Selection of the optimum electrospray voltage for gradient elution LC-MS measurements

  • Ioan Marginean
  • Ryan T. Kelly
  • Ronald J. Moore
  • David C. Prior
  • Brian L. LaMarche
  • Keqi Tang
  • Richard D. Smith


Changes in liquid composition during gradient elution liquid chromatography (LC) coupled to mass spectrometry (MS) analyses affect the electrospray operation. To establish methodologies for judicious selection of the electrospray voltage, we monitored in real time the effect of the LC gradient on the spray current. The optimum range of the electrospray voltage decreased as the concentration of organic solvent in the eluent increased during reversed-phase LC analyses. These results and related observations provided the means to rationally select the voltage to ensure effective electrospray operation throughout gradient-elution LC separations. For analyses in which the electrospray was operated at constant voltage, a small run-to-run variation in the spray current was observed, indicating a changing electric field resulting from fouling or degradation of the emitter. Algorithms using feedback from spray current measurements that can maintain the electrospray voltage within the optimum operating range throughout gradient elution LC-MS were evaluated. The electrospray operation with voltage regulation and at a constant, judiciously selected voltage during gradient elution LC-MS measurements produced data with similar reproducibility.


Anal Ysis Electrospray Voltage Valco Instrument Spray Current Electrospray Ionization Fourier Transform 


