Journal of Non-Crystalline Solids 356 (2010) 747–753

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Journal of Non-Crystalline Solids

journal homepage: www.elsevier .com/ locate/ jnoncrysol

Structural and spectroscopic studies of DMPC/cationic surfactant system

Maciej Kozak a,*, Aleksandra Wypych a, Kamil Szpotkowski a, Stefan Jurga a, Andrzej Skrzypczak b

a Department of Macromolecular Physics, Faculty of Physics, A. Mickiewicz University, ul. Umultowska 85, 61-614 Poznan, Polandb Faculty of Chemical Technology, Poznan University of Technology, Pl. M.Skłodowskiej-Curie 2, Poznan, Poland

a r t i c l e i n f o a b s t r a c t

Article history:Available online 28 January 2010

Keywords:Biological systemPhospholipidsBicellar phaseNanostructureSynchrotron radiationDielectric spectroscopy

0022-3093/$ - see front matter � 2010 Elsevier B.V. Adoi:10.1016/j.jnoncrysol.2009.07.043

* Corresponding author. Tel.: +48 61 8295266; fax:E-mail address: mkozak@amu.edu.pl (M. Kozak).

Aqueous suspension of a mixture of 1,2-dimyrilstoyl-sn-glycero-3-phosphocholine (DMPC) and (dodecyl-oxymethyl)dodecydimethylammonium (DDMDDAC) has been investigated by Fourier transform infraredspectroscopy (FTIR), small angle scattering of synchrotron radiation (SAXS) and dielectric spectroscopy(DS). The introduction of surfactant to DMPC water solution probably induces formation of discoidal(bicellar) phase, as revealed by SAXS studies. The presence of the surfactant has been found to decreasethe main phase transition temperature, i.e., from rippled gel to liquid crystalline phase, as indicated bythe DS and FTIR results. The small steps in the real part of dielectric permittivity plot vs. temperaturecan be related to the: planar gel to rippled gel (Lb0 ? Pb0 ) and rippled gel–liquid crystal (Pb0 ? La) phasetransitions, which is supported by the temperature dependencies of the symmetric and anti-symmetricCH2 band frequencies from FTIR measurements.

� 2010 Elsevier B.V. All rights reserved.

1. Introduction

Phospholipids are widespread amphiphilic molecules being themain components of biological membranes [1]. In water environ-ment phospholipids form many different structural phases [2].The saturated diacyl phosphatidylcholine derivatives, like 1,2-dimyrilstoyl-sn-glycero-3-phosphocholine (DMPC), in water solu-tion form four lamellar phases: the subgel phase (Lc), planar gel(Lb0 , and rippled gel (Pb0 ) and liquid–crystalline phase (La) [3].The lamellar crystalline (subgel Lc) phase is observed in low tem-peratures, below 279 K.

Increasing temperature induces the first structural transforma-tion from the subgel phase to the lamellar gel phase (Lb0 ), the so-called subtransition [2]. The second phase transition, i.e., the pre-transition from the planar gel Lb0 to the rippled gel Pb0 phase is ob-served at ca. 288 K. The main phase transition (Pb0 ? La) takesplace at 297 K [2].

In water solutions, fully hydrated phospholipids can form lipo-somes, which are multilamellar or unilamellar vesicles. Such vesi-cles are promising as drug-carriers in drug delivery systems [1,4,5].Mixtures of phosphatidylcholines with short and long acyl chainsor mixtures of phosphatidylcholines with surfactants can formthe discoidal phase (bicelles) [4,5]. Bicelles are intermediate mor-phological forms between the classical bilayers and sphericalmicelles (liposomes). Bicelles are useful model systems for

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structural investigation of membrane protein and lipid proteininteractions [5,6].

The characteristic structural phases of phospholipids depend ondifferent factors such as: pH, ionic strength or additives (i.e., cho-lesterol or surfactants). Surface acting agents (surfactants) arewidely used as molecular tools in membrane biology (i.e., purifica-tion of membrane proteins) and membrane mediated drug delivery(i.e., preparation of homogenous lipid vesicles) [7].

