Langmuir 1994,10,3507-3511
3507
Efficiency of Alkyl Chain Packing in Dialkyl Phosphate Vesicles in Aqueous Solutions Michael J. Blandamer,* Barbara Briggs, and Paul M. Cullis Department of Chemistry, The University, Leicester, LE1 7RH, U.K.
Jan B. F. N. Engberts, Anno Wagenaar, and Elly Smits Department of Organic & Molecular Inorganic Chemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands
Dick Hoekstra Department of Physiological Chemistry, University of Groningen, Bloemsingel 10, 9712 KZ Groningen, The Netherlands
Anna Kacperska Department of Physical Chemistry, University of U d i , Pomorska 18, 91-416 Udi, Poland Received March 14, 1994. I n Final Form: June 14, 1994@ The temperature T m characterizing the gel t o liquid crystalline transition in vesicles formed from dialkyl phosphates increases with an increase in alkyl chain length for phosphates with two identical alkyl groups. The temperature T, also depends on the counter cation. Calorimetric data are interpreted in terms of cooperative melting in domains characterized by patch numbers. The scans recorded by differential scanning microcalorimetry show two or more extrema where the alkyl chain lengths differ.
An interesting characteristic of vesicles in aqueous is the melting temperature T,, associated with a gel to liquid crystalline t r a n ~ i t i o n .Moreover, ~ this temperature is characteristic of the amphiphile forming vesicles and both concentration and nature of added solutes. This temperature is conveniently determined using a differential scanning microcalorimeter,4being the temperature a t which the configurational (relaxational) heat capacity is a maximum. The gel to liquid crystalline transition is further characterized by a standard enthalpy of melting A S and by a patch number n which describes the number of monomers involved in the cooperative melting process. R e ~ e n t l ywe , ~ confirmed the importance of developing a satisfactory protocol for preparing (sodium)di-n-dodecyl phosphate (DDP) vesicles and hence in obtaining reproducible and reversible scans using a differential scanning microcalorimeter. Further, we showed5that dilute solutions of sodium DDP [aq; 8.4 x (monomer mol) dm-31 produce a single extremum in the dependence of heat capacity on temperature where T, = 35 "C, A J P = 3.9 kcal (monomer mol)-l, and patch number n equals 168. In the (sodium) DDP vesicles the phosphate head groups are exposed to the aqueous medium. Sodium ions exist in close proximity to the vesicle-water interface and in the bulk aqueous phase. When the vesicles undergo a gel to liquid crystalline transition, it is therefore anticipated that the melting temperature T, depends on the chain length of the alkyl groups. But such gross reorganization
is accompanied by a change in the organization of head groups, prompting the idea that T, will also depend on the counter cation. In fact, this suggestion is supported by recent fluorescence depolarization studies.6 In the case of vesicles prepared from ( C ~ ~ H ~ ~ O ) Z P OT,~ -depends M+, on the counter cation M+:T, = 66 (Na+),54 (K+),and 39 (Me,") "C, according to the fluorescence studies.6 This pattern is confirmed here using scanning microcalorimetry. In addition,we comment on the associated standard enthalpy of melting. We report that, with an increase in alkyl chain length for vesicles formed from symmetrical monomers, T, and A J P increase whereas the patch numbers decrease. However, when vesicles are formed from nonsymmetric sodium di-n-alkyl phosphates, the scan is more complicated, but overall, T, is shifted to a lower temperature. The latter pattern is observed, for example, in the case of vesicles formed from the monomer where R1 = CloHzl and Rz = C14H29 relative to vesicles formed from monomers where R1 = R1 = C12H25, i.e. for the same total number of carbon atoms in the monomers.
