carboxylic anhydride or aromatic substituent, a branched or unbranched C.sub.2-10 alkenyl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one hydroxyl, carbonyl, carboxyl, carboxylic anhydride or aromatic substituent, a C.sub.6-20 aryl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and at least one hydroxyl, carbonyl, carboxyl, or carboxylic anhydride substituent, and a C.sub.4-20 heteroaryl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and at least one hydroxyl, carbonyl, carboxyl, or carboxylic anhydride substituent, and a hydroxy, thiol or amino group.

18. The multimodal adsorption medium as claimed in claim 17, wherein G denotes a branched or unbranched C.sub.3-10 alkenyl group that contains one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one hydroxyl, carbonyl, carboxyl, carboxylic anhydride or aromatic substituent.

19. The multimodal adsorption medium of claim 9, wherein R is selected respectively from the group composed of hydrogen, F, Cl, Br, I, OH, NH.sub.2, SH, CO2H, a branched or unbranched C.sub.1-10 alkyl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and/or optionally at least one hydroxyl, carbonyl, carboxyl, carboxylic anhydride or aromatic substituent, a branched or unbranched C.sub.2-10 alkenyl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one hydroxyl, carbonyl, carboxyl, carboxylic anhydride or aromatic substituent, a C.sub.6-20 aryl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and/or at least one hydroxyl, carbonyl, carboxyl, or carboxylic anhydride substituent, and a C.sub.4-20 heteroaryl group containing one or a plurality of heteroatoms selected from O, S, N and halogens, and at least one hydroxyl, carbonyl, carboxyl, or carboxylic anhydride substituent, wherein A is a C6 aryl group or a five-membered or six-membered heteroaromatic group, and wherein m is a whole number from 1 to 3.

Description

EXAMPLES

Methods

M1: Determination of Ligand/Charge Density of Amine-Functionalized Adsorption Media

[0072] Three membrane layers were clamped into a membrane holder. The membrane stack had a membrane area of 15 cm.sup.2, an inflow area of 5 cm.sup.2 and a bed height (thickness of the membrane stack) of 750 m in the membrane holder. The membranes in the membrane holder were flooded with 20 mM TRIS/HCl buffer at pH=7.4 in order to displace the air and then connected to an kta Explorer 100 FPLC unit from the firm General Electric Health Care. The membranes or the membrane stack were then tested for charge density using a test program comprising four steps.

[0073] The four steps of the test program are given below:

[0074] 1. conditioning of the membrane with 6 ml of 1 M NaCl solution in 20 mM TRIS/HCl at pH=7.4

[0075] 2. regeneration of the membrane with 6 ml of a 1 M solution of NaOH in RO water

[0076] 3. washing of the membrane with 100 ml of RO water and

[0077] 4. loading of the membrane with 135 ml of 10 mM HCl.

[0078] All of the steps were carried out with a flow rate of 10 mL/min. In all of the steps, conductivity was measured in the detector behind the membrane unit. The area above the curve thus recorded was integrated after subtracting the dead volume, and the charge density was calculated therefrom.

M2: Determination of Ligand/Charge Density of Cation Exchange Adsorption Media

[0079] Three membrane layers were clamped into a membrane holder. The membrane stack had a membrane area of 15 cm.sup.2, an inflow area of 5 cm.sup.2 and a bed height (thickness of the membrane stack) of 750 m in the membrane holder. The membranes in the membrane holder were flooded with 20 mM KPi-Puffer at pH=7 in order to displace the air and then connected to an kta Explorer 100 FPLC unit from the firm General Electric Health Care. The membranes or the membrane stack were then tested for charge density using a test program comprising four steps. The four steps of the test program are given below:

[0080] 1. conditioning of the membrane with 6 ml of 1 M NaCl solution in 20 mM KPi at pH=7.0

[0081] 2. regeneration of the membrane with 6 ml of a 1 M solution of HCl in RO water

[0082] 3. washing of the membrane with 88 ml of RO water and

[0083] 4. loading of the membrane with 135 ml of 10 mM NaOH.