Low concentration of surfactant in the phospholipid systemcauses weakening of interactions between molecules into bilayerand also changes hydration degree of the polar heads of phospho-lipids. The high concentration of surfactant in phospholipids sys-tems leads to lysis and fusion of vesicles [7,8].

The aim of our study was to establish the influence of additionof a cationic surfactant in a low concentration on the phase transi-tion of DMPC as seen by the FTIR, SAXS and DS methods. Determi-nation of changes in the phase transitions temperature andknowledge of the dielectric properties of the system investigatedappear to be crucial to the understanding of: the transport proper-ties through bilayers, the immunological activity of the living cellas well as drug-membrane interactions.

Dielectric spectroscopy is a powerful technique for investiga-tion of heterogeneous systems. It brings information not onlyabout the structure, but also about electrical properties of materi-als investigated, thus allowing characterization of systems studiedat molecular and macroscopic levels [9].

In the case of biological cells or proteins, DS allows evaluation ofthe macroscopic dielectric properties or the temperature inducedtransitions [10,11]. It has been evidenced that the dielectric

748 M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753

measurements in radio frequency range are particularly useful forthe studies of biological materials and their phase transitions, likefor example: protein denaturation [7,8] as well as relaxation pro-cesses of polar head dipoles of phospholipids in the MHz range[12–14] or bound water in lipid/water mixtures in higher frequen-cies like: 109–1010 Hz [14–16].

On the basis of radiowave dielectric measurements it was pos-sible to associate the pretransition of DMPC to a thermally fluctu-ating rippled deformation of the phospholipid bilayer, which maybe decomposed in spatial modes called ‘ripplons’, whose are drivenby the viscoelastic properties of the lipid–water interface [17,18].

2. Experimental

2.1. Materials

2.1.1. Sample preparationSurfactant – (dodecyloxymethyl)dodecydimethylammonium

chloride (DDMDDAC) was prepared in our laboratory [19]. 1,2-Dimyristoyl-sn-glycero-3-phosphocholine (DMPC) was purchasedfrom Avanti Polar Lipids (Alabaster, AL). Schematic structures ofboth compounds are presented in Fig. 1(A) DMPC and Fig. 1(B)for DDMDDAC, respectively. The samples investigated were com-posed of: 10% DMPC in 20 mM K2HPO4 in D2O (for FTIR and DSmeasurements) and in H2O (for SAXS measurements). DMPC/DDMDDAC systems studied contained: 0.1%, 0.5%, 1% and 5% ofDDMDDAC. All samples were prepared by 10 cycles of successivesonication at 303 K (30 min) and cooling at 278 K. D2O was usedas a solvent in preparation of the DMPC/surfactant mixture for bet-ter visualization of the carbonyl region in the FTIR spectroscopy[20]. Sonication at a temperature higher than the phase transitionpoint prevents phase separation [21] and allows better incorpora-tion of the surfactant molecules into phospholipids bilayers [22].

2.2. The methods

2.2.1. FTIR measurementsFTIR spectra of DMPC and DMPC/DDMDDAC systems were col-

lected using a BRUKER IFS 66 FTIR-RAMAN Spectrometer with aSPECAC Variable Temperature Cell. Samples of 20 ll volume wereplaced between KRS-5 windows with a Teflon spacer (0.2 mm).One hundred and twenty eight scans in the spectral range of4000–600 cm�1 were collected over the temperatures rangingfrom 273 to 323 K with the resolution of 1 cm�1.

2.2.2. Dielectric measurementsDS measurements were performed using Hewlett–Packard

impedance analyzer model 4291B. The DMPC/surfactant sampleswere placed between two electrodes (diameter 10 mm) and sur-

Fig. 1. Chemical structure of: DM

rounded by a Teflon spacer of the thickness of 50 lm. For the cal-ibration of the system some standards available in the calibrationkit, whose dielectric properties are very well known, were used.

