Langmuir 2001, 17, 6577-6584
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Reversible Stepwise Formation of Mono- and Bilayers of a Fluorocarbon/Hydrocarbon Diblock on Top of a Phospholipid Langmuir Monolayer. A Case of Vertical Phase Separation Marie Pierre Krafft,*,† Franc¸ oise Giulieri,‡ Philippe Fontaine,§ and Michel Goldmann§,| Chimie des Syste` mes Associatifs. Institut Charles Sadron (UPR CNRS 22). 6 rue Boussingault, 67083 Strasbourg Cedex, France, Unite´ de Chimie Mole´ culaire, Universite´ de Nice, 06108 Nice Cedex, France, Laboratoire pour l’Utilisation du Rayonnement Electromagne´ tique (LURE, UMR CNRS 130), BP 34, 91 898 Orsay Cedex, France, and Laboratoire des Objets Complexes et Interfaces d’Inte´ reˆ t Biologique (FRE CNRS 2303), Universite´ Rene´ Descartes, 45 rue des Saints-Pe` res, 75270 Paris Cedex 06, France Received April 23, 2001. In Final Form: June 29, 2001 Compression of mixed Langmuir monolayers made from combinations of dipalmitoyl phosphatidylethanolamine (DPPE) and the semi-fluorinated alkane C8F17C16H33 (F8H16) results in expulsion of the diblock molecule at high pressure. Depending on the DPPE/F8H16 molar ratio, either a monolayer or a bilayer of F8H16 is formed on top of a DPPE-only monolayer. The structure of the hydrocarbon and fluorocarbon layers that constitute the resulting vertically separated bi- or trilayer was determined using grazing incidence X-ray diffraction. The phenomenon is reversible and represents a novel case of vertical, pressure-induced phase separation.
Introduction Fluorinated chains are extremely hydrophobic and lipophobic as well, the latter feature being illustrated by the fact that fluorocarbons and hydrocarbons form highly nonideal mixtures.1 This unique combination of characters, usually considered as antinomic, can be exploited to induce macro- or microphase separations in 2D and 3D molecular systems. For example, segregated micelles,2 multicompartment polymeric micelles,3 and liposomes containing separated fluorinated domains in their bilayer membrane4 were obtained by combining fluorinated and hydrogenated surfactants. We report here the first example of a reversible, pressure-induced vertical phase separation in a Langmuir monolayer made from a lipid and a semi-fluorinated alkane. This situation is different from the lateral phase separations observed in monolayers composed by fluorinated and hydrogenated amphiphiles.4,5 The driving forces for the phase separation reported here include the reduced miscibility of the two monolayer components, the propensity of fluorinated chains to self-assemble in an orderly manner, and the decrease in surface tension resulting from the expulsion of the semi-fluorinated compound when its fluorinated segment points toward the air phase above * To whom correspondence should be addressed. Tel: (33) 3 88 41 40 60. Fax: (33) 3 88 40 41 99. E-mail:
[email protected]. † Institut Charles Sadron. ‡ Universite ´ de Nice. § Laboratoire pour l’Utilisation du Rayonnement Electromagne´tique. | Universite ´ Rene´ Descartes. (1) Hildebrandt, J. H.; Prausnitz, J. M.; Scott, R. L. Regular and Related Solutions; Van Nostrand-Reinhold: New York, 1970. (2) Mukerjee, P.; Yang, A. Y. S. J. Phys. Chem. 1976, 80, 1388. (3) Sta¨hler, K.; Selb, J.; Candau, F. Langmuir 1999, 15, 7565. (4) Elbert, R.; Folda, T.; Ringsdorf, H. J. Am. Chem. Soc. 1984, 106, 7687. (5) Zhang, L. H.; Zhu, B. Y.; Zhao, G. X. J. Colloid Interface Sci. 1991, 144, 483. Zhang, L. H.; Zhao, G. X.; Zhu, B. Y. J. Colloid Interface Sci. 1991, 144, 491.
