阅读说明：本技术 基于聚烷基多胺聚合物层的阴离子交换固定相 (Anion exchange stationary phase based on polyalkyl polyamine polymer layer ) 是由 C·A·波尔 陈锦华 于 2019-07-30 设计创作，主要内容包括：用于分离多种碳水化合物的阴离子交换包括带负电的基材颗粒。基础聚合物层包括第一多个季胺。聚烷基多胺聚合物层共价连接到基础缩聚物层上。聚烷基多胺聚合物层包括聚合物分支结构,其包括第二多个季胺。第二多个季胺的密度在远离基础缩聚物层的方向上增加。阴离子交换固定相不具有通过乙基与第一或第二多个季胺中的任何一个间隔开的羟基。(Anion exchange for the separation of various carbohydrates includes negatively charged substrate particles. The base polymer layer includes a first plurality of quaternary amines. The layer of polyalkylpolyamine polymer is covalently linked to the base layer of polycondensation. The polyalkylpolyamine polymer layer includes a polymeric branch structure including a second plurality of quaternary amines. The density of the second plurality of quaternary amines increases in a direction away from the base polycondensation layer. The anion exchange stationary phase does not have hydroxyl groups spaced apart from any of the first or second plurality of quaternary amines by ethyl groups.)

1. An anion exchange stationary phase for separating a plurality of carbohydrates comprising:

a) negatively charged substrate particles;

b) a base polycondensation layer attached to the negatively charged substrate particles, the base polycondensation layer comprising a first plurality of quaternary amines, wherein the first plurality of quaternary amines are separated by a first spacer or a second spacer, wherein the base polycondensation layer does not have hydroxyl groups separated from one of the first plurality of quaternary amines by ethyl groups;

c) a polyalkyl polyamine condensate layer covalently attached to the base condensate layer, the polyalkyl polyamine condensate layer comprising a polymeric branching structure comprising a second plurality of quaternary amines, wherein the second plurality of quaternary amines are spaced apart by the first spacer or the second spacer, wherein the density of the second plurality of quaternary amines increases in a direction away from the base condensate layer, wherein the polyalkyl polyamine condensate layer does not have hydroxyl groups spaced apart from the second plurality of quaternary amines by ethyl groups.

2. The anion exchange stationary phase of claim 1, wherein the negatively charged substrate particles comprise a crosslinked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises sulfonate groups.

3. The anion exchange stationary phase of claim 1, wherein the negatively charged substrate particles comprise a crosslinked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises carboxylate groups.

4. The anion exchange stationary phase of claim 1, wherein the base polycondensate layer is positively charged and ionically attached to the negatively charged substrate particles.

5. The anion exchange stationary phase of claim 1, wherein the first spacer comprises a formula (-CH) ₂-) _xAnd the second gapThe spacer group comprises the formula (-CH) ₂-) _yWherein x and y are each independently 3 to 6.

6. The anion exchange stationary phase of claim 1, wherein the first spacer comprises a first alkyl group and the second spacer comprises a second alkyl group, wherein the first alkyl group and the second alkyl group are both linear and unsubstituted alkyl groups.

7. The anion exchange stationary phase of claim 1, wherein the first spacer comprises a linear and unsubstituted alkyl group and the second spacer comprises an aralkyl group.

8. The anion exchange stationary phase of claim 1, wherein the first spacer is selected from the group consisting of alkyl, dialkyl ether, cycloalkyl, aralkyl, and combinations thereof.

9. The anion exchange stationary phase of claim 1, wherein the first spacer comprises an alkyl group and the second spacer is selected from the group consisting of an alkyl group, an aralkyl group, and combinations thereof.

10. An anion exchange stationary phase for separating a plurality of carbohydrates, the anion exchange stationary phase formed by a process comprising:

reacting a polyhalogenated hydrocarbon with a polyalkyl polyamine to form a base polycondensation layer on negatively charged substrate particles; and

reacting the base polycondensation layer for a number of reaction cycles to form a polyalkyl polyamine polycondensation layer, wherein the number of reaction cycles is in the range of about three to about ten, and each reaction cycle comprises a polyhalogenated hydrocarbon treatment and a polyalkyl polyamine treatment.

11. The anion exchange stationary phase of claim 10, further comprising:

reacting the polyalkyl polyamine condensate layer with a monohaloalkane treatment.

12. The anion exchange stationary phase of claim 10, wherein the negatively charged substrate particles are contained as a packed bed in a column, wherein the reaction of the polyhalogenated hydrocarbon with the polyalkylpolyamine comprises: flowing a solution of the polyhalogenated hydrocarbon and the polyalkylpolyamine through the column to form the base polycondensation layer on the negatively charged substrate particles.

13. The anion exchange stationary phase of claim 12, wherein the polyhalogenated hydrocarbon treatment comprises: flowing a solution of the polyhalogenated hydrocarbon through the column; the polyalkylpolyamine treatment comprises: flowing the solution of polyalkylpolyamine through the column; the monohaloalkane treatment comprises: flowing a solution of the monohaloalkane through the column.

14. The anion exchange stationary phase of claim 10, wherein the number of reaction cycles ranges from about 3 to about 4.

15. The anion exchange stationary phase of claim 10, wherein the negatively charged substrate particles comprise a cross-linked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises sulfonate groups.

16. The anion exchange stationary phase of claim 10, wherein the negatively charged substrate particles comprise a crosslinked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises carboxylate groups.

17. The anion exchange stationary phase of claim 10, wherein the polyhalogenated hydrocarbon comprises a material selected from the group consisting of dihaloalkanes, dihalodialkylethers, dihalocycloalkanes, trihaloaralkanes, and combinations thereof.

18. The anion exchange stationary phase of claim 10, wherein the polyalkyl polyamine comprises a material selected from the group consisting of polyalkyltriamines, polyalkyldiamines, and combinations thereof.

19. The anion exchange stationary phase of claim 10, wherein all amines of the polyalkylpolyamine are tertiary amines.

20. The anion exchange stationary phase of claim 10, wherein the polyhalogenated hydrocarbon is selected from dibromobutane, dibromopentane, dibromohexane, tribromomethylbenzene, and combinations thereof.

21. The anion exchange stationary phase of claim 10, wherein the polyalkyl polyamine is selected from pentamethyl dipropyl triamine, pentamethyl dihexyl triamine, permethylated spermine, permethylated spermidine, and combinations thereof.

22. The anion exchange stationary phase of claim 10, wherein the polyhalogenated hydrocarbon is dibromobutane and the polyalkylpolyamine is pentamethyldihexyltriamine.

23. The anion exchange stationary phase of claim 10, wherein the polyhalogenated hydrocarbon is a trihaloalkylaryl group and the polyalkylpolyamine is an alkyldiamine.

24. The anion exchange stationary phase of claim 10, wherein the polyhalogenated hydrocarbon is tribromomethylbenzene and the polyalkylpolyamine is tetramethylhexamethylenediamine.