The measured complex dielectric spectrum (e*) is:

e� ¼ e0 � ie00; ð1Þ

where e0 and e00 are the real and imaginary (loss part) components,respectively, and i ¼

ffiffiffiffiffiffiffi

�1p

. These parameters were then calculateddirectly after each frequency sweep, while the errors were within1% for both of them. The measurements were performed in the tem-perature range from 273 to 323 K, with a step of 1 K in the 106–1.8 � 109 Hz frequency range. Part of the dielectric data was ana-lysed by WinFIT impedance analysis software assuming Cole–Colerelaxation functions.

2.2.3. SAXS measurementsSAXS data were collected at the beamline X33 [23] of DORIS

storage ring at DESY (Hamburg, Germany). The measurementswere performed in 1 mm cells with mica windows using MarRe-search 345 mm imaging plate scanner as a detector (MarResearchGmbH). 180-s frames were collected at temperatures from therange: 277–311 K. The scattering vector range was 0.12 < s <5.5 nm�1 (s = 4psinh/k, where k = 0.15 nm is the X-ray wavelengthand 2h is the scattering angle). The normalization of the data, cor-rection for non-homogenous detector response, absorption correc-tion and subtraction of the background scattering were performedusing the program PRIMUS [24]. The pair-distance distributionfunctions p(r) were calculated from the experimental scatteringby indirect Fourier transformation using the program GNOM[25,26].

3. Results

Fourier transform infrared spectroscopy is a method suitable tostudy the structural changes in phospholipids caused by surfactantaddition as a function of temperature. Asymmetric (vas) and sym-metric (vs) stretching vibration of CH2 groups and asymmetricstretching vibration of PO�2 groups were analyzed. Selected FTIRspectra of masCH2 and masPO2 are presented in Fig. 2. Monitoringof the stretching modes of PO�2 groups allows determination ofthe changes in hydration of the outer part of the phospholipidbilayers [27]. Increasing hydration of phospholipid head groups re-sulted in a shift of the wavenumbers characterizing the vibrationsof PO�2 moieties toward lower values (Fig. 2(a)).

The symmetric (vs) and asymmetric vibrations (vas) of CH2

groups are sensitive indicators of phase transitions. Fig. 2(b) and(c) presents the exemplary parts of FTIR spectra (in region of CH2

stretching vibrations) collected for pure DMPC and selectedDMPC/DDMDDAC. For pure DMPC the gel phase is characterized

PC (a) and DDMDDAC (b).

Fig. 2. Exemplary FTIR spectra of fully hydrated DMPC and DMPC/5% DDMDDAC systems in the P–O stretching (a) and in the CH2 anti-symmetric (b) CH2 symmetric (c)vibration regions.

M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753 749

by masCH2 2918 cm�1 and msCH2 2850 cm�1, whereas the liquidcrystal phase is characterized by masCH2 2922 cm�1 and msCH2

2852 cm�1 (Fig. 3). The temperature dependent changes in thewavenumbers characterizing the anti-symmetric and symmetricCH2 stretching vibrations for DMPC and DMPC/surfactant systemsare shown in Fig. 2(b) and (c), and Fig. 3, respectively.

The experimental SAXS data recorded for fully hydrated DMPCand DMPC/DDMDDAC mixtures are presented in Fig. 4. ForDMPC/0.1% DDMDDAC, weak diffraction peaks are still visible.Introduction of the DDMDDAC surfactant at a concentration of

0.5% or higher, results in the total disappearance of the diffractionpeaks corresponding to the lamellar structure of pure DMPC. Thescattering curves obtained for the DMPC/DDMDDAC systems con-taining the surfactant at a concentration of 0.5% or higher exhibitwide maxima characteristic of the bicellar phase. Such a scatteringpattern was observed also for other systems with single phospho-lipid bilayers in the absence of stacking [28,29].

Dielectric spectroscopy was used to verify the dielectric permit-tivity response of the DMPC/DDMDDAC systems in the tempera-ture range in which the phase transitions were expected. The

Fig. 3. Temperature dependence of masCH2 stretching vibrations for DMPC andselected DMPC/DDMDDAC systems.

