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Spectroscopic studies of D-α-tocopherol concentration-induced transformation in egg phosphatidylcholne vesicles

Cellular & Molecular Biology Letters volume 12, pages 51–69 (2007)Cite this article

Abstract

The effects of embedding up to 60 mol% of α-tocopherol (α-Toc) on the morphology and structure of the egg phosphatidylcholine (PC) membrane were studied using spectroscopic techniques. The resulting vesicles were subjected to turbidometric and dynamic light scattering measurements to evaluate their size distribution. The α-Toc intrinsic fluorescence and its quenching was used to estimate the tocopherol position in the membrane. Optical microscopy was used to visualize morphological changes in the vesicles during the inclusion of tocopherol into the 2 mg/ml PC membrane. The incorporation of up to 15 mol% of tocopherol molecules into PC vesicles is accompanied by a linear increase in the fluorescence intensity and the simultaneous formation of larger, multilamellar vesicles. Increasing the tocopherol concentration above 20 mol% induced structural and morphological changes leading to the disappearance of micrometer-sized vesicles and the formation of small unilamellar vesicles of size ranging from 30 to 120 nm, mixed micelles and non-lamellar structures.

Abbreviations

α-Toc:

D-α-tocopherol

ACR:

acrylamide

DLS:

dynamic light scattering

DMPC:

dimyristoyl phosphatidylcholine

DPPC:

dipalmitoyl phosphatidylcholine

GUV:

giant unilamellar vesicle

GMV:

giant multilamellar vesicle

GLV:

giant lamellar vesicle

KI:

potassium iodide

KD :

dynamic fluorescence quenching constant

KSV :

Stern-Volmer quenching constant

LMV:

large multilamellar vesicle

NMR:

nuclear magnetic resonance

PC:

L-α-phosphatydylcholine

SSBV:

small single-bilayer vesicle

SUV:

small unilamellar vesicle

V:

static fluorescence quenching constant

References

  1. Ortiz, A., Aranda, F.J. and Gomez-Fernandez, J.C. A differential scanning calorimetry study of the interaction of alpha-tocopherol with mixtures of phospholipids. Biochim. Biophys. Acta 898 (1987) 214–222.

    PubMed  Article  CAS  Google Scholar 

  2. Ortiz, A., Villalain, J. and Gomez-Fernandez, J.C. Interaction of diacylglycerols with phosphatidylcholine vesicles as studied by differential scanning calorimetry and fluorescence probe depolarization. Biochemistry 27 (1988) 9030–9036.

    PubMed  Article  CAS  Google Scholar 

  3. Walke, M., Beckert, D. and Lasch, J. Interaction of UV light-induced alphatocopherol radicals with lipids detected by an electron spin resonance prooxidation effect. Photochem. Photobiol. 68 (1998) 502–510.

    PubMed  Article  CAS  Google Scholar 

  4. Aranda, F.J., Sanchez-Migallon, M.P. and Gomez-Fernandez, J.C. Influence of alpha-tocopherol incorporation on Ca(2+)-induced fusion of phosphatidylserine vesicles. Arch. Biochem. Biophys. 333 (1996) 394–400.

    PubMed  Article  CAS  Google Scholar 

  5. Grau, A. and Ortiz, A. Dissimilar protection of tocopherol isomers against membrane hydrolysis by phospholipase A2. Chem. Phys. Lipids 91 (1998) 109–118.

    PubMed  Article  CAS  Google Scholar 

  6. Gutierrez, M.E., Garcia, A.F., Africa de Madariaga, M., Sagrista, M.L., Casado, F.J. and Mora, M. Interaction of tocopherols and phenolic compounds with membrane lipid components: evaluation of their antioxidant activity in a liposomal model system. Life Sci. 72 (2003) 2337–2360.

    PubMed  Article  CAS  Google Scholar 

  7. Chen, C.S., Patterson, M.C., Wheatley, C.L., O’Brien, J.F. and Pagano, R.E. Broad screening test for sphingolipid-storage diseases. Lancet 354 (1999) 901–9051.

