Insight into polymorphism of the ethosuximide (ETX)
Journal of Thermal Analysis and Calorimetry
Insight into polymorphism of the ethosuximide (ETX)
Natalia Osiecka 0 1 2 3
Ewa Juszyn´ ska-Gał a˛zka 0 1 2 3
Zbigniew Galewski 0 1 2 3
Teresa Jaworska-Goła˛ b 0 1 2 3
Aleksandra Deptuch 0 1 2 3
Maria Massalska-Arodz´ 0 1 2 3
0 Faculty of Chemistry, University of Wrocław , Joliot-Curie 14, 50-383 Wrocław , Poland
1 The Henryk Niewodniczan ́ski Institute of Nuclear Physics, Polish Academy of Sciences , E. Radzikowskiego 152, 31-342 Krako ́w , Poland
2 & Natalia Osiecka
3 Institute of Physics, Jagiellonian University , S. Łojasiewicza 11, 30-348 Krako ́w , Poland
Rich polymorphism of ethosuximide compound (ETX) is described in detail using complementary thermal analysis methods. The paper shows as well the results of differential scanning calorimetry (DSC), of polarizing microscope observations (POM) and of X-ray diffraction (XRD) measurements taken using horizontal rotating capillary method. Molecular structure of ethosuximide favors appearance of a conformationally disordered (CONDIS) crystal phase CrI in its polymorphism. Ethosuximide is a good glass former, and glass of the CrI phase was observed even for 5 C min-1 rate of cooling. Moreover, monotropic plastic crystal CrII phase was observed during heating above the CrI temperature range.
Ethosuximide; Polarizing microscopy; Phase transition vitrification; Thermooptical analysis
Macroscopically, matter can exist in solid, melt and gas
states. The large amplitude motions as translations and
rotations of molecules, internal rotational and/or
conformational changes determine the differences between fluid
(IL) and solid states. In fact, some solid-state phases are not
fully ordered crystal and not all large amplitude motions
are frozen there. In liquid crystal (LC) phases, molecules
act as orientationally (and in some cases even positionally)
ordered but they are mobile to various extents. In plastic
crystal phase (ODIC), molecules exhibit orientational
mobility and local rotational disorder, but their centers of
masses are positionally ordered. In conformationally
disordered (CONDIS) crystal phase, molecules are on
average positionally and orientationally ordered, but they
have partial (or full) conformational freedom [
]. It is
well known that any disordered phase may be supercooled
to form glass, and in case of LC, ODIC and CONDIS
phases partially ordered glassy phases are formed [
When a compound exists in various solid-state forms, the
following important questions should be asked: 1/ what is
their thermodynamic stability, 2/ what are thermodynamic
conditions in which any transformation can occur, and 3/
how long it lasts to have new phase in equilibrium state.
Answers for those questions are given by thermal analysis
Studying solid-state polymorphism of pharmaceutical
compounds is a crucial issue, as each polymorph may have
different bioactivities. In amorphous state, pharmaceuticals
are more advantageous in therapy, so to gain knowledge of
complexity of phase diagram and its evolution during
storage may help in usage of smaller dose of medicine.
Ethosuximide or 3-ethyl-3-methylpyrrolidine-2,5-dione
(ETX) is a well-known substance used in epilepsy disease
]. In the literature, one can find publications on
its bioactivity studies [
], while there is less information
about ETX polymorphism and physicochemical properties.
Most of physicochemical parameters were obtained at
room temperature [
]. In this paper, results of
differential scanning calorimetry (DSC), polarizing microscope
observation (POM) and TOA thermooptical analysis are
presented. This approach allowed to show rich solid-state
polymorphism of ETX compound. To define structure of
ETX polymorphs, XRD method was applied.
Figure 1 illustrates chemical formula of ETX, which
consists of ethyl chain, methyl groups and imide ring. Such
molecular structure allows to anticipate that ETX
compound may exhibit a conformationally disordered crystal
(CONDIS) in its polymorphism. It is known that
conformational flexibility causes a reduced crystallization
tendency and favors glass transition [
As we are going to present, complementary methods
should be used in polymorphism investigation even for the
material of such simple molecule as ETX. DSC method is
one of the most commonly used techniques allowing to
estimate the thermodynamic functions of the phase
transition and its temperature. Microscopic texture observations
together with thermooptical analysis (TOA) help in
identification of liquid-like and solid-like phases found.
