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E. M. BARRALL, 11, R. S. PORTER, AND J. F. JOHNSON
temperature plot for all three anomalies (232, 209, and 256°K). Determination of the position of the “meshed” methyl hydrogens in trimethylacetonitrile by neutron diffraction might provide definitive evidence for the distinguishability of the methyl groups. Further cryogenic calorimetric studies on the molecules of the types (CH&CX2 and (CH&CX together with X-ray diffraction determinations would be very useful
in the determination of the structure, the molecular freedom, and the nature of the interaction existing in the crystalline state of these materials.
Acknowledgment. The authors thank the U. S. Atomic Energy Commission for partial financial support and Mr. J. T. S. Andrews and Mrs. C. M. Barber for cooperation in the experimental aspects of the work.
Heats of Transition for Some Cholesteryl Esters by Differential
Scanning Calorimetry1
by Edward M. Barrall, 11, Roger S. Porter, and Julian F. Johnson Chevron Research Company, Richmond, California (Received July 27,1966)
The thermal behavior of liquid crystal compounds has been compared by several authors to the thermal behavior of certain groups of high polymers. The esters of cholesterol are especially interesting, since between the true solid and isotropic liquid a t least two “liquid crystal” mesophases are possible-smectic and cholesteric. Although the temperatures of these transitions have been reported previously, no data have been published on the heats of these phase changes. Such information is necessary before any thermodynamic evaluation of these transformations can be attempted. The heats of transition have been determined for nine cholesteryl esters from formic to stearic by differential scanning calorimetry (dsc). The thermally large transition (smectic, cholesteric, isotropic liquid) varied depending on the ester, quite unlike the regular behavior of nematic liquid crystal compounds which have been studied. Several previously unreported solid phases depending on conditions of crystallization have been found by dsc. The reproducibility of phase formation has been studied.
The transition from solid to isotropic liquid is usually a simple, single-step phase transformation. The esters of cholesterol and over 500 other compounds do not undergo simple melting or freezing, but proceed from .the solid to the isotropic liquid via a single step or sequence of intermediate, distinct liquid crystal or mesophase states.2 The term liquid crystal or mesophase refers to a phase which is liquid in mobility but with molecules in domains arranged in a structured The Journal of Physical Chemistry
order similar to a solid.2 The esters of cholesterol have been described in context with other liquid crystals by Usol’tseva and Chistyakov3 and by Gray.‘ ( 1 ) Part VI11 of a series on order and flow of liquid crystals. (2) (a) J. L. Fergason, Sci. Am., 211, 76 (1964); (b) G. H. Brown and W. G. Shaw, Chem. Rev., 57, 1049 (1957). (3) V. A. Usol’tseva and I. G. Chistyakov, Russ. C h m . Rev., 32, 495 (1963). (4) G. W. Gray, “Molecular Structure and the Properties of Liquid Crystals,” Academic Press Inc., New York, N. Y., 1962.
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HEATSOF TRANSITION FOR CHOLESTERYL ESTERS
An important practical significance of such mesophases is that they have been reported in mammalian tissue and have been suggested to be of causal importance in atheroscler~sis.~Calorimetric studies on the esters of cholesterol have not been reported in the literature previously. The results are of value for comparison with published calorimetry available on the two other major classes of liquid crystal, the nematic and smectic mesophase~.~ Liquid crystals also frequently undergo monotropic transitions.2 That is, the phases observed depend on the conditions of the experiment, eriz., cooling from the melt or heating from the solid. Previously, the temperatures of transition on heating for nine cholesteryl esters6 have been reported as determined by differential thermal analysis (dta). I n this study of transition heats, the recently developed technique of differential scanning calorimetry (dsc)' was used. Experimental Section Cholesteryl Esters. The source, purification, and characterization of these esters have been fully given elsewhere.6 All materials were established as having less than 0.1% impurities. The ethanol recrystallization solvent was removed by drying overnight at 30" and 2 mm pressure. Instrumentation. A differential scanning calorimeter, DSC-1, Perkin-Elmer Corp., Norwalk, Conn., was used. The samples were contained in closed aluminum planchets under nitrogen. The dsc was purged continuously at a slow and constant flow rate with dry nitrogen gas. The calorimeter was calibrated over a broad temperature range using stearic acid, indium, @-tin,sodium nitrate, and lead. These standard materials have well-defined transition heats and temperatures in the range 30-327". Both heating rate and temperature of the transition have an effect on the constant. The 2-mg standards and samples were weighed on a Cahn microbalance to the nearest 0.001 mg. The chart areas of the dsc curves, which are proportional to the number of calories involved in transitions, were measured with an optical integrator. The accuracy and precision of this instrument have been described e l s e ~ h e r e . * Each ~~ sample was heated and cooled three times a t 5"/min. I n addition, duplicate runs were made on fresh samples of each ester. The first run, in any case, corresponds to the fusion from ethanol-recrystallised material; the second and third runs correspond to fusion from melt-recrystallized material. The heat capacity changes concurrent with the formation of the liquid crystal phases were eliminated from consideration in this study of transition heats by