750 M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753

motions of the zwitterionic phospholipid head group in DMPC sus-pension can be observed and described as Cole–Cole [10] or Havr-iliak–Negami approximated to Debye process [30] in radiowavefrequency range.

In Fig. 5 the real part (e0) and imaginary part (e0) of the complexdielectric permittivity are plotted vs. frequency for DMPC suspen-sion at temperature 298 K (liquid crystalline phase). In the case ofphospholipids suspension three types of dispersions have beengenerally reported: (1) in the frequency regions below 40 MHz, re-lated to the ionic species around the phospholipids vesicle surface;(2) between 20 MHz and 300 MHz, caused by the rotational diffu-sion of the zwitterionic head groups of lipid molecules and (3) from6 GHz to 80 GHz due to bound water [31,32]. In the frequency win-dow available in our experiment the first two dispersions weremeasured, thus the real and imaginary parts of dielectric permit-tivity were analyzed assuming two Cole–Cole relaxation time dis-tributions. This function was previously found as a most suitablefor analysis of similar systems [32]. A broad relaxation at low fre-quency is considered to be related to the restricted motions of elec-trically charged spices, i.e., contribution from counterions

Fig. 4. SAXS curves recorded for DMPC and DMPC/DDMDDAC systems at temperature 282277 to 310.6 K (b). Intramolecular pair-distance distribution function p(r) calculated for

surrounded DMPC vesicles [33]. The second smaller dispersion inFig. 5 was found for the frequency of about 3 � 107 Hz and is attrib-uted to the rotational diffusion of the zwitterionic head groups ofDMPC. The relaxation time of this process was found to be equal6 ns while its dielectric strength (De) was about 11 as assessedby the fitting procedure at 298 K. These dielectric parameters arein a good agreement with the data reported previously [14,30].

At a frequency 3 � 107 Hz, at which the relaxation ascribed toDMPC head motions is observed (see Fig. 5(b)), 80% of the loss partof dielectric permittivity signal is due to conductivity contribution.For this reason the best fits of dispersion parts presented can becharged with an error of about 20% of parameter calculated. There-fore, in the further studies we will analyze the real part of thedielectric permittivity (e0) of the materials investigated. e0 is anindication of the polarizability of the molecules and their abilityto store electric energy, so that evolution of e0 can give informationabout the phase transition of the systems investigated, as evi-denced earlier on the example of determination of protein dena-turation [10] and transformations of phospholipids [18]. The realpart of the complex dielectric spectrum of DMPC suspension inD2O, as a function of frequency at a few selected temperaturesfrom the range 273 K to 323 K, is presented in Fig. 6.

At lower frequencies one can observe the value of e0 to increasewith increasing temperature (from bottom to top). This frequencyregion is assigned to restricted counterion motions whose give thepredominant contribution to overall dielectric response over thedielectric signal from zwitterionic DMPC head group (whoseamount in the mixture studied is only 10%). Nevertheless, analysisof this frequency range allows characterizing the phospholipidsmobility, i.e., to probe transformations of the phospholipids inves-tigated [18]. Fig. 7(a)), shows the real part of dielectric permittivitye0 (at 1.5 MHz), while Fig. 7(b)) presents the conductivity of DMPCsuspension measured in the temperature range from 273 to 315 K.In such a graph a small step increase in the e0 value at about 286 Kcan be connected with the pretransition and an increase in themobility of the DMPC aliphatic chains, while the second step ob-served at about 298 K can be assigned to the main transition in

K (a). SAXS curves recorded for DMPC/5% DDMDDAC systems at temperatures fromselected DMPC/DDMDDAC systems at temperature 282 K (c).

106 107 108

100

200

300

400

Frequency (Hz)

ε'

T=273K T=283K T=293K T=303K T=313K T=323K

Fig. 6. Real part of complex dielectric spectrum (e0) of the DMPC suspension vs.frequency measured at different temperatures.