25. An anion exchange stationary phase for separating a plurality of carbohydrates, the anion exchange stationary phase comprising:

A) negatively charged substrate particles;

B) a base polycondensation layer attached to the negatively charged substrate particles, the base polycondensation layer comprising the reaction product of

i) A first polyhalogenated hydrocarbon, and

ii) a first polyalkylpolyamine;

C) a first alkyl condensation reaction product covalently attached to the base polycondensation layer, the first alkyl condensation reaction product comprising the reaction product of

i) Amine groups of the base polycondensation product layer, and

ii) a second polyhalogenated hydrocarbon, wherein said amine groups of said base polycondensation layer comprise a positive charge, such that said base polycondensation layer is ionically coupled to said negatively charged substrate particles;

D) a first polyalkyl polyamine condensation reaction product covalently attached to said first alkyl condensation reaction product, said first polyalkyl polyamine condensation reaction product comprising the reaction product of

i) A halo group of said second polyhalogenated hydrocarbon, and

ii) a second polyalkylenepolyamine; and

E) a second alkyl condensation reaction product covalently attached to the first polyalkyl polyamine condensation reaction product, the second alkyl condensation reaction product comprising the reaction product of

i) The amine groups of said first polyalkylpolyamine condensation reaction product, and

ii) a third polyhalogenated hydrocarbon;

F) a second polyalkylenepolyamine CRP covalently linked to the second alkylpolyamine CRP, the second polyalkylenepolyamine CRP comprising a reaction product of

i) The halogen group of the third polyhalogenated hydrocarbon and

ii) a third polyalkylenepolyamine;

G) a third alkyl CRP covalently linked to the second polyalkylenepolyamine CRP, the third alkyl CRP comprising a reaction product of

i) The amine group of the second polyalkylenepolyamine CRP, and

ii) a fourth polyhalogenated hydrocarbon; and

H) a third polyalkylenepolyamine CRP covalently linked to a third polyalkylenepolyamine CRP comprising a reaction product of

i) The halo group of said fourth polyhalogenated hydrocarbon, and

ii) a fourth polyalkylpolyamine.

26. The anion exchange stationary phase of claim 25, wherein the first polyhalogenated hydrocarbon, the second polyhalogenated hydrocarbon, the third polyhalogenated hydrocarbon and the fourth polyhalogenated hydrocarbon comprise dihaloalkanes; the first, second, third, and fourth polyalkylpolyamines comprise polyalkyltriamines.

27. A method of separating multiple carbohydrates in a sample using an anion exchange stationary phase comprising:

a) negatively charged substrate particles;

b) a base polycondensation layer attached to the negatively charged substrate particles, the base polycondensation layer comprising a first plurality of quaternary amines, wherein the first plurality of quaternary amines are separated by a first spacer or a second spacer, wherein the base polycondensation layer does not have hydroxyl groups separated from one of the first plurality of quaternary amines by ethyl groups;

c) a polyalkyl polyamine condensate layer covalently attached to the base condensate layer, the polyalkyl polyamine condensate layer comprising a polymeric branching structure comprising a second plurality of quaternary amines, wherein the second plurality of quaternary amines are spaced apart by the first spacer or the second spacer, wherein the density of the second plurality of quaternary amines increases in a direction away from the base condensate layer, wherein the polyalkyl polyamine condensate layer does not have hydroxyl groups spaced apart from the second plurality of quaternary amines by ethyl groups, the method comprising:

flowing an eluent through a chromatography column containing the anion exchange stationary phase, wherein the eluent comprises a hydroxide;

injecting a sample comprising a plurality of carbohydrates into a chromatography column;

separating at least one carbohydrate from a sample injected into a chromatography column; and

detecting the at least one carbohydrate at a detector.

28. The method of claim 27, wherein the at least one carbohydrate is a branched glycan.

Technical Field

The present invention relates generally to anion exchange stationary phases based on a polyalkyl polyamine layer for separating samples comprising anions, in particular combinations of carbohydrates, more particularly combinations of branched glycans, using for example chromatography.

Background

Chromatography is a widely used analytical technique for chemical analysis and separation of molecules. Chromatography involves the separation of one or more analyte species from other matrix components present in a sample. The stationary phase of the chromatography column is typically selected so that there is an interaction with the analyte. Such interactions may be ionic, hydrophilic, hydrophobic, or a combination thereof. For example, the stationary phase can be derivatized with an ionic moiety that desirably binds to the ionic analyte and a matrix component having different levels of affinity. The mobile phase permeates through the stationary phase and competes with the analyte and matrix components for binding to the ionic portion. Mobile phase or eluent is a term used to describe a liquid solvent or buffer solution that is pumped through a chromatography column. During this competition, the analyte and matrix components will elute from the stationary phase over time and then subsequently be detected at the detector. Examples of some typical detectors are conductivity detectors, UV-VIS spectrophotometers and mass spectrometers. Chromatography has developed over the years into an effective analytical tool suitable for creating healthier, cleaner and safer environments where complex sample mixtures can be isolated and analyzed for various industries such as water quality, environmental monitoring, food analysis, medicine and biotechnology.

U.S. Pat. No. 7,291,395, "coated ion exchange substrate and method of formation," describes hyperbranched anion exchange materials based on a crosslinked layer comprising quaternary amine groups and hydroxyl groups in one embodiment, anion exchange materials are formed using a reaction between a diepoxide reagent and an amine group a ring opening reaction produces hydroxyl groups spaced from quaternary amine groups by two carbon spacers the hydroxyl groups produced in this type of reaction can be referred to as β hydroxyl groups over the season, which makes the hydroxyl groups more acidic and adversely affects the anion binding properties of the anion exchange resin during anion exchange chromatography separation, hydroxide eluents are typically used β hydroxyl groups can be deprotonated when the pH of the hydroxide eluent is sufficiently high, resulting in the formation of zwitterionic pairs with the quaternary amines, which reduces the binding of anions to the quaternary amine ion exchange sites, the zwitterionic pairs have positively charged quaternary amines that are stabilized by the proximity of negatively charged and deprotonated β hydroxyl groups, the proximity of the positively charged quaternary amines and the deprotonated hydroxyl groups forms relatively stable zwitterionic pairs, which reduces the binding strength of the anionic exchange resins, thus making the anion exchange resins available for a variety of anion exchange tests based on the pH of eluents.

However, in some cases, applicants have found that carbohydrates retain little to no hyperbranched anion exchange material containing β hydroxyl groups and are therefore not useful for this application.

Disclosure of Invention

A first embodiment of an anion exchange stationary phase for separating a plurality of carbohydrates comprises a) negatively charged substrate particles, b) a base polycondensation product layer, and c) a polyalkyl polyamine polycondensation product layer. The base polycondensation layer is attached to negatively charged substrate particles. The base polycondensation layer includes a first plurality of quaternary amines, where the first plurality of quaternary amines are spaced apart by a first spacer or a second spacer. The base polycondensation layer has no hydroxyl groups spaced apart from one of the first plurality of quaternary amines by ethyl groups. The layer of polyalkylpolyamine condensate is covalently linked to the base condensate layer. The polyalkylpolyamine polycondensation layer includes a polymer branched structure comprising a second plurality of quaternary amines, wherein the second plurality of quaternary amines are separated by a first spacer or a second spacer. The density of the second plurality of quaternary amines increases in a direction away from the base polycondensation layer. The polyalkylpolyamine condensate layer has no hydroxyl groups spaced apart from one of the second plurality of quaternary amines by ethyl groups.

With respect to any of the first embodiments, the first spacer can be an alkyl, dialkyl ether, cycloalkyl, arylalkyl, and combinations thereof.

With respect to any of the first embodiments, the second spacer can be an alkyl, dialkyl ether, cycloalkyl, arylalkyl, and combinations thereof.

With respect to the first embodiment wherein the first spacer comprises a first alkyl group and the second spacer comprises a second alkyl group, wherein both the first alkyl group and the second alkyl group are straight chain and unsubstituted alkyl groups.

With respect to the first embodiment, the first spacer comprises a linear and unsubstituted alkyl group and the second spacer comprises an aralkyl group.

With respect to the first embodiment, the first spacer comprises the formula (-CH) ₂-) _xThe second spacer comprises a formula (-CH) ₂-) _yWherein x and y are each independently 3 to 10, preferably 3 to 6.