the lipid layer. This work therefore also differs from papers that report the formation of multilayers upon compression from a monolayer made from one single component. Examples of such molecules forming multilayers include the ferroelectric liquid crystal p-hexyloxybenzylidene-p'amino-2-chloro-R-propylcinnamate (HOBACPC),6 the thermotropic liquid crystal 4'-n-octyl-4-cyanobiphenyl (8CB),7 discotic8 and pyramidic 9 liquid crystals, polysiloxanes,10 and R-helical polyglutamates.11 In the latter cases, the growth of multilayers is essentially driven by the weakness of the lateral interactions (collapse). One paper reports the monolayer behavior of a mixture of a lipid and a liquidcrystal-forming molecule.12 Upon compression, the liquid crystal is expelled under the lipid monolayer. No structural information was provided, however, on the organization of the segregated layers. We present here the monolayer behavior of combinations of dipalmitoyl phosphatidylethanolamine (DPPE) and semi-fluorinated alkane C8F17C16H33 (F8H16) in various ratios as studied by grazing incidence X-ray diffraction (GIXD). Use of such lipid/diblock combinations was reported to provide highly stable fluorocarbon emulsions and liposomes with high thermal stability and reduced membrane permeability.13-16 The determination of the diffraction patterns (in-plane (qxy) and out-of-plane (qz)) (6) Rapp, B.; Gruler, H. Phys. Rev. A. 1990, 42, 2215. (7) Xue, J.; Jung, C. S.; Kim, M. W. Phys. Rev. Lett. 1992, 69, 474. (8) Auweraer, M. V.d.; Catry, C.; Feng Chi, L.; Karthaus, O.; Knoll, W.; Ringsdorf, H.; Sawodny, M.; Urban, C. Thin Solid Films 1992, 210/211, 39. (9) El Abed, A.; Daillant, J.; Peretti, P. Langmuir 1993, 9, 3111. (10) Fang, J. Y.; Dennin, M.; Knobler, C. M.; Godovsky, Yu. K.; Makarova, N. N.; Yokoyama, H. J. Phys. Chem. B 1997, 101, 3147. (11) Tsukruk, V. V.; Einloth, T. L.; van Esbroeck, H.; Frank, C. W. Supramol. Sci. 1995, 2, 219. (12) Fang, J. Y.; Uphaus, R. A. Langmuir 1994, 10, 1005. (13) Corne´lus, C.; Krafft, M. P.; Riess, J. G. J. Colloid Interface Sci. 1994, 163, 391. (14) Trevino, L.; Fre´zard, F.; Rolland, J. P.; Postel, M.; Riess, J. G. Colloids Surf. 1994, 88, 223. (15) Krafft, M. P.; Riess, J. G. Biochimie 1998, 80, 489. Krafft, M. P. Adv. Drug Delivery Rev. 2001, 47, 209.
10.1021/la010587a CCC: $20.00 © 2001 American Chemical Society Published on Web 09/20/2001
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Figure 1. In-plane structure showing the rectangular cell and the a and b parameters in the case of a tilted NN phase (L2).
allowed us to establish the structure of the lattice formed by the hydrophobic chains of the amphiphiles in the alternate fluorinated and hydrogenated layers. Experimental Section Materials. F8H16 was synthesized according to reference 17 and thoroughly purified by column chromatography. Its chemical purity (>99%) was determined using GC, TLC, NMR, and elemental analysis. DL-R-DPPE was obtained from Sigma. Spreading solutions of DPPE and F8H16 (0.5 mmol L-1 for minitrough experiments and 1 mmol L-1 for GIXD experiments) were prepared in chloroform (analytical grade)/methanol (9:1). Water was purified using a Millipore system (pH ) 5.5, surface tension: 72.1 mN m-1 at 20 °C, resistivity: 18 MΩ cm). Monolayer Isotherms. Surface pressure versus molecular area isotherms were recorded on a Langmuir minitrough (Riegler & Kirstein, Germany) equipped with a movable single barrier (speed barrier: 4.0 mm min-1). The surface pressure π was measured using the Wilhelmy plate method. Temperature was regulated at 20.0 ( 0.5 °C. 25 µL of DPPE or F8H16 solutions were spread on the water surface. For each DPPE/F8H16 molar ratio, DPPE and F8H16 were co-solubilized, and appropriate volumes of the mixture were deposited so that the number of DPPE molecules that were spread was constant. As a consequence, the isotherms of the mixed monolayers are shifted toward large molecular areas when the amount of diblock is increased. The difference in extrapolated molecular area between the pure DPPE curve and that of any of the mixed monolayer curves corresponds to the area occupied by diblocks on the water surface. Grazing Incidence X-ray Diffraction (GIXD). GIXD experiments were achieved