2112
Langmuir 1993,9, 2112-2118
Comparison of Enantiomeric and Racemic Monolayers of N-Stearoylserine Methyl Ester by Fluorescence Microscopy Keith J. Stine,' Jack Y.-J. Uang,and Sean D. Dingman Department of Chemistry, University of Missouri-St. Louis, St. Louis, Missouri 63121 Received February 17, 1993. In Final Form: May 14, 1993
Monolayersof L-, D-, and racemicN-stearoylserine methyl ester (SSME)were examinedusing fluorescence microscopy and surface pressure isotherm measurements. Fluorescence microscopy reveals distinct differences in the condensed phase domain structures between the enantiomeric and racemic monolayers. The racemic SSME monolayer exhibits isotropic growth of the condensed phase, but with the formation of counterclockwise and clockwise hooks at the periphery of the domains. In contrast, the condensed phase of the L-SSME or D-SSME monolayers exhibits dendritic growth. The formation of condensed phase domains with the macroscopic curvature dependent upon the headgroup chirality is also observed. In the racemic monolayer, domains with both clockwise and counterclockwise curvature are observed; however, the L-SSME monolayer shows only domains with counterclockwise curvature and the D-SSME monolayer shows only domains with clockwise curvature. Fluorescence anisotropy is observable within the condensed phase at lower temperature, suggesting the presence of long-range tilt order.
Introduction The oriented two-dimensional geometry of a Langmuir monolayer1 provides an opportunity to study the dependence of molecular packing and phase behavior on molecular interactions. The importance of chiralitydependent interactions is manifested in numerous areas of biochemistry and analytical chemistry. Studies of mixed monolayers of the D and L enantiomers of a surfactant with a single chiral carbon in the headgroup allow for investigationof chiral discrimination effects.2 Such effects are well-known for the solid state for melting points and enthalpies of f ~ s i o n . The ~ phenomenon of chiral discrimination arises from a nonequivalence of the interaction potential between two molecules of the same chirality (D:D or L:L) versus that between two molecules of opposite chirality (D:L). If the D:D or L:L interaction is more favorable than the D:L interaction, one has homochiral discrimination, while a more favorable D:L interaction indicates heterochiral discrimination. In monolayers, chirality-dependent differences are directly observable as differences in the surface pressure versus area per molecule isotherms of enantiomeric as compared to racemic monolayers. Differences arise at temperatures and surface densities where the monolayer exhibits a condensed phase in which the surfactant headgroups are packed closely and chirality-dependent interactions become significant. The differences in the surface pressure isotherms imply differences in all the thermodynamic and macroscopic properties of the monolayer, and would seem to inevitably arise from structural differences in the condensed phase at the molecular level. A statistical mechanical model for surfactants with a single chiral center near the headgroup has been pre~ented.~ In this model, the hydrocarbon chain points away from the surface, while the chemically distinct groups A, B, and C of the headgroup are arranged on the water surface at the corners of a triangle. The N-acyl derivatives of amino acids and amino acid methyl esters are readily prepared5 amphiphiles for (1) Gaines, G. L. Insoluble Monolayers at Liquid-Gas Interfaces; Wiley: New York, 1966. (2) Stewart, M.; Amett, E. M. In Topics in Stereochemistry; Eliel, E. L., Allinger, N. L., Eds.; Wiley: New York, 1982; Vol. 13. (3) Jacques, J.; Collet, A.; Wilen, S. H. Enantiomers, Racemates, and Resolutions; Wiley: New York, 1981. (4)Andelman, D. J. Am. Chem. SOC.1989,111,6536. (5) Zeelen, F.J.; Havinga, E. Red. Trav. Chim.Pays-Bas 1958,77,267.
