Vibrational, calorimetric, and molecular conformational study on calcein interaction with model lipid membrane

  • Behnoush Maherani
  • Elmira Arab-Tehrany
  • Ewa Rogalska
  • Beata Korchowiec
  • Azadeh Kheirolomoom
  • Michel Linder
Research Paper


Nanoliposomes are commonly used as a carrier in controlled release drug delivery systems. Controlled release formulations can be used to reduce the amount of drug necessary to cause the same therapeutic effect in patients. One of the most noticeable factors in release profiles is the strength of the drug-carrier interaction. To adjust the pharmacokinetic and pharmacodynamic properties of therapeutic agents, it is necessary to optimize the drug-carrier interaction. To get a better understanding of this interaction, large unilamellar liposomes containing calcein were prepared using 1,2-dioleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, and 1,2-palmitoyl-sn-glycero-3-phosphocholine, and a mixture of them; calcein was chosen as a model polar molecule of biological interest. The thermodynamic changes induced by calcein and its location in lipid bilayers were determined by differential scanning calorimetry and Raman spectroscopy, respectively. The results reveal that calcein has no significant influence on thermotropic properties of the lipid membrane, but causing the abolition of pre-transition. The decreasing of the pre-transition can be ascribed to the presence of calcein near the hydrophilic cooperative zone of the bilayer. The change in intensity of the Raman peaks represents the interaction of calcein with choline head groups. Moreover, the impact of calcein on phosphoglyceride Langmuir layers spread at the air–water interface was studied using surface pressure-area and surface potential-area isotherms, as well as polarization-modulation infrared reflection–absorption spectroscopy and Brewster angle microscopy. The results obtained indicate that calcein introduce no major modification on the systems prepared with pure lipids.


Hydrophilic drugs Mechanical properties Compressibility modulus Surface pressure Thermodynamic changes 



Molecular area


Brewster angle microscopy


Compressibility modulus








Differential scanning calorimetry


Transition enthalpy


High sensitivity differential scanning calorimetry




Lactate dehydrogenase




Large unilamellar vesicles


Mixture design of experiments


Multilamellar vesicles


Poly dispersity index




Polarization modulation infrared reflection–absorption spectrometry


Transmission electron microscopy


Cooperativity of the bilayer


Temperature at which the transition is half completed


Phase transition temperature


Surface potential


Surface pressure



The authors would like to thank Bruno J. Beccard and Karine Gorin-Ninat for their excellent technical support in the joint service of Raman spectroscopy/Thermo Fisher Scientific-Paris. In particular, the authors also would like to thank Dr. Terry Wagner for her assistance in English corrections and her valuable suggestions for the improvement of article.