[0084] All of the steps were carried out with a flow rate of 10 mL/min. In all of the steps, conductivity was measured in the detector behind the membrane unit. The area above the curve thus recorded was integrated after subtracting the dead volume, and the charge density was calculated therefrom.

M3: Determination of Binding Capacity for Lysozyme of Modified Membranes by Means of Breakthrough Curve

[0085] Three membrane layers were clamped into a membrane holder. The membrane stack had a membrane area of 15 cm.sup.2, an inflow area of 5 cm.sup.2 and a bed height (thickness of the membrane stack) of 900 m in the membrane holder. The membranes in the membrane holder were flooded with 10 mM KR-Puffer at pH=7 in order to displace the air and then connected to an kta Explorer 100 FPLC unit from the firm General Electric Health Care. The membranes or the membrane stack were then tested with a test program comprising three steps with respect to lysozyme-binding capacity. The three steps of the test program are given below:

[0086] 1. conditioning of the membrane with 20 ml of 1 M NaCl solution in 10 mM KPi at pH=7.0

[0087] 2. equilibration of the membrane with 20 ml of binding buffer (10 mM KPi, pH=7.0)

[0088] 3. loading of the membrane with 250 ml of lysozyme solution (0.20% lysozyme in binding buffer).

[0089] All of the steps were carried out with a flow rate of 10 mL/min. In all of the steps, absorption at 280 nm was measured in the detector behind the membrane unit. The area above the curve thus recorded was integrated after subtracting the dead volume, and the amount of bound lysozyme was calculated therefrom.

M4: Determination of Binding Capacity for -Globulin of Modified Membranes by Means of Breakthrough Curve

[0090] Three membrane layers were clamped into a membrane holder. The membrane stack had a membrane area of 15 cm.sup.2, an inflow area of 5 cm.sup.2 and a bed height (thickness of the membrane stack) of 900 m in the membrane holder. The membranes in the membrane holder were flooded with 20 mM NaAc solution at pH=5 in order to displace the air and then connected to an kta Explorer 100 FPLC unit from the firm General Electric Health Care. The membranes or the membrane stack were then tested with a test program comprising three steps with respect to -globulin-binding capacity. The three steps of the test program are given below:

[0091] 1. conditioning of the membrane with 20 ml of 1 M NaCl solution in 20 mM NaAc at pH=5.0

[0092] 2. equilibration of the membrane with 20 ml of binding buffer (25 mM NaCl in 20 mM NaAc at pH=5.0)

[0093] 3. loading of the membrane with 250 ml of 1 mg/mL -globulin solution in binding buffer.

[0094] All of the steps were carried out with a flow rate of 10 mL/min. In all of the steps, absorption at 280 nm was measured in the detector behind the membrane unit. The area above the curve thus recorded was integrated after subtracting the dead volume, and the amount of bound y-globulin was calculated therefrom. The measurement was repeated using a fresh membrane sample with 150 mM NaCl and 300 mM NaCl.

Modification Protocol for the Immobilization of Carboxylic Anhydrides on Amine-Modified Starting Matrices

Modification of Cellulose Hydrate Membranes

[0095] 1. Polyamine Immobilization

[0096] 1a) Polyallylamine (PAA)

[0097] The spacer immobilization is based on a known protocol, which was described in DE 10 2008055 821 A1 (examples 21 and 22). In this case, spacers with a molar mass of 15,000 g/mol to 150,000 g/mol are used. In a typical reaction, the cellulose acetate (CA) membrane (3 m pore size, Sartorius Stedim Biotech GmbH) was saponified in a 0.6 M aqueous sodium hydroxide solution (4 g/cm.sup.2) for 30 min at room temperature and then rinsed three times for 10 min in a 0.25 M sodium hydroxide solution (0.5 g/cm.sup.2), The membrane obtained was treated for 30 min with a solution composed of 15% 1,4-butanediol diglycidyl ether and 85% 0.25 M aqueous sodium hydroxide solution (0.5 g/cm.sup.2) and then stored for 18 h in a sealed container at room temperature. Finally, rinsing was carried out for 30 min with running water.