0743-746319312409-2112$04.OO/Q
investigating chirality-dependent effects in monolayers. Interestingly, extensive studies comparing the properties of enantiomeric and racemic phosphatidylcholine monolayers did not reveal any thermodynamic differences! although striking differences in domain structures were observed, especially when mixed with cholesterol.7 A number of isotherm studies of surfactants with amino acid headgroups have revealed significant chiral discrimination.2>@-11 Arnett and co-workers investigated monolayers of N-stearoylserine methyl ester (SSME) using surface pressure measurements, surface shear viscosity measurements, and some microscopy techniques.12 Their study included a pair of images of an enantiomeric and a racemic monolayer of SSME obtained by using fluorescence microscopy, as well as a pair of transmission electron micrographs of collapsed Langmuir-Blodgett (LB) films, and a pair of scanning tunneling micrographs of collapsed LB films. Surface pressure isotherms showed the racemic SSME monolayer to be more expanded than the enantiomeric monolayer. This indicates that the SSME monolayer exhibits homochiral behavior. Compressionexpansion cycles through the transition to the condensed phase revealed substantial hysteresis. The STM images of the LB films of collapsed domains showed the enantiomeric film to have a striated pattern which was absent in the racemic film. Studies of mixtures of SSME with palmitic acid showed that the chiral discrimination was only evident when the mixture was more than 50% SSME.13 In this paper, we present a more detailed study of the domain morphology of the condensed phases in monolayers of SSME over a range of temperature. As SSME monolayers exhibit homochiral behavior in their surface pressure (6) Amett, E. M.; Gold, J. M. J. Am. Chem. SOC.1982,104, 636. (7) McConnell, H. M. In Annual Reviews of Physical Chemistry; Straws, H. L., Babcock, G. T., Leone, S. R., Eds.; Annual Reviews Inc.: Pal0 Alto, CA, 1991; Vol. 42, pp 171-195. (8)Marr-Leisy, D.; Neumann, R.; Ringsdorf, H. Colloid Polym. Sci. 1985,263,791. Dupeyrat, M. Biochim. Biophys. Acta 1988, 938, (9) Bouloussa, 0.; 395. (10) Harvey, N. G.; Rose, P. L.; Mirajovsky, D.; Amett, E. M. J.Am. Chem. SOC.1990,112,3547. (11) Heath, J. F.;Amett, E. M. J. Am. Chem. SOC.1992, 114, 4500. (12) Harvey, N. G.; Mirajovsky, D.; Rose, P. L.; Verbiar, R.; Arnett, E. M. J. Am. Chem. SOC.1989, 111,1115. (13) Harvey, N. G.; Arnett, E. M. Langmuir 1989,5, 998.
1993 American Chemical Society
Langmuir, Vol. 9,No. 8, 1993 2113
Monolayers of N-Stearoylserine Methyl Ester
isotherms, we were particularily interested in determining if evidence for chiral segregation could be observed in the racemic monolayer by direct observation of the condensed phase domains. We report here evidence for chiral segregation obtained by using fluorescence microscopy. In addition to our microscopy observations of this and other domain phenomena, surface pressure isotherm measurements are presented.
25
Experimental Section 1. Fluorescence Microscopy. A Teflon trough was fitted onto the stage of an Olympus BH-2 microscope attached to a Dage-MTI SIT-68 camera. The monolayers were observed through 20X and 40X ultralong working distance objectives (Olympus CDPLANPO and CDPLAN40). A Panasonic UP-850 video printer was used to obtain copies of monolayer images. Temperature was measured by a fine thermocouple in a Teflon sheath positioned just below the water surface. The trough coverglasswas conductivelycoated (Delta Technologies)and was heated by a Variac to burn off condensation. The monolayer area was adjusted by moving two small Teflon barriers which fit underneath the coverglass. Fluorescence was excited by either a mercury lamp or the 488-nm line of an Omnichrome multiline multimode argon ion laser. 2. Isotherm Measurements. Isotherms were measured in a Teflon trough enclosed in a Plexiglas housing. Surface pressure was measured using a Cahn Model 25 electrobalance using filter paper as the Wilhelmy plate. The BCD output from the electrobalance was read by a PCL-720 32 bit digital 1 / 0 card (B&C Microsystems) in a 386 computer. A Teflon barrier on a threaded rod attached to a universaljoint was driven by a stepper motor using a home-built circuit. During an isotherm run,force versus time data were recorded and subsequently converted to surface pressure versus area per molecule. 