at the D41B beamline of the LUREDCI synchrotron source (Orsay, France). The X-ray wavelength λ ) 1.646 Å of the incoming X-ray beam was selected using a focusing Ge (111) monochromator. The grazing angle of incidence θi ) 2.08 mrad was fixed slightly below the critical angle for total external reflection of X-rays at the air/water interface (about 2.8 mrad at 1.646 Å). The intensity of the X-ray beam scattered by the monolayer was monitored vertically using a vertical gasfilled (Ar-CO2) position sensitive detector (PSD) (13°) as a function of the in-plane component of the scattering wave vector qxy selected by means of a Soller slits collimator. The resulting qxy resolution was 0.007 Å-1 for the q-range explored here. The study of the qxy pattern integrated over the vertical wave vector component qz allowed us to determine the 2D lattice of the hydrogenated and fluorinated chains, while the shape of the Bragg rods gave information about the tilt angle t and tilt azimuth φ.18 In this study, two distinct phases have been observed: the untilted rotator hexagonal phase (LS) (t ) 0, φ = 0) and the nearest neighbor tilted phase (L2d) (t * 0, φ ) 0), according to the nomenclature introduced in reference19. In the following, we will use the rectangular description of the chain lattice described in Figure 1. For the LS phase, the diffraction pattern exhibits a single, degenerate peak corresponding to the [1h 1], [11], and [02] reflections which intensity along the Bragg rod is located in plane (qz ) 0 Å-1). For the L2d phase, the diffraction pattern exhibits two peaks. The peak located at low qxy corresponds to the [1 h 1] and [11] reflections, and the other peak corresponds to the [02] reflection. Since the chains are tilted to the nearest (16) Krafft, M. P.; Riess, J. G.; Weers, J. G. In Submicronic Emulsions in Drug Targeting and Delivery; Benita, S., Ed.; Harwood Academic Publ.: Amsterdam, 1998; p 235. (17) Brace, N. O. J. Org. Chem. 1973, 38, 3167. (18) Als-Nielsen, J.; Jacquemain, D.; Kjaer, K.; Leveiller, F.; Lahav, M.; Leiserowitz, L. Phys. Rep. 1994, 246, 251. (19) Kaganer, V. M.; Mo¨hwald, H.; Dutta, P. Rev. Mod. Phys. 1999, 71, 779 and references therein.
Figure 2. Molecular area vs surface pressure isotherms of monolayers made from F8H16, DPPE, and DPPE/F8H16 mixtures in various ratios (2:1, 1:1, 1:1.3, 1:1.5, 1:2). neighbor in the L2d phase, the maximum of intensity along the Bragg rods [1h 1], [11] is located out of the plane and in the scattering plane along the Bragg rod [02]. The cell parameters (a, b) were determined from the integrated diffraction data: a
x
2
) 4π/ 4q211-q 02and b ) 4π/q02. In the case of the LS phase, note that b ) x3a ) {4π}/{q02}. The d spacing of the corresponding diffracting planes was derived from the diffraction peak position by d ) 2π/qxy. Under the assumption that the peaks, corrected for resolution effects, may be fitted by a Lorentzian shape function, the positional correlation length ξ may be extracted from experimental data using the well-known relation: ξ ) 2/fwhm, where fwhm is full width at half-maximum. The chain cross section A0 was given by A0 ) Axy cos t. The molecular area per chain was given by Axy ) ab/2. The tilt angle was extracted from the position of the maximum along the Bragg rods qz[11] (or qz[1h 1]). The maximum intensity along the Bragg rod is obtained when the scattering wave factor transfer q is perpendicular to the molecular axis, whose direction is given by the director n. From the relation q.n ) 0, we get in the case of the NN tilted structure q11z ) q11xy tan(t). The fwhm along qz gives the thickness of the monolayer. The Teflon Langmuir trough mounted on the diffractometer was equipped with a movable single barrier. The surface pressure π, measured using the Wilhelmy plate method, was kept constant during a given scan. The trough was enclosed in a gastight box, with Kapton windows, that was filled with water-saturated helium. The temperature was regulated to 20 ( 0.5 °C. Approximately 100 µL of F8H16, DPPE, or DPPE/F8H16 mixtures were spread on the water surface. The film was compressed step by step, and Bragg peaks were recorded at each fixed pressure step. The total duration of a scan was typically 1.5 h.