  1. Bae SJ, Kitamura S, Herbette LG, Sturtevant JM (1989) The effects of calcium channel blocking drugs on the thermotropic behavior of dimyristoylphosphatidylcholine. Chem Phys Lipids 51(1):1–7CrossRefGoogle Scholar
  2. Blume A, Huebner W, Messner G (1988) Fourier transform infrared spectroscopy of 13C=O-labeled phospholipids hydrogen bonding to carbonyl groups. Biochemistry 27:8239–8249CrossRefGoogle Scholar
  3. Bonora S, Di Foggia M, Iafisco M (2008) DSC and Raman study on the interaction of DDT [1,1,1-trichloro-2,2-bis(p-chlorophenyl)-ethane] with liposomal phospholipids. Pesticide Biochem Physiol 92(3):144–149CrossRefGoogle Scholar
  4. Bower DI, Maddams WF (1989) The vibrational spectroscopy of polymers. Cambridge University Press, New YorkCrossRefGoogle Scholar
  5. Calvagno MG, Celia C, Paolino D, Cosco D, Iannone M, Castelli F, Doldo P, Fresta M (2007) Effects of lipid composition and preparation conditions on physical-chemical properties, technological parameters and in vitro biological activity of gemcitabine-loaded nanoliposomes. Curr Drug Deliv 4(1):89–101CrossRefGoogle Scholar
  6. Colas JC, Shi W, Rao VSNM, Omri A, Mozafari MR, Singh H (2007) Microscopical investigations of nisin-loaded nanoliposome prepared by Mozafari method and their bacterial targeting. Micron 38(8):841–847CrossRefGoogle Scholar
  7. Cornell JA (2002) Experiments with mixtures. Wiley, New YorkCrossRefGoogle Scholar
  8. Corvis Y, Barzyk W, Brezesinski G, Mrabet N, Badis M, Hecht S, Rogalska E (2006a) Interactions of a fungistatic antibiotic, griseofulvin, with phospholipid monolayers used as models of biological membranes. Langmuir 22(18):7701–7711CrossRefGoogle Scholar
  9. Corvis Y, Brezesinski G, Rink R, Walcarius A, Van Der Heyden A, Mutelet F, Rogalska E (2006b) Analytical investigation of the interactions between SC3 hydrophobin and lipid layers: elaborating of nanostructured matrixes for immobilizing redox systems. Anal Chem 78(14):4850–4864CrossRefGoogle Scholar
  10. Csiszár A, Koglin E, Meier RJ, Klumpp E (2006) The phase transition behavior of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) model membrane influenced by 2,4-dichlorophenol—An FT-Raman spectroscopy study. Chem Phys Lipids 139(2):115–124CrossRefGoogle Scholar
  11. Czapla K, Korchowiec B, Orlof M, Magnieto JR, Rogalska E (2011) Enzymatic probing of model lipid membranes: phospholipase A2 activity toward monolayers modified by oxicam NSAIDs. J Phys Chem B 115(29):9290–9298CrossRefGoogle Scholar
  12. Davies JT, Rideal EK (1963) Interfacial phenomena. Academic Press, New YorkGoogle Scholar
  13. Dicko A, Bourque H, Pézolet M (1998) Study by infrared spectroscopy of the conformation of dipalmitoylphosphatidylglycerol monolayers at the air-water interface and transferred on solid substrates. Chem Phys Lipids 96(1–2):125–139CrossRefGoogle Scholar
  14. El Maghraby GMM, Williams AC, Barry BW (2005) Drug interaction and location in nanoliposomes: correlation with polar surface areas. Int J Pharm 292(1–2):179–185CrossRefGoogle Scholar
  15. Fan R, Gan L, Liu M, Zhu D, Chen L, Xu Z, Hao Z, Chen L (2011) An interaction of helicid with nanoliposome biomembrane. Appl Surf Sci 257:2102–2106CrossRefGoogle Scholar
  16. Gardikis K, Hatziantoniou S, Viras K, Demetzos C (2006a) Effect of a bioactive curcumin derivative on DPPC membrane: a DSC and Raman spectroscopy study. Thermochim Acta 447(1):1–4CrossRefGoogle Scholar
