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Langmuir 1997, 13, 2490-2493
Droplet Structure in Phosphocholine Microemulsions Julian Eastoe,* Karen J. Hetherington, and Donal Sharpe School of Chemistry, University of Bristol, Bristol BS8 1TS, U.K.
David C. Steytler School of Chemical Sciences, University of East Anglia, Norwich NR4 7TJ, U.K.
Stefan Egelhaaf ILL, BP 156-X, Grenoble, F-38042 Cedex, France
Richard K. Heenan ISIS Facility, Rutherford Appleton Laboratory, Chilton, Oxon OX11 0QX, U.K. Received November 25, 1996. In Final Form: February 18, 1997X The small-angle neutron scattering (SANS) measurements reported here, for Winsor II microemulsions containing water, phosphocholines (PC’s), and cyclohexane, are shown to give a wealth of information about droplet and interfacial structure. The surfactant chains are either well defined di-C18 cis or trans isomers or a mixture of alkyl groups in naturally occurring Epikuron 200. For each surfactant and concentration three different neutron contrasts (core-shell-drop) were studied, and these data were analyzed together in terms of a Schultz distribution of core-shell particles. Although this is a simple model, it works well, and for the cis and trans isomers, differences in surfactant film thicknesses and oil mixing into the film can be discerned.
Introduction The drive to make biocompatible microemulsions containing natural or synthetic phosphocholines (PC’s) is an important and challenging area.1-7 There have been numerous phase behavior studies1-7 but fewer on the internal structures (e.g. ref 6). This paper describes smallangle neutron scattering (SANS) experiments on some model ternary water-in-oil (w/o) microemulsions consisting of water/PC/cyclohexane and three different surfactants: synthetic C18:1 cis or trans isomers and natural soy bean lecithin (Epikuron 200). The scattering can be accounted for by a polydisperse core-shell particle model. This allows differences in droplet and film structures to be quantified so that natural and synthetic PC analogues can be compared. Initial studies of PC reversed micelle and w/o microemulsion formation were in C6H6 and CCl4 solvents,1 and for a long chain PC (e.g. C18) the solubilization of water w ()[water]/[PC]) is quite small. However, the w/o region can be enlarged by adding medium chain length alkanols. * To whom correspondence should be addressed. E-mail:
[email protected]. Telephone: U.K. +117-9289180. Fax: U.K. +117-9250612. X Abstract published in Advance ACS Abstracts, April 1, 1997. (1) Kumar, V. V.; Raghumathan, P.; Kumar, C. J. Colloid Interface Sci. 1984, 99, 315. (2) Peng, Q.; Luisi, P.-L. Eur. J. Biochem. 1990, 188, 471. (3) Shinoda, K.; Araki, M.; Sadaghiani, A.; Khan, A.; Lindman, B. J. Phys. Chem. 1991, 95, 989. (4) Schurtenberger, P.; Peng, Q.; Leser, M. E.; Luisi, P.-L. J. Colloid Interface Sci. 1993, 156, 43. (5) Shinoda, K.; Shibata, Y.; Lindman, B. Langmuir 1993, 9, 1254. (6) (a) Schurtenberger, P.; Scartazzini, R.; Magid, L. J.; Leser, M. E.; Luisi, P.-L. J. Phys. Chem. 1990, 94, 3695. (b) Schurtenberger, P.; Jerke, G.; Cavaco, C.; Pederson, J. S. Langmuir 1996, 12, 2433. (7) (a) Kahlweit, M.; Busse, G.; Faulhaber, B. Langmuir 1995, 11, 1576. (b) Kahlweit, M.; Busse, G.; Faulhaber, B.; Eibl, H. Langmuir 1995, 11, 4185. (c) Kahlweit, M.; Busse, G.; Faulhaber, B. Langmuir 1996, 12, 861. (d) Aboofazeli, R.; Lawrence, M. J. Int. J. Pharm. 1993, 93, 161. (e) Aboofazeli, R.; Lawrence, M. J. Int. J. Pharm. 1994, 106, 51. (f) Aboofazeli, R.; Lawrence, M. J.; Lawrence, C. B.; Wicks, S. R. Int. J. Pharm. 1994, 111, 63.