Usually, DSC results are in good agreement with TOA
]. By DSC alone, it is difficult to detect phase
transitions characterized by small changes of heat capacity.
TOA is not only very sensitive to any changes of phase
structure but it does not suffer from thermal relaxation
behavior after cooling/heating, what permits using fast
rates of temperature changes during experiment [
XRD results show difference in detail of ETX crystal
ETX sample was purchased in Sigma-Aldrich Company
and studied on cooling and heating the samples with
various rates of temperature changes in 0.2–50 C min-1
Polarizing microscope textures were observed using
Biolar PI polarized microscope (PZO Warsaw) with the
scanning rates 5, 10, 20 and 50 C min-1. The temperature
was stabilized by Linkam THM 600 silver heating/cooling
stage and TMS 90 temperature controller. Substance was
placed between two glass plates at the temperature above
melting point. Temperature was measured by platinum
resistance thermometer with 0.1 C accuracy.
Thermooptical analysis (TOA) was performed by
TOApy program [
] based on digitalized images of ETX
microscopic textures observed on cooling and heating
DSC measurements were taken using TA Instruments,
Q2500. The mass of sample was equal to 13.56 mg. The
sample was placed in aluminum TA Tzero pan and TA
Tzero hermetic lid. During DSC experiment, the nitrogen
purge was on the level 1.3 bar. The cooling/heating rate
was 0.2, 2, 5, 8, 10, 15 and 20 C min-1.
XRD measurements were taken in horizontal rotating
capillaries made by borosilicate glass, with outside
diameter 0.3 mm on Empyrean 2 (PANalytical) diffractometer
with CuKa anode, parabolic mirror on the incident beam,
slit for capillaries and PIXcel detector working in 1D
scanning mode. The temperature was controlled with the
help of Cryostream 700 Plus (Oxford Cryosystems). The
data were collected in temperature range between - 90 C
and 50 C at several chosen temperature points during
heating and subsequent cooling the sample. The XRD data
were analyzed using XCell program from Material Studio
software package. Fitting the XRD data was made using
the Pawley refinement. The Rwp uncertainty parameter was
on level 12% and Rp vary around 9%.
Results and discussion
Results of POM observations and TOA analysis based on
intensity changes of the light transformed through
microscopic textures on cooling and heating the ETX samples
are presented in Figs. 2–4. As it is shown in Fig. 2, during
Fig. 2 TOA curve of light intensity transmitted through the
microscopic textures of ETX sample during POM observed on cooling with
rate 10 C min-1
cooling only one crystal phase and glass of that phase were
observed. In IL phase, no texture was observed and due to
dark image the line of low-light intensity in TOA plot is
visible. At 25 C, the light intensity jumps to higher value
due to IL—CrI transition. In CrI it is on a stable level until
- 20 C. Below this temperature cracks began to appear
on CrI texture (see Fig. 3c) what is a well-known signature
of glass transition identified in [
]. The tendency of
vitrification is a characteristic feature of many ODIC phases.
The observed glass is glass of plastic crystal CrI phase
During heating glass of CrI of the ETX compound, the
first metastable CrI was identified at the TOA curve and
then evidence of additional crystal CrII phase appearance
was found. Softening of glass gCrI is observed until 18 C
as a slightly growing light intensity due to a process of
cracks shrinkage. The CrI crystal phase is illustrated by a
plateau in the TOA plot, and then, increase in intensity due
to a solid–solid CrI–CrII transition is visible (see Fig. 4).
Just below the isotropization point, a maximum is visible
corresponding to a new crystal CrII phase with narrow
temperature range of 48 C–50 C.
Comparing plots in Figs. 2 and 4, one can see that ETX
compound exhibits tendency to supercooling IL and CrI
phases, which we found to be dependent on the thermal
history in POM observations. It was established that
crystallization temperature decreases with increasing of the
cooling rate. Moreover, for the sample cooled with slow
temperature change rates (i.e., 2–0.2 C min-1)
crystallization was observed after 1–2 min. For higher cooling
rates (i.e., 10–50 C min-1), crystallization was not
observed at, for example, 30 C, when the waiting time
was below 2 h.