Experimental Section Materials. The phosphate vesicles used in this study have the following general formula RIO
\
P=o
/ \0- M' RzO where we use the following symbolism:
Abstract published in Advance ACS Abstracts, September 1, 1994. (1)Kunitake, T. Angew. Chem., Int. Ed. Engl. 1992,13, 709. (2) Fendler, J. H. ACC.Chem. Res. 1980,13, 7. (3)Carmona-Ribeiro, A.M. Chem. SOC.Rev. 1992,21,209. (4)Blandamer, M.J.;Briggs, B.; Cullis, P. M.; Green, J. A.; Waters, M.; Soldi, G.; Engberts, J. B. F. N.; Hoekstra, D. J.Chem. SOC., Faraday Trans. 1992,88,3431. ( 5 ) Blandamer, M. J.;Briggs, B.; Butt, M. D.; Cullis, P. M.; Engberts, J. B. F. N.; Hoekstra, D.; Mohanty, R. K. Submitted for publication. @
0743-746319412410-3507$04.5010
(i) di-n-dodecyl phosphate, DDP-
= (R, = R,
(ii) di-n-tetradecyl phosphate, DTDP-
= Cl2HZ,)
= (R, = Rz = Cl4H29)
~~
~
(6) Wagenaar, A.; Streefland, L.; Hoekstra, D.; Engberts, J. E.F. N. J.Phys. Org. Chem. 1992,5, 451.
0 1994 American Chemical Society
3508 Langmuir, Vol. IO, No.IO, 1994
Blandamer et al.
Figure 1. Scanning electron microscopyof solutions of DDP (sodium) vesicles prepared by dispersion of monomer in hot aqueous solution by mechanical stirring: magnification 36 K; sample prepared for EM using negative staining with uranyl acetate solution; JEOL JEMlOOCX (Electron Microscope Laboratory, School of Biological Sciences). CalorimetricMeasurements. Following preparation of the (iii)di-n-hexadecyl phosphate, DHDP- = solution and loading into the cell of the microcalorimeter, the (R1= R, = C16H33) solution was held at temperature TIfor at least 1h, the scan was recordedfrom to Thigh, the solution was cooled to temperature (iv) di-n-octadecyl phosphate, DODPTI,,, and the scan was again recorded from TI,, to Thigh. The solution was then cooled to temperature TI,, and held at that temperature for a time t. The scan was then again recorded from Tiowto Thigh. (v)n-decyl-n-tetradecyl phosphate, DETDP- = (a)For DDP-M+,DTDP-M+,and DHDP-Me&J+ solutions, 2'1 (R, = C10H2,and R, = C14Hm) = 60 "C. (b) For DHDP-K+ and DHDP-Na+ solutions, TI= 70 "C. (c) For DODP-Na+ solutions, TI= 75 "C. (vi) n-decyl-n-octadecyl phosphate, DEODP- = (d) For K+and Na+ alkyl phosphates, TI,, = 15 "C and Thigh (R1= ClOH21and R, = c&j7) = 90 "C, where t > 6 h. The preparation and purification of the above alkyl phosphates (e) For DDP-Me&J+,TI,, = 2 "C and Thigh = 90 "C, where t > were described previously.6-8 The essential step involved 6 h. alkylation of the appropriate alkyl dihydrogen phosphate. As (f) For DTDP-Me&J+,,I'! = 10 "cand Thigh = 90 "c,where described in ref 7, the yields were satisfactory,being for the most t>6h. part greater than 70%. (g) For DHDP-Mea+, Tiow= 15 "C and Thigh = 90 "C, where Vesicle Preparation. The appropriate mass of dialkyl t = 11h. phosphate was dissolvedin water (2.2 cm3)to produce a solution (h) For DETDP-Na+, TI= 60 "C, TI,,= 2 "C, and Thigh = 90 where the concentration was 8.4 x (mol monomer) dm-3. "C, where t = 6 h. In all cases, the aqueous solutions were prepared by dissolving (i)For DEODP-Na+,I'! = 5 "C, TI,,= 5 "C, and Thigh = 90 "C, the solid surfactant in water at a temperature r" with stirring where t = 3 h. for approximately 10 min: for DDP-M+, DTDP-M+ (where M+ Analysis of Calorimetric Data. For a system in which a = Na" and K&),DDP-Me&J+,DTDP-Me4N+, DHDP-Me&J+, chemicalequilibrium existsbetween substancesXand