Results and Discussion Figure 2 shows surface pressure/molecular area (π/A) isotherms for F8H16, DPPE, and DPPE/F8H16 (2:1, 1:1, 1:1.3, 1:1.5, and 1:2) monolayers (characterized by the corresponding molar ratios 2, 1, 0.77, 0.67, and 0.5, respectively). DPPE Monolayer. DPPE forms at 20 °C and at a nonzero pressure a monolayer in the liquid condensed (LC) state (Figure 2) that is stable up to ∼47 mN m-1. Figure 3 shows the diffracted integrated intensity (I/I0) as a function of qxy, and the corresponding contour plots of I/I0 as a function of qxy and qz at 10 and 38 mN m-1. For pressures lower than 35 mN m-1 (Figure 3a), the two characteristic diffraction peaks of the tilted L2d phase are
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Figure 3. Grazing incidence X-ray scattered intensity (I/I0) as a function of the in-plane (qxy) component of the scattering wave vector q (left), and contour plots (I/I0 as the function of qxy and of the out-of-plane component of q (qz)) (right), for pure DPPE at 10 mN m-1 (a) and 38 mN m-1 (b). Table 1. Unit Cell Dimensions (a, b) and Projected Molecular Areas (Axy) as Derived from In-Plane Diffraction Data at Different Surface Pressures for DPPEa unit cell surface cross dimensions molecular mono- pressure area Axy tilt struc- section A0 -1 2 layer (mN m ) a (Å) b (Å) (Å ) t (°) ture (Å2) DPPE
2 10 20 30 38
5.22 5.12 5.00 4.90 4.79
8.46 8.43 8.38 8.33 8.30
44.2 43.2 41.9 40.8 39.7
26 23 18 12 0
NN NN NN NN H
39.7 39.6 39.8 39.9 39.7
a Tilt angle (t) and cross section (A ) were derived as described 0 in the Experimental Section.
observed. As pressure is increased, the DPPE molecules straighten progressively on the water surface. A transition, that is hardly visible on the isotherm, occurs at 35 mN m-1, after which the diffraction pattern is characteristic of the LS untilted phase (one peak at 1.52 Å-1) (Figure 3b). Table 1 collects the lattice parameters and tilt values for this monolayer as a function of pressure. The results are in agreement with those of Bo¨hm et al.20 F8H16 Monolayer. Compression isotherms of some FnHm diblocks (F12H8, F10H12, F12H18) were first reported by Gaines, who demonstrated that these molecules have π-A isotherms similar to those of Langmuir monolayers made from conventional amphiphiles.21 F8H16 forms a monolayer that remains stable up to ∼13 mN m-1 (Figure 2) with a limiting area of about 31 Å2, that is, significantly larger than the cross section (ca. 20 Å2) of a (20) Bo¨hm, C.; Mo¨hwald, H.; Leiserowitz, L.; Als-Nielsen, J.; Kjaer, K. Biophys. J. 1993, 64, 553. (21) Gaines, G. L., Jr. Langmuir 1991, 7, 3054.
typical hydrogenated chain, but corresponding to the reported cross section of a perfluorinated chain.22 Because the van der Waals radius of the fluorine atom is larger than half the C-C distance, steric hindrance causes the chain to adopt a 15/7 helical conformation, which confers added rigidity to the carbon backbone and enlarges the chain’s cross-sectional area.23 The isotherm is reversible, without hysteresis upon compression-expansion cycles. GIXD scans were taken, mostly in the qxy range, from 0.8 to 1.6 Å-1, where one would expect to find diffraction peaks resulting from fluorinated chain-fluorinated chain (1.25 Å-1) and hydrogenated chain-hydrogenated chain close packing. Figure 4 shows the scattered intensity as a function of qxy and contour plot at 8 mN m-1. A Bragg peak is obtained, showing that the diblock monolayer has an ordered structure. The peak is broad, however, revealing a low positional correlation length (ξ ) 20 Å). As mentioned by Huang et al24 for F12H18, the peak position (1.25 Å-1) is consistent with the location of the first-order diffraction peak from monolayers of perfluorinated acids22 and perfluoro-n-eicosane,25 which suggests that in the F8H16 monolayer, as in the F12H18 one, the perfluoroalkyl segments (F8) form a hexagonal lattice. The contour plot shows that the F8H16 molecules are only slightly tilted (10° at maximum). No peak was obtained for the alkyl segments (H16) of F8H16, indicating that they are likely in the liquid state. An important question was to (22) Acero, A. A.; Li, M.; Lin, B.; Rice, S. A.; Goldmann, M.; Azouz, I. Z.; Goudot, A.; Rondelez, F. J. Chem. Phys. 1993, 99, 7214. (23) Bunn, C. W.; Howell, E. R. Nature 1954, 174, 549. (24) Huang, Z.; Acero, A. A.; Lei, N.; Rice, S. A.; Zhang, Z.; Schlossmann, M. L. J. Chem. Soc., Faraday Trans. 1996, 92, 545. (25) Li, M.; Acero, A. A.; Huang, Z.; Rice, S. A. Nature (London) 1994, 367, 151.
