초록 ▼

The invention provides novel ion-exchange media and related methods for their preparation and use. Ion-exchange stationary phases according to the invention are suitable for chromatographic separation of a variety of biomolecules. Distinguishing characteristics of ion-exchange media according to thi

The invention provides novel ion-exchange media and related methods for their preparation and use. Ion-exchange stationary phases according to the invention are suitable for chromatographic separation of a variety of biomolecules. Distinguishing characteristics of ion-exchange media according to this invention includes, for example, their ability to separate variants of monoclonal antibodies via cation-exchange liquid chromatography using porous substrates with particle sizes 5 μm. The ion-exchange stationary media include a hydrolytically stable layer, which inhibits surface degradation of the particles in 100% aqueous media. Another unique feature is low molecular weight building blocks used to functionalize the particles with ion-exchange groups.

대표청구항 ▼

1. An ion-exchange medium, comprising: a particulate substrate having an average particle size of about 0.5 μm to about 100 μm in diameter;a hydrolytically stable monolayer of a bi- or poly-functional organosilane covalently attached to a surface of the particulate substrate and comprising a plurali

1. An ion-exchange medium, comprising: a particulate substrate having an average particle size of about 0.5 μm to about 100 μm in diameter;a hydrolytically stable monolayer of a bi- or poly-functional organosilane covalently attached to a surface of the particulate substrate and comprising a plurality of cation-exchange sites, the bi- or poly-functional organosilane having the structural formula of (I) and having a molecular weight less than about 20,000, wherein: each of R1, R2, R3, R4, R5 and R6 are independently H, a C1-C6 alkyl group, a Si atom of the surface of the particulate substrate, or a Si atom of another bi- or poly-functional organosilane,each of Q1 and Q2 is independently selected from a linear or branched, acyclic or cyclic, saturated aliphatic moiety, optionally with one or more carbon atoms substituted by oxygen, nitrogen or a sulfur heteroatom,L is an aliphatic spacer group, optionally with one or more carbon atoms substituted by oxygen, nitrogen or sulfur heteroatoms,R7 is H or a C1-C6 alkyl group,R9 is H or an alkyl group,l is 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12, andY is —(C═O)—(CH2)j—(C═O)—OR10 or —(C═O)—(CH2)j—U—(CH2)j—(CH═O)—OR10, wherein R10 is H, U is —NH—, oxygen or a sulfur heteroatom and j is an integer from about 1 to about 6. 2. The ion-exchange medium of claim 1, wherein the particulate substrate is porous. 3. The ion-exchange medium of claim 1, wherein the particulate substrate comprises a non-porous core and a porous shell. 4. The ion-exchange medium of claim 1, wherein the particulate substrate is non-porous. 5. The ion-exchange medium of claim 2, wherein the particulate substrate has an average pore size of about 60 Å to about 2,000 Å. 6. The ion-exchange medium of claim 1, wherein Q1 is —(CH2)m—CH2— or —(CH2)m-G-(CH2)n—, and Q2 is —(CH2)m—CH2— or —(CH2)m-G′-(CH2)n— each of m and n is an integer from about 1 to about 6, G and G′ are independently —NH—, oxygen, or a sulfur heteroatom. 7. The ion-exchange medium of claim 6, wherein L is —(CH2)k—, k is an integer from about 1 to about 6, and R7 is H. 8. The ion-exchange medium of claim 1, wherein the bi- or poly-functional organosilane has the structural formula of (II): wherein each of R1, R2, R3, R4, R5 and R6 is independently H, a C1-C3 alkyl group, a Si atom of the surface of the particulate substrate, or a Si atom of another bi- or poly-functional organosilane,R7 is H or C1-C3 alkyl group,R9 is a H or an alkyl group,each of m, n and k is an integer from about 1 to about 3,l is 3, 4, 5, 6, 7, 8, 9, 10, 11, andY is —(C═O)—(CH2)j—(C═O)—OR10 or —(C═O)—(CH2)j—U—(CH2)j—(C═O)—OR10, wherein R10 is H, U is —NH—, oxygen, or a sulfur heteroatom and j is an integer from about 1 to about 6. 9. The ion-exchange medium of claim 1, wherein l is 3, 4, 5, or 6. 