    PubMed  Article  CAS  Google Scholar 

  8. Nacka, F., Cansell, M., Meleard, P. and Combe, N. Incorporation of alphatocopherol in marine lipid-based liposomes: in vitro and in vivo studies. Lipids 36 (2001) 1313–1320.

    PubMed  Article  CAS  Google Scholar 

  9. Stillwell, W., Ehringer, W. and Wassall, S.R. Interaction of alpha-tocopherol with fatty acids in membranes and ethanol. Biochim. Biophys. Acta 1105 (1992) 237–244.

    PubMed  Article  CAS  Google Scholar 

  10. Chen, H.W., Chiang, T., Wang, C.Y. and Lii, C.K. Inhibition of tert-butyl hydroperoxide-induced cell membrane bleb formation by alpha-tocopherol and glutathione. Food Chem. Toxicol. 38 (2000) 1089–1096.

    PubMed  Article  CAS  Google Scholar 

  11. Sezer, A.D., Bas, A.L. and Akbuga, J. Encapsulation of enrofloxacin in liposomes I: preparation and in vitro characterization of LUV. J. Liposome Res. 14 (2004) 77–86.

    PubMed  Article  CAS  Google Scholar 

  12. Quinn, P.J. Characterisation of clusters of alpha-tocopherol in gel and fluid phases of dipalmitoylglycerophosphocholine. Eur. J. Biochem. 233 (1995) 916–925.

    PubMed  Article  CAS  Google Scholar 

  13. Wang, X., Semmler, K., Richter, W. and Quinn, P.J. Ripple phases induced by alpha-tocopherol in saturated diacylphosphatidylcholines. Arch. Biochem. Biophys. 377 (2000) 304–314.

    PubMed  Article  CAS  Google Scholar 

  14. Massey, J.B. Interfacial properties of phosphatidylcholine bilayers containing vitamin E derivatives. Chem. Phys. Lipids 109 (2001) 157–174.

    PubMed  Article  CAS  Google Scholar 

  15. Naumowicz, M. and Figaszewski, Z.A. Impedance analysis of phosphatidylcholine/alpha-tocopherol system in bilayer lipid membranes. J. Membr. Biol. 205 (2005) 29–36.

    PubMed  Article  CAS  Google Scholar 

  16. Kaasgaard, T., Mouritsen, O.G. and Jorgensen, K. Lipid domain formation and ligand-receptor distribution in lipid bilayer membranes investigated by atomic force microscopy. FEBS Lett. 515 (2002) 29–34.

    PubMed  Article  CAS  Google Scholar 

  17. Leidy, C., Mouritsen, O.G., Jorgensen, K. and Peters, G.H. Evolution of a rippled membrane during phospholipase A2 hydrolysis studied by timeresolved AFM. Biophys. J. 87 (2004) 408–418.

    PubMed  Article  CAS  Google Scholar 

  18. Pedersen, T.B., Kaasgaard, T., Jensen, M.O., Frokjaer, S., Mouritsen, O.G. and Jorgensen, K. Phase behavior and nanoscale structure of phospholipid membranes incorporated with acylated C14-peptides. Biophys. J. 89 (2005) 2494–2503.

    PubMed  Article  CAS  Google Scholar 

  19. Menger, F.M. and Keiper, J.S. Chemistry and physics of giant vesicles as biomembrane models. Curr. Opin. Chem. Biol. 2 (1998) 726–732.

    PubMed  Article  CAS  Google Scholar 

  20. Asai, Y. Structural differences in aqueous dispersions of alpha-tocopheryl acetate and phosphatidylcholine upon varying their molar fractions. Pharmazie 59 (2004) 849–853.

    PubMed  CAS  Google Scholar 

  21. Gramlich, G., Zhang, J. and Nau, W.M. Diffusion of alpha-tocopherol in membrane models: probing the kinetics of vitamin E antioxidant action by fluorescence in real time. J. Am. Chem. Soc. 126 (2004) 5482–5492.