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Figure 4. X-ray scattered intensity as the function of qxy (left) and contour plot (right) for pure F8H16 at 8 mN m-1.
determine the configuration of the FnHm molecules at the air/water interface. GIXD and X-ray reflectivity (GIXR) studies24 on F12H18 supported Gaines’ conclusions that the most probable model was a monolayer with the Fn segments extending up from the surface, the Hm segments being in contact with water. Huang et al.24 insisted that the F12H18 monolayer undergoes slow relaxation, accompanied by transient increasing order and increasing macroscopic inhomogeneity. It was recently suggested, however, that the configuration of FnHm diblocks at the air/water interface may depend on the relative length of the perfluoroalkyl and alkyl segments.26 For F8H18, analysis of X-ray reflectivity measurements suggested a bilayer model in which the diblocks are antiparallel, with tilted F8 segments outward and interleaved H18 segments inward.26 This model is closely related to one of the crystallization models proposed for FnHm in the bulk. These molecules crystallize in a large number of different stable smectic phases, dependent on temperature and on the block lengths n and m.27-30 Results based on X-ray scattering and Raman studies indicated that F12Hm diblocks with m e 6 form monolayer crystals in which the molecules are tilted with respect to the lamella’s surface below the solid-solid transition and perpendicular to the lamella’s surface above the solid-solid transition.28-30 Bilayer lamella crystals were reported to form for diblocks with longer Hm segments.27-29 The observed lamella spacings indicated either a tilted structure or the existence of double-layered lamella with indented Hm segments. An alternative structural model depicted the molecular packing of F12H20 as concentric tubular lamella of uniform diameter.31 In our case, the fact that we have found that the F8H16 monolayer was loosely ordered and that, in contrast, a highly ordered F8H16 bilayer was formed on top of the DPPE monolayer for DPPE/F8H16 ratios < 0.77 (see below) supports the model of a monolayer formed by F8H16 molecules with F8 segments up and H16 segments down. Mixed DPPE/F8H16 Monolayers with Molar Ratios g 0.77: Two Superposed Monolayers at High (26) El Abed, A.; Pouzet, E.; Faure´, M.-C.; Sanie`re, M.; Abillon, O. Phys. Rev. E 2000, 62, R5895. (27) Mahler, W.; Guillon, D.; Skoulios, A. Mol. Cryst. Liq. Cryst. Lett. 1985, 2, 111. (28) Rabolt, J. F.; Russell, T. P.; Twieg, R. Macromolecules 1984, 17, 2786. (29) Russell, T. P.; Rabolt, J. F.; Twieg, R.; Siemens, R. L.; Farmer, B. L. Macromolecules 1986, 19, 1135. (30) Marczuck, P.; Lang, P. Macromolecules 1998, 31, 9013. (31) Ho¨pken, J.; Mo¨ller, M. Macromolecules 1992, 25, 2482.
Pressure. A transition (∼10 mN m-1) is observed on the isotherms of the mixed monolayers (2:1, 1:1, and 1:1.3 molar ratios) (Figure 2). At this point, two features are important to note. First, all the isotherms are reversible (with hysteresis), indicating that no significant amount of amphiphiles, in particular no diblocks, has been solubilized in the sub-phase during compression. Second, when pressure is increased, the difference in area between the DPPE-only monolayer and any of the mixed monolayers, that is, the area occupied by the diblock, decreases, and at high pressure, this difference becomes very small. These two points strongly suggest the formation of multilayers. Figure 5 shows the diffraction patterns and contour plots at 10 and 38 mN m-1 for DPPE/F8H16 (1:1). It can be seen that at low (10 mN m-1, Figure 5a) and intermediate pressure (20 mN m-1, not shown), the presence of the diblock has a strong impact on the ordering of the monolayer. The peak corresponding to the signal of alkyl chains (in the 1.40-1.55 Å-1 range), which may originate either from DPPE chains or from H16 segments, is broader and weaker than in the case of pure DPPE (Figure 3a). The resulting lattice is still rectangular, (Table 2) but the molecules are more vertical (t ) 13.4° at 10 mN m-1 for the mixed monolayer vs 23° for DPPE). This indicates that DPPE and F8H16 are mixed at the molecular level in the monolayer in such a way that the H16 segments of the diblocks are inserted between the DPPE chains, strongly modifying their organization. This leads to a more dense, but less organized packing. At 10 mN m-1, which corresponds to the F8H16 collapse pressure, the