Applied Magnetic Resonance

, Volume 46, Issue 8, pp 885–895 | Cite as

Spin-Labeled Small Unilamellar Vesicles with the T 1-Sensitive Saturation-Recovery EPR Display as an Oxygen-Sensitive Analyte for Measurement of Cellular Respiration

  • Laxman Mainali
  • Jeannette Vasquez-Vivar
  • James S. Hyde
  • Witold K. SubczynskiEmail author


This study validated the use of small unilamellar vesicles (SUVs) made of 1-palmitoyl-2-oleoylphosphatidylcholine with 1 mol % spin label of 1-palmitoyl-2-(16-doxylstearoyl)phosphatidylcholine (16-PC) as an oxygen-sensitive analyte to study cellular respiration. In the analyte, the hydrocarbon environment surrounds the nitroxide moiety of 16-PC. This ensures high oxygen concentration and oxygen diffusion at the location of the nitroxide as well as isolation of the nitroxide moiety from cellular reductants and paramagnetic ions that might interfere with spin-label oximetry measurements. The saturation-recovery electron paramagnetic resonance approach was applied in the analysis since this approach is the most direct method to carry out oximetric studies. It was shown that this display (spin–lattice relaxation rate) is linear in oxygen partial pressure up to 100 % air (159 mmHg). Experiments using a neuronal cell line in suspension were carried out at X-band for closed-chamber geometry. Oxygen consumption rates showed a linear dependence on the number of cells. Other significant benefits of the analyte are: the fast effective rotational diffusion and slow translational diffusion of the spin-probe is favorable for the measurements, and there is no cross-reactivity between oxygen and paramagnetic ions in the lipid bilayer.


Electron Paramagnetic Resonance Electron Paramagnetic Resonance Spectrum Nitroxide Electron Paramagnetic Resonance Signal Spin Label 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



This work was supported by Grants EY015526, EB002052, EB001980, and NS081936 from the National Institutes of Health.