3. Compound Preparation. D,L-Serinemethyl ester, L-serine methyl ester, and stearoyl chloride (97 %) were obtained from Sigma Chemicals (St.Louis, MO) and used as received. &Serine methyl ester was obtained from Advanced Chemtech (Louisville, KY) and used as received. The procedures given by %elens and Arnett12 were followed. The structures of the resulting compounds were checked by '9c and proton NMR using a Varian XL-300 instrument. IR spectra were recorded with a PerkinElmer 1600 series FTIR. L-SSME L-serine methyl ester hydrochloride (1.23 g) was dissolved in a solution of 1.4 g of potassium carbonate in 12 mL of water and 10 mL of chloroform was added. A solution of 2.00 g of stearoyl chloride in 12 mL of chloroform was added and stirring was continued for 1h. The chloroformlayer was separatd and dried and the solvent removed by rotary evaporation. The remaining solid was recrystallized 3 times from pentane/ chloroform (53), yielding 2.05 g of product. The melting point of the L compound was 89.5-90.5 "C. The characteristic IR absorption bands were (in cm-1) 3540 (OH), 1725 (ester C-0), 1652 (amide band I), and 1550 (amide band 11). D-SSME: The same procedure as used to prepare the L-SSME was followed using D-serine methyl ester hydrochloride ae a starting material. The melting point of the D compound was 89.5-90.3 "C, and the same IR bands as for the L compound were observed. D,L-SSME D,L-Serine methyl eater hydrochloride (0.918 g) was dissolved in a solution of 1.4 g of potassium carbonate in 10 mL of water and 10 mL of chloroform was added. A solution of 1.0 g of stearoyl chloride in 10 mL of chloroform was added and stirring was continued for 1h. The chloroformlayer was separated and dried and the solvent removed by rotary evaporation. The remaining solid was recrystallized 3 times from ethyl acetate yielding 1.65 g of product. The melting point of the racemic compound was 98.0-99.0 "C. The characteristic IR bands were (in cm-1) 3485 (OH), 1736(ester C=O), 1630(amide band I),and 1550 (amide band 11). 4. Monolayer Preparation. The compounds were dissolved in 9 1 hexane/ethanol at concentrations of (0.66-1.0) X 109 M. Solutions for microscopy contained 0.3-1.0 mol % of the fluorescentprobe 4-(hexadecylamino)-7-nitrobenzoxa-1,3-diazole
O L 20 24
28
32
36
40
44
48
52
SQUARE ANGSTROMS PER MOLECULE 35,
,I
20
30
,
,
,
,
,
I
I
40
50
60
70
80
90
100
SQUARE ANGSTROMS PER MOLECULE 3 5 , ,
,
,
1
,
I
I
I
I
30
40
50
60
70
80
90
100
"
20
SQUARE ANGSTROMS PER MOLECULE
Figure 1. Surface pressure versus area per molecule isotherm measurements for stearoylserine methyl ester on pure water: (A) 20 OC; (B)24 "C;(C) 31 "C. The solid l i e represents the data for the racemic monolayer, while the dashed line represents the data for the monolayer of the L enantiomer. The isotherms were measured on compression followed by expansion at a rate of 26 A2 molecule-' min-1, except for the racemic f i at 24 "C, for which the rate was 4 AZmolecule-' min-I. obtained from Molecular Probes (Eugene, OR). Monolayers of the probe itaelfwerefound by fluorescencemicroscopy and surface pressure isotherm measurements to undergo a direct transition from the gas phase to a condensed phase in the temperature range of these studies. The monolayers were spread from a 25p L microsyringe onto a MilliQ water subphase.
Results 1. Isotherms. Figure 1 shows isotherms measured on compression/expansioncycles at 20 "C,24 OC,and 31 OC for enantiomeric L and racemic monolayers of SSME. Isotherms of D-SSMEat these temperatures were equiv-
Stine et al.
2114 Langmuir, Vol. 9, No. 8, 1993 40
I
I
I
I
1
time (sec)
Figure 2. Surface pressure relaxation observed for monolayers of stearoylserine methyl ester a t 31 "C. The surface pressure was monitored in each case after halting a compression carried out a t 26 A2 molecule-' min-1.