Figures 5–8 present results of DSC experiments. As one
can see the unusual heat flow response to temperature
decrease was detected (see inset in Fig. 5), while DSC plot
transformed to heat flow vs time (see Fig. 6) illustrates a
typical phase transition signature. The atypical DSC plot
results from a phenomenon accompanying crystallization
of supercooled IL phase [
] what will be explained in
further part of this publication. All analyses were
performed on heat flow DSC curves in function of time. The
maxima corresponding to phase transitions were observed
on cooling at 25.4 C and during heating at 47.5 C (with
rate ± 10 C min-1). That observation seems to suggest
the ETX has only one crystal phase. However, the value of
full width at half maximum observed during heating is
much higher than the value recorded during cooling. This
information implies that during heating in fact two
transitions may occur, at temperatures close to each other.
Crystallization of CrI on cooling and CrI-CrII transition
and melting of CrII on heating were already given by TOA
analysis. The anomaly recorded during cooling
corresponds to enthalpy change DHcool = 10.03 kJ mol-1, while
for that observed during heating DHheat = 11.78 kJ mol-1.
Those values are related to entropy change DScool = 33.6 J
K-1 mol-1 and DSheat = 36.7 J K-1 mol-1, respectively.
Knowledge of magnitude of entropy change at phase
transition is helpful to determine a type of solid phase
which crystallizes/melts on cooling/heating run. In case of
first order of phase transition, change of entropy DSfusion is
characterized by equation:
DSfusion ¼ DSc þ DSo þ DSp;
where DSc, DSo and DSp correspond to entropy drop/jump
at phase transition temperature given by freezing/activation
of various degrees of freedom of molecules:
conformational rotatable parts, orientational and positional,
respectively. Usually DSp varies from 7 to 14 J K-1 mol-1, DSo
occurs to have value in range 20–50 J K-1 mol-1, while
DSc = n 9 (7 – 12) J K-1 mol-1, where n is a number of
rotatable parts of the molecule [
]. Based on this
information, one may suggest that crystal phase observed
during cooling is a CONDIS crystal due to zero value of
DSc estimated. Registered entropy change DSfusion = 33.6
± 0.1 J K-1 mol-1 of IL-CrI phase transition seems to be
the sum of DSo = 23 ± 3 J K-1 mol-1 and DSp = 10 ±
3 J K-1 mol-1. No contribution of DSc means that
conformational disorder of ETX molecules in IL phase is
probably the same as in the CONDIS CrI phase obtained.
One can see that in case of heating experiment, values of
entropies suggest that during heating rather two solid
phases appeared, i.e., in addition to CrI identified on
cooling an extra CrII was found only on heating ETX
compound. The value of difference between DSheat and
DScool is equal around 3 J K-1 mol-1. It is similar as the
value of difference that was found between smectic and
nematic phases for a liquid crystal compound . This
observation suggests that differences between CrII and CrI
crystal phases correspond to different orientational orders
of ETX molecules in crystalline lattices. It seems that both
crystalline phases are CONDIS crystals.
The peak separation corresponding to two phase
transitions, which overlap to each other, may be observed, if
the DSC experiment is performed with smaller temperature
change. Figure 7 shows result of DSC plot obtained for
ETX compound with heating range of 0.2 C min-1. As
one can see the DSC peak is asymmetric and derivative of
DSC plot points clearly two phase transitions near 48 C.
The DSC experiment did not show evidence of glass
transition, but the POM observation clearly shows cracks
on the texture, which are signature of glass transition.
During heating, cracks start to shrink until they disappear,
what is a sign of glass softening. In Fig. 8, the temperature
of glass transition Tg registered on cooling during POM
observation is presented. Vitrification of phases with some
degrees of disorder, i.e., of plastic crystals and
conformationally disordered CONDIS crystals, is a well-known
behavior. On changing the cooling rate from 50 C min-1
transition temperature a small amount of the sample
undergoes to CrII. This exudes a heat detected as the
complex curve of heat flow vs T observed on cooling.
