Dynamic response on a nanometer scale of binary phospholipid-cholesterol vesicles: Low-frequency Raman scattering insight

Low-frequency Raman spectroscopy was used to study the dynamic response on a nanometer scale of aqueous suspensions of two-component lipid vesicles. Binary mixtures of saturated phospholipid (1,2-dipalmitoyl-sn-glycero-3-phosphocholine, DPPC) and cholesterol are interesting for possible coexistence of solidlike and liquid-ordered phases, while the phase coexistence was not reported for unsaturated phospholipid (1,2-dioleoyl-sn-glycero-3-phosphocholine, DOPC) and cholesterol mixtures. The DOPC-DPPC mixtures represent the well-documented case of coexisting domains of solidlike and liquid-disordered phases. These three series of lipid mixtures are studied here. A broad peak with the maximum in the range of $30–50\mathrm{c}{\mathrm{m}}^{–1}$ and a narrow peak near $10\mathrm{c}{\mathrm{m}}^{–1}$ are observed in the Raman susceptibility of the binary mixtures and attributed to the acousticlike vibrational density of states and layer modes, respectively. Parameters of the broad and narrow peaks are sensitive to lateral and conformational hydrocarbon chain ordering. It was also demonstrated that the low-frequency Raman susceptibility of multicomponent lipid bilayers allows one to determine the phase state of lipid bilayers and distinguish the homogeneous distribution of molecular complexes from coexisting domains with sizes above several nanometers. Thus, the low-frequency Raman spectroscopy provides unique information in studying phase coexistence in lipid bilayers.

Figure 1

Raman susceptibility spectra for various DOPC-DPPC mixtures. The spectra are normalized to the amplitude of the C-N stretching peak near $720\mathrm{c}{\mathrm{m}}^{–1}$. The spectra are vertically shifted for clarity.

Figure 2

Raman susceptibility spectra for various DPPC-Chol mixtures. The spectra are normalized to the amplitude of the C-N stretching peak near $720\mathrm{c}{\mathrm{m}}^{–1}$. The spectra are vertically shifted for clarity.

Figure 3

Raman susceptibility spectra for various DOPC-Chol mixtures. The spectra are normalized to the amplitude of the C-N stretching peak near $720\mathrm{c}{\mathrm{m}}^{–1}$. The spectra are vertically shifted for clarity.

Figure 4

Amplitude of the broad peak in the low-frequency Raman susceptibility (bottom) and its integral over $5–200\mathrm{c}{\mathrm{m}}^{–1}$ (top) versus the compositions of the binary phospholipid-phospholipid and phospholipid-cholesterol series.

Figure 5

Frequency of the maximum (circles) and right HWHM (triangles) of the broad peak in the low-frequency Raman susceptibility versus the binary composition. The open symbols correspond to the evaluation implying the smoothing, and the solid symbols to a quadratic polynomial fitting near the maximum.

Figure 6

Raman susceptibility spectra for the So phase (DPPC95-Chol05 mixture, thick line), the Ld phase (DOPC, squares), liquidlike phase of DOPC60-Chol40 (gray line), and Lo phase of DPPC60-Chol40 (thin line). The inset shows the low-frequency part of the DPPC60-Chol40 spectrum (circles) with the linear extrapolation of the broad peak contribution (dotted line) and Gaussian description of the layer mode (solid line).

Figure 7

Amplitude (top), position of the maximum (bottom, circles), and width (bottom, triangles) of the layer mode versus the binary composition.

Figure 8

(a) Raman susceptibility spectra for the DOPC-DPPC mixtures. The thick lines are DOPC and the mixture with 5 mol % of DOPC. The gray lines show the spectra for the mixtures with 10, 15, 20, 23, 30, 35, 40, 45, 50, 55, 65, 80, 90 mol % of DOPC. (b) Difference spectra between a particular spectrum and one of the mixture with 5 mol % of DOPC, the difference spectra were normalized to its amplitude. The thick line is the difference spectrum for DOPC, and the gray lines are for the mixtures with 20, 23, 30, 35, 40, 45, 50, 55, 65, 80, 90 mol % of DOPC.

Figure 9

(a) Raman susceptibility spectra for the DPPC-Chol mixtures. The thick lines correspond to the mixtures with 5 and 40 mol % of Chol. The gray lines show the spectra for the mixtures with 10, 12, 15, 20, 25, 30, 35 mol % of Chol. (b) Difference spectra between a particular spectrum and one of the mixture with 5 mol % of Chol, the difference spectra were normalized to its amplitude. The thin lines are the difference spectrum for the mixtures with 12, 15, 20 mol % of Chol, the gray lines for the mixtures with 25, 30, 35 mol % of Chol, the thick solid line corresponds to the mixture with 40 mol % of Chol.

Figure 10

(Top) Raman susceptibility at minimum in the normalized difference spectra of Figs. 8 and 9 versus the binary composition. (Bottom) The amplitude of the difference spectra used for the normalization in Figs. 8 and 9 (triangles) versus the binary composition. The lines describe the composition dependences in the supposed coexistence range.