Analytical and Bioanalytical Chemistry

, Volume 408, Issue 30, pp 8681–8689 | Cite as

Surfactant-free microemulsion electrokinetic chromatography (SF-MEEKC) with UV and MS detection - a novel approach for the separation and ESI-MS detection of neutral compounds

  • Urška Mohorič
  • Andrea Beutner
  • Sebastian Krickl
  • Didier Touraud
  • Werner Kunz
  • Frank-Michael MatysikEmail author
Research Paper
Part of the following topical collections:
  1. Fundamental Aspects of Electromigrative Separation Techniques


Microemulsion electrokinetic chromatography (MEEKC) is a powerful tool to separate neutral species based on differences in their hydrophobic and hydrophilic properties. However, as a major drawback the conventionally used SDS based microemulsions are not compatible with electrospray ionization mass spectrometry (ESI-MS). In this work, a surfactant-free microemulsion (SFME) consisting of water, ethanol, and 1-octanol is used for surfactant-free microemulsion electrokinetic chromatography (SF-MEEKC). Ammonium acetate was added to the SFME enabling electrophoretic separations. The stability of SFMEs containing ammonium acetate was investigated using small-angle X-ray scattering and dynamic light scattering. A method for the separation of a model system of hydrophobic and hydrophilic neutral vitamins, namely the vitamins B2 and D3, and the cationic vitamin B1 was developed using UV/VIS detection. The influence of the ammonium acetate concentration on the separation performance was studied in detail. The method was characterized concerning reproducibility of migration times and peak areas and concerning the linearity of the calibration data. Furthermore, SF-MEEKC was coupled to ESI-MS investigating the compatibility between SFMEs and the ESI process. The signal intensities of ESI-MS measurements of the model analytes were comparable for SFMEs and aqueous systems. Finally, the vitamin D3 content of a drug treating vitamin D3 deficiency was determined by SF-MEEKC coupled to ESI-MS using 25-hydroxycholecalciferol as an internal standard.

Graphical abstract

The concept of surfactant-free microemulsion electrokinetic chromatography coupled to electrospray ionization mass spectrometry


Surfactant-free microemulsion MEEKC Electrospray ionization mass spectrometry Hydrophobic and hydrophilic neutral compounds Electrokinetic chromatography 



The authors thank Tobias Lopian from the Institut de Chimie Séparative Marcoule (ICSM) for SAXS measurements. S.K. thanks the Fonds der chemischen Industrie (FCI) for a scholarship.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

216_2016_57_MOESM1_ESM.pdf (1.7 mb)
ESM 1 (PDF 1.72 mb)


