Skip to main content
Log in

Epoxy-based monoliths for capillary liquid chromatography of small and large molecules

  • Original Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

A versatile epoxy-based monolith was synthesised by polycondensation polymerisation of glycidyl ether 100 with ethylenediamine using a porogenic system consisting of polyethylene glycol, M w = 1000, and 1-decanol. Polymerisation was performed at 80 °C for 22 h. A simple acid hydrolysis of residual epoxides resulted in a mixed diol-amino chemistry. The modified column was used successfully for hydrophilic interaction liquid chromatography (HILIC) of small molecule probes such as nucleic acid bases and nucleosides, benzoic acid derivatives, as well as for peptides released from a tryptic digest of cytochrome c. The mixed-mode chemistry allowed both hydrophilic partitioning and ion-exchange (IEX) interactions to contribute to the separation, providing flexibility in selectivity control. Residual epoxide groups were also exploited for incorporating a mixed IEX chemistry. Alternatively, the surface chemistry of the monolith pore surface rendered hydrophobic via grafting of a co-polymerised hydrophobic hydrogel. The inherent hydrophilicity of the monolith scaffold also enabled high performance separation of proteins under IEX and hydrophobic interaction modes and in the absence of non-specific interactions.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Hosoya K, Hira N, Yamamoto K, Nishimura M, Tanaka N (2006) High-performance polymer-based monolithic capillary column. Anal Chem 78(16):5729–5735. doi:10.1021/ac0605391

    Article  CAS  Google Scholar 

  2. Liu K, Tolley HD, Lee ML (2012) Highly crosslinked polymeric monoliths for reversed-phase capillary liquid chromatography of small molecules. J Chromatogr A 1227:96–104. doi:10.1016/j.chroma.2011.12.081

    Article  CAS  Google Scholar 

  3. Nischang I, Bruggemann O (2010) On the separation of small molecules by means of nano-liquid chromatography with methacrylate-based macroporous polymer monoliths. J Chromatogr A 1217(33):5389–5397. doi:10.1016/j.chroma.2010.06.021

    Article  CAS  Google Scholar 

  4. Nischang I, Teasdale I, Bruggemann O (2011) Porous polymer monoliths for small molecule separations: advancements and limitations. Anal Bioanal Chem 400(8):2289–2304. doi:10.1007/s00216-010-4579-6

    Article  CAS  Google Scholar 

  5. Lubbad SH, Buchmeiser MR (2009) Highly cross-linked polymeric capillary monoliths for the separation of low, medium, and high molecular weight analytes. J Sep Sci 32(15–16):2521–2529. doi:10.1002/jssc.200900188

    Article  CAS  Google Scholar 

  6. Greiderer A, Trojer L, Huck CW, Bonn GK (2009) Influence of the polymerisation time on the porous and chromatographic properties of monolithic poly(1,2-bis(p-vinylphenyl))ethane capillary columns. J Chromatogr A 1216(45):7747–7754. doi:10.1016/j.chroma.2009.08.084

    Article  CAS  Google Scholar 

  7. Urban J, Svec F, Frechet JM (2010) Efficient separation of small molecules using a large surface area hypercrosslinked monolithic polymer capillary column. Anal Chem 82(5):1621–1623. doi:10.1021/ac100008n

    Article  CAS  Google Scholar 

  8. Causon TJ, Shellie RA, Hilder EF (2009) High temperature liquid chromatography with monolithic capillary columns and pure water eluent. Analyst 134(3):440–442. doi:10.1039/b815886j

    Article  CAS  Google Scholar 

  9. Greiderer A, Ligon SC Jr, Huck CW, Bonn GK (2009) Monolithic poly(1,2-bis(p-vinylphenyl)ethane) capillary columns for simultaneous separation of low- and high-molecular-weight compounds. J Sep Sci 32(15–16):2510–2520. doi:10.1002/jssc.200900211

    Article  CAS  Google Scholar 

  10. Urban J, Svec F, Frechet JM (2010) Hypercrosslinking: new approach to porous polymer monolithic capillary columns with large surface area for the highly efficient separation of small molecules. J Chromatogr A 1217(52):8212–8221. doi:10.1016/j.chroma.2010.10.100

    Article  CAS  Google Scholar 

  11. Chambers SD, Svec F, Frechet JM (2011) Incorporation of carbon nanotubes in porous polymer monolithic capillary columns to enhance the chromatographic separation of small molecules. J Chromatogr A 1218(18):2546–2552. doi:10.1016/j.chroma.2011.02.055

    Article  CAS  Google Scholar 

  12. Bui TN, Verhage JJ, Irgum K (2010) Tris(hydroxymethyl)aminomethane-functionalized silica particles and their application for hydrophilic interaction chromatography. J Sep Sci 33(19):2965–2976. doi:10.1002/jssc.201000154