A second embodiment of an anion exchange stationary phase for separating a plurality of carbohydrates is formed by a process comprising reacting a polyhalogenated hydrocarbon with a polyalkyl polyamine to form a base polycondensation layer on negatively charged substrate particles. The base polycondensation layer is reacted with a plurality of reaction cycles to form a layer of a polyalkylamine polycondensation product. The number of reaction cycles ranges from about 3 to about 10, each reaction cycle including polyhalogenated hydrocarbon treatment and polyalkylpolyamine treatment. After the reaction cycle, the layer of polyalkylpolyamine condensate is reacted with the monohaloalkane treatment. An example of a monohaloalkane is methyl iodide.

With respect to the second embodiment, the negatively charged substrate particles are contained in the chromatography column as a packed bed. The reaction of the polyhalogenated hydrocarbon with the polyalkyl polyamine comprises flowing a solution of the polyhalogenated hydrocarbon and the polyalkyl polyamine through the column to form a base polycondensation layer on the negatively charged substrate particles.

With respect to the second embodiment, polyhalogenated hydrocarbon treatment comprises flowing a solution of polyhalogenated hydrocarbon through the column. The polyalkylpolyamine treatment comprises flowing a solution of the polyalkylpolyamine through a column. The monohaloalkane treatment comprises flowing a solution of monohaloalkane through the column.

With respect to any of the second embodiments, the number of reaction cycles ranges from about 3 to about 4.

With respect to any of the second embodiments, the polyhalogenated hydrocarbon may be a dihaloalkane, a dihalodialkyl ether, a dihalocycloalkane, a trihaloalkane, and combinations thereof.

With respect to any of the second embodiments, the polyalkylpolyamine can include ether groups, cycloalkane groups, arylalkanes, and combinations thereof.

With respect to any of the second embodiments, the polyalkyl polyamine can be a polyalkyltriamine, a polyalkyldiamine, and combinations thereof.

With respect to the second embodiment, all of the amines of the polyalkylpolyamines may be tertiary amines (e.g., polyalkyl poly-tertiary amines).

With respect to the second embodiment, the polyhalogenated hydrocarbon may be dibromobutane, dibromopentane, dibromohexane, tribromomethylbenzene, and combinations thereof.

With respect to the second embodiment, the polyalkyl polyamine may be pentamethyl dipropyl triamine, pentamethyl dihexyl triamine, permethylated spermine, permethylated spermidine, and combinations thereof.

With respect to the second embodiment, the polyhalogenated hydrocarbon is dibromobutane and the polyalkyl polyamine is pentamethyldihexyltriamine.

With respect to the second embodiment, the polyhalogenated hydrocarbon is a trihaloalkylaryl and the polyalkylpolyamine is an alkyldiamine.

With respect to the second embodiment, the polyhalogenated hydrocarbon is tribromomethylbenzene and the polyalkyl polyamine is tetramethylhexamethylenediamine.

A third embodiment of an anion exchange stationary phase for separating a plurality of carbohydrates comprises a) negatively charged substrate particles, B) a base polycondensation layer 300, C) a first alkyl Condensation Reaction Product (CRP)400, D) a first polyalkyl polyamine CRP500, E) a second alkyl CRP 600, F) a second alkyl CRP 700, G) a third alkyl CRP, and H) a third alkyl CRP. The base polycondensation layer 300 can be attached to negatively charged substrate particles. The base polycondensation layer includes the reaction product of i) a first polyhalogenated hydrocarbon and ii) a first polyalkylpolyamine. First alkyl CRP 400 is covalently attached to base polycondensation layer 300. First alkyl CRP 400 comprises the reaction product of i) the amine groups of base polycondensation layer 300 and ii) a second polyhalogenated hydrocarbon. The amine groups of the base polycondensation layer 300 include a positive charge such that the base polycondensation layer is ionically coupled to the negatively charged substrate particles. The first polyalkylpolyamine CRP500 is covalently linked to the first alkyl CRP 400. The first polyalkylpolyamine CRP500 comprises the reaction product of i) a halogen group of a second polyhalogenated hydrocarbon and ii) a second polyalkylpolyamine. The second alkyl CRP 600 is covalently linked to the first polyalkylpolyamine CRP 500. The second alkyl CRP 600 comprises the reaction product of i) the amine groups of the first polyalkylpolyamine CRP500 and ii) a third polyhalogenated hydrocarbon. The second polyalkylenepolyamine CRP 700 is covalently linked to a second alkyl CRP 600. The second polyalkylenepolyamine CRP 700 comprises the reaction product of i) a halogen group of a third polyhalogenated hydrocarbon and ii) a third polyalkylenepolyamine. The third alkyl CRP is covalently attached to the second polyalkylpolyamine CRP 700. The third alkyl CRP comprises the reaction product of i) the amine group of the second polyalkylpolyamine CRP and ii) a fourth polyhalogenated hydrocarbon. The third polyalkylenepolyamine CRP is covalently linked to the third alkyl CRP. The third polyalkylenepolyamine CRP comprises the reaction product of i) a halogen group of a fourth polyhalogenated hydrocarbon and ii) a fourth polyalkylenepolyamine.

With respect to the third embodiment, the first polyhalogenated hydrocarbon, the second polyhalogenated hydrocarbon, the third polyhalogenated hydrocarbon and the fourth polyhalogenated hydrocarbon may comprise dihaloalkanes. The first, second, third, and fourth polyalkylpolyamines may include polyalkyltriamines.

A fourth embodiment of an anion exchange stationary phase for separating a plurality of carbohydrates comprises a) negatively charged substrate particles, B) a base polycondensation layer 1500, C) a first polyalkyl polyamine 1600, D) a first polyalkylaryl CRP1700, E) a second polyalkyl polyamine CRP1800, F) a second polyalkylaryl CRP1900, G) a third polyalkyl polyamine CRP, and H) a third polyalkylaryl CRP. The base polycondensation layer 1500 is attached to the negatively charged substrate particles. The base polycondensation layer includes the reaction product of i) a first halogenated hydrocarbon and ii) a first polyalkylpolyamine. The amine groups of the base polycondensation layer 1500 include a positive charge such that the base polycondensation layer is ionically coupled to the negatively charged substrate particles. The first polyalkyl polyamine condensation reaction product 1600 is covalently attached to the base polycondensation layer 1500. The first polyalkyl polyamine condensation reaction product 1600 includes the reaction product of i) the halide groups of the base polycondensation layer 1500 and ii) the second polyalkyl polyamine. First polyalkylaryl condensation reaction product 1700 is covalently linked to first polyalkylpolyamine condensation reaction product 1600. First polyalkylaryl condensation reaction product 1700 comprises the reaction product of i) the amine groups of first polyalkylpolyamine condensation reaction product 1600 and ii) a second polyhalohydrocarbon. Second polyalkylpolyamine condensation reaction product 1800 is covalently linked to first polyalkylaryl condensation reaction product 1700. The second polyalkylpolyamine condensation reaction product 1800 comprises the reaction product of i) the halogen groups of the first polyalkylaryl condensation reaction product 1700 and ii) a third polyalkylpolyamine. The second polyalkylaryl CRP1900 is covalently linked to the second polyalkylpolyamine CRP 1800. The second polyalkylaryl CRP1900 includes the reaction product of i) the amine groups of the second polyalkylpolyamine CRP1800 and ii) a third polyhalogenated hydrocarbon. The third polyalkylpolyamine CRP is covalently linked to the second polyalkylaryl CRP 1900. The third polyalkylpolyamine CRP comprises the reaction product of i) a halogen group of a second polyalkylaryl CRP1900 and ii) a fourth polyalkylpolyamine. The third polyalkylaryl CRP is covalently linked to the third polyalkylpolyamine CRP. The third polyalkylaryl CRP includes the reaction product of i) the amine group of the third polyalkylpolyamine and ii) a fourth polyhalogenated hydrocarbon.