  1. 1.
    Aleksandrov, M. L.; Gall, L. N.; Krasnov, V. N.; Nikolaev, V. I.; Pavlenko, V. A.; Shkurov, V. A.; Baram, G. I.; Gracher, M. A.; Knorre, V. D.; Kusner, Y. S. Direct Coupling of a Microcolumn Liquid Chromatograph and a Mass Spectrometer. Bioorg. Khim. 1984, 10, 710–712.Google Scholar
  2. 2.
    Whitehouse, C. M.; Dreyer, R. N.; Yamashita, M.; Fenn, J. B. Electrospray Interface for Liquid Chromatographs and Mass Spectrometers. Anal. Chem. 1985, 57, 675–679.CrossRefGoogle Scholar
  3. 3.
    Aebersold, R.; Mann, M. Mass Spectrometry-Based Proteomics. Nature 2003, 422, 198–207.CrossRefGoogle Scholar
  4. 4.
    Domon, B.; Aebersold, R. Mass Spectrometry and Protein Analysis. Science 2006, 312, 212–217.CrossRefGoogle Scholar
  5. 5.
    Marginean, I.; Kelly, R. T.; Page, J. S.; Tang, K. Q.; Smith, R. D. Electrospray Characteristic Curves: In Pursuit of Improved Performance in the Nanoflow Regime. Anal. Chem. 2007, 79, 8030–8036.CrossRefGoogle Scholar
  6. 6.
    Jackson, G. S.; Enke, C. G. Electrical Equivalence of Electrospray Ionization with Conducting and Nonconducting Needles. Anal. Chem. 1999, 71, 3777–3784.CrossRefGoogle Scholar
  7. 7.
    Ramanathan, R.; Zhong, R.; Blumenkrantz, N.; Chowdhury, S. K.; Alton, K. B. Response Normalized Liquid Chromatography Nanospray Ionization Mass Spectrometry. J. Am. Soc. Mass Spectrom. 2007, 18, 1891–1899.CrossRefGoogle Scholar
  8. 8.
    Valaskovic, G. A.; Murphy, J. P.; Lee, M. S. Automated Orthogonal Control System for Electrospray Ionization. J. Am. Soc. Mass Spectrom. 2004, 15, 1201–1215.CrossRefGoogle Scholar
  9. 9.
    Staats, S. L. T.; Fogiel, A. J.; Suna, A. Active Spray Control with Electric Field Optimization for Online NanoLC with Polymeric Spray Tips. In Proceedings of the 56th ASMS Conference on Mass Spectrometry, Denver, CO, June 1–5, 2008.Google Scholar
  10. 10.
    Gapeev, A.; Berton, A.; Fabris, D. Current-Controlled Nanospray for the Analysis of Less Than Ideal Samples. In Proceedings of the 56th ASMS Conference on Mass Spectrometry, Denver, CO, June 1–5, 2008.Google Scholar
  11. 11.
    Marginean, I.; Kelly, R. T.; Prior, D. C.; LaMarche, B. L.; Tang, K.; Smith, R. D. Analytical Characterization of the Electrospray Ion Source in the Nanoflow Regime. Anal. Chem. 2008, 80, 6573–6579.CrossRefGoogle Scholar
  12. 12.
    Kinter, M. M.; Sherman, N. E. Protein Sequencing and Identification Using Tandem Mass Spectrometry. New York: Wiley-Interscience.Google Scholar
  13. 13.
    Shen, Y. F.; Tolic, N.; Zhao, R.; Pasa-Tolic, L.; Li, L. J.; Berger, S. J.; Harkewicz, R.; Anderson, G. A.; Belov, M. E.; Smith, R. D. High-Throughput Proteomics Using High Efficiency Multiple-Capillary Liquid Chromatography with On-line High-Performance ESI FTICR Mass Spectrometry. Anal. Chem. 2001, 73, 3011–3021.CrossRefGoogle Scholar
  14. 14.
    Shen, Y. F.; Zhao, R.; Belov, M. E.; Conrads, T. P.; Anderson, G. A.; Tang, K. Q.; Pasa-Tolic, L.; Veenstra, T. D.; Lipton, M. S.; Udseth, H. R.; Smith, R. D. Packed Capillary Reversed-Phase Liquid Chromatography with High-Performance Electrospray Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometry for Proteomics. Anal. Chem. 2001, 73, 1766–1775.CrossRefGoogle Scholar
  15. 15.
    Kelly, R. T.; Page, J. S.; Luo, Q. Z.; Moore, R. J.; Orton, D. J.; Tang, K. Q.; Smith, R. D. Chemically Etched Open Tubular and Monolithic Emitters for Nanoelectrospray Ionization Mass Spectrometry. Anal. Chem. 2006, 78, 7796–7801.CrossRefGoogle Scholar
  16. 16.
    Jaitly, N.; Mayampurath, A.; Littlefield, K.; Adkins, J. N.; Anderson, G. A.; Smith, R. D. Decon2LS: An Open-Source Software Package for Automated Processing and Visualization of High Resolution Mass Spectrometry Data. BMC Bioinformatics 2009, in press.Google Scholar
  17. 17.
    Jaitly, N.; Monroe, M. E.; Petyuk, V. A.; Clauss, T. R. W.; Adkins, J. N.; Smith, R. D. Robust Algorithm for Alignment of Liquid Chromatography-Mass Spectrometry Analyses in an Accurate Mass and Time Tag Data Analysis Pipeline. Anal. Chem. 2006, 78, 7397–7409.CrossRefGoogle Scholar
  18. 18.
    Livesay, E. A.; Tang, K. Q.; Taylor, B. K.; Buschbach, M. A.; Hopkins, D. F.; LaMarche, B. L.; Zhao, R.; Shen, Y. F.; Orton, D. J.; Moore, R. J.; Kelly, R. T.; Udseth, H. R.; Smith, R. D. Fully Automated Four-Column Capillary LC-MS System for Maximizing Throughput in Proteomic Analyses. Anal. Chem. 2008, 80, 294–302.CrossRefGoogle Scholar
  19. 19.
    Nilsson, S.; Svedberg, M.; Pettersson, J.; Bjorefors, F.; Markides, K.; Nyholm, L. Evaluations of the Stability of Sheathless Electrospray Ionization Mass Spectrometry Emitters Using Electrochemical Techniques. Anal. Chem. 2001, 73, 4607–4616.CrossRefGoogle Scholar
  20. 20.
    Chen, M. L.; Cook, K. D. Oxidation Artifacts in the Electrospray Mass Spectrometry of A β Peptide. Anal. Chem. 2007, 79, 2031–2036.CrossRefGoogle Scholar
  21. 21.
    Geromanos, S.; Freckleton, G.; Tempst, P. Tuning of an Electrospray Ionization Source for Maximum Peptide-Ion Transmission into a Mass Spectrometer. Anal. Chem. 2000, 72, 777–790.CrossRefGoogle Scholar
  22. 22.
    Marginean, I.; Kelly, R. T.; Page, J. S.; Tang, K. Q.; Smith, R. D. Toward a Stable Electrospray. Proceedings of the 55th ASMS Conference on Mass Spectrometry, Indianapolis, IN, June 3–7, 2007.Google Scholar
  23. 23.
    Evans, C. A.; Hendricks, C. D. Electrohydrodynamic Ion-Source for Mass-Spectrometry of Liquids. Rev. Sci. Instrum. 1972, 43, 1527–1530.CrossRefGoogle Scholar
  24. 24.
    Chen, D. R.; Pui, D. Y. H.; Kaufman, S. L. Electrospraying of Conducting Liquids for Monodisperse Aerosol Generation in the 4 nm to 1.8 µm Diameter Range. J. Aerosol Sci. 1995, 26, 963–977.CrossRefGoogle Scholar
  25. 25.
    Old, W. M.; Meyer-Arendt, K.; Aveline-Wolf, L.; Pierce, K. G.; Mendoza, A.; Sevinsky, J. R.; Resing, K. A.; Ahn, N. G. Comparison of Label-Free Methods for Quantifying Human Proteins by Shotgun Proteomics. Mol. Cell. Proteomics 2005, 4, 1487–1502.CrossRefGoogle Scholar
  26. 26.
    Ono, M.; Shitashige, M.; Honda, K.; Isobe, T.; Kuwabara, H.; Matsuzuki, H.; Hirohashi, S.; Yamada, T. Label-Free Quantitative Proteomics Using Large Peptide Data Sets Generated by Nanoflow Liquid Chromatography and Mass Spectrometry. Mol. Cell. Proteomics 2006, 5, 1338–1347.CrossRefGoogle Scholar
  27. 27.
    Kelly, R. T.; Page, J. S.; Marginean, I.; Tang, K. Q.; Smith, R. D. Nanoelectrospray Emitter Arrays Providing Interemitter Electric Field Uniformity. Anal. Chem. 2008, 80, 5660–5665.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2009

Authors and Affiliations

  • Ioan Marginean
    • 1
  • Ryan T. Kelly
    • 1
  • Ronald J. Moore
    • 1
  • David C. Prior
    • 1
  • Brian L. LaMarche
    • 1
  • Keqi Tang
    • 1
  • Richard D. Smith
    • 1
  1. 1.Pacific Northwest National LaboratoryBiological Sciences DivisionRichlandUSA

Personalised recommendations