diblocks are ejected from the monolayer toward the air phase. At high pressure (38 mN m-1), a peak with strong intensity is observed (1.52 Å-1) (Figure 5b). The lattice formed at high pressure is the same as the DPPE lattice (LS), showing that the diblock is completely expelled from the now DPPE-only monolayer (Table 2). The transition from the NN-tilted to the untilted phase occurs at lower pressure than in pure DPPE (∼25 mN m-1 vs 35 mN m-1). Figure 6 shows diffraction spectra for the (1:1.3) DPPE/ F8H16 molar ratio. The main difference with the above 1:1 molar ratio case is that, at low pressure, the molecules are less tilted (t ) 8° vs. 13.4° at 10 mN m-1), because more diblocks are present in the DPPE. Ejection of F8H16 started at 10 mN m-1. Complete expulsion of the diblocks was also observed at high pressure (Table 2). For molar ratios g 0.77, no peaks were observed for the H16 segments at any pressure, indicating that they are likely in the
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Figure 5. X-ray scattered intensity as a function of qxy (left) and contour plots (right) for the mixed DPPE/F8H16 monolayer (1:1) at 10 mN m-1 (a) and 38 mN m-1 (b). Table 2. Unit Cell Dimensions (a, b) and Projected Molecular Areas (Axy) as Derived from In-Plane Diffraction Data at Different Surface Pressures for DPPE/F8H16 Mixtures in Various Molar Ratiosa
monolayer DPPE/F8H16 (1:1)
DPPE/F8H16 (1:1.3) DPPE/F8H16 (1:2)
a
unit cell dimensions
surface pressure (mN m-1)
a (Å)
b (Å)
molecular area Axy (Å2)
2 10 20 30 38 45 10 20 45 2 10 20 30 38 45
5.13 5.04 4.85 4.80 4.77 4.77 4.96 4.94 4.77 5.00 5.00 5.00 4.85 4.83 4.83
8.51 8.49 8.55 8.32 8.27 8.26 8.26 8.27 8.27 8.79 8.67 8.39 8.36 8.28 8.25
43.7 42.8 41.4 40.0 39.5 39.4 41.0 40.8 39.4 44.6 44.4 41.9 40.5 39.9 39.8
Tilt t (°)
structure
22.0 13.4 13.1 0 0 0 10.0 11.0 0
NN NN NN H H H NN NN H
b b
19.1 13.0 0 0
cross section A0 (Å2) 40.5 41.6 40.4 40.0 39.5 39.4 41 40 39 b b
NN NN H H
39.6 39.5 40 39.8
Tilt angle (t) and cross section (A0) were derived as described in the Experimental Section. b Peak intensity too weak to measure qz.
liquid state. For both (1:1) and (1:1.3) molar ratios, a broad Bragg peak was observed at 1.25 Å-1 at high pressure, showing that the F8 segments forming the upper monolayer, on top of the DPPE monolayer, do present order. The positional correlation length (ξ ) 25 Å) is, however, rather low, and very similar to the ordering of the F8 segments in the monolayer of pure F8H16 (ξ ) 20 Å). As in the case of the monolayer of pure F8H16, the molecules are not, or only very slightly, tilted (t < 10°). At low and intermediate pressures, the peak of the F8 segments is much weaker, suggesting solubilization of F8H16 in the DPPE monolayer. Taking these results into account, we propose that at high pressure a monolayer of F8H16, with ordered F8 segments up and liquid H16 segments down, is formed on top of the hexagonal DPPE monolayer (Scheme 1). In this configuration, owing to their respective cross sections, 1.3 molecules of F8H16 for 1 molecule of
DPPE is the maximal ratio for which a F8H16 monolayer can be accommodated on top of a close-packed DPPE monolayer. Comparing π-A isotherms of the DPPE/hexadecane mixtures studied by Brezesinski et al.32 and the DPPE/ F8H16 mixtures investigated here allows determination of the influence of the F8 segment on the ejection of F8H16. The insertion of hexadecane in the DPPE monolayer resulted in a more compact monolayer, as shown by a reduction of the lipid tilt.32 On the other hand, our results show that F8H16 diblocks disorganize the DPPE monolayer at low pressure, owing to the larger cross section of the F8 segments, which hampers their insertion into the DPPE monolayer. As a consequence, a larger number of F8H16 molecules than hexadecane can be accommodated (32) Brezesinski, G.; Thoma, M.; Struth, B.; Mo¨hwald, H. J. Phys. Chem. 1996, 100, 3126.
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Figure 6. X-ray scattered intensity as a function of qxy (left) and contour plots (right) for the mixed DPPE/F8H16 monolayer (1:1.3) at 10 mN m-1 (a) and 38 mN m-1 (b). Scheme 1. Schematic Representation of a Langmuir Monolayer Made from Combinations of C8F17C16H33 Diblock (F8H16) and DPPE (DPPE/F8H16 molar ratios g 0.77)*
* At low pressure, the two components are partially miscible (a). At high pressure, F8H16 is expelled toward the air phase and forms an additional monolayer on top of a DPPE-only monolayer (b).