With respect to the fourth embodiment, it may also include the reaction product of i) the halo group of the third polyalkylarylCRP and ii) the tertiary amine.

With respect to the fourth embodiment, the first, second, third, and fourth polyhalogenated hydrocarbons may include trihaloalkylaryl groups, and the first, second, third, and fourth polyalkylpolyamines may include polyalkyldiamines.

A fifth embodiment is a method of separating a plurality of carbohydrates in a sample from an anion exchange stationary phase of any of the first, second, and third embodiments using the anion exchange stationary phase. The method comprises passing the eluent through a chromatographic column containing an anion exchange stationary phase. The eluent comprises a hydroxide. A sample comprising a plurality of carbohydrates is injected into a chromatography column. Separating at least one carbohydrate from the sample injected into the chromatography column. At least one carbohydrate is detected at a detector.

With respect to the fifth embodiment, the at least one carbohydrate is a branched glycan.

With respect to the above embodiments, the negatively charged substrate particles can comprise a crosslinked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises sulfonate groups.

With respect to the above embodiments, the negatively charged substrate particles comprise a crosslinked divinylbenzene and ethylvinylbenzene copolymer, wherein at least one surface of the negatively charged substrate particles comprises carboxylate groups.

With respect to the above embodiment, wherein the base polycondensate layer is positively charged and ionically attached to the negatively charged substrate particles.

Drawings

The accompanying drawings, which are incorporated herein and constitute part of this specification, illustrate presently preferred embodiments of the invention and, together with the general description given above and the detailed description given below, serve to explain the features of the invention (with like numerals designating like elements).

FIG. 1 shows various chemical structures of a polyalkyl polyamine reagent that can be used to form the polymers and reaction products of an anion exchange resin.

Figure 2 illustrates various chemical structures of polyhalogenated hydrocarbon reagents that can be used to form polymers and reaction products of anion exchange resins.

FIG. 3 illustrates a schematic representation of a base polymer layer formed from a first polyalkyltriamine and a first dihalide, wherein the base polymer layer is attached to a negatively charged substrate particle.

FIG. 4 shows a schematic representation of a first alkyl reaction product having pendant halogen groups and covalently attached to a base polymer. The first alkyl reaction product is the reaction product of an amine group of a base polymer and a second dihaloalkane.

FIG. 5 shows a schematic of a first alkylpolyamine reaction product covalently attached to a first alkyl reaction product. The first polyalkylpolyamine reaction product is the reaction product of a pendant halide group of a second dihaloalkane and an amine group of a second polyalkyltriamine.

FIG. 6 shows a schematic representation of a second alkyl reaction product having a pendant halogen group and covalently linked to a first polyalkyl polyamine reaction product. The second alkyl reaction product is the reaction product of the amine group of the first polyalkylpolyamine reaction product and a third dihaloalkane.

Fig. 7 shows a schematic of a second polyalkylenepolyamine covalently attached to a second alkyl reaction product. The second polyalkylenepolyamine reaction product is the reaction product of a pendant halogen group of a third dihaloalkane and an amine group of a third polyalkylenepolyamine.

Figure 8 shows six chromatograms, each using a different NaOH eluent concentration and standard solution containing five different carbohydrates. Chromatograms were performed using an anion exchange resin containing pentamethyldihexyltriamine PMDHTA and 1-4-dibromobutane DBB.

Figure 9 shows two chromatograms using an anion exchange resin comprising PMDHTA and DBB. The standard solution was injected into a column containing five carbohydrates (upper chromatogram) or two carbohydrates (lower chromatogram).

Figure 10 shows three chromatograms, each using a different NaOH eluent concentration and standard solution containing five different carbohydrates. Chromatography was performed using an anion exchange resin containing N, N' -tetramethyl-1, 6-hexanediamine, TMHDA, and 1,3, 5-tris (bromomethyl) benzene, TBMB.

Figure 11 shows two chromatograms using anion exchange resins including TMHDA and TBMB. The standard solution was injected into a column containing five carbohydrates (upper chromatogram) or two carbohydrates (lower chromatogram).

Figure 12 shows a chromatogram using an anion exchange resin comprising PMDHTA and DBB, showing 38 peaks. The sample solution is injected into a chromatographic column containing fetuin aldol.

Fig. 13 shows a chromatogram using an anion exchange resin comprising TMHDA and TBMB, showing 28 peaks. The sample solution is injected into a chromatographic column containing fetuin aldol.

Figure 14 shows an ion chromatography system suitable for analyzing samples using an ion exchange chromatography column and an electrochemical detector using a quadruple voltage waveform.

Fig. 15 shows a schematic representation of the formation of a base polymer layer from N, N' -tetramethyl-1, 6-hexanediamine TMHDA and 1,3, 5-tris (bromomethyl) benzene TBMB, wherein the base polymer layer is attached to negatively charged substrate particles.

FIG. 16 shows a schematic of a first polyalkyl polyamine reaction product having pendant tertiary amine groups and covalently attached to a base polymer. The first polyalkyl polyamine reaction product is the first polyalkyl polyamine reaction product of the halide groups of the base polymer and the second TMHDA.

FIG. 17 shows a schematic representation of a first polyalkylaryl reaction product covalently attached to a first polyalkylpolyamine reaction product. The first polyalkylaryl reaction product is the reaction product of a pendant tertiary amine of a second TMHDA and a halide group of a second TBMB.

FIG. 18 shows a schematic representation of a second polyalkylpolyamine reaction product having a pendant tertiary amine group and covalently attached to a first polyalkylaryl reaction product. The second polyalkylpolyamine reaction product is the reaction product of the halide group of the first polyalkylaryl reaction product and the third TMHDA.

FIG. 19 shows a schematic of the covalent attachment of a second polyalkylaryl group to a second polyalkylpolyamine reaction product. The second polyalkylaryl reaction product is the reaction product of a pendant tertiary amine group of a third TMHDA and a halogen group of a third TBMB.

Detailed Description

The following detailed description should be read with reference to the drawings, in which like elements in different drawings are numbered the same. The drawings, which are not necessarily to scale, depict selected embodiments and are not intended to limit the scope of the invention. The detailed description illustrates by way of example, and not by way of limitation, the principles of the invention. This description will clearly enable one skilled in the art to make and use the invention, and describes several embodiments, adaptations, variations, alternatives and uses of the invention, including what is presently believed to be the best mode of carrying out the invention. As used herein, the terms "about" or "approximately" for any numerical value or range indicate a suitable dimensional tolerance (dimensional tolerance) that allows a portion or collection of components to function for their intended purpose, as described herein.

Unless otherwise specified, the term "alkyl" alone or as part of another substituent means herein a straight or branched chain or cyclic hydrocarbon group or combination thereof, which may be fully saturated, mono or polyunsaturated and may include a number of the specified carbon atomsZi (i.e., C) ₁-C ₁₀Meaning one to ten carbons) of a divalent or polyvalent group. Examples of saturated hydrocarbon groups include (but are not limited to) the following: such as methyl, ethyl, n-propyl (e.g., -CH) ₂-CH ₂-CH ₃、-CH ₂-CH ₂-CH ₂-), isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, cyclohexyl, (cyclohexyl) methyl, cyclopropylmethyl, homologs and isomers of, for example, n-pentyl, n-hexyl, n-heptyl, n-octyl and the like. Unsaturated alkyl is alkyl having one or more double or triple bonds. Examples of unsaturated alkyl groups include, but are not limited to, ethenyl, 2-propenyl, crotyl, 2-isopentenyl, 2- (butadienyl), 2, 4-pentadienyl, 3- (1, 4-pentadienyl), ethynyl, 1-and 3-propynyl, 3-butynyl, and higher homologs and isomers. Unless otherwise indicated, the term "alkyl" is also meant to include those alkyl derivatives defined in more detail below, such as "heteroalkyl. Alkyl groups limited to hydrocarbyl groups are referred to as "higher alkyl groups". The term "alkyl" may also mean "alkylene" or "alkyldiyl" and alkylidene in those instances where alkyl is a divalent group.