the method of base line construction. The instrument base line was ignored, and the best straight line was constructed from the onset to the conclusion of the endotherm as noted by its departure from the straight line running into and out of the transition. This method is common in dta.lo Minor heat capacity effects may remain even in this interpretation, but this should be smaller than instrumental and other systematic errors. An accuracy of *4% and a reproducibility of d=2% on the same sample were established from measurements on known materials and multiple measurements on aliquots of individual esters. Results The results of the calorimetric measurements by dsc are shown in Table I. These are correlated with the previously published transition temperatures as measured by dta. The dsc curves are not given since they are essentially equivalent to the dta curves which have appeared previously.s Cholesteryl Fomnate. Gray's tabulation of the cholesteryl esters indicates that the formate should have a single melting point on heating a t 97.5"." This single transition is easily seen by dsc. On cooling from the melt, however, two first-order transitions are observable-the liquid-cholesteric transition with a heat of 0.2 cal/g and the cholesteric-solid transition with a heat of 8.6 cal/g. The cooling data are somewhat less accurate than the heating data. The freezing process occurs under dynamic conditions of selfnucleation, and the resultant flow of heat from the sample to the dsc control sensor is rapid. This introduces out-of-balance errors in the recorder and differential power controller. Although these errors are balanced out over-all as cooling progresses, a residual factor exists which results in an uncertainty of (68%) in the fiual calorimetric measurement from cooling data. In cases where the freezing process is slow and the exotherm broad, better accuracy is obtained on freezing. In addition, the formate ester does not rapidly re-form the solid phase. Long standing a t room temperature (4-6 hr) was apparently neces-
*
(6) G. T. Stewart, Nature, 192, 624 (1961). (6) E. M. Barrall, 11, R. S.,J'orter, and J. F. Johnson, J . Phye. c h m . . 70, 385 (1966). . . (7) E. 5. Watson, M. J. O'Neill, J. Justin, and N. Brenner, A&. Chem*f361 1233 (1964)@)
A.
*'
Gray* private communication^
(9) K. W. Gardiner, R. F. Klaver, F. Baumann, and J. F. Johnson, a a C~omatography,~~ ~ ~ ~ N, B ~J. E.callen, ~ and ~ M, ~ Weiss, , Ed., Academic Press Inc., New York, N. Y., 1962, Chapter 24, pp 349-361. (10) E. M. Barrall, 11, R. 8. Porter, and J. F. Johnson, J . Phys. them., a,2810 (1964). (11) G. w. Gray, J . chm. Soc., 3733 (1966).
Volume 71, Number 6 April 1967
E.M. BARRALL, 11, R. S. PORTER, AND J. F. JOHNSON
1226
I
.. .. :. , 2 2 $ B 0000
I
: *
sary to effect complete re-formation of the solid phase aa indicated by reheating. Cholesteryl Acetate. The thermographic behavior of cholesteryl acetate is complex, as has been discussed previously.6 The data are listed in Table I. On cooling, a single exotherm is observed a t 95O, which absorbs 10.7 cal/g of sample. The apparent temperature of freezing varies over a *3" range depending on cooling rate. On further cooling to 0", no additional transition is noted. Reheating of the solid formed from the melt produces a single endotherm at 110.9' which requires 10.7 cal/g. This calorimetric evidence supports the monotropic nature of the acetate ester phases which have been discussed previously.11 Ethanol recrystallization of a fused sample previously thermographed results in the reappearance of the 4.89cal/g endotherm at 81-87", The ethanol-recrystallized solid is definitely not in the same phase as the meltcrystallized material. C h i s t y a k o ~ ~has ~ ~ 'reported ~ two and Kofler14 has suggested three different solid forms for cholesteryl acetate. Cholesteryl n-Propionate. Three transitions are observed on heating ethanol-recrystallized material at 99, 110, and 115.3"; the heats of transition at these points are listed in Table I. The 99" transition agrees with that reported by Gray." Only two endotherms appear on heating melt-recrystallized material at 101.6 and 115.2". The first transition, solid-cholesteric, requires 13.0 cal/g, and the second transition requires 0.23 cal/g. The heating of melt- and ethanol-crystallized samples produces a different number of transitions, but the total heat is essentially the same: 13.8 vs. 13.2 cal/g. For the two cases, the nominal solidcholesteric transition is observed a t temperatures which differ by 1.6". Cooling from the isotropic liquid gives two freezing exotherms at 108.7", 0.43 cal/g, and 66.3", 11.1 cal/g. The difference in total heat required for melting and freezing is due in part to the calorimetric error previously discussed. Choleskryl n-Heptanoate. A single endotherm was observed on heating ethanol-recrystallized material a t 114.1". This solid-isotropic liquid transition required 13.8 cal/g. Reheating the melt-recrystallized material produced an endotherm at the same temperature, but about one-fourth larger, 17.0 cal/g. An endotherm of this magnitude persists as long as a meltrecrystallized sample is used. If the sample fused in the dsc is redissolved in ethanol and the alcohol evaporated without melting, the 13.8-cal/g transition reap(12) I. G. Chistyakov, Soviet Phy8. Cryst., 8, 67 (1963). (13) I. G.Chistyakov,$ b k L 5, 917 (1961). (14)A. Kofler, Arch. P h a m . , 281, 8 (1943).