d e n t to those of L-SSME. The transition pressures and the magnitude of the hysteresis curves observed are consistent with those previously reported.12 We believe that our compounds are of the same high purity as those used in the work of Arnett and co-workers. Surface pressure relaxation is observed for both the enantiomer and racemic monolayers. If the compression of the monolayer is halted above a temperature-dependent stability limit,12then relaxation of the surface pressure is observed as shown in Figure 2. The time dependence of these curves indicates a faster relaxation for the surface
pressure of the racemic monolayer than for the enantiomeric monolayer. The surface pressure of the enantiomeric monolayer becomes close to flat later than the curve for the racemic monolayer. An isotherm measured on the racemic monolayer a t 25 "C a t our slowest available compression/expansion rate of 4 A2 molecule-' min-1 exhibits hysteresis of the same magnitude as that observed at a rate of 25A2molecule-' min-l. Isotherm measurements at slower compression/expansion rates would clearly be of interest to further study the hysteresis. The SSME monolayers were found to collapse on compression to a surface pressure above 40 dyn cm-l. 2. Fluorescence Microscopy. At 25 "C, the racemic monolayer is in the LE + G coexistence region on deposition a t an area of 150 A2molecule-l. On compression, the monolayer first enters the uniformly fluorescent LE phase. Further compression results in the formation of dark condensed phase domains. Compression to low areas shows a surface covered by LC phase domains with some small drops of LE phase, which may be probe or impurity enriched, between the LC phase regions. Further compression into the collapsed regime shows striated patterns under the microscope. Compressionswere carried out a t a rate of about 15 A2 molecule-' min-l and the monolayer observed with the barrier stopped. Very similar domains were observed on cooling from the LE phase at 0.5 "C min-l. The domains nucleate and grow as shapes resemblingthe tip-splitting type of fractal growth observed in Hele-Shaw cell experiments,14as seen in Figure 3a. The growth of such a shape indicates an isotropic line tension between the racemic LC phase and the LE phase.I5 The
Figure 3. Growth of condensed phase domains observed on compression of racemic SSME monolayer from the LE phase a t 24 OC, 50 A* molecule-': (a) initially observed fractal growth; (b) counterclockwise curved hook formed at periphery of a condensed phase domain; (c) clockwise curved hook formed a t the periphery of a condensed phase domain; (d) filament-like structures evolved from curved hook after 10 min. The monolayer contains 0.3 mol % fluorescent probe. The bar in the lower left represents 250 pm.
Monolayers of N-Stearoylserine Methyl Ester
Langmuir, Vol. 9, No. 8, 1993 2115
&
P
I
Figure 4. Fluorescence micrographsof SSMEcondensed phase domains observed to form on a second compression of a racemic SSME monolayer after the initial compression in which the domains shown in Figure 3 were observed: (a) domains which curve both clockwise and counterclockwise;(b) domains which appear to exhibit twinning. Images taken at 24 "C, 35 A2 molecule-'. The monolayercontains0.3mol 96 fluorescentprobe. The bar in the lower left represents 250 pm.
Figure 5. Fluorescence micrographs of L-SSME monolayers: (a) dendritic growth at 38 "C, 38 A2molecule-' where the barrier motion is stopped as soon as the domain nucleation is observed; (b) dendritic growth at 38 "C as seen when the initial monolayer compressionstops at 33 A2molecule-'. The monolayer contains 0.3 mol 96 fluorescentprobe. The bar in the lower left represents 250 pm.