During further cooling vitrification of CrI occurs. While the
sample is heated, softening of gCrI to CrI takes place. At
the higher temperature range GCrII - GCrI \ 0 so at some
temperature spontaneous transition of supercooled CrI to
more stable CrII phase occurs (small vertical red arrow).
The results of XRD experiment are presented in
Figs. 10–12. These data corroborate well polymorphism of
ETX compound established above. Diffractograms
presented in Fig. 10 show difference between data recorded at
temperature 49 C for CrII phase and at 20 C for CrI
phase. The temperature XRD patterns suggest that CrI
crystallize in monoclinic P2 space group, while CrII in
monoclinic C2/c. The temperature dependence of a, b and c
Fig. 8 Temperature of glass transition (Tg) temperature obtained
from POM observation with various cooling rates. The uncertainty of
Tg temperature for experiment obtained for 5 C min-1 and for
10 C min-1 is smaller than the size of data point
to 5 C min-1, the Tg temperature shifts toward lower
values (see Fig. 8). Usually, substances lose ability to
vitrification if the cooling rate is lower than 8 C min-1. ETX
seems to be good glass former, as it shows good
vitrification tendency even for 5 C min-1 cooling rate.
The thermodynamic conditions for equilibrium between
phases and the directions of the possible phase
transformations for an ETX compound at constant pressure are
shown in schematic Gibbs free energy plot (Fig. 9). Since
DG = GCrI - Gsupercooled is \ 0, at some temperature
spontaneous crystallization occurs in supercooled liquid
(vertical blue arrow). Unusual DSC plot on cooling (see
inset Fig. 5) points to conclusion that just above IL-CrI
unit cell parameters of CrI phase is presented during
cooling in Fig. 11 and on heating in Fig. 12, while the b
parameter varies from 92.67 to 92.14 . A slight decrease
in unit cell parameters around – 40 C for cooling run and
increase around 20 C for heating run were found, which
stay in good agreement with vitrification and glass
softening temperatures obtained from thermal analysis
methods. For the CrII unit cell parameters are
a = 6.7641 ± 0.0016, b = 28.0308 ± 0.0039,
c = 21.4885 ± 0.0061, b = 98.4363 ± 0.0023.
ETX molecule compound occurs to show interesting
solidstate polymorphism with the CrI, gCrI and the CrII phases.
POM observations and the TOA analysis have allowed to
obtain isotropic CrI phase transition and vitrification of
CrI/ softening of glass of CrI and have clearly shown
monotropic crystal CrII phase on heating. Based on the
results of thermooptical methods, free energy Gibbs
diagram of ETX has been proposed. The XRD measurements
confirm the presence of two crystal phases and vitrification
of CrI phase. Analysis of entropy changes at phase
transitions given by DSC results points to conclusion that both
crystalline phases are conformationally disordered. Thanks
to analysis of derivative of heat flow vs temperature, the
montropic CrII phase has been detected also by DSC
method. The unusual DSC result found during
crystallization of CrI on cooling was explained as accompanied by a
weak process of crystallization of the CrII phase. The DSC
experiment alone could not evidence of
vitrification/softening phenomenon in ETX compound.
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1. Wunderlich B. A classification of molecules, phases, and transitions as recognized by thermal analysis . Termochim. Acta . 1999 ; 340 - 341 : 37 - 52 .
2. Inaba A , Massalska-Arodz´ M , Suzuki H , Krawczyk J. (S ) -4-(2- methylbutyl)-4'-cyanobiphenyl (5*CB) glass former: are the crystalline polymorphs ordered? Mol Cryst Liq Cryst . 2011 ; 540 : 102 - 10 .
3. Juszyn´ ska E, Jasiurkowska M , Massalska-Arodz´ M , Takajo D , Inaba A . Phase transition and structure studies of a liquid crystalline schiff-base compound (4O.8) . Mol Cryst Liq Cryst . 2011 ; 540 : 127 - 34 .