  1. 1.
    J.M. Backer, V.G. Budker, S.I. Eremenko, Y.N. Molin, Biochim. Biophys. Acta 460, 152–156 (1977)CrossRefGoogle Scholar
  2. 2.
    C.-S. Lai, L.E. Hopwood, J.S. Hyde, S. Lukiewicz, Proc. Natl. Acad. Sci. USA 79, 1166–1170 (1982)ADSCrossRefGoogle Scholar
  3. 3.
    A. Ligeza, A. Wisniewska, W.K. Subczynski, Curr. Topics in Biophys. 16, 92–98 (1992)Google Scholar
  4. 4.
    A. Kusumi, W.K. Subczynski, J.S. Hyde, Proc. Natl. Acad. Sci. USA 79, 1854–1858 (1982)ADSCrossRefGoogle Scholar
  5. 5.
    J.-J. Yin, J.S. Hyde, Z. Phys. Chem. 153, 57–65 (1987)CrossRefGoogle Scholar
  6. 6.
    W.K. Subczynski, J.S. Hyde, A. Kusumi, Proc. Natl. Acad. Sci. USA 86, 4474–4478 (1989)ADSCrossRefGoogle Scholar
  7. 7.
    W.K. Subczynski, J.S. Hyde, Biochim. Biophys. Acta 643, 283–291 (1981)CrossRefGoogle Scholar
  8. 8.
    C. Altenbach, W. Froncisz, J.S. Hyde, W.L. Hubbell, Biophys. J. 56, 1183–1191 (1989)CrossRefGoogle Scholar
  9. 9.
    W. Froncisz, C.-S. Lai, J.S. Hyde, Proc. Natl. Acad. Sci. USA 82, 411–415 (1985)ADSCrossRefGoogle Scholar
  10. 10.
    H.S. Mchaourab, J.S. Hyde, J. Magn. Reson. 101, 178–184 (1993)CrossRefzbMATHGoogle Scholar
  11. 11.
    H.S. Mchaourab, J.S. Hyde, J.B. Feix, Biochemistry 33, 6691–6699 (1994)CrossRefzbMATHGoogle Scholar
  12. 12.
    J.S. Hyde, W.K. Subczynski, J. Magn. Reson. 56, (1984)Google Scholar
  13. 13.
    W.K. Subczynski, J.S. Hyde, Biophys. J. 45, 743–748 (1984)CrossRefGoogle Scholar
  14. 14.
    J.S. Hyde, W.K. Subczynski, in Biological Magnetic Resonance, vol. 8, ed. by L.J. Berliner, J. Reuben (Plenum Press, New York, 1989), pp. 399–425Google Scholar
  15. 15.
    D.A. Windrem, W.Z. Plachy, Biochim. Biophys. Acta 600, 655–665 (1980)CrossRefGoogle Scholar
  16. 16.
    C.E. St Denis, C.J. Fell, Can. J. Chem. Eng. 49, 885 (1971)CrossRefGoogle Scholar
  17. 17.
    Y.N. Molin, K.M. Salikhov, K.I. Zamaraev, Spin Exchange (Springer-Verlag, New York, 1980), pp. 111–115CrossRefGoogle Scholar
  18. 18.
    M.-K. Ahn, J. Magn. Reson. 22, 289–293 (1976)ADSGoogle Scholar
  19. 19.
    W.K. Subczynski, L.E. Hopwood, J.S. Hyde, J. Gen. Physiol. 100, 69–87 (1992)CrossRefGoogle Scholar
  20. 20.
    W.F. Linke, Solubilities. Inorganic and Metal Organic Compounds II, 4th edn. (American Chemical Society, Washington, DC, 1965), pp. 1233–1236Google Scholar
  21. 21.
    L. Mainali, M. Raguz, W.K. Subczynski, J. Phys. Chem. B 117, 8994–9003 (2013)CrossRefzbMATHGoogle Scholar
  22. 22.
    J.-J. Yin, W.K. Subczynski, Biophys. J. 71, 832–839 (1996)CrossRefGoogle Scholar
  23. 23.
    W.K. Subczynski, J. Widomska, A. Wisniewska, A. Kusumi, Methods in Molecular Biology, Lipid Rafts, vol. 398 (Humana Press, Totowa, 2007), pp. 143–157CrossRefGoogle Scholar
  24. 24.
    M.L. Hitchman, Measurement of Dissolved Oxygen (John Wiley & Sons Inc, New York, 1978)Google Scholar
  25. 25.
    H.M. Swartz, K. Chen, M. Pals, M. Sentjurc, P.D. Morse 2nd, Magn. Reson. Med. 3, 169–174 (1986)CrossRefGoogle Scholar
  26. 26.
    W.K. Subczynski, H.M. Swartz, in Biomedical ESR Part A: Free Radicals, Metals, Medicine, and Physiology, Biological Magnetic Resonance, vol. 23, ed. by S.S. Eaton, G.R. Eaton, L.J. Berliner (Kluwer, Boston, 2005), pp. 229–282CrossRefGoogle Scholar
  27. 27.
    M. Raguz, L. Mainali, J. Widomska, W.K. Subczynski, Chem. Phys. Lipids 164, 819–829 (2011)CrossRefGoogle Scholar
  28. 28.
    H. Träuble, H. Eibl, Proc. Natl. Acad. Sci. USA 71, 214–219 (1974)ADSCrossRefGoogle Scholar
  29. 29.
    D. Papahadjopoulos, Biochim. Biophys. Acta 163, 240–254 (1968)CrossRefGoogle Scholar
  30. 30.
    M. Egret-Charlier, A. Sanson, M. Ptak, FEBS Lett. 89, 313–316 (1978)CrossRefGoogle Scholar
  31. 31.
    A. Sanson, M. Ptak, J.L. Rigaud, C.M. Gary-Bobo, Chem. Phys. Lipids 17, 435–444 (1976)CrossRefzbMATHGoogle Scholar
  32. 32.
    A. Kusumi, W.K. Subczynski, J.S. Hyde, Fed. Proc. 41, 1394 (1982)Google Scholar
  33. 33.
    A. Ligeza, H. Swartz, W.K. Subczynski, Curr. Topics in Biophys. 18, 29–38 (1994)Google Scholar
  34. 34.
    K.J. Liu, M.W. Grinstaff, J. Jiang, K.S. Suslick, H.M. Swartz, W. Wang, Biophys. J. 67, 896–901 (1994)CrossRefGoogle Scholar
  35. 35.
    W. Froncisz, T. Oles, J.S. Hyde, Rev. Sci. Instrum. 57, 1095–1099 (1986)ADSCrossRefGoogle Scholar
  36. 36.
    J.W. Sidabras, R.R. Mett, W. Froncisz, T.G. Camenisch, J.R. Anderson, J.S. Hyde, Rev. Sci. Instrum. 78, 034701 (2007)ADSCrossRefGoogle Scholar
  37. 37.
    J.S. Hyde, J.-J. Yin, W.K. Subczynski, T.G. Camenisch, J.J. Ratke, W. Froncisz, J. Phys. Chem. B 108, 9524–9529 (2004)CrossRefGoogle Scholar
  38. 38.
    W.K. Subczynski, L. Mainali, T.G. Camenisch, W. Froncisz, J.S. Hyde, J. Magn. Reson. 209, 142–148 (2011)ADSCrossRefGoogle Scholar
  39. 39.
    L. Mainali, J.W. Sidabras, T.G. Camenisch, J.J. Ratke, M. Raguz, J.S. Hyde, W.K. Subczynski, Appl. Magn. Reson. 45, 1343–1358 (2014)CrossRefzbMATHGoogle Scholar
  40. 40.
    L. Mainali, M. Raguz, T.G. Camenisch, J.S. Hyde, W.K. Subczynski, J. Magn. Reson. 212, 86–94 (2011)ADSCrossRefGoogle Scholar
  41. 41.
    L. Mainali, J.S. Hyde, W.K. Subczynski, J. Magn. Reson. 226, 35–44 (2013)ADSCrossRefGoogle Scholar
  42. 42.
    W. Froncisz, T.G. Camenisch, J.J. Ratke, J.R. Anderson, W.K. Subczynski, R.A. Strangeway, J.W. Sidabras, J.S. Hyde, J. Magn. Reson. 193, 297–304 (2008)ADSCrossRefGoogle Scholar
  43. 43.
    J.-J. Yin, J.S. Hyde, J. Chem. Phys. 91, 6029–6035 (1989)ADSCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Wien 2015

Authors and Affiliations

  • Laxman Mainali
    • 1
  • Jeannette Vasquez-Vivar
    • 1
  • James S. Hyde
    • 1
  • Witold K. Subczynski
    • 1
    Email author
  1. 1.Department of BiophysicsMedical College of WisconsinMilwaukeeUSA

Personalised recommendations