    Article  CAS  Google Scholar 

  13. Svec F (2010) Porous polymer monoliths: amazingly wide variety of techniques enabling their preparation. J Chromatogr A 1217(6):902–924. doi:10.1016/j.chroma.2009.09.073

    Article  CAS  Google Scholar 

  14. Tsujioka N, Hira N, Aoki S, Tanaka N, Hosoya K (2005) A new preparation method for well-controlled 3D skeletal epoxy resin-based polymer monoliths. Macromolecules 38(24):9901–9903. doi:10.1021/ma051409h

    Article  CAS  Google Scholar 

  15. Tsujioka N, Ishizuka N, Tanaka N, Kubo T, Hosoya K (2008) Well-controlled 3D skeletal epoxy-based monoliths obtained by polymerization induced phase separation. J Polymer Sci, Part A: Polymer Chem 46(10):3272–3281. doi:10.1002/pola.22665

    Article  CAS  Google Scholar 

  16. Li Y, Lee ML (2009) Biocompatible polymeric monoliths for protein and peptide separations. J Sep Sci 32(20):3369–3378. doi:10.1002/jssc.200900478

    Article  CAS  Google Scholar 

  17. Xin P, Shen Y, Qi L, Yang G, Chen Y (2011) Preparation of poly(N-isopropylacrylamide)-grafted well-controlled 3D skeletal monolith based on E-51 epoxy resin for protein separation. Talanta 85(2):1180–1186. doi:10.1016/j.talanta.2011.05.037

    Article  CAS  Google Scholar 

  18. Liu J, Ren L, Liu Y, Li H, Liu Z (2012) Weak anion exchange chromatographic profiling of glycoprotein isoforms on a polymer monolithic capillary. J Chromatogr A 1228:276–282. doi:10.1016/j.chroma.2011.08.079

    Article  CAS  Google Scholar 

  19. Hosoya K, Mori T, Sakamoto M, Kubo T, Kaya K (2009) Properties of a non-aromatic epoxy polymer-based monolithic capillary column for μ-HPLC. Chromatographia 70(5):699–704. doi:10.1365/s10337-009-1260-3

    Article  CAS  Google Scholar 

  20. Krenkova J, Gargano A, Lacher NA, Schneiderheinze JM, Svec F (2009) High binding capacity surface grafted monolithic columns for cation exchange chromatography of proteins and peptides. J Chromatogr A 1216(40):6824–6830. doi:10.1016/j.chroma.2009.08.031

    Article  CAS  Google Scholar 

  21. Dinh NP, Cam QM, Nguyen AM, Shchukarev A, Irgum K (2009) Functionalization of epoxy-based monoliths for ion exchange chromatography of proteins. J Sep Sci 32(15–16):2556–2564. doi:10.1002/jssc.200900243

    Article  CAS  Google Scholar 

  22. Peters EC, Svec F, Fréchet JMJ (1997) Thermally responsive rigid polymer monoliths. Adv Mater 9(8):630–633. doi:10.1002/adma.19970090807

    Article  CAS  Google Scholar 

  23. Courtois J, Byström E, Irgum K (2006) Novel monolithic materials using poly(ethylene glycol) as porogen for protein separation. Polymer 47(8):2603–2611. doi:10.1016/j.polymer.2006.01.096

    Article  CAS  Google Scholar 

  24. Jiang Z, Smith NW, Ferguson PD, Taylor MR (2009) Novel highly hydrophilic zwitterionic monolithic column for hydrophilic interaction chromatography. J Sep Sci 32(15–16):2544–2555. doi:10.1002/jssc.200900130

    Article  CAS  Google Scholar 

  25. Chen ML, Li LM, Yuan BF, Ma Q, Feng YQ (2012) Preparation and characterization of methacrylate-based monolith for capillary hydrophilic interaction chromatography. J Chromatogr A 1230:54–60. doi:10.1016/j.chroma.2012.01.065

    Article  CAS  Google Scholar 

  26. Jiang Z, Smith NW, Ferguson PD, Taylor MR (2007) Hydrophilic interaction chromatography using methacrylate-based monolithic capillary column for the separation of polar analytes. Anal Chem 79(3):1243–1250. doi:10.1021/ac061871f

    Article  CAS  Google Scholar 

  27. Alpert AJ (2008) Electrostatic repulsion hydrophilic interaction chromatography for isocratic separation of charged solutes and selective isolation of phosphopeptides. Anal Chem 80(1):62–76. doi:10.1021/ac070997p

    Article  CAS  Google Scholar 

  28. Gilar M, Yu YQ, Ahn J, Fournier J, Gebler JC (2008) Mixed-mode chromatography for fractionation of peptides, phosphopeptides, and sialylated glycopeptides. J Chromatogr A 1191(1–2):162–170. doi:10.1016/j.chroma.2008.01.061