HEATSOF TRANSITION FOR CHOLESTERYL ESTERS
pears on the first heating. Probably a more highly ordered crystal is formed from the melt than from solution. This unusual melt recrystallization has been discussed elsewhere.6 The cooling exotherm shows a shoulder at 95" and a principal peak at 87". The total amount of heat involved in the cooling exotherm is 16.8 cal/g. The cooling exotherm and second heating endotherm agree closely owing to the temperature broadness of the transitions and identical solid states formed. Cholesteryl n-Nonanoate. The n-nonanoate ester shows three endotherms on heating: at 74.0, 80.8, and 93.0'. These have been previously identified as the solid-smectic, smectic-cholesteric, and cholestericisotropic liquid transitions.s The 74.0' endotherm appeared as a shoulder on the 80.8" endotherm and was not separated for calorimetric evaluation. The combined heats of the solid-smectic and smectic-cholesteric transitions are 10.2 cal/g. The cholesteric-isotropic liquid transition required only 0.22 cal/g. The nnonanoate ester is unusual among this group of esters in exhibiting a small, large, small sequence of thermal transitions. Cooling to room temperature produces two exotherms at 86 and 66' which evolve 0.23 and 0.11 cal/g. The sample can remain liquid at room temperature for several hours before the major caloric event occurs. Reheating melt-recrystallized ester produces essentially the same thermogram as obtained on first heating. Cholesteryl n-Decanoate. Two endothermal transitions at 85.7 and 91.2' were noted. The 85.7' solidcholesteric transition is much the larger, 13.5 cal/g vs. the 91.2' cholesteric-isotropic liquid transition, 0.27 cal/g. I n a previous work, the relative magnitudes of these two transitions were inadvertently reversed in the Discussion.6 However, the previously published dta thermograms support the relative heats given here. The small transition appears as a shoulder on the larger endotherm. The magnitude of the transitions was independent of the origin of the sample, viz., prepared from the melt or by ethanol recrystallization. Cooling produced two freezing exotherms, isotropic liquidcholesteric transition at 81.9' releasing 0.30 cal/g and the cholesteric-smectic transition at 68.5' releasing 0.18 cal/g. The smectic-solid transition supercooled to below room temperature and was not measured. The sample remained in the smectic phase for 30 min at 25' before final freezing. This crystallization was complete, however, as essentially the same transition temperatures and heats were observed on second heating; see Table I. Cholesteryl Myristate. This ester shows three sharp endothermal transitions on heating. These are solidsmectic, 73.6' and 18.7 cal/g; smectic-cholesteric,
1227
79.7 ' and 0.52 cal/g; and cholesteric-isotropic liquid, 85.5' and 0.41 cal/g. The magnitude of these transitions w&s unaffected by the origin of the sample, whether melt or ethanol recrystdlized. The narrow temperature range over which the transitions are observed is remarkable. Dr. A. P. Gray examined the same ester sample in his dsc apparatus at PerkinElmer Corp. and obtained the results given in Table I1 on two separate aliquots of 4.64 and 6.64 mg. The agreement between laboratories and instruments is good.