Y,namely DETDP-Na+, r" I60 "C; for DHDP-K+, DHDP-Na+, r" I70 X Y, the dependence on temperature of the relaxational "C; for DODP-Na+, F I75 "C. The "hot water + stirring" (configurational) heat capacitylO forms a bell-shaped plot if it procedure was shown using electron microscopy Figure 1in the can be assumed that the partial molar heat capacities of X(aq) case of, for example, DDP(Na+)to yield spherical vesicles rather and Y(aq)are negligiblysmall. The dependenceon temperature than ill-defined bilayer structures. The reproducibility of the of the molar relaxational heat capacity is given4J1by eq 1. DSC scans offered supporting evidencefor well-definedstructures C,,,,(aq; config) = A,,,w(c~) A J W V H ) HRP(I [11 which undergo characteristic thermal transitions. Scanning Calorimetry. An MC-2 differential scanning where K is the temperature-dependent equilibrium constant. microcalorimeter (MicroCal Ltd.) recordedgthe dependence on The area under the "bell" is, for dilute solutions, a directmeasure temperature of the differential isobaric heat capacities of vesicle of the calorimetric standard enthalpy of melting A,H"(cal). The solutions relative to that of water. The volumes of solutions and maximum in Cp,m (aq; config) at T m , where K = 1, yields an reference were approximately 1.2 cm3. The aqueous solutions estimate of the van't Hoff standard enthalpy of melting, AJ-Pwere loaded into the sample cell of the calorimeter and scanned (vH). In the cases considered here, symbolsXand Y refer to two against water as a reference at a scan rate of approximately 60 statesof the vesicles-gel state and liquid crystalline, respectively. "C h-l. Although the monomer concentration is known a priori, the number of monomers involved in the transitions, the patch (7) Wagenaar, A; Rupert, L. A. M.; Engberts, J. B.F. N.; Haekstra, number, is unknown. However, an estimate of this number is D.J . Org. Chem. 1989,54,2638.
.-
+ m2
(8) Streefland, L.; Yuan, F.; Rand, P.; Hoekstra, D.; Engberts, J. B. 1992,8, 1715. (9) Blandamer, M. J.; Briggs, B.;Burgess, J.; Cullis, P. M.; Eaton, G. J . Chem. Sm., Faraday Trans. 1991,87, 1169.
F.N. Langmuir
(10) Prigogine, I.; Defay, R. Chemical Thermody"zmics; Longmans Green: London, 1954; p 293, (trans. Everett, D. H.). (11) Sturtevant, J. M. Annu. Rev. Phys. Chem. 1987,38,463.
Alkyl Chain Packing in Dialkyl Phosphate Vesicles 0.08
1
IC1
20
Langmuir, Vol. 10, No. 10, 1994 3509
I
60
40
Temperature/Celsius
TemperaturelCelsius
Figure 2. Dependences on temperatureof the differentialheat
capacities (reference= water) for aqueous solutions containing vesicles prepared from dialkyl phosphates [aq; 8.4 x (monomer mol) dm-31; sodium salts formed by (a) DDP-, (b) DTDP-, and (c) DHDP-. [For clarity the scans have been displaced on the heat capacity axis.]
Figure 4. Dependence on temperature of the configurational isobaric heat capacity C,,(aq; config) for DDP-K+ [aq; 8.4 x
(monomer mol) dm-31; comparison between observed and calculated dependencesusing patch number n equal to 153 and A,H”(cal)equal to 670 kcal (patch mol)-’ or 4.4 kcal (monomer mol)-l. 200
150 C
b n
5
Z
c
2 n
100 M,N+
50 DDP~
DTDP~
DHDP~
DODP~
1
I
4
I
12
14
16
18
Carbon atoms in each alkyl chain
Figure 3. Dependences of melting temperature T,,, characterizing the gel to liquid crystalline transitions on alkyl chain length for systems where the counter cations are Na+, K+, and
Mea+.