size shown in Figure 3a, after which growth is observed a t the periphery of the domains in the form of small hooks (Figure 3b,c) which increase in size and ramify into filaments (Figure 3d) over a period of about 5 min. The hooks curve clockwise or counterclockwise in apparently equal numbers. Domains which did not have curved hooks form on their periphery anneal over 10-20 min toward a compact shape,while the filament structures do not anneal. The majority of the domains exhibit the formation of hooks around the periphery. Many of the domains which nucleate and grow on further compression of the monolayer appear as the curved shapes shown in Figure 4a, which resemble the hooks that nucleated around the periphery of the large region of the domains. Other domains that form appear to exhibit twinning, with two arms curved in opposite direction, as shown in Figure 4b. Domains such as those in Figure 4b will anneal slowly and develop a fringe similar to the filaments shown in Figure 3d. A slower compression rate of about 4 A2 molecule-' min-I shows domains like those in Figure 4 to form during the first appearance of domains along with domains like those shown in Figure 3. The same phenomena are also observed near 30 "C in the racemic monolayer. Observations a t slower controlled rates would be informative and will be
a part of our study of domain behavior as a function of enantiomeric composition in D-SSME and L-SSME mixtures followingtechnical improvements in our microscope trough. Compressionof monolayersof L-SSMEor D-SSMEfrom the LE phase results in tip-stable or dendritic type of fractal growth14J6as shown in Figure 5. Tip-stable growth requires an anisotropy in the line tension of the LC ~ h a s e , ' ~which J ~ is found here for an enantiomeric SSME monolayer. The dendrites nucleate and grow with long straight main axes if the monolayer compression is stopped at the point where the dendrites are first observed. If the monolayer is compressed past this point during the initial growth, then the dendrites will tend to be curved (Figure 5b). The dendritic shapes anneal very slowly,and over an hour the sidearms shrink toward the main axis so that the .side of the dendrite looks like the edge of a saw blade. Compressionalso shows the formation of individual curved LC domains, as seen in Figure 6. These domains are found to curve clockwise in the D-SSME monolayer (Figure 6a) and counterclockwise (Figure 6b) in the L-SSME monolayer. In Figure 6c, examples of L-SSMEdomains with more than one arm are shown. Experiments in which the enantiomeric SSME monolayer was warmed slowly (0.2 "C min-l) from below the triple point at about 20 "C into the coexistence region, or
(14) Langer, J. S. Science 1989,243,1150. (15) Suresh, K. A.; Nittmann,J.; Rondelez, F. h o g . Colloid Polym. Sci. 1989, 79,184.
(16) Akamatau, S.; Boulosea, 0.;To, Kiwing; Rondelez, F. Phys. Reu. A 1992,46,4504. (17) Martin,0.;Goldenfeld, N. Phys. Rev. A 1987,35,1382.
LC domains expand in size over about 30 s and reach the
2116 Langmuir, Vot. 9, No.8, 1993
Stine et at.
Figure 7. Fluorescence micrographs of SSME monolayers containing 0.3 mol rh fluorescent probe: (a) L-SSME at 37 "C, 40 A2 molecule-'; (b) racemic SSME at 29 "C, 40 Hi2molecule-*. The bar in the lower left represents 125 pm. I
c'
,
c
Figure 6. Fluorescence micrographs of SSME monolayers containing 0.3 mol 5% fluorescent probe: (a) D-SSMEat 37 "C, 37 A2 molecule-*;(b) L-SSME at 37 "C, 36 A2 molecule-'; (c) L-SSME at 37 "C, 36 A2 molecule-'. The bar in the lower left represents 125 pm.
cooled from near 40 O C into the coexistence region, and then slowly (4 A2 molecule-' min-I) compressed showed the formation of additional domain shapes for the condensed phase. The triple point temperature is taken as the lowest temperature at which an LE-LC transition can be detected in the isotherm and is 27 "C in the enantiomeric SSME monolayer and 21 O C in the racemic SSME monolayer. The enantiomeric LC domains form as small dark squares (Figure 7a). If the monolayer is subjected to an additional compression, then curved LC domains will form, either independently or as a fringe around the square domains. The individual curved domains grow in the regions of the monolayer where there was a small patch of the LE phase and all curve counterclockwise (L-SSME) or clockwise (D-SSME). If the racemic monolayer is
warmed up slowly from 15 "C, then the condensed phase domains which form are small triangles as shown in Figure 7b. Compression of the racemic monolayer a t this stage will result in the growth of some domains like those depicted in Figure 4, either independently or as a fringe around the triangle shapes. Again, the individual curved domains form in regions of the monolayer where there was a small patch of LE phase and are found to curve in either direction. The domain structures of the enantiomeric SSME monolayer and of the racemic SSME monolayer exhibit small but consistent differences when observed a t 10,15, and 20 OC. Representative fluorescence micrographs are shown in Figure 8. A t these temperatures, which are below the monolayer triple point, the condensed phase is present along with the gas phase and a small amount (about 5%) of the LE phase. The phase rule indicates that for a monolayer in true equilibrium, only the LC and gas phases should exist below the triple point temperature. Fluorescence microscopyexperiments on fatty acids have shown the persistence of a substantial amount of LE phase below the triple point temperature,18 although its area fraction was reported to decrease with time a t lower temperat~re9.l~ These results indicate a nonequilibrium effect most likely related to surface pressure gradients, which in itself warrants further study in monolayer systems by comparing the LE phase fraction present below the triple point under preparations of the monolayer which induce surface (18)Moore, B. G.; Knobler, C. M.; Akamatsu, S.;Rondelez, F. J. Phys. Chem. 1990,94,4588. (19) Qiu, X.; Ruiz-Garcia, J.; Knobler, C. M. Mater. Res. Symp. Ser. 1992,237, 263.
