Four new polymorphic forms of suplatast tosilate

Article abstract of DOI:10.1016/j.ijpharm.2013.10.049

We found four new polymorphic forms (γ-, E?-, ζ-, and η-forms) of suplatast tosilate (ST) by recrystallization and seeding with ST-analogous compounds; three polymorphic forms (α-, β-, and δ-forms) of ST have been previously reported. The physicochemical properties of these new forms were investigated using infrared (IR) spectroscopy, solid-state nuclear magnetic resonance (NMR) spectroscopy, differential scanning calorimetry, and powder X-ray diffractometry. The presence of hydrogen bonds in the new forms was assessed from the IR and solid-state NMR spectra. The crystal structures of the E?- and η-forms were determined from their powder X-ray diffraction data using the direct space approach and the Monte Carlo method, followed by Rietveld refinement. The structures determined for the E?- and η-forms supported the presence of hydrogen bonds between the ST molecules, as the IR and solid-state NMR spectra indicated. The thermodynamic characteristics of the seven polymorphic forms were evaluated by determining the solubility of each form. The α-form was the most insoluble in 2-propanol at 35 C, and was thus concluded to be the most stable form. The E?-form was the most soluble, and a polymorphic transition from the E?- to the α-form was observed during solubility testing.

Full text of DOI:10.1016/j.ijpharm.2013.10.049

International Journal of Pharmaceutics 460 (2014) 83–91
Contents lists available at ScienceDirect
International Journal of Pharmaceutics
journal homepage: www.elsevier.com/locate/ijpharm
Four new polymorphic forms of suplatast tosilate
a,e,∗
b
c
d
Keiko Nagai
, Takanori Ushio , Hidenori Miura , Takashi Nakamura ,
e
e
Kunikazu Moribe , Keiji Yamamoto
a
Taiho Pharmaceutical Co. Ltd., Ebisuno, Hiraishi, Kawauchi-cho, Tokushima 771-0194, Japan
Ace Japan Co. Ltd., Higashine-koh, Higashine, Yamagata 999-3701, Japan
Taiho Pharmaceutical Co. Ltd., Kamikawa-machi, Kodama-gun, Saitama 367-0241, Japan
RIKEN, Wako, Saitama 351-0198, Japan
b
c
d
e
Graduate School of Pharmaceutical Sciences, Chiba University, Inohana, Chuo-ku, Chiba 260-8675, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 16 August 2013
Received in revised form 20 October 2013
Accepted 29 October 2013
Available online 6 November 2013
We found four new polymorphic forms (␥-, ␧-, ␨-, and ␩-forms) of suplatast tosilate (ST) by recrystal-
lization and seeding with ST-analogous compounds; three polymorphic forms (␣-, ␤-, and ␦-forms) of
ST have been previously reported. The physicochemical properties of these new forms were investigated
using infrared (IR) spectroscopy, solid-state nuclear magnetic resonance (NMR) spectroscopy, differen-
tial scanning calorimetry, and powder X-ray diffractometry. The presence of hydrogen bonds in the new
forms was assessed from the IR and solid-state NMR spectra. The crystal structures of the ␧- and ␩-
forms were determined from their powder X-ray diffraction data using the direct space approach and the
Monte Carlo method, followed by Rietveld refinement. The structures determined for the ␧- and ␩-forms
supported the presence of hydrogen bonds between the ST molecules, as the IR and solid-state NMR
spectra indicated. The thermodynamic characteristics of the seven polymorphic forms were evaluated
Keywords:
Suplatast tosilate
Polymorphs
Seeding analogous compounds
Physicochemical properties
Solubility
◦
by determining the solubility of each form. The ␣-form was the most insoluble in 2-propanol at 35 C,
Rietveld refinement
and was thus concluded to be the most stable form. The ␧-form was the most soluble, and a polymorphic
transition from the ␧- to the ␣-form was observed during solubility testing.
©
2013 Elsevier B.V. All rights reserved.
1
. Introduction
Tozuka et al., 2002). Most polymorphs can be identified using these
analytical techniques, but some polymorphs are still difficult to
identify.