S0743-7463(96)02039-2 CCC: $14.00
Shinoda et al.3,5 have shown how n-propanol can be used to tune the phase behavior of water/C18 soybean lecithin/ n-hexadecane systems. A minimum in the oil/water interfacial tension γo/w occurs at about 15 wt % alcohol,5 and this is related to Winsor I f Winsor III f Winsor II transitions with decreasing alcohol content, the same pattern as for anionic surfactants on adding electrolyte.8 The evolution of structure from oil droplets in water (WI), through a bicontinuous region (WIII), to water droplets in oil (WII) was demonstrated by NMR self-diffusion measurements.3 Schurtenberger et al.4 have studied phase behavior, and light scattering (LS), for quaternary water/alkanol/di-CnPC/isooctane w/o systems with m ) 5 f 8 and n ) 5 f 18, where m is the number of carbons in the alkanol. With hexanol a maximum in water solubility as a function of PC alkyl chain length was observed, and at n ) 8 wmax was around 60. For PC chain lengths between n ) 5 and 16 the LS data were consistent with spherical droplets so that the hydrodynamic radius Rh (Å) ∼ 2.0w. This simple behavior contrasts with that of di-C18PC’s, for which the rheology, SANS, and LS data were consistent with long polymer-like micelles at low w, but with spherical droplets close to the wmax WII boundary.6 SANS is useful for resolving microemulsion structure, since different domains can be readily contrasted by selective deuteration of components (e.g. refs 9 and 10). A good way to find a self-consistent picture for the internal droplet and film structure is to analyze data from a coreshell-drop (CSD) contrast series simultaneously.9,10 The key data sets are as follows: core, D-water/H-surfactant/ H-oil (D/H/H); shell, D-water/H-surfactant/D-oil (D/H/D); (8) Aveyard, R.; Binks, B. P.; Clark, S.; Mead, J. J. Chem. Soc., Faraday Trans. 1 1986, 82, 125. (9) Eastoe, J.; Dong, J.; Hetherington, K. J.; Steytler, D. C.; Heenan, R. K. J. Chem. Soc., Faraday Trans. 1996, 92, 65. (10) Eastoe, J.; Dong, J.; Hetherington, K. J.; Steytler, D. C.; Heenan, R. K. Langmuir 1996, 12, 3876.
© 1997 American Chemical Society
Droplet Structure in Phosphocholine Microemulsions
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Figure 1. Scattering length density profiles used in simultaneous analyses of SANS data. The F values and volume fractions for water and cyclohexane were fixed at their known values, whereas the water core radius Rc, film thickness ts, and apparent film scattering length density Ffilm (- - -) were adjusted.
drop, H-water/H-surfactant/D-oil (H/H/D). Together these contain information on the core radius Rc, the effective thickness of the film ts, and the overall droplet radius Rd ()Rc + ts). The apparent film scattering length density Ffilm can also be obtained, and this can tell about any mixing of solvent into the layer.9 For a range of different surfactants, including Aerosol-OT, DDAB, and some PC’s, it has been shown that a sharp-step interface profile (Figure 1) gives better fits than either linear or exponential F(r) gradients.10 Although no evidence was found for a distinct hydration layer, it was possible to pick out the mean level of hydrocarbon associated with the interface (homogeneous mixing). The ways that polydispersity and surfactant/solvent mixing contribute to I(Q), as well as other limitations of the model, have been discussed elsewhere.9,10 Here this global CSD approach is used to investigate ternary Winsor II droplet systems formed by C18PC’s with different chain conformations and structures. This paper expands on previous work10 because natural and synthetic PC’s are compared for the first time and a wider Q-range has been studied. Experimental Section Chemicals. The two synthetic C18:1PC’s, 1,2-dioleoyl-snglycero-3-phosphocholine (cis-PC) and 1,2-di-elaidoyl-sn-glycero3-phosphocholine (trans-PC) were purchased from Avanti Polar Lipids. These isomers (MW ) 786.1) have a CdC bond halfway along the C18 chains. 