  1. 1.
    Terabe S, Otsuka K, Ichikawa K, Tsuchiya A, Ando T. Electrokinetic separations with micellar solutions and open-tubular capillaries. Anal Chem. 1984;56:111–3.CrossRefGoogle Scholar
  2. 2.
    Altria KD, Mahuzier PE, Clark BJ. Background and operating parameters in microemulsion electrokinetic chromatography. Electrophoresis. 2003;24(3):315–24.CrossRefGoogle Scholar
  3. 3.
    Pyell U, ed. Electrokinetic chromatography: theory, instrumentation and applications. John Wiley & Sons; 2006.Google Scholar
  4. 4.
    Tripodi V, Flor S, Carlucci A, Lucangioli S. Simultaneous determination of natural and synthetic estrogens by EKC using a novel microemulsion. Electrophoresis. 2006;27:4431–8.CrossRefGoogle Scholar
  5. 5.
    Vomastova L, Mikšik I, Deyl Z. Microemulsion and micellar electrokinetic chromatography of steroids. J Chromatogr B Biomed Sci Appl. 1996;681:107–13.CrossRefGoogle Scholar
  6. 6.
    Mahuzier PE, Altria KD, Clark BJ. Selective and quantitative analysis of 4-hydroxybenzoate preservatives by microemulsion electrokinetic chromatography. J Chromatogr A. 2001;924(1–2):465–70.CrossRefGoogle Scholar
  7. 7.
    Huang HY, Lai YC, Chiu CW, Yeh JM. Comparing micellar electrokinetic chromatography and microemulsion electrokinetic chromatography for the analysis of preservatives in pharmaceutical and cosmetic products. J Chromatogr A. 2003;993(1–2):153–64.CrossRefGoogle Scholar
  8. 8.
    Hsi-Ya H, Chia-Ling C, Chen-Wen C, Jui-Ming Y. Application of microemulsion electrokinetic chromatography for the detection of preservatives in foods. Food Chem. 2005;89(2):315–22.CrossRefGoogle Scholar
  9. 9.
    Xie JP, Zhang JY, Liu HX, et al. Microemulsion electrokinetic chromatography with laser-induced fluorescence detection: as tested with amino acid derivatives. Biomed Chromatogr. 2004;18(8):600–7.CrossRefGoogle Scholar
  10. 10.
    Javor T, Buchberger W, Tanzcos I. Determination of low-molecular-mass phenolic and non-phenolic lignin degradation compounds in wood digestion solutions by capillary electrophoresis. Microchim Acta. 2000;135(1–2):45–53.CrossRefGoogle Scholar
  11. 11.
    Terabe S, Matsubara N, Ishihama Y, Okada Y. Microemulsion electrokinetic chromatography: comparison with micellar electrokinetic chromatography. J Chromatogr A. 1992;608(1–2):23–9.CrossRefGoogle Scholar
  12. 12.
    Subirats X, Yuan HP, Chaves V, Marzal N, Roses M. Micoemulsion electrokinetic chromatography as a suitable tool for lipophilicity determination of acidic, neutral, and basic compounds. Electrophoresis 2016;37(14):2010–2016.Google Scholar
  13. 13.
    Zemann AJ. Conductivity detection in capillary electrophoresis. Trends Anal Chem. 2001;20(6–7):346–54.CrossRefGoogle Scholar
  14. 14.
    Gennaro LA, Salas-Solano O, Ma S. Capillary electrophoresis-mass spectrometry as a characterization tool for therapeutic proteins. Anal Biochem. 2006;355(2):249–58.CrossRefGoogle Scholar
  15. 15.
    Olivares JA, Nguyen NT, Yonker CR, Smith RD. On-line mass spectrometric detection for capillary zone electrophoresis. Anal Chem. 1987;59(9):1230–2.CrossRefGoogle Scholar
  16. 16.
    Yu LS, Xu XQ, Huang L, Ling JM, Chen GN. Separation and determination of flavonoids using microemulsion EKC with electrochemical detection. Electrophoresis. 2008;29(3):726–33.CrossRefGoogle Scholar
  17. 17.
    Bytzek AK, Reithofer MR, Galanski M, Groessl M, Keppler BK, Hartinger CG. The first example of MEEKC-ICP-MS coupling and its application for the analysis of anticancer platinum complexes. Electrophoresis. 2010;31(7):1144–50.CrossRefGoogle Scholar
  18. 18.
    Hoffmann E, Stroobant V. Mass spectrometry. Principles and applications. John Wiley & Sons; 2007.Google Scholar
  19. 19.