Polymorphism has long been a subject of great concern in
the pharmaceutical industry because different solid pharmaceu-
tical forms can impart different physical and chemical properties
that can occasionally affect the bioavailability, dissolution rate,
stability, and manufacturability of solid dosage forms (Brittain,
Polymorphism is defined as the occurrence of different arrange-
ments and/or conformations of molecules within the crystal lattice.
Elucidating the arrangement and/or conformation of the molecules
is often crucial to understanding the solid-state chemistry of active
pharmaceutical ingredients.
2
012; Grunenberg et al., 1996; Jones, 1997; Kawakami, 2007;
Thirunahari et al., 2010; Yamamoto et al., 2011). The character-
ization of solid pharmaceuticals is therefore very important to
ensure that pharmaceuticals with standardized qualities are sup-
plied to the consumer. There have been many studies in which
solid pharmaceuticals have been physicochemically characterized,
and techniques such as differential scanning calorimetry (DSC),
infrared (IR) spectroscopy, microscopy, powder X-ray diffractome-
try (PXRD), Raman spectroscopy, and solid-state nuclear magnetic
resonance (NMR) spectroscopy have been used (Ikeda et al., 2010;
The crystal structure is one of the most crucial pieces of
information relating to polymorphism; however, it is not always
possible to determine the crystal structure of a metastable crys-
talline form because single crystals that are suitable for X-ray
crystallographic analysis may be unavailable. Such difficulties have
recently been overcome by advances in the procedures available
for solving structures from powder X-ray diffraction data mea-
sured using laboratory X-ray sources (Engel et al., 1999; Harris
et al., 1994, 2001; Neumann, 2003; Rietveld, 1969; Stephenson,
2000; Young and Ed, 1993), although there still are some
limitations.
Suplatast tosilate (ST) [(± )-[2-[4-(3-ethoxy-2-hydroxypropoxy)
∗ Corresponding author at: Taiho Pharmaceutical Co. Ltd., Ebisuno, Hiraishi,
Kawauchi-cho, Tokushima 771-0194, Japan. Tel.: +81 88 665 3571;
fax: +81 88 665 7225.
phenylcarbamoyl]ethyl]dimethylsulfonium p-toluenesulfonate] is
an excellent IgE antibody production suppressor, and is a use-
ful therapeutic agent for various allergic diseases. ST is a glycerol
E-mail address: kei-nagai@taiho.co.jp (K. Nagai).
0
378-5173/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.ijpharm.2013.10.049

8
4
K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
derivative with one chiral carbon atom, which has been developed
as a racemate.
variable torsional angles are shown in Fig. 1. All other chemicals
used were of special reagent grade.
It has been reported that changing the crystallization condi-
tions, such as the solvent(s) and temperature, pseudo-seeding the
supersaturated solution with a crystal with the desired structure
2.2. Preparation of ˛-, ˇ-, and ı-form ST crystals
(
following an epitaxy protocol), or adding impurities or additives to
The ␣-, ␤-, and ␦-forms of ST were prepared following the previ-
ously reported procedures (Miura et al., 2003; Tamura et al., 1997;
Ushio et al., 1996c).
inhibit the nucleation and/or crystal growth of an undesired poly-
morph can be used to give a desired polymorph (Maruyama and
Ooshima, 2000; Blagden et al., 1998). Using this technique, poly-
morphs of ST and ST analogous compounds were also prepared
(
2
Takahashi et al., 1998a, 1998b, 2002; Tamura et al., 1997, 1998,
001a, 2001b, 2011).
2.3. Preparation of ꢀ-form ST crystals
Three polymorphic forms (␣-, ␤-, and ␦-forms) of ST were pre-
Suplatast tosilate (5 g) was dissolved in chloroform (20 mL) and
the solvent was evaporated under reduced pressure. Crystals pre-
pared in this way were designated as the ␥-form.
viously reported, and their crystal structures were determined
Miura et al., 2003; Takahashi et al., 2001; Ushio et al., 1996a, 1996b,
996c, 2002; Ushio and Yamamoto, 1994). We have found four new
(
1
polymorphic forms (␥-, ␧-, ␨-, and ␩-forms) by recrystallizing and
seeding with ST-analogous compounds.