275 280 285 290 295 300 305 310 315

120

160

200

240

Die

lect

ric

prem

itivi

tty ( ε

')

(a)

0.10

0.15

0.20

0.25

Con

duct

ivity

(S/m

)

Temperature (K)

(b)

275 280 285 290 295 300 305 310 315

Fig. 7. The real part of dielectric permittivity e0 (at 1.5 MHz) (a) and theconductivity (b) measured for DMPC suspension in the temperature range from273 to 315 K. Rectangles are drawn as a guide for eyes to show the approximatetemperature range of transitions in both representations.

106 107 108100

101

102

103

ε"

Frequency (Hz)

(b)

106 107 108

0,0

5,0x102

1,0x103

1,5x103

2,0x103

2,5x103

3,0x103

ε"

Frequency (Hz)

106 107 108

102

(a)

ε'

Frequency (Hz)

Fig. 5. Real part e0 (a) and imaginary part e0 0 (b) of the complex dielectricpermittivity plotted vs. frequency for DMPC suspension at 298 K. The squaresymbols in the e0 and e00 plots vs. frequency correspond to measured values, whilethe thick lines represent the best fitting curves evaluated with assumption of twodispersion contributions (dashed curves) in investigated system. The thin solid linein imaginary part representation (b) denotes contribution from the electricconductivity. In the inset, e00 vs. frequency is shown without logarithmic scale inorder to exposure predominant contribution of electrical conductivity in imaginarypart of dielectric spectra.

M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753 751

phospholipids, called the ‘chain melting’. It is important to stressthat rectangles, which are drawn as a guide for eyes in Fig. 7(a)and (b) shows the approximate temperature range of transitionsin both graphs, however the pretransition is less visible in the con-ductivity plot. Fig. 8 presents the dielectric spectra of e0 vs. temper-ature, measured at 1.5 MHz, for DMPC/DDMDDAC systems withdifferent amounts of the surfactant. As a result of the surfactantaddition, the temperature of the main phase transition of DMPCslightly decreases and the interval over which the transition takesplace broadens even down to 278 K, in the presence of 5% ofDDMDDAC.

Finally, it is important to emphasise that DS measurements canbe applied as a appropriate and sensitive tool for phase transfor-mation temperature determinations in phospholipids, even in di-lute systems. A comparison of the temperature dependence ofthe real part of dielectric permittivity and that of the asymmetricstretching vibrations of PO�2 from FTIR measurements, presentedin Fig. 9, clearly shows that in these two plots the temperaturescorresponding to the pre- and the main transitions in 1,2-dimyri-stoyl-sn-glycero-3-phosphocholine are the same.

4. Discussion

The wavenumbers observed for PO�2 vibrations in pure fully hy-drated DMPC are 1212 cm�1 and 1209 cm�1, respectively, for thegel and the liquid–crystalline phase. For the system of DMPC/0.1% DDMDDAC, a shift of vs from 1212 cm�1 (in gel phase) to1210 cm�1 (in liquid–crystalline phase) was observed. However,for the systems containing 0.5–5% DDMDDAC the wavenumbershift was observed from 1217 cm�1 to 1212 cm�1. In the gel phase,the trans conformers of acyl chains dominate and therefore, thesystem is characterized by a high degree of packing. For this phasethe mas CH2 and ms CH2 vibrations occur at lower wavenumbers [34].Increasing temperature of the system induces the conformationalisomerisation, which results in formation of gauche conformers,whose presence reduces the strength of interaction between thealkyl chains of phospholipids. This reduction is manifested as anincrease in the wavenumber values [35]. The observed conforma-tional disorder is caused by the interactions with polar head

280 290 300 3101206

1209

1212

1215

1218

1221

1224

1227 Lα

Pβ'

Temperature [K]

Wav

enum

ber [

cm-1]

Lβ'

100

120

140

160

180

200

220

240

ε'

Fig. 9. The real part of dielectric permittivity e0 (at 1.5 MHz) and the asymmetricstretching vibrations of PO�2 of DMPC suspension in the temperature range from273 to 315 K. Phase transitions are visible as steps in presented curves. Lb0 , Pb0 , La

denote: planar gel, rippled gel, liquid–crystalline phases, respectively.