Table II Transition temp, O C
73.6 79.7 85.5
Heat of transition, cal/g This laboratory Perkin-Elme+
18.5, 18.7, 18.8 0.49, 0.53, 0 . 5 5 0.41, 0.42, 0.41
18.8, 18.8, 18.8, 18.9 0.57, 0.58, 0.54 0.42, 0.40, 0.47
Cooling the sample produces three freezing exotherms: 76.4', 0.44 cal/g; 70.0', 0.56 cal/g; and 36.7', 16.8 cal/g, on an average of four determinations. After standing for 1 hr at 20°, the solid-smectic transition was 18.69 cal/g on reheating. The melt crystallization is apparently slow but complete. Cholesteryl Palmitate. The palmitate ester exhibits a single, broad endotherm centering at 79.7', 23.2 cal/g. This means that the endotherm is a combination of the solid-cholesteric and the cholesteric-isotropic transitions. The mesophase for this ester is extant over only a few degrees. The shape of the endotherm as recorded by both dta and dsc instruments indicates a temperature range for interpenetration of the solid and cholesteric transitions.6 A lower heating rate was unsuccessful in improving the resolution between solid-cholesteric and cholesteric-isotropic liquid transitions. The solid-cholesteric transition requires the largest amount of heat as indicated by a long tail on the endotherm in the direction of increasing temperature (see figure in ref 6). Cooling thermograms show three freezing exotherms. The liquidcholesteric transition at 70.0' liberates 0.46 cal/g, the cholesteric-smectic transition at 64' liberates 0.57 cal/g, and the smectic-solid transition at 47' evolves 17.5 cal/g. Some supercooling is evident. Cooling to room temperature resulted in no further transitions. Immediate reheating of the sample produced a single, broad endotherm again at 79' which required only 18.5 cal/g-4.7 cal/g less than that required on first Voluma 71, Number 6 April 1967
1228
E. M. BARRALL, 11, R. S. PORTER, AND J . F. JOHNSON
heating. If the sample was allowed to cool to room temperature and stand overnight, reheating resulted in a 22.7-cal/g endotherm. Apparently, a solid transition, not previously reported, occurs after the smecticsolid transition. Interpenetration of the smectic and solid phases is another possibility. Chistyakov’s data indicate a similar situation between the solid and the isotropic liquid under some conditions of heating. Cholesteryl Stearate. This ester exhibits a single endotherm on heating at 85.1’ which requires 25.5 cal/g. On cooling, a sharp exotherm occurs at 71’ which corresponds to an evolution of 0.54 cal/g. A broad exotherm occurs at 67’ with the liberation of 25.8 cal/g. The 71’ exotherm corresponds to the isotropic liquid-cholesteric transition, and the broad exotherm includes the unresolved sum of the cholestericsmectic and smectic-solid transitions reported by Gray.” The temperature range for mesophases in this ester is again small, 7.5’. Reheating resulted in the reproduction of the first observed endotherm (in both temperature and heat). There was no distinction between ethanol and melt recrystallization of ester.
nematic mesophase forming materials.1° It appears now to be empirically established that, for one-component mesophase systems of the three basic types, the highest of the multiple first-order transitions is generally the smallest in heat and entropy. The change in order at this transition represents only a few per cent of the heat normally required for a firstorder transition. Investigations of heat capacity as a function of temperature for nematic and cholesteric materials currently in progress indicate that the liquid crystal phase is more “liquid,” i.e., has higher specific heat, than “solid” in this physical characteristic as well. It appears that the total heats or entropies of fusion are about the same, with no regular trend, for all straight-chain esters up to about n-nonanoate, after which the totals increase rapidly with n-alkyl length. This could mean that the basic cholesteric layer structure is usually the same at low molecular weights and that some additional ordering occurs with long chains.16e1e I n some cases reported here, the difference between heats of fusion and freezing is larger than expected from experimental error. The final slow crystallization of the sample as evidenced by the smaller total heat of freezing compared to the total heat of fusion could be due to the slow freezing of liquid droplets trapped in the vacuoles which can be seen on microscopic examination of a melt-crystallized preparation. Transition supercooling and the known high viscosities for cholesteric and smectic me so phase^'^ are also consistent with incomplete crystallization. When a slide preparation is allowed to stand for several hours, many of the dark vacuoles observable with polarized light grow appreciably smaller. The crystal structure and heat content for cholesteryl esters depend in some cases on the medium from which they are crystallized. This feature may be of major significance in the deposition of cholesterol derivatives from living tissue.
Conclusions A calorimetric study of nine saturated straightchain hydrocarbon esters of cholesterol has indicated many interesting features. The mesophase structures, although highly optically active and quite easily distinguished by optical microscopy,2 are actually separated by less than 1 cal/g from one another and from the corresponding isotropic liquid of the ester. The heat required to go from the solid to the isotropic liquid or mesophase ranges from 12 to 25 cal/g-not unusual for normal first-order transitions. I n general, total heats and entropies of transition increase as the length of the ester carbon chain increases. Individual transition heats and the total for each ester vary widely with the carbon number of the ester. With the possible exception of the acetate ester, the calorically large event is the solid-mesophase (smectic or cholesteric) transition. This behavior is similar to that previously demonstrated for typical
The J o u d of Physical Chamistry
~~~~
(16) R:S. Porter and J. F. Johnson, J. A p p l . Phys., 34, 55 (1963). (16) R. Bchenck, “Crystalline Liquids and Liquid Crystals,” W. Engelmann, Leipzig, 1905, p 88.