2.4. Preparation of ␧-form ST crystals
We investigated the physicochemical properties of the new
polymorphic forms of ST by DSC, IR and solid-state NMR spectros-
copies, and PXRD. We also determined the solubilities of the new
polymorphic forms, and compared them with the solubilities of the
three crystals that were previously characterized.
We determined the crystal structures of the ␧- and ␩-forms from
their PXRD data using the direct space approach in conjunction with
the Monte Carlo method, followed by Rietveld refinement.
Suplatast tosilate (1 g) was dissolved with heating in 2-
propanol (4 mL). After cooling the solution to room temper-
ature, (± )-[2-[4-(3-ethoxy-2-hydroxypropoxy)phenylcarbamoyl]-
ethyl]trimethylammonium p-toluenesulfonate (2) (approximately
mg) was added to act as pseudo-seed crystals. The wet crystals
obtained from the solution were added to another solution of ST
5 g) in 2-propanol (20 mL) to act as seed crystals. The crystals that
formed were filtered and dried, and classed as the ␧-form.
5
(
2
. Materials and methods
2.5. Preparation of ␨-form ST crystals
2.1. Materials
Suplatast tosilate (1 g) was dissolved with heating in 2-propanol
Suplatast tosilate [(± )-[2-[4-(3-ethoxy-2-hydroxypropoxy)
(4 mL). After cooling the solution to room temperature, (± )-[2-[4-
(3-propoxy-2-hydroxypropoxy)phenylcarbamoyl]ethyl]dimethyl-
sulfonium p-toluenesulfonate (1b) (approximately 2 mg) was
added to act as pseudo-seed crystals. The wet crystals obtained
from the solution were added to another solution of ST (5 g) in
2-propanol (20 mL) to act as seed crystals. The crystals that formed
were filtered and dried, and classed as the ␨-form.
phenylcarbamoyl]ethyl]
dimethylsulfonium
p-toluenesulfonate]
(1a),
which
has
been developed as
a
racemate, was provided by Taiho
Pharmaceutical Co. Ltd. (Tokyo, Japan). Analogous com-
pounds of ST,
(± )-[2-[4-(3-propoxy-2-hydroxypropoxy)
phenylcarbamoyl]ethyl]dimethylsulfonium p-toluenesulfonate
(
1b) and (± )-[2-[4-(3-ethoxy-2-hydroxypropoxy)phenylcarba-
moyl]ethyl]trimethylammonium p-toluenesulfonate (2), were
provided by the Graduate School of Human and Environmental
Studies, Kyoto University (Japan), and Taiho Pharmaceutical Co.
Ltd., respectively. The chemical structures of ST, the ST-analogous
compounds and the structure formula of (± )-1a with the six
2.6. Preparation of ꢁ-form ST crystals
Suplatast tosilate (5 g) was dissolved with heating in a mixture
of acetone (30 mL) and water (0.75 mL). After cooling the solution
◦
to 5 C, the ␣-form of ST crystals (1a) (approximately 5 mg) was
added to act as seed crystals. The crystals that formed were filtered
and dried, and classed as the ␩-form.
⋅
⋅
2.7. Powder X-ray diffraction measurements
Powder X-ray diffraction patterns of each polymorphic form of
ST were determined on a Spectris X’Pert instrument (Spectris Co.,
Ltd., Tokyo, Japan) with Cu K␣ radiation at 40 kV and 30 mA, passed
through a nickel filter. The analysis was performed at a continu-
◦
◦
ous scanning rate of 2 /min over a 2ꢂ angular range of 5–35 . The
diffractograms obtained were analyzed using Spectris diffraction
software.
2.8. Differential scanning calorimetry
Differential scanning calorimetry analysis was performed using
a Rigaku 8230 instrument (Rigaku, Tokyo, Japan). A sample (5 mg)
was weighed in an open aluminum pan and subjected to a
Fig. 1. The chemical structures of suplatast tosilate (1a), the suplatast tosilate analo-
gous compounds (1b and 2), and the structure formula of (± )-1a with the six variable
torsional angles.