As used herein, the term "alkylene" or "alkyldiyl", alone or as part of another substituent, means a divalent group derived from an alkyl group, such as from-CH ₂CH ₂CH ₂- (propylene or propane-1, 3-diyl) is exemplified by, but not limited to, and further includes those described below, such as "heteroalkylene". Typically, the alkyl (or alkylene) group will have from 1 to about 30 carbon atoms, preferably from 1 to about 25 carbon atoms, more preferably from 1 to about 20 carbon atoms, even more preferably from 1 to about 15 carbon atoms and most preferably from 1 to about 10 carbon atoms. "lower alkyl", "lower alkylene" or "lower alkyldiyl" is a shorter chain alkyl, alkylene or alkyldiyl group, typically having about 10 or less carbon atoms, about 8 or less carbon atoms, about 6 or less carbon atoms or about 4 or less carbon atoms.

As used herein, the term "alkylidene" alone or as part of another substituent means a divalent group derived from an alkyl group, such asCH ₃CH ₂CH ₂Is exemplified by (propylidene) but not limited thereto. Typically, the alkylidene group will have from 1 to about 30 carbon atoms, preferably from 1 to about 25 carbon atoms, more preferably from 1 to about 20 carbon atoms, even more preferably from 1 to about 15 carbon atoms and most preferably from 1 to about 10 carbon atoms. "lower alkyl" or "lower alkyl" is a shorter chain alkyl or alkylidene group, typically having about 10 or fewer carbon atoms, about 8 or fewer carbon atoms, about 6 or fewer carbon atoms, or about 4 or fewer carbon atoms.

As used herein, the terms "alkoxy", "alkylamino" and "alkylthio" (or thioalkoxy) are used in their conventional sense and refer to those alkyl groups attached to the remainder of the molecule via an oxygen atom, an amino group or a sulfur atom, respectively.

Unless otherwise specified, the term "heteroalkyl," alone or in combination with another term, is intended herein to mean a stable straight or branched chain or cyclic hydrocarbon group, or combinations thereof, consisting of the specified number of carbon atoms and at least one heteroatom selected from the group consisting of: o, N, Si, S and B, and wherein the nitrogen and sulfur atoms may optionally be oxidized, and the nitrogen heteroatom may optionally be quaternized. The heteroatom O, N, B, S and Si can be located at any internal position of the heteroalkyl group or at the position where the alkyl group is attached to the remainder of the molecule. Examples include, but are not limited to, -CH ₂-CH ₂-O-CH ₃、-CH ₂-CH ₂-NHCH ₃、-CH ₂-CH ₂-N(CH ₃)-CH ₃、-CH ₂-S-CH ₂-CH ₃、-CH ₂-CH ₂,-S(O)-CH ₃、-CH ₂-CH ₂-S(O)2-CH ₃、-CH＝CH-O-CH ₃、-Si(CH ₃) ₃、-CH ₂-CH＝N-OCH ₃and-CH ═ CH-N (CH) ₃)-CH ₃. Up to two heteroatoms may be consecutive, e.g. -CH ₂-NH-OCH ₃and-CH ₂-O-Si(CH ₃) ₃. Similarly, the term "heteroalkylene" alone or as part of another substituent means a divalent radical derived from a heteroalkyl group, such as represented by the formula-CH ₂-CH ₂-S-CH ₂-CH ₂-and-CH ₂-S-CH ₂-CH ₂-NH-CH ₂By way of illustration and not limitation. For heteroalkylene groups, heteroatoms can also occupy either or both of the chain ends (e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, and the like). Optionally, for alkylene and heteroalkylene linking groups, the written direction of the formula for the linking group does not indicate an orientation of the linking group. For example of the formula-CO ₂R ' -optionally represents-C (O) OR ' and-OC (O) R '.

Unless otherwise indicated, the terms "cycloalkyl" and "heterocycloalkyl", alone or in combination with other terms, herein denote the cyclic forms of "alkyl" and "heteroalkyl", respectively. In addition, for heterocycloalkyl, a heteroatom may occupy a position where the heterocycle is attached to the rest of the molecule. Examples of cycloalkyl groups include, but are not limited to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like. Examples of heterocycloalkyl groups include (but are not limited to): 1- (1,2,5, 6-tetrahydropyridinyl), 1-piperidyl, 2-piperidyl, 3-piperidyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl, tetrahydrothien-2-yl, tetrahydrothien-3-yl, 1-piperazinyl, 2-piperazinyl and the like.

Unless otherwise specified, the terms "halo" or "halogen" herein, alone or as part of another substituent, mean a fluorine, chlorine, bromine or iodine atom. Additionally, terms such as "haloalkyl" are intended to include monohaloalkyl and polyhaloalkyl. For example, the term "halo (C) ₁-C ₄) Alkyl "is intended to include, but is not limited to, trifluoromethyl, 2,2, 2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

Unless otherwise indicated, the term "aryl" herein means a polyunsaturated aromatic substituent which can be a single ring or multiple rings (preferably 1 to 3 rings) which are fused together or linked covalently. The term "heteroaryl" refers to aryl (or ring) containing one to four heteroatoms selected from N, O, S, Si and B, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atom is optionally quaternized. The heteroaryl group may be attached to the rest of the molecule through a heteroatom. Non-limiting examples of aryl and heteroaryl groups include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl, 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-benzothiazolyl, purinyl, 2-benzimidazolyl, 5-indolyl, 1-isoquinolyl, 5-isoquinolyl, 2-quinoxalyl, 5-quinoxalyl, 3-quinolyl and 6-quinolyl. The substituents for each of the above-indicated aryl and heteroaryl ring systems are selected from the group of acceptable substituents described below.

For brevity, when used herein, the term "aryl" in combination with other terms (e.g., aryloxy, arylsulfenoxy, aralkyl) includes both aryl and heteroaryl rings as defined above. Thus, the term "aralkyl" is intended to include those groups in which an aryl group is attached to an alkyl group (e.g., benzyl, phenethyl, pyridylmethyl and the like), including those alkyl groups in which a carbon atom (e.g., methylene) is replaced by, for example, an oxygen atom (e.g., phenoxymethyl, 2-pyridyloxymethyl, 3- (1-naphthyloxy) propyl, and the like).

Each of the above terms (e.g., "alkyl," "heteroalkyl," "aryl," and "heteroaryl") is intended to include both substituted and unsubstituted forms of the indicated group. Preferred substituents for each type of group are provided below.