obtained through the data analysis by effectively seeking agreement between the two standard enthalpies, AI1Jl”(cal) and A a ( v H ) . Therefore, the ratio of the standard enthalpy of melting AI1Jl”(cal)to the patch number yields the standard enthalpy of melting per monomer on going from gel to liquid crystalline states.l
Results In the followingpresentation of our results we describe the scans recorded by the differential scanning microcalorimeter and the results of analyses based on eq 1.We have previously shown that some of the generally accepted methods for preparing solutions of vesicles show variable and irreversible behavior in studies by differential scanning microcalorimetry. In this regard DSC provides a particularly critical criterion of a defined and reproducible system. The most convenient and reliable method of preparation of vesicle solutions involves dispersion into hot aqueous solution with mechanical stirring and is described above in the Experimental Section and in our
v\ DDP~
12
,-\,
DTDPO
14
DODP~ DHDP~
18
16
Carbon atoms in each alkyl chain
Figure 5. Dependences of patch numbers characterizing the gel to liquid crystalline transitions on alkyl chain length for systems where the counter cations are Na+, K+, and Me4N+.
previous publication^.^ Reproducibility alone does not confirm the presence of vesicles; however, independent studies by electron microscopy have shown that, for the different systems that we have studied (DOAB,dialkyl phosphates), closed vesicle structures of relatively uniform size are formed. Figure 1shows the EM for sodium dodecyl phosphate (DDP). A typical set of scans for three sodium dialkyl phosphates is shown in Figure 2 a t a common monomer concentration, 8.4x (monomer mol) dm-3. In each set the scans are reversible following the heatcool-heat-cool ... cycles described in the Experimental Section. This reproducibility of the scans was a common feature for all the systems reported here, supporting the assignment of the transitions to well-defined vesicular structures. Furthermore, the reproducibility ofthe results for independent, freshly prepared samples further supports this protocol as the method of choice. The melting temperatures characterizing these systems are summarized in Table 1 and in Figure 3. The T,,, for the sodium-based systems is slightly higher than for the systems where the counter cation is K+.There
Blandamer et al.
Langmuir, Vol. 10, No. 10, 1994
Table 1. Characteristic Melting Temperatures Tm (Recorded in Celsius) for Vesicles Formed in Aqueous (monomer mol) dm-s) by Dialkyl Solution (8.4 x Phosphates
/*
phosphate anion
DDP DTDP DHDP
DODP DETDP
DEODP a
DTDP~
DDPO
12
DHDP~
DODP~
I
I
I
14
16
18
I
Carbon atoms in each alky chain
Figure 6. Dependences of standard enthalpies of melting characterizing the gel to liquid crystalline transitions on alkyl chain length for systems where the counter cations are Na+, K+, and Me4N+.
is a marked shift in T, to lower temperature when the counter cation is Me&+ for systems containing DDP-, DTDP-, and DHDP- vesicles. Similarlyfor a given counter cation, there is a shift in T , to lower temperatures with a decrease in the number of carbon atoms in each alkyl chain within the symmetrical di-n-alkyl phosphates. The dependence on temperature of the molar configurational isobaric heat capacity C,, (aq; config) for DDP-K+(aq) is satisfactorily accounted for using eq 1 (Figure 4). Similar satisfactory agreement between experimental and calculated curves was obtained for eight systems, leading to the patch numbers and enthalpies of melting summarized in Figures 5 and 6, respectively. In both cases, the derived parameters for K+ and Na+ systems are slightly different. For a given cation, the patch number
Na+ 34.8 f 0.2 52.2 f 0.1 66.3 3z 0.1 11.1 0.2 14.4 f 0.1 20.9 f 0.3
*
cation K+
Me4N+
33.3 0.1 52.0 i 0.1 65.2 & 0.1
12.1 f 0.1 29.4 f 0.1 a
Two maxima at 48.5 and 53.8 "C.