    CAS  Google Scholar 

  29. Liu X, Pohl CA (2010) HILIC behavior of a reversed-phase/cation-exchange/anion-exchange trimode column. J Sep Sci 33(6–7):779–786. doi:10.1002/jssc.200900645

    Article  CAS  Google Scholar 

  30. Zhang K, Dai L, Chetwyn NP (2010) Simultaneous determination of positive and negative pharmaceutical counterions using mixed-mode chromatography coupled with charged aerosol detector. J Chromatogr A 1217(37):5776–5784. doi:10.1016/j.chroma.2010.07.035

    Article  CAS  Google Scholar 

  31. Skerikova V, Jandera P (2010) Effects of the operation parameters on Hydrophilic Interaction Liquid Chromatography separation of phenolic acids on zwitterionic monolithic capillary columns. J Chromatogr A 1217(51):7981–7989. doi:10.1016/j.chroma.2010.07.061

    Article  CAS  Google Scholar 

  32. Alpert AJ (1990) Hydrophilic-interaction chromatography for the separation of peptides, nucleic acids and other polar compounds. J Chromatogr 499:177–196

    Article  CAS  Google Scholar 

  33. Guo Y, Gaiki S (2005) Retention behavior of small polar compounds on polar stationary phases in hydrophilic interaction chromatography. J Chromatogr A 1074(1–2):71–80

    CAS  Google Scholar 

  34. Hemstrom P, Irgum K (2006) Hydrophilic interaction chromatography. J Sep Sci 29(12):1784–1821

    Article  Google Scholar 

  35. Dean JA (1999) Lange’s Handbook of Chemistry, 15th edn. McGraw-Hill, New York

    Google Scholar 

  36. Hatambeygi N, Abedi G, Talebi M (2011) Method development and validation for optimised separation of salicylic, acetyl salicylic and ascorbic acid in pharmaceutical formulations by hydrophilic interaction chromatography and response surface methodology. J Chromatogr A 1218(35):5995–6003. doi:10.1016/j.chroma.2011.06.009

    Article  CAS  Google Scholar 

  37. Horvath CG, Preiss BA, Lipsky SR (1967) Fast liquid chromatography: an investigation of operating parameters and the separation of nucleotides on pellicular ion exchangers. Anal Chem 39(12):1422–1428

    Article  CAS  Google Scholar 

  38. Hao Z, Xiao B, Weng N (2008) Impact of column temperature and mobile phase components on selectivity of hydrophilic interaction chromatography (HILIC). J Sep Sci 31(9):1449–1464. doi:10.1002/jssc.200700624

    Article  CAS  Google Scholar 

  39. Padivitage NL, Armstrong DW (2011) Sulfonated cyclofructan 6 based stationary phase for hydrophilic interaction chromatography. J Sep Sci 34(14):1636–1647. doi:10.1002/jssc.201100121

    Article  CAS  Google Scholar 

  40. Causon TJ, Cortes HJ, Shellie RA, Hilder EF (2012) Temperature pulsing for controlling chromatographic resolution in capillary liquid chromatography. Anal Chem 84(7):3362–3368. doi:10.1021/ac300161b

    Article  CAS  Google Scholar 

  41. Yang Y, Boysen RI, Hearn MT (2009) Hydrophilic interaction chromatography coupled to electrospray mass spectrometry for the separation of peptides and protein digests. J Chromatogr A 1216(29):5518–5524. doi:10.1016/j.chroma.2009.05.085

    Article  CAS  Google Scholar 

  42. Chen X, Tolley HD, Lee ML (2011) Weak cation-exchange monolithic column for capillary liquid chromatography of peptides and proteins. J Sep Sci. doi:10.1002/jssc.201100156

  43. Li Y, Tolley HD, Lee ML (2010) Monoliths from poly(ethylene glycol) diacrylate and dimethacrylate for capillary hydrophobic interaction chromatography of proteins. J Chromatogr A 1217(30):4934–4945. doi:10.1016/j.chroma.2010.05.048

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by the Australian Research Council’s Linkage Infrastructure and Equipment Fund and Discovery Projects Schemes. EFH is the recipient of an ARC Future Fellowship. Technical support from Dr Karsten Gömann (Central Science Laboratory, UTAS) for SEM is gratefully acknowledged.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Emily F. Hilder.

Additional information

Published in the topical collection Monolithic Columns in Liquid Phase Separations with guest editor Luis A. Colon.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM 1

(PDF 86 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Talebi, M., Arrua, R.D., Gaspar, A. et al. Epoxy-based monoliths for capillary liquid chromatography of small and large molecules. Anal Bioanal Chem 405, 2233–2244 (2013). https://doi.org/10.1007/s00216-012-6486-5

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-012-6486-5

Keywords

Navigation