  17. Gardikis K, Hatziantoniou S, Viras K, Wagner M, Demetzos C (2006b) A DSC and Raman spectroscopy study on the effect of PAMAM dendrimer on DPPC model lipid membranes. Int J Pharm 318(1–2):118–123CrossRefGoogle Scholar
  18. Gregoriadis G (2007) Nanoliposome technology, nanoliposome preparation and related techniques. Informa Healthcare Inc., New YorkGoogle Scholar
  19. Hata T, Matsuki H, Kaneshina S (2000) Effect of local anesthetics on the bilayer membrane of dipalmitoylphosphatidylcholine: interdigitation of lipid bilayer and vesicle-micelle transition. Biophys Chem 87(1):25–36CrossRefGoogle Scholar
  20. Hernandez-Borrell J, Mas F, Puy J (1990) A theoretical approach to describe monolayer-nanoliposome lipid interaction. Biophys Chem 36(1):47–55CrossRefGoogle Scholar
  21. Huang CH, Levin IW (1983) Effect of lipid chain length inequivalence on the packing characteristics of bilayer assemblies. Raman spectroscopic study of phospholipid dispersions in the gel state. J Phys Chem 87(9):1509–1513CrossRefGoogle Scholar
  22. Khatri L, Taylor KMG, Craig DQM, Palin K (2006) High sensitivity differential scanning calorimetry investigation of the interaction between nanoliposomes, lactate dehydrogenase and tyrosinase. Int J Pharm 322(1–2):113–118CrossRefGoogle Scholar
  23. Leitch J, Kunze J, Goddard JD, Schwan AL, Faragher RJ, Naumann R, Knoll W, Dutcher JR, Lipkowski J (2009) In situ PM-IRRAS studies of an archaea analogue thiolipid assembled on a au(111) electrode surface. Langmuir 25(17):10354–10363CrossRefGoogle Scholar
  24. Leite VBP, Cavalli A, Oliveira ON Jr (1998) Hydrogen-bond control of structure and conductivity of Langmuir films. Phys Rev E 57(6):6835–6839CrossRefGoogle Scholar
  25. Levin IW, Keihn E, Harris WC (1985a) A Raman spectroscopic study on the effect of cholesterol on lipid packing in diether phosphatidylcholine bilayer dispersions. Biochim Biophys Acta 820(1):40–47CrossRefGoogle Scholar
  26. Levin IW, Thompson TE, Barenholz Y, Huang C (1985b) Two types of hydrocarbon chain interdigitation in sphingomyelin bilayers. Biochemistry 24(22):6282–6286CrossRefGoogle Scholar
  27. Lewis RNAH, McElhaney RN, Pohle W, Mantsch HH (1994) Components of the carbonyl stretching band in the infrared spectra of hydrated 1,2-diacylglycerolipid bilayers: a re-evaluation. Biophys J 67(6):2367–2375CrossRefGoogle Scholar
  28. Li XM, Zhao B, Zhao DQ, Ni JZ, Wu Y, Xu WQ (1996) Interaction of La3+ and cholesterol with dipalmitoylphosphatidylglycerol bilayers by FT-Raman spectroscopy. Thin Solid Films 284–285:762–764CrossRefGoogle Scholar
  29. MacDonald RC, MacDonald RI, Menco BPM, Takeshita K, Subbarao NK, Hu L (1991) Small-volume extrusion apparatus for preparation of large unilammelar vesicles. Biochim Biophys Acta 1061:297–303CrossRefGoogle Scholar
  30. MacPhail RA, Strauss HL, Snyder RG, Eiliger CA (1984) C–H stretching modes and the structure of n-alkyl chains. 2. Long, all-trans chains. J Phys Chem 88(3):334–341CrossRefGoogle Scholar
  31. Maherani B, Arab-Tehrany E, Linder M (2011) Mechanism of bioactive transfer through liposomal bilayers. Curr Drug Targets 12(4):531–545CrossRefGoogle Scholar
  32. Maherani B, Arab-Terhrani E, Kheirolomoom A, Stebe MJ, Linder M (2012) Optimization and characterization of nanoliposome formulation by mixture design. Analyst 137(3):773–786CrossRefGoogle Scholar
  33. Mason JT, Huang CH (1981) Chain length dependent thermodynamics of saturated symmetric-chain phosphatidylcholine bilayers. Lipids 16(8):604–608CrossRefGoogle Scholar