    PubMed  Article  CAS  Google Scholar 

  22. Pandey, B.N. and Mishra, K.P. Effect of radiation induced lipid peroxidation on diphenylhexatriene fluorescence in egg phospholipid liposomal membrane. J. Biochem. Mol. Biol. Biophys. 6 (2002) 267–272.

    PubMed  CAS  Google Scholar 

  23. Naguib, Y.M. A fluorometric method for measurement of peroxyl radical scavenging activities of lipophilic antioxidants. Anal. Biochem. 265 (1998) 290–298.

    PubMed  Article  CAS  Google Scholar 

  24. Di Giulio, A., Saletti, A., Oratore, A. and Bozzi, A. Monitoring by cisparinaric fluorescence of free radical induced lipid peroxidation in aqueous liposome suspensions. J. Microencapsul. 13 (1996) 435–445.

    PubMed  Google Scholar 

  25. LaLonde, R.T. and Xie, S. Glutathione and N-acetylcysteine inactivations of mutagenic 2(5H)-furanones from the chlorination of humics in water. Chem. Res. Toxicol. 6 (1993) 445–451.

    PubMed  Article  CAS  Google Scholar 

  26. Benezra, C., Sigman, C.C., Perry, L.R., Helmes, C.T. and Maibach, H.I. A systematic search for structure-activity relationships of skin contact sensitizers: methodology. J. Invest. Dermatol. 85 (1985) 351–356.

    PubMed  Article  CAS  Google Scholar 

  27. Kuksis, A. Animal lecithins. J. Am. Oil Chem. Soc. USA 12 (1985) 105–162.

    CAS  Google Scholar 

  28. Gennis, R.B. Biomembranes: molecular structure and function in Membranes. Springer-Verlag: New York, 1989.

    Google Scholar 

  29. Berne, R. and Pecora, R. Dynamic Light Scattering. New York: Wiley 1976.

    Google Scholar 

  30. Eastman, S.J., Hope, M.J. and Cullis, P.R. Transbilayer transport of phosphatidic acid in response to transmembrane pH gradients. Biochemistry 30 (1991) 1740–1745.

    PubMed  Article  CAS  Google Scholar 

  31. Moro, F., Goni, F.M. and Urbaneja, M.A. Fluorescence quenching at interfaces and permaetion of acrylamide and iodide across phospholipid bilayers. FEBS 330 (1993) 129–132.

    Article  CAS  Google Scholar 

  32. Eftink, M.R. and Ghiron, C.A. Fluorescence quenching of indole and model micelle systems J. Phys. Chem. 80 (1976) 486–492.

    Article  CAS  Google Scholar 

  33. Kalinin, S.V. and Molotkovsky, J.G. Anion binding to lipid bilayers: determination using fluorescent membrane probe by direct quenching or by competitive displacement approaches. J. Biochem. Biophys. Methods 46 (2000) 39–51.

    PubMed  Article  CAS  Google Scholar 

  34. Zheng, B., Wu, J.N., Schober, W., Lewis, D.E. and Vida, T. Isolation of yeast mutants defective for localization of vacuolar vital dyes. Proc. Natl. Acad. Sci. USA 95 (1998) 11721–11726.

    PubMed  Article  CAS  Google Scholar 

  35. Fukuzawa, K., Ikebata, W. and Sohmi, K. Location, antioxidant and recycling dynamics of alpha-tocopherol in liposome membranes. J. Nutr. Sci. Vitaminol. (Tokyo) 39 Suppl (1993) S9–22.

    CAS  Google Scholar 

  36. Ferezou, J., Nguyen, T.L., Leray, C., Hajri, T., Frey, A., Cabaret, Y., Courtieu, J., Lutton, C. and Bach, A.C. Lipid composition and structure of commercial parenteral emulsions. Biochim. Biophys. Acta 1213 (1994) 149–158.