decreases and the enthalpy change per monomer increases with an increase in the number of carbon atoms in each of the dialkyl chains. Turning to the tetramethylammonium di-n-alkyl phosphates, only for the DDP-Me4Nf (aq) system was it possible to analyze the data using eq 1. In the case of DTDP-Me4N+ (aq), T, shifted to higher temperatures (Figure 3), but the dependence of dC, on temperature was not bell-shaped. This hint of complexity was borne out by the even more complicated dependence of heat capacity on temperature recorded for DHDP-Me4N+(aq). In the latter case, two rather strangely shaped components were recorded near 50 "C. Nevertheless, the scans produced by the symmetric sodium di-n-alkyl phosphates were relatively straightforward in the sense of producing a single extremum. It was interesting, therefore, to discover that the scans for asymmetric sodium di-n-alkyl phosphates showed two features (Figure 7). Replacing the two Clz chains in DDP-Na+ with Cl0 and C14 chains to form DETDP-Na+ causes the scan with the single T , a t 34.8 "C to be replaced (Figure 7 and cf. Figure 1)by a scan with two close extrema a t 14.3and 15.5"C. Similarly, replacingthe two C14chains in DTDP-Na+ with Clo and C18 chains causes a single T, to be replaced by two close extrema at 20.7 and 22.0 "C. In both cases, the scans are fully reversible. At temperatures over the range 40-90 "C, there are no other extrema in the dependence on temperature of the differential heat capacities.
Ib 0.010
c
I
y
.
o
(d
0
8
-O.O4
-0.08 -0.010
I
20
t-
is
20
-
40
Temperature / Celsius Temperature/ Celsius Figure 7. Dependences on temperature of the differential isobaric heat capacities (reference = water) for aqueous solutions containing vesicles prepared from sodium dialkyl phosphates [aq; 8.4 x (monomer mol) dm-31. Scans recorded for (a) DETDP-Na+(aq)where T, = 14 "C and (b) DETOP-Na+(aq)where T, = 21 "C.
Alkyl Chain Packing in Dialkyl Phosphate Vesicles
Discussion The key property ofvesicles probed in the study reported here is the gel to liquid crystalline transition a t temperature T,. An important discovery concerns the underlyingpatterns described in Figures 3,5,and 6, which are derived from studies such as those shown in Figure 2. The model which emerges envisages a patchwork structure of vesicles in which the melting process involves a group of n (patch number) monomers. Interestingly, increasing the chain length leads to a n expected increase in the standard enthalpy of melting (Figure 6) but a decrease in the patch number; i.e. the size of the cooperative unit decreases with an increase in chain length, possibly related to vesicle size. The data also show that T, and related parameters are sensitive to the counter cation. The changes in these parameters show that reorganization of the alkyl chains during the melting process is sensitive to the replacement of Na+ or K+ by Me4N+ in the boundary layers. The dramatic change in T,, n, and A X when Na+ is replaced by Me4N+ in the DDP- system is attributed to the modest hydrophobic character of M e a + cations which might penetrate into the surface of the vesicular aggregate. The scans for the DTDP-Me4N+system are not reported here; suffice it to say they are complex and not reversible. We
Langmuir, Vol. 10, No. 10, 1994 3511 surmise that for this system the patch number n is perhaps not constant because penetration of the cations into the vesicle surface is not constant across the vesicle and is dependent on temperature. The dependenceon chain length for Na+and K+ systems is consistent with a pattern in which n decreases, A f l increases, and T, increases with an increase in chain length. The gross shift in T, to lower temperatures when the alkyl chains in the same surfactant have different lengths points to the importance of satisfactorypacking12J3 of these chains in controlling the melting process. For the CldCls and C1dC14 systems it is tempting to assign the two extrema (Figure 7) to two processes or melting in two different types of patches. The overall standard enthalpy change calculated from the integrated area is smaller for the CldC14 than for the C1dC12 systems.
Acknowledgment. We thank the SERC for their support through the Molecular Recognition Initiative. We thank the British Council for a grant to A.K. and the University of E6di for granting a leave of absence for A.K. (12)Jaeger, D. A.; Subotkowski,W.; Mohebalian, J.; Sayed, Y. M.; Sanyal, B. J.; Heath, J.; Amett, E.M. Langmuir 1991,7, 1935. (13)Harvey, N.G.;Rose, P. L.; Mirajovsky, D.; Amett, E. M. J.Am. Chem. SOC.1990,112,3547.