  34. Mason JT, Huang C, Biltonen RL (1981) Calorimetric investigations of saturated mixed-chain phosphatidylcholine bilayer dispersions. Biochemistry 20(21):6086–6092CrossRefGoogle Scholar
  35. Matsui H, Pan S (2001) Distribution of DNA in cationic nanoliposome complexes probed by Raman microscopy. Langmuir 17(3):571–573CrossRefGoogle Scholar
  36. Matti V, Säily J, Ryhänen SJ, Holopainen JM, Borocci S, Mancini G, Kinnunen PKJ (2001) Characterization of mixed monolayers of phosphatidylcholine and a dicationic gemini surfactant SS-1 with a langmuir balance: effects of DNA. Biophys J 81(4):2135–2143CrossRefGoogle Scholar
  37. Mozafari MR (2007) Nanomaterials and nanosystems for biomedical applications. Springer, DordrechtCrossRefGoogle Scholar
  38. Mozafari MR (2010) Nanoliposome: preparation and analysis. In: Weissig V (ed) Nanoliposomes: methods and protocols pharmaceutical nanocarriers, vol 1 methods in molecular biology, vol 605. Humana Press, GlendaleGoogle Scholar
  39. Mozafari MR, Mortazavi MS (2005) Nanoliposome: from fundamentals to recent developments. Trafford Publishing Ltd, OxfordGoogle Scholar
  40. Painter PC, Coleman MM, Koenig JL (1982) The theory of vibrational spectroscopy and its application to polymeric materials. Wiley, New YorkGoogle Scholar
  41. Pétriat F, Roux E, Leroux JC, Giasson S (2004) Study of molecular interactions between a phospholipidic layer and a pH-sensitive polymer using the langmuir balance technique. Langmuir 20(4):1393–1400CrossRefGoogle Scholar
  42. Shimanouchi T, Ishii H, Yoshimoto N, Umakoshi H, Kuboi R (2009) Calcein permeation across phosphatidylcholine bilayer membrane: effects of membrane fluidity, nanoliposome size, and immobilization. Coll Surf B Biointerf 73(1):156–160CrossRefGoogle Scholar
  43. Smith E, Dent G (2005) Modern Raman spectroscopy: a practical approach. Wiley, ChesterGoogle Scholar
  44. Snyder RG, Strauss HL, Elliger CA (1982) C–H stretching modes and the structure of n-alkyl chains. 1. Long, disordered chains. J Phys Chem 86(26):5145–5150CrossRefGoogle Scholar
  45. Taylor KMG, Morris RM (1995) Thermal analysis of phase transition behaviour in nanoliposomes. Thermochim Acta 248(C):289–301CrossRefGoogle Scholar
  46. Vankann M, Möllerfeld J, Ringsdorf H, Höcker H (1996) Amphiphilic model peptides: circular dichroism measurements and investigations by a langmuir balance. J Colloid Interface Sci 178(1):241–250CrossRefGoogle Scholar
  47. Zhao L, Feng SS, Go ML (2004) Investigation of molecular interactions between paclitaxel and DPPC by langmuir film balance and differential scanning calorimetry. J Pharm Sci 93(1):86–98CrossRefGoogle Scholar
  48. Zhao LY, Feng SS, Kocherginsky N, Kostetski I (2007) DSC and EPR investigations on effects of cholesterol component on molecular interactions between paclitaxel and phospholipid within lipid bilayer membrane. Int J Pharm 338(1–2):258–266CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Behnoush Maherani
    • 1
  • Elmira Arab-Tehrany
    • 1
  • Ewa Rogalska
    • 2
  • Beata Korchowiec
    • 3
  • Azadeh Kheirolomoom
    • 4
  • Michel Linder
    • 1
  1. 1.Laboratoire d’Ingénierie des Biomolécules (LIBio)Université de LorraineVandoeuvre lès NancyFrance
  2. 2.Faculté des SciencesGEVSM, UMR, SRSMC UMR 7565, CNRS/Université de LorraineVandoeuvre lès NancyFrance
  3. 3.Department of Physical Chemistry and ElectrochemistryFaculty of Chemistry, Jagiellonian UniversityCracowPoland
  4. 4.Department of Biomedical EngineeringUniversity of CaliforniaDavisUSA

Personalised recommendations