    PubMed  CAS  Google Scholar 

  37. Chungcharoenwattana, S. and Ueno, M. Size control of mixed egg yolk phosphatidylcholine (EggPC)/oleate vesicles. Chem. Pharm. Bull. (Tokyo) 52 (2004) 1058–1062.

    Article  CAS  Google Scholar 

  38. Wang, X. and Quinn, P.J. Preferential interaction of alpha-tocopherol with phosphatidylcholines in mixed aqueous dispersions of phosphatidylcholine and phosphatidylethanolamine. Eur. J. Biochem. 267 (2000) 6362–6368.

    PubMed  Article  CAS  Google Scholar 

  39. Wang, X. and Quinn, P.J. The interaction of alpha-tocopherol with bilayers of 1-palmitoyl-2-oleoyl-phosphatidylcholine. Biochim. Biophys. Acta 1567 (2002) 6–12.

    PubMed  Article  CAS  Google Scholar 

  40. Nakajima, K., Utsumi, H., Kazama, M. and Hamada, A. Alpha-tocopherol-induced hexagonal HII phase formation in egg yolk phosphatidylcholine membranes. Chem. Pharm. Bull. (Tokyo) 38 (1990) 1–4.

    CAS  Google Scholar 

  41. Chudinova, V.V., Zakharova, E.I., Alekseev, S.M., Chupin, V.V. and Evstigneeva, R.P. [Study of the interaction of alpha-tocopherol with phospholipids, fatty acids, and their oxygenated derivatives by (31)P-NMR spectroscopy]. Bioorg. Khim. 19 (1993) 243–249.

    PubMed  CAS  Google Scholar 

  42. Bagatolli, L.A. and Gratton, E. Two photon fluorescence microscopy of coexisting lipid domains in giant unilamellar vesicles of binary phospholipid mixtures. Biophys. J. 78 (2000) 290–305.

    PubMed  CAS  Article  Google Scholar 

  43. Erin, A.N., Spirin, M.M., Tabidze, L.V. and Kagan, V.E. [Formation of alpha-tocopherol complexes with fatty acids. Possible mechanism of biomembrane stabilization by vitamin E]. Biokhimiia 48 (1983) 1855–1861.

    PubMed  CAS  Google Scholar 

  44. Erin, A.N., Spirin, M.M., Tabidze, L.V. and Kagan, V.E. Formation of alpha-tocopherol complexes with fatty acids. A hypothetical mechanism of stabilization of biomembranes by vitamin E. Biochim. Biophys. Acta 774 (1984) 96–102.

    PubMed  Article  CAS  Google Scholar 

  45. Kagan, V.E. and Quinn, P.J. The interaction of alpha-tocopherol and homologues with shorter hydrocarbon chains with phospholipid bilayer dispersions. A fluorescence probe study. Eur. J. Biochem. 171 (1988) 661–667.

    PubMed  Article  CAS  Google Scholar 

  46. Urano, S., Shichita, N. and Matsuo, M. Interaction of vitamin E and its model compounds with unsaturated fatty acids in homogeneous solution. J. Nutr. Sci. Vitaminol. (Tokyo) 34 (1988) 189–194.

    CAS  Google Scholar 

  47. Chudinova, V.V., Zakharova, E.I., Alekseev, S.M. and Evstigneeva, R.P. [Interaction of vitamin E (alpha-tocopherol) with oxygenated fatty acid derivatives]. Bioorg. Khim. 19 (1993) 505–511.

    PubMed  CAS  Google Scholar 

  48. Salgado, J., Villalain, J. and Gomez-Fernandez, J.C. Alpha-tocopherol interacts with natural micelle-forming single-chain phospholipids stabilizing the bilayer phase. Arch. Biochem. Biophys. 306 (1993) 368–376.

    PubMed  Article  CAS  Google Scholar 

  49. Bourgeois, C.F., George, P.R. and Cronenberger, L.A. Automated determination of alpha-tocopherol in food and feed. Part 2. Continuous flow technique. J. Assoc. Anal. Chem. 67 (1984) 631–634.