in DPPE. Above 10 mN m-1, F8H16 is more rapidly expelled than hexadecane. The strong hydrophobic interactions and low van der Waals interactions that characterize fluorinated chains dramatically increase the tendency of fluorinated amphiphiles to self-assemble in water and to collect at interfaces, displaying interfacial activity.15 The two main driving forces that promote the ejection of F8H16 may be assigned to the decrease in monolayer surface tension33 induced by F8 segments pointing to the air phase and their strong tendency to auto-associate. The ability of semi-fluorinated alkanes to form liquid crystalline phases in bulk27-31 that mainly results from the rigidity of fluorinated chains is also a (33) Reed, T. M. In Fluorine Chemistry; Simmons, J. H., Ed.; Academic Press: New York, 1964; p 133.
strong driving force. It was reported that amphiphiles that exhibit a strong tendency to form smectic LC phase also have a strong tendency to form multilayers.7 Mixed DPPE/F8H16 Monolayers with Molar Ratios < 0.77: A Bilayer on Top of a Monolayer. A second transition appears on the isotherms of the (1:1.5) and (1: 2) DPPE/F8H16 monolayers (20 mN m-1, 60 Å2 molecule-1 and 25 mN m-1, 65 Å2 molecule-1, respectively), in addition to the transition (∼10 mN m-1) already found for molar ratios g 0.77. Here again, the reversibility of the isotherms shows that no diblocks were desorbed upon compression, and the area extrapolated at high pressure corresponds, within experimental errors, to the area of pure DPPE, indicating that the diblocks are expelled upward. Before the first transition (at 2 mN m-1, for example), the diffraction data show that the positional correlation length ξ is larger than in the case of a F8H16 monolayer or of mixed monolayers with molar ratios g 0.77 (ξ ) 40 Å, compared to 20 and 25, respectively), indicating that F8 segments are more ordered, probably because of mixing of DPPE and H16 segments. No additive order was evidenced for the H16 segments. Until 20 mN m-1, by contrast with what occurred for molar ratios g 0.77 (for which all F8H16 were progressively ejected), the DPPE peaks (1.43 and 1.48 Å-1) are strongly depressed (Figure 7a), indicating that DPPE is still disorganized by F8H16 diblocks that have remained on the surface. This is explained by the fact that at ∼25 mN m-1, approximately half of the F8H16 are still on the water surface (molecular area ∼ 68 Å2). Figure 7bc shows diffraction data at 20 and 38 mN m-1. The F8H16 that have remained on the water surface at this pressure are ejected, which allows DPPE to regain some order. As pressure is increased, the peaks shift toward higher qxy values and they are narrower,
A Case of Vertical Phase Separation
Figure 7. X-ray scattered intensity as a function of qxy (left) and contour plots (right) for the mixed DPPE/F8H16 monolayer (1:2) at 10 mN m-1 (a) and 20 mN m-1 (b) and 38 mN m-1 (c).
indicating that the organization of DPPE progressively increases. At 45 mN m-1, DPPE is again fully organized in its hexagonal lattice (Table 2), showing that all the F8H16 diblocks are ejected, as seen in the case of molar ratios g 0.77. Another proof of the diblock expulsion is given by the rod-scan profiles (Figure 8), which show that the thickness of the mixed layer is significantly larger than that of pure DPPE (33 Å vs 21 Å) and that this thickness increases upon compression.
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These observations show that, up to the second transition, the mechanism of ejection of F8H16 is similar to that occurring for DPPE/F8H16 molar ratios g 0.77. The system is similarly composed of a lower DPPE monolayer covered by an upper F8H16 monolayer with the F8 segments being almost untilted. However, when the DPPE/F8H16 ratio exceeds the critical 1:1.3 value, the F8H16 surface concentration becomes too high to allow all the F8H16 diblock molecules to be engaged in one single upper monolayer. The upper monolayer is saturated while there are still F8H16 molecules embedded in the DPPE monolayer. The isotherm (Figure 2) shows that, upon further compression, a second transition occurs, after which pressure rises sharply again. This indicates that there exists another stable phase at smaller molecular area. The F8H16 molecules are forced to form an additional layer on top of the two already existing monolayers. We have investigated the structure of this new layer by GIXD. Figure 7 shows diffraction data for the (1:2) DPPE/ F8H16 molar ratio at 10, 20, and 38 mN m-1. The highly structured pattern of the Bragg peaks of F8 segments (in the 1.0-1.3 Å-1 range) shows that the new layer formed by F8H16 on top of the DPPE layer is highly organized. It can be seen on Figure 7bc that these peaks increase in intensity but do not shift when pressure is increased. This indicates the formation of crystalline self-clustering, the number of clusters increasing with the increased number of F8H16 molecules expelled from the DPPE monolayer. The F8 segments, untilted until 20 mN m-1, are now strongly tilted (t ) 33°). We attribute the four peaks at 1.11, 1.16, 1.22, and 1.27 Å-1 to the F8 segments. They correspond to two different tilted rectangular lattices (with molecules oriented toward their nearest neighbor (NN) or next nearest neighbor (NNN) with a tilt of about 33°). Three peaks, at 1.33, 1.38 Å-1, and 1.52 Å-1 (peak hidden under the DPPE’s peak) are attributed to the H16 segments. The presence of the peak hidden under the DPPE’s peak is inferred from the rod-scan profile (Figure 8) that shows that the film is much thicker (33 Å) for the DPPE/F8H16 mixture than for DPPE (21 Å). The 21 Å thickness corresponds to the calculated length of the 16carbon chain DPPE in an extended, all trans configuration. The 12 Å-difference (that corresponds to 9 CH2) measured for the DPPE/F8H16 mixture is then attributed to nine CH2 units of the H16 segment. These CH2 form a hexagonal lattice (19.5 Å2). The diffraction peaks at 1.33 and 1.38
Figure 8. Rod-scans profiles for pure DPPE at 38 mN m-1 (2), and 1:2 DPPE/F8H16 mixture at 38 mN m-1 ([) and 45 mN m-1 (9).