[0098] The membrane thus obtained was treated for 1 h with a 20% solution of polyallylamine in RCS water (1 g/cm.sup.2) at 50 C. The membrane was then treated for 5 min at room temperature with 5% sulfuric acid solution and finally rinsed for 10 min with running water.

[0099] The amino group density on the membrane was determined by titration.

TABLE-US-00001 Carrier material Amino group density PAA-modified cellulose hydrate membranes 450-550 mol/mL

[0100] 1b) Polyethyleneimine (PEI)

[0101] Spacer immobilization is carried out based on a known protocol, which was described in DE 102008055 821 A1 (examples 15, 16 and 17). In a typical reaction, the CA membrane (3 pm pore size, Sartorius Stedim Biotech GmbH) was saponified in a 0.6 M aqueous sodium hydroxide solution (4 g/cm.sup.2) for 30 min at room temperature and then rinsed three times for 10 min in a 0.25 M sodium hydroxide solution (0.5 g/cm.sup.2). The membrane obtained was treated for 30 min with a solution composed of 15% 1,4-butanediol diglycidyl ether and 85% 0.25 M aqueous sodium hydroxide solution (0.5 g/cm.sup.2) and then stored for 18 h in a sealed container at room temperature. Finally, rinsing was carried out for 30 min with running water. The membrane thus obtained was treated for 2 h with a 30% solution of Lupasol WF (polyethyleneimine from BASF AG, molecular mass 25000 g/mol) in RO water (1 g/cm.sup.2) at 50 C. The membrane was then rinsed for 30 min with running water, treated for 10 min with 5% sulfuric acid solution, and finally rinsed for 10 min with running water.

[0102] The amino group density on the membrane was determined by titration.

TABLE-US-00002 Carrier material Amino group density PEI-modified cellulose hydrate membranes 600-650 mol/mL

[0103] 2. Ligand Immobilization

[0104] In a typical reaction, 16 g of carboxylic anhydride was dissolved in 64 g of DMSO (20 wt %) and the solution was heated to 60 C. The PAA-modified cellulose hydrate membrane was placed in the reaction solution (0.5 g/cm.sup.2) and agitated at 60 C. for 1 h. The reaction solution was then filtered off, and the membrane was washed with ethanol (0.5 g/cm.sup.2) and a large excess of RO water.

[0105] Ligand Structures:

[0106] The cation exchangers listed here were produced according to the above-described method, wherein the following carboxylic anhydrides were used. The results are shown in Tables 1 through 3 and FIGS. 1 through 3 below.

TABLE-US-00003 Example Anhydride 1 Succinic anhydride 2 Glutaric anhydride 3 Malic anhydride 4 Itaconic anhydride 5 Maleic anhydride 6 Phthalic anhydride 7 Quinolinic anhydride 8 Trimellitic anhydride 9 Pyromellitic anhydride 10 4((2,5-dioxotetrahydrofuran-3-yl)thio)benzoic acid 11 N-(2,5-dioxotetrahydrofuran-3-yl)acetamide 12 N-(2,5-dioxotetrahydrofuran-3-yl)-2,2,2-trifluoroacetamide 13 Maleic anhydride 14 Maleic anhydride 15 Succinic anhydride 16 Maleic anhydride C-1* N-benzoyl-L-aspartic anhydride *Comparison example 1

[0107] As a comparison, a strong cation exchanger-membrane adsorber known in the prior art, Sartobind S (strong cation exchanger of cellulose hydrate with sulfonic acid ligands, Sartorius Stedim Biotech GmbH), was tested. The results are marked with Ref in Table 4 and FIGS. 1 through 3 below.