    CAS  Google Scholar 

  50. Tiurin, V.A., Kagan, V.E., Serbinova, E.A., Gorbunov, N.V. and Erin, A.N. [The interaction of alpha-tocopherol with phospholipid liposomes: the absence of transbilayer mobility]. Biull. Eksp. Biol. Med. 102 (1986) 689–692.

    PubMed  CAS  Google Scholar 

  51. Urano, S., Iida, M., Otani, I. and Matsuo, M. Membrane stabilization of vitamin E; interactions of alpha-tocopherol with phospholipids in bilayer liposomes. Biochem. Biophys. Res. Commun. 146 (1987) 1413–1418.

    PubMed  Article  CAS  Google Scholar 

  52. Urano, S., Yano, K. and Matsuo, M. Membrane-stabilizing effect of vitamin E: effect of alpha-tocopherol and its model compounds on fluidity of lecithin liposomes. Biochem. Biophys. Res. Commun. 150 (1988) 469–475.

    PubMed  Article  CAS  Google Scholar 

  53. Wassall, S.R., Phelps, T.M., Wang, L.J., Langsford, C.A. and Stillwell, W. Membrane stabilization by vitamin E: magnetic resonance studies of the interaction of alpha-tocopherol with fatty acid acyl chains in phospholipid model membranes. Prog. Clin. Biol. Res. 292 (1989) 435–444.

    PubMed  CAS  Google Scholar 

  54. Wang, X. and Quinn, P.J. Vitamin E and its function in membranes. Prog. Lipid. Res. 38 (1999) 309–336.

    PubMed  Article  CAS  Google Scholar 

  55. Srivastava, S., Phadke, R.S. and Govil, G. Effect of incorporation of drugs, vitamins and peptides on the structure and dynamics of lipid assemblies. Mol. Cell. Biochem. 91 (1989) 99–109.

    PubMed  Article  CAS  Google Scholar 

  56. Bellemare, F. and Fragata, M. Transmembrane distribution of alphatocopherol in single-lamellar mixed lipid vesicles. J. Membr. Biol. 58 (1981) 67–74.

    PubMed  Article  CAS  Google Scholar 

  57. Kagan, V.E., Bakalova, R.A., Serbinova, E.E. and Stoytchev, T.S. Fluorescence measurements of incorporation and hydrolysis of tocopherol and tocopheryl esters in biomembranes. Methods Enzymol. 186 (1990) 355–367.

    PubMed  CAS  Article  Google Scholar 

  58. Bally, M.B., Mayer, L.D., Loughrey, H., Redelmeier, T., Madden, T.D., Wong, K., Harrigan, P.R., Hope, M.J. and Cullis, P.R. Dopamineaccumulation in large unilamellar vesicle systems induced by transmembrane ion gradients. Chem. Phys. Lipids 47 (1988) 97–107.

    PubMed  Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Physics, ul. Wojska Polskiego 38/42, 60-637, Poznań, Poland

    Krzysztof Dwiecki, Paweł Górnas & Krzysztof Polewski

  2. Department of Biochemistry and Food Analysis, ul. Mazowiecka 48, 60-623, Poznań, Poland

    Małgorzata Nogala-Kałucka

  3. August Cieszkowski Agricultural University, Poznań, Poland

    Małgorzata Nogala-Kałucka

  4. Institute of Physics, A. Mickiewicz University, 61-614, Poznań, Poland

    Agnieszka Wilk

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  1. Krzysztof Dwiecki
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Dwiecki, K., Górnas, P., Wilk, A. et al. Spectroscopic studies of D-α-tocopherol concentration-induced transformation in egg phosphatidylcholne vesicles. Cell Mol Biol Lett 12, 51–69 (2007). https://doi.org/10.2478/s11658-006-0059-6

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  • DOI: https://doi.org/10.2478/s11658-006-0059-6

Key words

  • Egg phosphatidylcholine
  • Tocopherol
  • Fluorescence
  • Vesicles
  • Optical microscopy
  • Dynamic light scattering

Cellular & Molecular Biology Letters

ISSN: 1689-1392

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