Substituents for alkyl and heteroalkyl (including those groups commonly referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl) are generally referred to as "alkyl substituents" and may be one or more of a variety of groups selected from (but not limited to) the following: substituted OR unsubstituted aryl, substituted OR unsubstituted heteroaryl, substituted OR unsubstituted heterocycloalkyl, -OR ', -O, ═ NR ', -N-OR '-NR 'R ", -SR', -halogen, -SiR 'R" R' ", -OC (O) R ', -C (O) R', -CO ₂R’、-CONR’R”、-OC(O)NR’R”、-NR”C(O)R’、-NR’-C(O)NR”R”’、-NR”C(O) ₂R’、-NR-C(NR’R”R’”)＝NR””、-NR-C(NR’R”)＝NR’”、-S(O)R’、-S(O) ₂R’、-OS(O) ₂R’、-S(O) ₂NR’R”、-NRSO ₂R', -CN and-NO ₂The number is in the range of zero to (2m '+ 1), where m' is the total number of carbon atoms in such group. R', R "and R" "each preferably independently refer to hydrogen, substituted or unsubstituted heteroalkyl, substituted or unsubstituted aryl (e.g., aryl substituted with 1-3 halogens), substituted or unsubstituted alkyl, alkoxy or thioalkoxy, or aralkyl. When a compound of the invention includes more than one R group, for example, each R group is independently selected, as are the R ', R ", R'" and R "" groups when more than one of the R, R ', R ", R'" and R "" groups is present. When R' and R "are attached to the same nitrogen atom, they may combine with the nitrogen atom to form a 5-, 6-or 7-membered ring. For example, -NR' R "is intended to include, but is not limited to, 1-pyrrolidinyl and 4-morpholinyl. From the above discussion of substituents, those skilled in the art will appreciate that the term "alkyl" is intended to include groups that are bonded to carbon atoms of groups other than hydrogen, such as haloalkyl (e.g., -CF) ₃and-CH ₂CF ₃) And acyl (e.g., -C (O) CH) ₃、-C(O)CF ₃、-C(O)CH ₂OCH ₃Etc.).

Similar to the substituents described for alkyl, aryl and heteroaryl groups are generally referred to as "aryl substituents". The substituents are selected from (for example): substituted OR unsubstituted alkyl, substituted OR unsubstituted aryl, substituted OR unsubstituted heteroaryl, substituted OR unsubstituted heterocycloalkyl, -OR ', -O, ═ NR ', -N-OR ', -NR ' R ", -SR ', -halo, -SiR ' R" R ' ", -OC (O) R ', -C (O) R ', -CO2R ', -CONR ' R", -OC (O) NR ' R ", -NR" C (O) R ', -NR ' -C (O) NR "R '", -NR "C (O) ' -C (O) (" R ' ") ₂R’、-NR-C(NR’R”R’”)＝NR””、-NR-C(NR’R”)＝NR’”、-S(O)R’、-S(O) ₂R’、-S(O) ₂NR’R”、-NRSO ₂R', -CN and-NO ₂、-R’、-N ₃-CH(Ph) ₂Fluorine (C) ₁-C ₄) Alkoxy and fluorine (C) ₁-C ₄) Alkyl groups ranging in number from zero to the total number of open valences on the aromatic ring system; and wherein R ', R ", R'" and R "" are preferably independently selected from the group consisting of hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted aryl and substituted or unsubstituted heteroaryl. When a compound of the invention includes more than one R group, for example, each R group is independently selected, as are the R ', R ", R'" and R "" groups when more than one of the R, R ', R ", R'" and R "" groups is present.

A combination of polyhalogen and polyamine reagents is described that produces hyperbranched polymers that do not contain β hydroxyl groups.

It is believed that the pKa of the β hydroxyl groups has a pKa of about 13.9, which is close to the pKa of the sugar subunit to which the glycan is attached, thus, pH values that result in ionization of the glycan moiety (as an anion) can also result in ionization of the β hydroxyl groups, resulting in low retention of the anion exchange resin.

In one embodiment, the hyperbranched polymer may be formed from dihalo alkyl and permethylated triamines as shown in FIGS. 1 and 2, respectively, first, these two reagents are brought together in a 1:1 molar ratio to form a bottom layer on a surface sulfonated ultramacroporous resin, then reacted alternately with 1, 4-dibromobutane, then reacted with pentamethyldipropylenetriamine to form a hyperbranched anion exchange polymer that does not have any β hydroxyl groups.

Applicants believe that the charge density of the quaternary ion exchange sites in the hyperbranched structure may be too high, which would result in overstressing the bond between the two ion exchange sites. In one embodiment, applicants believe that the spacer length between the quaternary amine group pair may be about equal to or greater than the propyl group so that the ion exchange site density is not too high. However, if the spacer group is too long, the ion exchange site density may be too low to effectively separate anions and provide a sufficiently high ion exchange capacity.

In another embodiment, permethylated amines such as pentamethyldibutylideriamine or pentamethyldihexamethyleneideriamine may be used in combination with 1, 4-dibromobutane, which will allow for the positioning of quaternary amine ion exchange sites separated by butyl or hexyl groups.

In another embodiment, the permethylated diamines may be used in combination with 1,3, 5-tribromomethylbenzenes to produce hyperbranched structures examples of permethylated diamines are tetramethyl-1, 4-butanediamine or tetramethyl-1, 6-hexanediamine 106 these agents may also form hyperbranched polymers that do not contain any β hydroxyl groups.

In one embodiment, a series of condensation reaction products or polymer layers may be formed on a substrate. The polymer formed in the polymerization reaction with the polyalkylpolyamine and the polyhalogenated hydrocarbon may be referred to as a polycondensate or polycondensate reaction product or polycondensate layer. Similarly, the condensation reaction product CRP may be a product from a condensation reaction between a polymer and a reagent (e.g., an alkyl halide or amine reagent chemical) that results in the loss of halide or halide and hydrogen. It should be noted that the reaction of the alkyl halide and the tertiary amine reagent results in a loss of halide and no loss of hydrogen. The reaction of alkyl halides with primary or secondary amine-based reagents results in the loss of halide and hydrogen, where the hydrogen produced can interfere with the polymerization process.

The polyalkyl polyamine may be a polyalkyltriamine or a polyalkyldiamine. FIG. 1 illustrates polyalkyltriamines in the form of 2,6, 10-trimethyl-2, 6, 10-triazaundecane (pentamethyldipropylenetriamine, PMDPTA 102) and 2,9, 13-trimethyl-2, 9, 13-triazaheptadecane (pentamethyldihexyltriamine, PMDHTA 104). Referring to fig. 1, the alkyl spacer for the polyalkyltriamine may have an n value of about 3 to about 10, preferably about 3 to about 6. FIG. 1 also illustrates polyalkyldiamines in the form of N, N, N ', N' -tetramethyl-1, 6-hexanediamine TMHDA 106.

The polyalkyl polyamine can be a reagent compound comprising two or more alkyl groups and two or more amine groups. In one embodiment, the polyalkyl polyamine may have some or all of the amine groups alkylated. The alkylated amine groups of the polyalkyl polyamine may have some or all of the amine groups as tertiary amines. Although fig. 1 illustrates only a polyalkyl polyamine reagent having an alkyl group attached to an amine, some or all of the alkyl groups can be alkyl ethers, cycloalkyl groups, aralkyl groups, and combinations thereof. In one embodiment, the alkane attached between two amine groups (i.e., spacer groups) can have a length of 3 to 10 atoms, and preferably has a length of 3 to 6 atoms.

The polyhalogenated hydrocarbon may be a polyhalogenated alkane or a polyhalogenated alkylaryl. The polyhalogenated alkane may be a dihalogenated alkane, such as 1-4-dibromobutane DBB202, as shown in figure 2. The polyhaloalkylaryl can be a trihaloalkylaryl, such as 1,3, 5-tris (bromomethyl) benzene TBMB204, as shown in FIG. 2. In one embodiment, the dihaloalkane may be represented by formula 206, as shown in figure 2. The value n associated with chemical formula 206 may be 3 to 10, preferably 3 to 6.