[0108] Moreover, a reaction of N-benzoyl-L-aspartic anhydride with the PAA-modified cellulose hydrate membrane (molar mass of PAA: 15,000 g/mol) was carried out as comparison example 1, thus obtaining a chromatography matrix with a 2-(benzoylamino)butanoic acid ligand in order to recreate the Capto MMC ligands known from prior art.

TABLE-US-00004 TABLE 1 Polyallylamine spacers (M = 15,000 g/mol) Ligand Ligand Capacity, Capacity, Capacity, density, density, globulin globulin globulin polyamine- anhydride- Capacity, 25 mM 150 mM 300 mM functionalized functionalized lysozyme NaCl NaCl NaCl membrane membrane Polyamine Structure [mg/mL] .sup.M3 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mol/mL] .sup.M1 [mol/mL] .sup.M2 1 PAA (15,000 g/mol) [00010] 137.1 60.0 20.0 0.3 494 431 2 PAA (15,000 g/mol) [00011] 191.7 97.2 30.6 0.8 494 438 3 PAA (15,000 g/mol) [00012] 87.5 31.3 21.9 9.4 494 290 4 PAA (15,000 g/mol) [00013] 94.3 22.9 31.4 14.3 550 574 5 PAA (15,000 g/mol) [00014] 123.5 23.5 42.4 23.5 494 441 6 PAA (15,000 g/mol) [00015] 123.7 15.0 15.7 22.0 524 525 7 PAA (15,000 g/mol) [00016] 81.2 29.7 37.1 19.4 494 425 8 PAA (15,000 g/mol) [00017] 61.1 16.0 24.5 25.3 494 760 9 PAA (15,000 g/mol) [00018] 83.3 16.7 30.6 22.2 494 833 10 PAA (15,000 g/mol) [00019] 76.8 13.0 13.3 18.3 546 640 11 PAA (15,000 g/mol) [00020] 111.4 57.1 14.3 16.9 546 437 12 PAA (15,000 g/mol) [00021] 83.4 39.1 40.7 5.4 546 534 C-1* PAA (15,000 g/mol) [00022] 55.3 15.3 17.1 24.0 494 473 *Comparison example 1

TABLE-US-00005 TABLE 2 Polyallylamine spacers (M = 100,000 g/mol and 150,000 g/mol) Ligand density, Ligand density, Capacity, Capacity, Capacity, polyamine- anhydride- Capacity, globulin globulin globulin functionalized functionalized lysozyme 25 mM NaCl 150 mM NaCl 300 mM NaCl membrane membrane Polyamine Structure [mg/mL] .sup.M3 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mol/mL] .sup.M1 [mol/mL] .sup.M2 13 PAA (150,000 g/mol) [00023] 150.0 26.7 56.7 33.3 843 506 14 PAA (100,000 g/mol) [00024] 120.0 31.4 54.3 25.7 667 345

TABLE-US-00006 TABLE 3 Polyethyleneamine spacers (M = 25,000 g/mol) Ligand density, Ligand density, Capacity, Capacity, Capacity, polyethyleneimin anhydride- Capacity, globulin globulin globulin e-functionalized functionalized lysozyme 25 mM NaCl 150 mM NaCl 300 mM NaCl membrane membrane Polyamine Structure [mg/mL] .sup.M3 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mol/mL] .sup.M1 [mol/mL] .sup.M2 15 PEI (25,000 g/mol) [00025] 90.7 6.7 3.3 1.1 650 373 16 PEI (25,000 g/mol) [00026] 61.9 35.5 9.7 2.1 650 548