    Matysik F-M, Neusüss C, Pelzing M. Fast capillary electrophoresis coupled with time-of-flight mass spectrometry under separation conditions of high electrical field strengths. Analyst. 2008;133(12):1764–6.CrossRefGoogle Scholar
  20. 20.
    Klossek ML, Touraud D, Zemb T, Kunz W. Structure and solubility in surfactant-free microemulsions. ChemPhysChem. 2012;13(18):4116–9.CrossRefGoogle Scholar
  21. 21.
    Smith GD, Donelan CE, Barden RE. Oil-continuous microemulsions composed of hexane, water, and 2-propanol. J Colloid Interface Sci. 1977;60(3):488–96.CrossRefGoogle Scholar
  22. 22.
    Diat O, Klossek ML, Touraud D, et al. Octanol-rich and water-rich domains in dynamic equilibrium in the pre-ouzo region of ternary systems containing a hydrotrope. J Appl Crystallogr. 2013;46(6):1665–9.CrossRefGoogle Scholar
  23. 23.
    Marcus J, Touraud D, Prévost S, Diat O, Zemb T, Kunz W. Influence of additives on the structure of surfactant-free microemulsions. Phys Chem Chem Phys. 2015;17(48):32528–38.CrossRefGoogle Scholar
  24. 24.
    Vitale SA, Katz JL. Liquid droplet dispersions formed by homogeneous liquid - liquid nucleation: “the ouzo effect”. Langmuir. 2003;19:4105–10.CrossRefGoogle Scholar
  25. 25.
    Schöttl S, Marcus J, Diat O, et al. Emergence of surfactant-free micelles from ternary solutions. Chem Sci. 2014;5(8):2909–3340.CrossRefGoogle Scholar
  26. 26.
    Zemb T, Klossek ML, Lopian T, et al. How to explain the occurrence of microemulsions formed by solvent mixtures without surfactants. Proc Natl Acad Sci U S A. 2015;113(16):4260–5.CrossRefGoogle Scholar
  27. 27.
    Fischer V, Marcus J, Touraud D, Diat O, Kunz W. Toward surfactant-free and water-free microemulsions. J Colloid Interface Sci. 2015;453:186–93.CrossRefGoogle Scholar
  28. 28.
    Hankel RF, Rojas PE, Cano-Sarabia M, et al. Surfactant-free CO2 -based microemulsion-like systems. Chem Commun. 2014;50(60):8215–8.CrossRefGoogle Scholar
  29. 29.
    Zoumpanioti M, Karali M, Xenakis A, Stamatis H. Lipase biocatalytic processes in surfactant free microemulsion-like ternary systems and related organogels. Enzym Microb Technol. 2006;39(4):531–9.CrossRefGoogle Scholar
  30. 30.
    Gawrys KL, Blankenship GA, Kilpatrick PK. Solvent entrainment in and flocculation of asphaltenic aggregates probed by small-angle neutron scattering. Langmuir. 2006;22(10):4487–97.CrossRefGoogle Scholar
  31. 31.
    Lopian T, Schöttl S, Prévost S, Pellet-Rostaing S, Horinek D, Kunz W, et al. Morphologies observed in unltraflexible microemulsions with and without the presence of a strong acid. ACS Cent Sci. 2016;2:467–75.CrossRefGoogle Scholar
  32. 32.
    Callmer K, Davies L. Separation and determination of vitamin B1, B2, B6 and nicotinamide in commercial vitamin preparations using high performance cation exchange chromatography. Chromatographia. 1974;7:644–50.CrossRefGoogle Scholar
  33. 33.
    Boso RL, Bellini MS, Miksik I, Deyl Z. Microemulsion electrokinetic chromatography with different organic modifiers: separation of water- and lipid-soluble vitamins. J Chromatogr A. 1995;709(1):11–9.CrossRefGoogle Scholar
  34. 34.
    Altria KD. Background theory and applications of microemulsion electrokinetic chromatography. J Chromatogr A. 2000;892(1–2):171–86.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Urška Mohorič
    • 1
  • Andrea Beutner
    • 1
  • Sebastian Krickl
    • 2
  • Didier Touraud
    • 2
  • Werner Kunz
    • 2
  • Frank-Michael Matysik
    • 1
    Email author
  1. 1.Institute of Analytical Chemistry, Chemo- and BiosensorsUniversity of RegensburgRegensburgGermany
  2. 2.Institute of Physical and Theoretical ChemistryUniversity of RegensburgRegensburgGermany

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