1H-NMR spectra (Jeol GX270, using CDCl3 as solvent) were consistent with the expected structures. Epikuron 200 (Lucas Meyer soybean lecithin) has a quoted chain composition as follows: n-C16 and n-C18, 16-20%; C18:1 cis, 8-12%; C18:2 cis, 62-66%; C18:3 cis, 6-8%. All the PC’s were stored at -20 °C and used as received. Cyclohexane-d12 and D2O were both >99% D-atom from CDN Isotopes. Cyclohexane-h12 was from Aldrich, and H2O was doubly distilled. Microemulsion phase equilibria were determined as described before.9,10 The Winsor II systems studied here consisted of a w/o microemulsion, at a maximum value wmax, with a small amount (∼5 vol %) of excess water. At 25 °C the wmax values for cis-PC, trans-PC, and Epikuron were 20.0, 22.0, and 22.0 (to (1), respectively. Measurements. The SANS measurements were made on the D17 and/or D22 diffractometers at ILL (Grenoble, France) using neutron wavelengths of λ ) 9.3 and 10.2 Å, respectively. The momentum transfer Q is (4π/λ) sin(θ/2) with θ the scattering angle. The Q-ranges were 0.025-0.29 Å on D17 and 0.00230.49 Å on D22. One millimeter of H2O was used as calibrant, and this gave an absolute intensity I(Q) (cm-1) to within (5%. Accepted procedures were used for the data treatment and background subtraction.11 The neutron beams impinged only on the upper w/o microemulsions (∼95 vol %) which were held at 25 ( 0.2 °C. Some supporting data were taken on the timeof-flight LOQ instrument (ISIS, U.K.). The reproducibility in the absolute values of I(Q) between different instruments and samples was no worse than (10% but typically (5%. (11) Information on SANS data treatment can be found at http:\\www.ill.fr.
Figure 2. SANS data from D22 for the cis-PC surfactant at wmax ) 20.0, φc ) 0.016, and T ) 25 °C. The error bars are shown, and the fit is to a Schultz core-shell model. Analysis. The least squares FISH program used here has been described before.9 The droplets were treated as spherical core-shell particles with a Schultz distribution in core radius Rc.12 By fitting the three contrast data sets simultaneously, only four adjustable parameters are required: the most probable water core radius and polydispersity width, Rcav and σ/Rcav, as well as the apparent film parameters ts and Ffilm. The solvent scattering length densities F (Figure 1) and volume fractions φ were fixed at known values. Literature values for solvent mass densities were used to calculate F, and for surfactants 1.0 g cm-3 was assumed. The amount of water present equals φc, the core volume fraction; adding in the surfactant gives φd for the droplets. Functions to approximate the effects of instrumental broadening (typically ∆Q/Q ∼ 0.08) were also included in the calculations. As the full scattering laws are given elsewhere,9,10 only a summary is needed here. For droplets at concentration φ, radius Ri, volume V, and scattering length density Fp dispersed in a medium of Fm, I(Q) (cm-1) may be written
I(Q) ) φ(Fp - Fm)2[
∑ V P(Q,R ) X(R )]S(Q,R ι
i
i
i
hs,φhs)
(1)
P(Q,Ri) is the single-particle form factor. The Schultz distribution X(Ri)12 is defined by an average radius Rav and a root mean squared deviation σ ) Rav/(Z + 1), with Z a width parameter. S(Q,Rhs,φhs) is the structure factor, and a polydisperse hardsphere model12 was used: the constraints were φhs ) φd and Rhs ) Rdav. Using the approach of Ottewill et al., eq 1 can be modified to allow for sharp-step shells built onto a spherical core.13
Results and Discussion SANS Data. Example SANS data from D22 for cis-PC microemulsions, and the model fits, are shown in Figure 2. The log-log representation helps discriminate between the data sets, especially at high Q. As can be seen, the I(Q) decays are over the intermediate range 0.02 < Q < 0.30 Å-1, meaning that the most important structural features have been well defined. At low Q the scaling of intensity is as expected for the different contrasts. At high Q the apparent increase in background is owing to incoherent scattering from the h-surfactant (shell) and also the H2O (drop). These levels were put into the model as constants by using an appropriate proportion of the scattering from 1 mm of H2O. The differences between the contrasts are further highlighted by Figure 3, which (12) (a) Kotlarchyk, M.; Chen, S.-H.; Huang, J. S.; Kim, M. W. Phys. Rev. A 1984, 29, 2054. (b) Hayter, J. B.; Penfold, J. Colloid Polym. Sci. 1983, 261, 1022. (13) Markovic, I.; Ottewill, R. H.; Cebula, D. J.; Field, I.; Marsh, J. Colloid Polym. Sci. 1984, 262, 648.