Fig. 8. Dielectric spectra of e0 vs. temperature for DMPC/DDMDDAC systems, withdifferent amounts of surfactant measured at 1.5 MHz. Rectangles are drawn as aguide for eyes to show the approximate temperature range of transitions.

752 M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753

groups [36]. Such interactions of cationic surfactants with DMPChead groups result in conformational disorder in the phospholipidbilayers and decrease the main phase transition temperature. Forfully hydrated pure DMPC the symmetric and anti-symmetricstretching modes are maCH2 2918 cm�1 and msCH2 2850 cm�1 inthe gel phase and maCH2 2922 cm�1 and msCH2 2852 cm�1 for theliquid–crystalline phase. The temperature range of the main phasetransition is quite narrow for pure DMPC. Introduction of the sur-factant to the system results also in an increase in the temperaturerange of the main phase transition from 4 K to 13 K for 0.1%DDMDDAC and 5% DDMDDAC, respectively. A similar situationwas observed for the interactions of dipalmitoylphosphatidylcho-line (DPPC) with (decyloxymethyl)dodecydimethylammoniumchloride (DMDDAC) and dodecyldimethyl(hexyloxymethyl)ammo-nium chloride (HMDDAC) [37].

The experimental SAXS data allow determination of structuralparameters and thermal stability of selected phospholipid/surfac-tant systems. The bicellar phase is very stable up to 295 K, whichis clearly visible from the superposition of the experimental SAXSdata collected at temperatures from 277 K to 310.6 K (Fig. 4(b)).The detail information about the structure of the lipid bilayer inthe systems studied can be derived from the pair-distance distribu-tion function p(r). The p(r) function is the distribution of the dis-tances r within a particle obtained by combining any volumeelement with any other volume element. The calculated p(r) func-tions for selected monodisperse data sets are presented in Fig. 4(c).The maximum dimension (Dmax) characterizing the bicelles deter-mined from the function p(r) increases with temperature and forDMPC/0.5% DDMDDAC varies from 40.5 nm (277 K) to 46 nm(295 K). The bicelles observed in DMPC/1% DDMDDAC system havegreater sizes ranging from 54 nm (277 K) to over 58 nm (290 K).Above 290 K (in the vicinity of the main phase transition in DMPC)the bicelles undergo transformation, most probably leading todifferentiation in their size and to increased polydispersity of thesystems studied. The addition of the surfactant in higher concen-tration (5%) resulted in destabilisation of bicellar phase. The p(r)function and Dmax for DMPC/5% DDMDDAC can be determined onlyup to 286 K and at which Dmax value reached nearly 55 nm. The ra-dii of gyration characterizing the bicellar systems studied werefrom 14.6 to 20.5 nm.

Dielectric analysis of the systems consisting of phospholipids isa complicated issue in which the amphiphilic nature of these mol-ecules is a crucial factor determining the interaction within themembrane, i.e., lipid double layer as well as interaction with sur-rounding medium (most often water). In a aqueous solutions phos-pholipids form spontaneously vesicles of different morphologylike: uni- or multi lamellar vesicles. In any biological or membranesystem one must consider complex interfacial polarization mecha-nism leading to the so-called multi-shell model of complex permit-tivity [9]. For this reason the complex dielectric spectrum stronglydepends on the morphology of phospholipids [9] and the shape ofexisting forms [38]. In general, besides the water relaxation de-tected at frequencies above 1 GHz (not visible in frequency rangeof our measurements) and reorientation of dipolar head groupsof DMPC (found at frequency of 3 � 107 Hz, presented in Fig. 5),the above-mentioned interfacial polarization of multi-vesiclesforms by DMPC (called also the Maxwell–Wagner polarization)and counterions displacement along the liposome surface mustbe taken into account in the systems studied. For the dispersionfound in low frequency range (Fig. 5(a) and (b)) it is difficult to

M. Kozak et al. / Journal of Non-Crystalline Solids 356 (2010) 747–753 753

asses if the interfacial polarization or ion diffusion around chargedDMPC interface bring a dominant contribution to this process. Anincrease in (e0) is visible in Fig. 5(a) and originates from one ofthese relaxations, rather than from the electrode polarization ef-fect, which was excluded in our analysis. It has already beenproved that the electrode polarization contribution is insignificantabove 1 MHz, i.e., in the frequency range chosen in the presentstudies [39,40].