The polyhalogenated hydrocarbon may be a reagent compound comprising two or more halogen groups attached to the hydrocarbon. The halo group may be a bromo or iodo group. The polyhalogenated hydrocarbon may have 2 to 10 halogen groups, preferably 2 to 3 halogen groups. In one embodiment, the spacer between the two halo groups of the polyhalogenated hydrocarbon may have a length of 3 to 10 atoms, and preferably has a length of 3 to 6 atoms. The hydrocarbon portion (spacer) of the polyhalogenated hydrocarbon between a pair of halo groups can include alkyl, cycloalkyl, aralkyl, dialkyl ether groups and combinations thereof.

Fig. 3 shows a schematic representation of a base polymer layer 300 formed from reaction with a first PMDPTA 102 and a first DBB202, wherein the base polymer layer 300 is attached to negatively charged substrate particles the negatively charged substrate particles may be contained as a packed bed in a chromatography column through which a solution of PMDPTA 102 and dihaloalkane 202 may flow to form the base polymer layer on the negatively charged substrate particles it is noted that the base polymer layer 300 does not have β hydroxyl groups because no epoxide reagent is used, as shown in fig. 3, the base polymer layer 300 includes quaternary amines and tertiary amines, in one aspect, the molar ratio may be 1:1 molar ratio of PMDPTA 102 and DBB 304 to form the base polymer layer, although the base polymer layer 300 is depicted as linear (by reaction of only the terminal amines of PMDPTA 102), as shown in fig. 3, it is possible that some of the intermediate amine groups (e.g. 302) are quaternized and form branched or crosslinked portions, as shown in fig. 3, the base layer 300 may be formed in the presence of negatively charged polymeric particles, wherein the base layer associates with negatively charged polymeric particles and/or the quaternary amine groups may be associated with a bromide salt, as shown in fig. 3, simply fig. 5, as shown in fig. 5.

The negatively charged polymeric particles can be any inert polymeric substrate particles that are chemically stable under the conditions of intended use (e.g., pH 0 to 14). The polymeric particles may be based on a Divinylbenzene (DVB) crosslinking monomer and a carrier resin monomer, wherein the carrier resin monomer may be an Ethylvinylbenzene (EVB) monomer, a styrene monomer, and combinations thereof. The mole percentage of DVB may be 55% and EVB may be 45%. The carrier resin particles may range in diameter from about 1 micron to about 20 microns, preferably from about 2 microns to about 10 microns, and more preferably from about 3 microns to about 7 microns. The surface area of the carrier resin particles may range from about 20m ²G to about 800m ²A/g, preferably of about 20m ²G to about 500m ²In g, more preferably about 20m ²G to about 100m ²A/g, and even more preferably about 20m ²G to about 30m ²(ii) in terms of/g. The carrier resin particles may have a pore size ranging from about 1000 angstroms to about 2000 angstroms.

In some embodiments, the negatively charged substrate particles can include one or more Super Macroporous Particles (SMPs). SMPs are available from commercial sources, including Agilent PLRP-s1000A and Waters Styragel HR4-HR 6. The diameter of the super-macroporous particles is 4-6 μm, and the surface area is 20-30m ²Per g, pore diameter of

And the crosslinking molar ratio of divinylbenzene was 55% and the molar ratio of ethylvinylbenzene was 45%.

The polymeric particles may alternatively be based on other vinyl aromatic monomers, such as α -methyl styrene, chlorostyrene, chloromethyl styrene, vinyl toluene, vinyl naphthalene, and combinations thereof.

The polymeric particles may be sulfonated to produce a negative charge at least on the surface of the particles. For example, particles made with 55% DVB and 45% EVB can be sulfonated by treating the particles with glacial acetic acid and concentrated sulfuric acid.

The base layer 300 may be reacted with a number of reaction cycles of reagents to form a layer of a polyalkyl polyamine condensate. In one embodiment, the number of reaction cycles may be from 1 to 10, preferably from 3 to 10, more preferably from 3 to 6. Each reaction cycle comprises a) a dihaloalkane treatment and b) a polyalkyltriamine treatment. After performing multiple reaction cycles, the polyalkyl polyamine condensate layer may be reacted with a monohaloalkane treatment to alkylate any remaining tertiary amines.

For the first step a) of the first cycle, the second DBB202 may react with the tertiary amine of the base layer 300 to form a first alkyl Condensation Reaction Product (CRP)400 having pendant epoxy groups 402, as shown in fig. 4. Further, at least a portion of the tertiary amine 302 of the base layer 300 is alkylated to form a quaternary amine having a positive charge. Notably, such positive charges of the quaternary amines are believed to assist the ionic binding of the base layer 300 to the negatively charged particles. The quaternary amines of the base polycondensation layer 300 are spaced apart by either the first spacer 406 or the second spacer 408. The first spacer 406 is derived from the PMDPTA 102 and the second spacer 408 is derived from the DBB202, as shown in fig. 3 and 4.

For the second step b) of the first cycle, the pendant halogen group 402 of the first alkyl CRP 400 can be reacted with a second PMDPTA (second polyalkyltriamine) to form a first polyalkylpolyamine CRP500, as shown in fig. 5. The first polyalkylpolyamine CRP500 is covalently linked to the first alkyl CRP 400. Each branch of the first polyalkylpolyamine CRP500 includes a quaternary amine 502 and two tertiary amines 504 and 506, as shown in fig. 5. Although fig. 5 shows PMDPTA as reacting with only one terminal amine group 502, it is possible that some PMDPTA compounds crosslink by reacting two or three amine groups with the halogen group of first alkyl CRP 400. Furthermore, it is possible that only the intermediate amine group 504 of PMDPTA reacts with the halide group of the first alkyl CRP 400.

Now that one reaction cycle of two steps has been performed, a second cycle of two steps can be performed to produce the hyperbranched structure. For the first step a) of the second cycle, the two tertiary amines 504 and 506 of the first polyalkylpolyamine CRP500 may each be reacted with two dihaloalkanes (third DBBs) to form a second dialkyl condensation reaction product 600, as shown in fig. 6. The second alkyl CRP 600 is covalently linked to the first polyalkylpolyamine CRP 500.

Although fig. 6 shows each dihaloalkane compound as reacting with only one halo group (of two halo groups), it is possible that some dihaloalkane compounds are cross-linked by 2 halo groups all reacting with the first polyalkylpolyamine CRP500 or one halo group of a dihaloalkane reacts with one portion of the first polyalkylpolyamine CRP500 while another halo group of the same dihaloalkane reacts with another portion of the first polyalkylpolyamine CRP 500.

For the second step b) of the second cycle, the side chain halogen groups of the second alkyl CRP 600 can then be reacted with a third PMDPTA to form a second polyalkylpolyamine CRP 700 having a quaternary amine 702 and two tertiary amines 704 and 706, as shown in fig. 7. A second polyalkylenepolyamine CRP 700 is covalently linked to a second alkyl CRP 600.

The third reaction cycle of the two steps may be performed with the second polyalkylenepolyamine CRP 700. For the first step a) of the third cycle, the two tertiary amines 704 and 706 of the first polyalkylpolyamine CRP 700 may each be reacted with two dihaloalkanes (fourth DBBs) to form a third alkyl CRP (not shown). It should be noted that the structure of the third alkyl CRP is similar to the structure of the second alkyl CRP 600.

For the second step b) of the third cycle, the side chain halogen groups of the third alkyl CRP can then be reacted with the fourth PMDPTA to form a third polyalkylenepolyamine CRP (not shown) having two tertiary amines. It should be noted that the structure of the third polyalkylenepolyamine CRP is similar to the structure of the second polyalkylenepolyamine CRP 700.