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Eastoe et al.
Table 1. Sample Compositions and Values Derived from Simultaneous Analyses of Core-Shell-Drop SANS Data at T ) 25 °Ca wmax cis 20 trans 22 Epikuron 22
a
φc
φd
Rcav/Å
σ/Rcav
2K + Kbar/kBT
ts/Å
10-10Ffilm/cm-2
ΦC6D12
as()3vww/Rc)/Å2
as(Porod)/Å2
0.008 0.016 0.024
0.033 0.065 0.10
27.1 29.6 29.7
0.18 0.20 0.20
1.70 1.40 1.37
17.3 15.6 15.9
0.80 1.01 1.05
0.08 0.10 0.10
66 61 64
51
0.009 0.018 0.027
0.033 0.066 0.098
31.4 31.4 32.9
0.20 0.21 0.21
1.45 1.30 1.27
21.9 26.1 25.6
1.35 1.38 1.34
0.15 0.15 0.15
63 63 60
53
0.009 0.018 0.027
0.033 0.066 0.10
33.3 33.5 36.3
0.18 0.18 0.19
1.68 1.63 1.47
16.5 14.3 16.7
1.12 0.95 0.96
0.10 0.08 0.08
59 59 54
53
Uncertainties: Rcav and ts, (1 Å; as(Porod), (7 Å2; as (3vww/Rc), (1 Å2; ΦC6D12, (0.025; σ/Rcav, (0.01; 2K + Kbar, (0.15kBT.
alkyl chain length as l3.15 The rigidities for the cis and Epikuron (predominantly cis-cis) surfactants seem to be slightly higher than those for trans chains, and this may be due to small changes in oil packing into the film, as discussed below. Surfactant Chain and Film Structures. The fully extended length of a di-C18:1PC molecule is approximately 26 Å.16 For the trans-PC the thickness ts is near this (Table 1), whereas for the two cis surfactants it is consistently lower. Moving from cis-PC to Epikuron introduces a second cis CdC, and this seems to have little effect on ts. Considering the rotational conformations of the isomers, these differences are to be expected, and the trans layers should be the thickest. The fitted values for Ffilm are given in Table 1. Assuming ideal mixing, they have been used to estimate volume fractions for solvent in the film ΦC6D12 ()xVC6D12/V) by solving eqs 3-5 Figure 3. SANS data for the same system as in Figure 2 but displayed as I(Q)‚Q2 vs Q and measured on D17. The fits are to the Schultz core-shell model.
shows D17 data, and fitted lines, on a I(Q)‚Q2 vs Q plot. Although the model is quite straightforward, and given that only four free parameters (Rcav, σ/Rcav, ts, Ffilm) describe three data sets, it provides a remarkably good picture of the droplet and film structure. Table 1 gives the sample compositions and results from the SANS analyses. It can be seen that there is little effect of concentration on the answers, which suggests the droplet structure remains largely unchanged, and S(Q) is well accounted for in the model. The polydispersities can be understood in terms of the film rigidity model of microemulsions. For droplets in dilute Winsor systems Gradzielski et al.14 have recently shown how the Schultz distribution width can be used to calculate a mean film rigidity using eq 2
(2K + Kbar) )
kBT 8π(σ/Rcav)
2
kBT {ln(φc) - 1} 4π
(2)
K is the mean bending modulus, which should always be positive, whilst Kbar is the Gaussian modulus, which for spheres should be negative. The values for 2K + Kbar calculated in this way are in Table 1. As predicted by eq 2, there seems to be a weak dependence on φc, but any changes are generally within the uncertainty of (0.15kBT. The 2K + Kbar values are all very similar, and the mean is 1.47kBT. This is to be expected for equal carbon number surfactants, since theory shows that K scales mainly with (14) Gradzielski, M.; Langevin, D.; Farago, B. Phys. Rev. E 1996, 53, 3900.