10. A method for purifying a biological analyte by ion-exchange chromatography, comprising: providing an ion-exchange chromatography column packed with an ion-exchange chromatography medium comprising: a particulate substrate having an average particle size of about 0.5 μm to about 100 μm in diameter,a hydrolytically stable monolayer of a bi- or poly-functional organosilane covalently attached to a surface of the particulate substrate and comprising a plurality of cation-exchange sites, the bi- or poly-functional organosilane having the structural formula of (I) and having a molecular weight less than about 20,000, wherein each of R1, R2, R3, R4, R5, and R6 are independently H, a C1-C6 alkyl group, a Si atom of the surface of the particulate substrate, or a Si atom of another bi- or poly- functional organosilane,each of Q1 and Q2 is independently selected from a linear or branched, acyclic or cyclic, saturated aliphatic moiety, optionally with one or more carbon atoms substituted by oxygen, nitrogen or sulfur heteroatoms,L is an aliphatic spacer group, optionally with one or more carbon atoms substituted by oxygen, nitrogen or sulfur heteroatoms,R7 is H or C1-C6 alkyl group,R9 is H or an alkyl group,l is 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12, andY is —(C═O)—(CH2)j—(C═O)—OR10 or —(C═O)—(CH2)j—U—(CH2)j—(C═O)—OR10, wherein R10 is H, U is —NH—, oxygen, or a sulfur heteroatom and j is an integer from about 1 to about 6.contacting a sample solution from which the biological analyte is to be purified with the ion-exchange chromatography medium allowing ion exchange between the sample solution and the ion-exchange chromatography medium; andeluting the biological analyte from the ion-exchange chromatography medium to purify the biological molecule. 11. The method of claim 10, wherein the particulate substrate is porous, comprises a non-porous core and a porous shell, or is non-porous. 12. The method of claim 10, wherein the biological analyte is selected from proteins, oligonucleotides, polynucleotides, and polysaccharides. 13. The method of claim 10, wherein Q1 is —(CH2)m—CH2— or —(CH2)m-G-(CH2)n—, and Q2 is —(CH2)m—CH2— or —(CH2)m-G′-(CH2)n— each of m and n is an integer from about 1 to about 6, G and G′ are independently —NH—, oxygen, or a sulfur heteroatom. 14. The method of claim 13, wherein L is —(CH2)k—, k is an integer from about 1 to about 6, and R7 is H. 15. The method of claim 10, wherein the bi- or poly-functional organosilane has the structural formula of (II): wherein each of R1, R2, R3, R4, R5 and R6 is independently H, a C1-C3 alkyl group, a Si atom of the surface of the particulate substrate, or a Si atom of another bi- or poly-functional organosilane;R7 is H or C1-C3 alkyl group;each of m, n and k is an integer from 1 to about 3;l is an integer from 3 to 6; andY is —(C═O)—(CH2)j—(C═O)—OR10 or —(C═O)—(CH2)j—U—(CH2)j—(C═O)—OR10, wherein R10 is H and U is —NH—, oxygen or a sulfur heteroatom and j is an integer from about 1 to about 6. 16. A cation-exchange chromatography stationary phase, comprising: a particulate substrate having an average particle size of about 0.5 μm to about 100 μm in diameter;a hydrolytically stable monolayer of a bi- or poly-functional organosilane covalently attached to a surface of the porous particulate substrate and comprising a plurality of cation-exchange sites, the bi- or poly-functional organosilane having the structural formula of (I) and having a molecular weight less than about 20,000, wherein each of R1, R2, R3, R4, R5 and R6 is independently H, a C1-C3 alkyl group, a Si atom of the surface of the particulate substrate, or a Si atom of another bi- or poly-functional organosilane,R7 is H or C1-C3 alkyl group,R9 is H or an alkyl group,each of m, n and k is an integer from 1 to about 3;1 is an integer from 3 to 6; andY is —(C═O)—(CH2)j—(C═O)—OR10 or —(C═O)—(CH2)j—U—(CH2)j—(C═O)—OR10, wherein R10 is H, U is —NH—, oxygen, or a sulfur heteroatom and j is an integer from about 1 to about 6. 17. The cation-exchange chromatography stationary phase of claim 16, wherein the particulate substrate is porous, comprises a non-porous core and a porous shell, or is non-porous.