In one embodiment, more than 3 cycles may be performed by repeating steps a) and b). It should be noted that the response using PMDPTA can be altered by replacing PMDPTA with PMDHTA, as shown in fig. 3 to 7. Anion exchange immobilization using PMDPTA has a higher density of anion exchange sites than anion exchange stationary phases using PMDHTA because the length of the spacer chain increases from C3 to C6. It should also be noted that fig. 3-7 are exemplary, and that PMDPTA may be substituted with other polyalkylpolyamines as described herein, and DBB may also be substituted with other polyhalogenated hydrocarbons as described herein.

In another embodiment, a hyperbranched ion-exchange stationary phase using a trihaloalkylaryl compound and a polyalkyldiamine will be described. Fig. 15 shows a schematic of a base polymer layer 1500 formed by reaction with the first TMHDA 106 and the first TBMB204, wherein the base polymer layer 1500 is attached to negatively charged substrate particles. As shown in fig. 15, the base polycondensation layer 1500 includes a quaternary amine. The term x in fig. 15 may be from about 5 to about 150.

The base layer 1500 can be reacted in several reaction cycles of reagents to form a layer of a polyalkyl polyamine condensate. In one embodiment, the number of reaction cycles may be from 1 to 10, preferably from 3 to 10, more preferably from 3 to 6. Each reaction cycle comprises a) a polyalkyldiamine treatment and b) a trihaloalkylaryl treatment. After performing multiple reaction cycles, the polyalkyl polyamine condensate layer may be reacted with a tertiary amine treatment to convert the remaining bromomethyl groups to quaternary amines.

For the first step a) of the first cycle, the second TMHDA 106 may react with the pendant halogen groups of the base polymer layer 1500 to form a first polyalkylpolyamine CRP 1600, as shown in fig. 16. The quaternary amines of the base polycondensation layer 1500 are spaced apart by either the first spacer 1602 or the second spacer 1604. The first spacer 1602 is derived from TBMB204 and the second spacer 1604 is derived from TMHDA 106, as shown in fig. 15 and 16.

For the second step b) of the first cycle, the side-chain tertiary amine 1606 of the first polyalkylpolyamine CRP 1600 can react with the second TBMB204 (second trihaloalkylaryl) to form a first polyalkylaryl CRP1700, as shown in fig. 17. The first polyalkylaryl CRP1700 is covalently linked to the first polyalkylpolyamine CRP 1600. The first polyalkylaryl CRP1700 includes two halo groups per benzyl ring as shown in FIG. 17.

Now that one reaction cycle of two steps has been performed, a second cycle of two steps can be performed to produce the hyperbranched structure. For the first step a) of the second cycle, two halogen groups in each benzyl ring of the polyalkylaryl CRP1700 can react with the third TMHDA 106 to form a second polyalkylpolyamine CRP1800, as shown in fig. 18. The second polyalkylpolyamine CRP1800 is covalently linked to the first polyalkylaryl CRP 1700.

For the second step b) of the second cycle, the pendant tertiary amine groups of the second polyalkylpolyamine CRP1800 may then be reacted with the third TBMB204 to form a second polyalkylaromatic CRP1900, as shown in fig. 19. The second polyalkyl-aromatic CRP1900 is covalently linked to the second polyalkylpolyamine CRP 1800.

A third reaction cycle of two steps may be performed with the second polyalkylaryl CRP 1900. For the first step a) of the third cycle, the two halogen groups in each benzyl ring of the second polyalkylaryl CRP1900 can be reacted with third TMHDA 106 to form a third polyalkylpolyamine CRP (not shown). The third polyalkylpolyamine CRP is covalently linked to the second polyalkylaryl CRP 1900. It should be noted that the structure of the third polyalkylenepolyamine CRP is similar to the structure of the second polyalkylenepolyamine CRP 1800.

For the second step b) of the third cycle, the pendant tertiary amine groups of the third polyalkylenepolyamine CRP may then react with the fourth TBMB204 to form a third polyalkylarylcrp (not shown). The third polyalkylaryl CRP is covalently linked to a third polyalkylpolyamine CRP (not shown). It should be noted that the structure of the third polyalkylaryl CRP is similar to that of the second polyalkylaryl CRP 1900.

In one embodiment, more than 3 cycles may be performed by repeating steps a) and b). When the number of reaction cycles is complete, the polyalkyl polyamine condensate layer may be reacted with a tertiary amine treatment to convert the remaining bromomethyl groups to quaternary amines. It should also be noted that fig. 15-19 are exemplary, TMHDA may be substituted with other polyalkylpolyamines having at least two amine groups and TBMB204 may also be substituted with polyhalohydrocarbons having three or more halo groups.

A chromatography system adapted to a waveform containing four voltages will be described below. Fig. 14 shows an embodiment of a chromatography system, which is an ion chromatography system 1400, comprising a pump 1402, an electrolytic eluent generation device 1403, a degassing assembly 1410, an injection valve 1412, a chromatographic separation device 1414, a detector 1416, and a microprocessor 1418. Recirculation line 1420 can be used to transfer liquid from the output of detector 1416 to the regeneration channel of degassing assembly 1410. The degassing assembly may also be in the form of a vacuum degasser.

The pump 1402 can be configured to draw liquid from a liquid source and fluidly connect with the electrolytic eluent generation device 1403. The electrolytic eluent generation device 1403 is configured to generate an eluent, such as NaOH or methanesulfonic acid. Details regarding electrolytic eluent generation devices (e.g., eluent generators) can be found in U.S. Pat. nos. 6,225,129 and 6,682,701. In one embodiment, residual gases such as carbon dioxide, hydrogen and oxygen may be generated or present in the eluent. The gas may be purged with a degas assembly 1410 using a recirculating liquid through a recirculating line 1420 located downstream of the detector 1416. Injection valve 1412 may be used to inject an aliquot of the sample into the eluate stream. Chromatographic separation device 1414, such as an ion exchange chromatography column, can be used to separate various matrix components present in a liquid sample from analytes of interest. The output of the chromatographic separation device 1414 may be fluidly connected to a detector 1416 to measure the presence of separated chemical components in the liquid sample. The chromatographic separation device 1414 may separate one or more analytes of the sample output from the chromatographic separation device 1414 at different retention times.

Detector 1416 can be in the form of an electrochemical detector that includes a flow channel configured to be in fluid contact with at least a working electrode, a reference electrode, and an optional counter electrode. Details regarding electrochemical detector flow cells and disposable working electrodes may be found in U.S. patent 8,925,374; 8,342,007, respectively; and 6,783,645, which are incorporated herein by reference in their entirety. The electrochemical detector also includes a potentiostat or analytical device to apply a voltage waveform between the working electrode and the reference electrode, and optionally to pass current between the counter electrode and the working electrode. Details regarding analytical devices employing quadrupole voltage waveforms can be found in U.S. patent 8,636,885, which is incorporated herein by reference in its entirety.

The electronic circuitry may include a microprocessor 1418 and a memory portion. The microprocessor 1418 may be used to control the operation of the chromatography system 1400. Microprocessor 1418 can be integrated into chromatography system 1400 or part of a personal computer in communication with chromatography system 1400. Microprocessor 1418 can be configured to communicate with and control one or more components of the chromatography system, such as pump 1402, electrolytic eluent generation device 1403, injection valve 1412, and detector 1416. The memory portion may contain instructions on how to apply the amplitude, polarity, and timing of one or more voltage waveforms. In terms of measurement, the memory portion may also contain instructions regarding the time period over which the present value is sampled to integrate the signal and/or measure the total charge for a particular time period.