Ffilm )
(∑ ) b V
film
(3)
∑b ) (∑b)surf + x(∑b)C6D12
(4)
V ) Vsurf + xVC6D12
(5)
where b is an isotope scattering length, V is an effective molecular volume in the film, and x is the number of C6D12 molecules present in the region ts. Values for ΦC6D12, which measure oil penetration into the curved monolayer, can be found in Table 1. These results suggest that a little more solvent is associated with the trans surfactant than either of the two cis ones. This may well be linked to the chain conformation; the ‘linear’ trans molecules are able to accommodate C6D12 better than the more ‘collapsed’ cis chains. These levels of oil penetration, ΦC6D12 ) 0.08 and 0.15, correspond approximately to between 0.5 and 1.5 C6D12 for each dichain surfactant molecule. Just how good a description of the film is this sharpstep model? Figure 4 shows the shell data and best fits for the three different PC’s together. The calculations reproduce the main oscillatory features in the SANS patterns well and, given the (5% variability in I(Q), are a good match to the absolute intensities. However, the model is not perfect, and this is highlighted by Figure 5, which is a Porod plot, I(Q)‚Q4 vs Q, of example trans-PC data. The most serious discrepancies are above 0.15 Å-1, where the intensities are lowest. In this region two factors (15) Szleifer, I.; Kramer, D.; Ben-Shaul, A.; Gelbart, W. M.; Roux, D. Phys. Rev. Lett. 1988, 60, 1966. (16) Small, D. M. In The Physical Chemistry of LipidssThe Handbook of Lipid Research; Plenum Press: New York and London, 1986; Vol. 4.
Droplet Structure in Phosphocholine Microemulsions
Langmuir, Vol. 13, No. 9, 1997 2493
Figure 4. Shell contrast data from D17 for the three different surfactants and fits to the Schultz core-shell model. φc ) 0.016 for the cis-PC and 0.018 for both the trans-PC and Epikuron samples. The cis-PC and trans-PC data/fits have been multiplied by a factor of 5 and 2, respectively.
Figure 6. Porod plot of core contrast D17 data and fits to the Schultz core-shell model. φc ) 0.016 for the cis-PC and 0.018 for both the trans-PC and Epikuron samples. Error bars are shown for the cis data only.
way are in Table 1. There is little difference between as for the synthetic cis and trans PC’s, but the Epikuron appears to occupy a slightly smaller area, and this may be because it is a mixture with about 4% of the head groups other than pure phosphocholine. These areas compare favorably with those for DDAB and AOT at similar w/o microemulsion interfaces.9,12a Another way for getting as is by using core contrast SANS data at high Q, and the Porod equation for sharp interfaces
{I(Q)‚Q4}Qf∞ ) 2π∆F2Σ
Figure 5. Porod plot of shell contrast D17 data for trans-PC samples and fits to the Schultz core-shell model. Error bars are shown.
conspire together in making the measurement difficult: (a) the inherent (5% uncertainty in I(Q) and (b) the exact subtraction of background. The best efforts have been made here to minimize these two effects, but a ‘perfect’ fit still remains elusive. The model may well be just too simplistic; nonetheless, introducing a step for the headgroups and/or using other types of interfacial profile always makes the fits across the Q-range worse!9,10 Lower residuals can be obtained if the data sets are taken individually, but this always results in a spread of fit parameters, whereas the CSD simultaneous method gives an overall picture and so is more representative. Between the two concentrations there are small changes in the Q values for maxima and minima on Figure 5, and these are linked to an increase in ts of about 4 Å (Table 1). Head Group Areas. The modeling indicates differences in the alkyl chain regions, but what can be said about the polar head groups? Estimates for the mean area per molecule as can be obtained in two ways. For spheres
as )
3vww Rc
(6)
where vw is the volume of a water molecule. With a Schultz distribution width of σ/Rcav ) 0.20 the effect of polydispersity is negligible and so the above equation is still a very good approximation. Values for as calculated this
(7)
where ∆F2 is the contrast step and Σ the total area per unit volume.17 If all N surfactant molecules were adsorbed, it would mean that as ≈ Σ/N. Porod plots for the three surfactants (Figure 6) show that eq 7 is a reasonable approximation over the range 0.175 < Q < 0.225 Å-1. Some values for as that were obtained in this way are also given in Table 1. The agreement here with eq 6 is reasonable given the assumptions involved (monodisperse spheres for eq 6 and sharp-step interfaces for eq 7). However, it is clear that as is essentially independent of alkyl chain type. Conclusions At low volume fractions (