The plots of dielectric permittivity (e0) and conductivity, mea-sured at 1.5 MHz, as a function of temperature (see Fig. 7) revealtwo steps at 287 K and 298 K, assigned to the pretransition andthe main phase transition of DMPC, respectively. The pretransitionis connected with in-phase modulation of phospholipid bilayersand affects the lipid–water interface [18]. The second step (a de-crease in e0 and conductivity at 288 K) is assigned to the main tran-sition of DMPC and the overall tendency of these parameters toincrease with temperature is obscured by the counteracting effectcoming from the increased number of ions trapped in vesicularcores of DMPC thus unable to move freely [32]. These findingsadditionally prove, that the increase in e0 (in Fig. 5(a)) originatesrather from the effect of counterions reorganization than fromelectrode polarization. The increase in DMPC hydration at the sub-sequent phase transitions was also confirmed by FTIR measure-ments as shown in Fig. 9. A perfect agreement was found for thephase transition temperatures in DMPC suspension, proving a highsensitivity of FTIR and dielectric spectroscopies in studies of trans-formations of phospholipids systems. After addition of DDMDDACsurfactant to DMPC suspension, the phase transition temperatureswere observed to decrease and the interval between the beginningand the end of the phase transition broadens as revealed in Figs. 3and 8.

Such changes can suggest that addition of even small amount ofsurfactant influences the organization of phospolipid structuredestroying the acyl chain arrangement typical of neat DMPC, thusmaking it easier to reorganise at the phase transition, i.e., it facili-tates the chain melting.

It is difficult to propose one certain mechanism of interactionbetween DMPC and surfactant molecules, since one must considerformation of complexes between surfactant and ions (present inbuffer), formation of surfactant dimers (or oligomers), either sur-factant incorporation into phospholipid bilayer due to hydrophobicinteractions [41,42]. In DMPC/surfactant systems investigated themost probable mechanism assumes the interaction between polarparts of phospholipids and surfactant, then gradual incorporationof DDMDDAC in DMPC connected with reorganization of sur-rounded moieties (as revealed by FTIR and DS measurements), to-gether with possible increase in hydrophobic interactions of acylchains of both components. Finally it leads to overall transforma-tion of the system studied from lamellar to more stable bicellarphase, as evidenced by SAXS data.

5. Conclusions

Our results have shown that the cationic surfactant DDMDDACstrongly interacts with DMPC. Surfactant molecules interactingwith phospholipid bilayer induce changes in hydration level ofphospholipids polar head groups and conformational disorder inthe hydrophobic part of bilayer. This effect is visible in FTIR resultsas decrease in the phase transition temperature and change inwavenumbers characteristic of the vibrations of CH2 and PO�2groups. As a result of addition of the surfactant studied, the SAXSscattering pattern characteristic of the lamellar structure is signif-icantly changed, which is manifested by disappearance of thestrong diffraction maxima. Dielectric spectroscopy in radio fre-quency range is a suitable method for characterization of biological

membrane systems such as phospholipids. The real part of dielec-tric permittivity vs. temperature shows broadens of the DMPCphase transition in the case of DMPC/DDMDDAC systems. Thephase transitions temperatures obtained from dielectric measure-ments are in good agreement with the FTIR data.

Acknowledgements

This work is supported in part by the Ministry of Science andHigher Education (MSHE) in Poland Grant No. NN202 248935.The data collection at beamline X33 (EMBL c/o DESY, Hamburg)was supported by European Community – EMBL Hamburg Outsta-tion, contract number: RII3-CT-2004-506008. A. Wypych acknowl-edges MSHE in Poland for a Postdoctoral Fellowship (POL-POSTDOC III).

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