TABLE-US-00007 TABLE 4 Membrane adsorber Sartobind S (strong cation exchanger of cellulose hydrate with sulfonic acid ligands, Sartorius Stedim Biotech GmbH) Capacity, Capacity, Capacity, Capacity, globulin globulin globulin Ligand lysozyme 25 mM NaCl 150 mM NaCl 300 mM NaCl density, Structure [mg/mL].sup.M3 [mg/mL].sup.M4 [mg/mL].sup.M4 [mg/mL].sup.M4 [mol/mL].sup.M2 Ref Sartobind S 46 37 14 2 96

Modification of Polyethylene Membranes

[0109] 1. Ligand Immobilization

[0110] The polyallylamine-functionalized polyethylene membrane Chromasorb (0.65 m pore size, EMD Millipore) was used as a starting material for ligand immobilization. In a typical reaction, 16 g of carboxylic anhydride was dissolved in 64 g of DMSO (20 wt %) and the solution was heated to 60 C. The polyallylamine-functionalized polyethylene membrane was placed in the reaction solution (0.5 g/cm.sup.2) and agitated at 60 C. for 1 h. The reaction solution was then filtered off, and the membrane was washed with ethanol (0.5 g/cm.sup.2) and a large excess of RO water.

[0111] The cation exchangers listed here were produced according to the above-described method, wherein the following carboxylic anhydrides were used. The results are shown in Table 5 below.

TABLE-US-00008 Example Anhydride 17 Succinic anhydride 18 Maleic anhydride 20

TABLE-US-00009 TABLE 5 Ligand density, Ligand density, Capacity, Capacity, Capacity, polyamine- anhydride- Capacity, globulin globulin globulin functionalized functionalized lysozyme 25 mM NaCl 150 mM NaCl 300 mM NaCl membrane membrane Structure [mg/mL] .sup.M3 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mg/mL] .sup.M4 [mol/mL] .sup.M1 [mol/mL] .sup.M2 17 [00027] 195.8 132.5 58.3 3.3 858 533 18 [00028] 141.7 47.5 71.7 33.3 858 500

Evaluation of Results

[0112] The results are summarized in FIGS. 1 through 3. As shown in FIG. 1, compared to the membrane obtained in comparison example 1, the membranes according to the invention with the multimodal ligands surprisingly show significantly higher binding capacity to small molecules, such as lysozyme, at comparable ligand density. The same applies for the strong cation exchanger-membrane adsorber Sartobind S known in the prior art.

[0113] In addition, all of the examples show favorable binding properties for larger molecules, such as globulin, over a wide salt range (25 mM to 300 mM NaCl). In order to better describe this binding capacity, a mean binding capacity is defined for globulin BC (globulin):

[00001]$\stackrel{\_}{\mathrm{BC}}\left(\mathrm{globulin}\right)=\frac{\begin{array}{c}\mathrm{BC}\left(\mathrm{globulin},25.Math..Math.\mathrm{mM}.Math..Math.\mathrm{NaCl}\right)+\\ \mathrm{BC}\left(\mathrm{globulin},150.Math..Math.\mathrm{mM}.Math..Math.\mathrm{NaCl}\right)+\\ \mathrm{BC}\left(\mathrm{globulin},300.Math..Math.\mathrm{mM}.Math..Math.\mathrm{NaCl}\right)\end{array}}{3}$

[0114] The results are summarized in FIG. 2.

[0115] In order to allow determination of a result with respect to the performance of the individual examples for numerous applications, the binding capacity for both small molecules (lysozyme) and large molecules (globulin) is taken into account below. For this purpose, a binding indicator BC (total) is defined:

[00002]$\stackrel{\_}{\mathrm{BC}}\left(\mathrm{total}\right)=\frac{\mathrm{BC}\left(\mathrm{lysozyme}\right)+\stackrel{\_}{\mathrm{BC}}\left(\mathrm{globulin}\right)}{2}$

[0116] The results are summarized in FIG. 3. Surprisingly, the membranes according to the invention show a significantly higher binding indicator BC (total) compared to the membrane obtained in comparison example 1. The same applies for the strong cation exchanger-membrane adsorber Sartobind S known in the prior art.