4. Rodriguez-Spong B , Price CP , Jayasankar A , Matzger AJ , Rodriguez-Hornedo N . Genaral principles of pharmaceutical solid polymorphism: a supermolecular perspective . Adv. Drug Deliv. Rev . 2004 ; 56 : 241 - 74 .
5. Sigler M , Strassburg HM , Boenigk HE . Effective and safe but forgotten: methsuximide in intractable epilepsies in childhood . Seizure . 2001 ; 10 : 120 - 4 .
6. He X , Zhong M , Zhang T , Wu W , Wu Z , Yang J , Xiao Y , Pan Y , Qiu G , Hu X . Synthesis and anticonvulsant activity of N-3-arylamide substituted 5,5-cyclopropanespirohydantoin derivatives . Eur J Med Chem . 2010 ; 45 : 5870 .
7. Kabra PM , Stafford BE , Marton LJ . Simultaneous measurement of phenobarbital, phenytoin, primidone, ethosuximide, and carbamazepine in serum by high-pressure liquid chromatography . Clin Chem . 1977 ; 23 : 1284 - 8 .
8. Krivoshein AV , Ordonez C , Khrustalev VN , Timofeeva TV . Distinct molecular structures and hydrogen bond patterns of a, a-diethyl-substituted cyclic imide, lactam, and acetamide derivatives in the crystalline phase . J Mol Struct . 2016 ; 1121 : 196 - 202 .
9. Lin PC , Su CS , Tang M , Chen YP . Micronization of ethosuximide using the rapid expansion of supercritical solution (RESS) process . J Supercrit Fluids . 2012 ; 72 : 84 - 9 .
10. Vijaya Chamundeeswari SP , Jebaseelan Samuel EJ , Sundaraganesan N. Quantum mechanical and spectroscopic (FT-IR, FTRaman, 13C, 1H and UV) investigations of antiepileptic drug Ethosuximide . Spectrochim Acta Part A . 2011 ; 83 : 478 - 89 .
11. Yu L , Reutzel-Edens SM , Mitchell CA. Crystallization and polymorphism of conformationally flexible molecules: problems, patterns, and strategies . Org Proc Res Dv . 2000 ; 4 : 396 - 402 .
12. Chachaj-Brekiesz A , Go´ rska N , Osiecka N , Dynarowicz-Ła˛tka P. Mesophases of non-conventional liquid crystalline molecules . J Therm Anal Calorim . 2016 ; 126 : 689 - 97 .
13. Chachaj-Brekiesz A , Go´rska N , Osiecka N , Makyła-Juzak K , Dynarowicz-Ła˛tka P. Surface and liquid-crystalline properties of FmHnFm triblock semifluorinated n-alkanes . J Mat Sci Eng C . 2016 ; 62 : 870 - 8 .
14. Chachaj-Brekiesz A , Go´rska N , Osiecka N , Mikuli E , Dynarowicz-Ła˛tka P. Synthesis and thermal behavior of triblock semifluorinated n-alkanes . J Therm Anal Calorim . 2016 ; 124 : 251 - 60 .
15. Osiecka N , Galewski Z , Massalska-Arodz´ M. TOApy program for the thermooptical analysis of phase transitions . Termochim Acta . 2017 ; 655 : 106 - 11 .
16. Jasiurkowska-Delaport M , Juszyn´ska E, Kołek Ł , Krawczyk J , Massalska-Arodz´ M , Osiecka N , Rozwadowski T. Signatures of glass transition in partially ordered phases . Liq Cryst . 2013 ; 40 : 1436 - 42 .
17. Bamezai RK , Godlewska M , Massalska-Arodz ´ M, S´ ciesin´ski J , Witko W. The adiabatic kalorymetry study of the polymorphism of solid 4,4'-di-n-butyloxyazoxybenzene . Phase Trans . 1990 ; 27 : 113 - 9 .
18. Horiuchi K , Yamamura Y , Pelka R , Sumita M , Yasuzuka S , Massalska-Arodz´ M , Saito K. Entropic contribution of flexible terminals to mesophase formation revealed by thermodynamic analysis of 4-alkyl-4'-isothiocyanatobiphenyl (nTCB) . J Phys Chem B . 2010 ; 114 : 4870 - 5 .