Monolayers of N-Stearoylserine Methyl Ester
Lcrngmuir, Vol. 9, No. 8, 1993 2117
Figure 8. Fluorescence micrographs of SSME monolayers: (a) L-SSME observed under oblique ppolarized laser illumination at 10 O C , 45 A2 molecule-'; (b) L-SSME under Hg lamp illumination at 10 "C,45 A2 molecule-l; (c) racemic SSME under oblique laser
illumination at 15 "C,40 A2 molecule-l; (d) racemic SSME under Hg lamp illumination at 15 "C, 40 A2 molecule-'. The monolayer contains 1.0 mol 96 fluorescent probe. The bar in the lower left represents 250 pm.
pressure gradients of different magnitude. The SSME monolayers shown in Figure 8 were spread at 150 A2 molecule-' and then compressed over a period of 10 min after 10 min for solvent evaporation. The area fraction of the LE phase found below the triple point in the SSME monolayers is very small (about 5%) and tends to be trapped between the LC domains (Figure 8d). The fluorescent probe molecule is incorporated into the condensed phase to a greater extent when the monolayer is spread below the triple point in this manner than when the monolayer is either compressed or cooled into the LELC coexistence region at temperatures above the triple point. The domains of the condensed phase in the enantiomeric SSME monolayer generally appear to have sharper edges and resemble broken plates, while the racemic domains appear to have smoother edges. Oblique illumination of the monolayer with a p-polarized argon laser beam reveals fluorescenceanisotropy, indicating that the molecules in this condensed phase have a long-range tilt ordering. The anisotropy pattern has a similar appearance in the both the enantiomeric and racemic monolayer (Figure 8a,c). The defect pattern within each domain radiates from the center and is brightest orthogonal to the illumination direction and darkest along the illumination direction. The pattern is found within the domains when the monolayer is first examined after deposition. Compression of the monolayer serves to squeeze out the gas phase and some of the LE phase. The pattern was observed to fade out gradually on heating, becoming unobservable near 23 "C in the racemic monolayer and becoming unobservable near 30 "C in the enantiomeric monolayer. Compression of the racemic
monolayer at 30 "C followed by slow cooling showed the sudden transformation of large regions of the LE phase into the kind of domains seen on deposition (Figure 8c,d) near 20 "C.
Discussion In addition to the thermodynamic chiral discrimination in SSME monolayers indicated by the differences in the isotherms, SSME monolayers exhibit domain structures which depend on the chirality of the headgroups in the monolayer. Compression from the LE phase yields condensed phase nonequilibrium growth forms similar to the tip-stable dendritic16p20p21and tip-splitting fractal stru~tures15J8*20~~~*~3 observed for monolayers of other materials, but with intriguing additional features. While the condensed phase of the enantiomeric monolayer exhibits the growth of fine dendritic structures, the condensed phase of the racemic monolayer initially nucleates in the form of more isotropicshapes. The growth behavior of the domains in the racemic monolayer is particularly interesting. The initial isotropic growth is followed by the formation of hooks around the edges of the domains. It appears that the chirality is not resolved during the initial period of nonequilibrium growth of these domains from the racemic monolayer, but that chiral (20) Knobler, C. M.; Moore, B. G.; Stine, K. J. In Dynamics and Patterns in Complex Fluids; Onuki, A., Kawasaki, K., Eds.; SpringerVerlag, Berlin, 1990; pp 131-140. (21) Dietrich, A.; M6hwald, H.; Rettig, W.; Brezesinski, G. Longmuir 1991, 7,539. (22) Miller, A.; Mehwald, H. J. Chem. Phys. 1987,86,4258. (23) Akamatsu, S.;Rondelez, F. J. Phys. II 1991, I , 1309.