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Scheme 2. For DPPE/F8H16 Molar Ratios < 0.77*
* The partially miscible mixed monolayer (a, low pressure) undergoes upon compression consecutive vertical phase separation processes that first lead to the formation of a F8H16 monolayer (b, intermediate pressure) and eventually to the formation of a F8H16 bilayer on top of a DPPE monolayer at high pressure (c).
Å-1 would then correspond to the seven remaining CH2, tilted by about 30°. Taking these results into account, we propose a model in which a bilayer of F8H16, with F8 segments pointing outward and interdigitated H16 segments inward, forms on top of the DPPE monolayer (Scheme 2). Approximately nine CH2 of the H16 segments are untilted, while about seven CH2, which are probably those directly connected to the F8 segments, are tilted by about 30°. The tilt of the F8 segments increases the surface area and allows interdigitation of untilted H16 segments, which favors the bilayer structure. This model is reminiscent to that proposed for the crystallization of semi-fluorinated alkanes in bulk.28-30 An alternative model in which H16 segments would point outward and F8 segments would stack inward could be proposed. Although attractive because the a priori unfavorable contact between F8 segments and DPPE chains would be avoided, this model is not consistent with
Krafft et al.
the diffraction data. In this model, it would be necessary that the H16 segments be strongly tilted to be organized, owing to their difference in cross section with F8 segments. Moreover, there would be no sound reason that could explain the tilt measured for the F8 chains. Additionally, this arrangement would not benefit from the more favorable fluorocarbon/air interaction as compared to the hydrocarbon/air interaction. A further possible model would be a trilayer arrangement of diblocks composed of a F8H16 monolayer intercalated between the F8H16 bilayer and the DPPE monolayer. We do not have absolute evidence from the diffraction data to refute it. However, an argument against this model is provided by a close examination of the 1:2 DPPE/ F8H16 isotherm. In this mixture, the number of F8H16 molecules is not sufficient to form a trilayer, that is, a complete monolayer surmounted by a complete bilayer. As a consequence, one would expect the molecular area at the end of the plateau of the second transition to be similar to that of the 1:1 mixture (F8H16 monolayer on top of DPPE monolayer). This is not borne out by the experimental data, the molecular area being about 52 Å2, that is, much larger than the value calculated for the 1:1 mixture (43 Å2). Further work, including the study of DPPE/F8H16 monolayers richer in F8H16, is underway to better understand the mechanism of expulsion of the semi-fluorinated diblock and the structure of the ejected layers. Conclusions Reversible vertical phase separation has been demonstrated in monolayers made from DPPE and F8H16. Depending on DPPE/F8H16 molar ratio and pressure, mixed monolayers, two vertically phase-separated superposed monolayers, or a bilayer on top of a monolayer were obtained. Considering that at high pressure the F8H16 molecules are located at the interface between air and an alkane (represented by the DPPE chains), it can be concluded that crystallization of F8H16 in a bilayer is favored at the air/alkane interface and that water hinders crystallization of F8H16. These results are believed to be relevant to understanding the structure and properties of lipid/FnHm stabilized fluorocarbon-in-water emulsions and other colloidal systems based on these components.15-16 Acknowledgment. The authors thank Prof. J. G. Riess (MRI Institute, Medical Center, University of California at San Diego) for advice. One of us (M.P.K.) gratefully acknowledges Alliance Pharmaceutical Corp. (San Diego, CA) for financial support. LA010587A