2118 Langmuir, Vol. 9, No. 8,1993 segregationinto domainswith an excess Of D or L is possible as the growth slows and is responsible for the growth of the clockwise and counterclockwise hooks. Domain shapes whose curvature depended on the chirality were observed in the studies of phospholipid monolayers' and are attributed to an asymmetry in the line tension between the two sides of a condensed phase domain and the LE phase. The domain would thus be expected to curve toward the side with the lower line tension. The independent nucleation and growth of small domains resembling the hooks, sometimes appearing as twins, either on further compression after the condensed phase has already formed or initially on a slower rate of compression supports this hypothesis. The observation of growth of domains of entirely counterclockwise curvature on compression of an L-SSME monolayer and of entirely clockwise curvature on compression of a DSSME monolayer provides further evidence for a connection between domain curvature and the chirality of the monolayer. The observation of dendritic growth on compression or cooling of the enantiomeric monolayer in contrast to the observation of isotropic growth on compression or cooling of the racemic monolayer suggests that the condensed phase of the racemic monolayer formed during the initial growth is a less ordered structure than the condensed phase of the enantiomeric monolayer. The observed dendrites are similar to those observed recently in monolayers of D-myristoylalanine.18 Our study reveals that chirality can effect the domain growth process, as evidenced by the nucleation of the hooks as the growth of the condensed phase domains of the racemic monolayer slows. There may also be some chirality-dependent details of the dendritic structure, such as in the pattern of the side branches (Figure 5). If one supposes that the domains observed in Figure 7 are equilibrium shapes, then there are highly ordered racemic domains and highly ordered enantiomeric domains that can be formed. The observation of smoother curvature initially in the hooks and related shapes formed in the racemic monolayer seem to imply a partial chiral resolution, as these shapes can be compared to the more angular shapes seen in Figure 6c. The observation of fluorescence anisotropy suggests that SSME monolayers exhibit a condensed phase with tilted hydrocarbon chains at lower temperatures. Fluorescence anisotropy has previously indicated the presence of longrange molecular tilt in phospholipids, fatty acids, and e~ters.~9~2~926 The fluorescence anisotropy is readily observed at low temperatures, where the probe incorporates
Stine et al.
into the condensed phase formed by spreading of the monolayer or by cooling of the LE phase. The observation of sharper edges for the domains of the enantiomeric monolayer than for those of the racemic monolayer is consistent with a more ordered structure in the enantiomeric monolayer, although both films have a similar fluorescence anisotropy pattern. The observed pattern indicates a rotation of the tilt order around a point defect at the center of the domain. We observe that domain nucleation and growth occurring on compression or cooling from the LE phase expels the fluorescent probe from the condensed phase. The anisotropy is readily observed in condensed phase domains that form from the LE phase when the monolayer is cooled below the triple point. Our study of SSME monolayers indicates that examination of these monolayers by fluorescence microscopy reveals domain morphology and growth phenomena for which chirality is an important factor. The origin of the hysteresis in the isotherms and likely dependence of secondary details of the domain behavior on compression rate are problems which require additional experimental work. The origin of the hysteresis is a particularly important issue to be resolved. The examination of mixtures of SSME of composition intermediate between the pure enantiomer and the racemic mixture and of mixturea of SSME with achiral components such as methyl palmitate, which exhibits a condensed phase near room temperature, and methyl oleate, which is in an entirely fluid state near room temperature, should provide further insight into the relation between chirality and domain structure. It seems to be appropriate to choose achiral additives which also have the methyl ester headgroup, and these studies are currently in progress. Mixtures with cholesterol may be interesting if the addition of cholesterol lowers the line tension for SSME as it does for phospholipids? The examination of the two-dimensional structure of these condensed phases by synchrotron X-ray methods would help to clarify the structural differences at the molecular level.
Acknowledgment. Acknowledgment is made to the donors of the Petroleum Research Fund, administered by the American Chemical Society, for support of this research. (24) Moy, V. T.; Keller, D.J.; Gaub, H.E.;McConnell, H.M. J. Phya. Chem. 1986.90.3198. (26) Qiu,X.; Ruiz-Garcia,J.;Stine, K.J.; Knobler, C.M.; Selinger,J. V. Phys. Rev. Lett. 1991, 67, 703.