◦
◦
3
0–130 C thermal scan at a heating rate of 4 C/min with a
100 mL/min dry nitrogen purge.

K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
85
2.9. Infrared spectroscopy
around all of the single bonds. The Monte Carlo calculations and
subsequent Rietveld refinements were repeated.
An infrared spectrum of each polymorphic form was recorded
using a JASCO FT/IR-420 instrument (JASCO International Co., Ltd.,
Tokyo, Japan) and the potassium bromide disk method.
3. Results and discussion
2
.10. 13C solid-state nuclear magnetic resonance spectroscopy
3.1. Polymorphic behavior
13C solid-state NMR spectra of the ␣-, ␤-, and ␥-forms of ST
were measured using a Chemagnetics CMX Infinity 400 instrument
Chemagnetics, Fort Collins, CO, USA) operated at 100.3 MHz with a
We obtained four new forms of ST by recrystallizing ST and
seeding with ST-analogous compounds, and investigated their
physicochemical properties.
(
cross-polarization/magic angle spinning (CP/MAS) probe. CP/MAS
with two-pulse phase modulated (TPPM) decoupling was used, and
the sample was placed in a 4 mm outer diameter sample tube that
was rotated at 12 kHz. The contact time and the repetition time
were 5.0 ms, and 5.0 s, respectively. The spectral width was 40 kHz,
and 3000 scans were recorded.
Visual observations of all seven crystalline forms are shown
in Fig. 2. The crystalline forms had different morphologies. The
␣-Form crystals were massive and solid. The ␤-, ␦-, and ␧-Form
crystals were relatively large but fragile. The ␥- and ␨-Form crystals
were fine, and the ␩-form crystals were needles.
The PXRD patterns of the seven crystalline ST forms are shown in
Fig. 3. We confirmed that the ␣-, ␤-, and ␦-form PXRD patterns coin-
cided with the patterns that were previously reported (Miura et al.,
2003; Takahashi et al., 2001; Ushio et al., 1996c, 2002). However,
the PXRD patterns of the four new crystals obtained in this study
were completely different from those of the other three forms, and
were also different from each other; each form showed a high
degree of crystallinity and distinct diffraction angles and inten-
sities. These results strongly suggested that the four new crystal
forms could be polymorphic forms of each other.
13C solid-state NMR spectra of the ␦-, ␧-, ␨-, and ␩-forms of ST
were measured using a Chemagnetics CMX Infinity 400 instrument
(
Chemagnetics, Fort Collins, CO, USA), operated at 100.4 MHz, using
ramped CP/MAS with TPPM decoupling. The sample was placed in
a 5 mm outer diameter sample tube that was rotated at 10 kHz.
The contact time and the repetition time were 3.0 ms, and 5.0 s,
respectively. The spectral width was 40 kHz, and 400 scans were
recorded.
2
.11. Solubilities
Adding a small amount of an ST analogous compound to act as
seed crystals may have induced three of the new ST polymorphs
(␦-, ␧-, and ␨-forms) to crystallize. It is noteworthy that additives,
which made up less than 0.5% of the recrystallization solution in all
of the cases, caused new polymorphic forms to crystallize.
An excess amount of a polymorph sample was added to 2-
propanol (2 mL). The sample was vigorously shaken for 30 s every
5
◦
min for 30 min, and then allowed to stand for 4 h at 35.0 C.
The saturated solution was filtered through a 0.45 ␮m mem-
brane filter, diluted with methanol, and analyzed by HPLC. HPLC
analyses were carried out using an octadecylsilanized silica gel
column (Shinwa chemical, ULTRON TDP, 4.6 mm × 150 mm), a mix-
ture of 2.5 mmol/L sodium 1-hexanesulfonate solution, acetonitrile,
and acetic acid (300:100:3) as the mobile phase at flow rate of
Kuroda et al. found a new polymorphic form of 6-
mercaptopurine, which was six to seven times more soluble
than the other known crystalline forms, by adding an analogous
compound as a seed (Kuroda et al., 1979). Kitamura and Ishizu
investigated the growth rate of l-glutamic acid in the presence of l-
phenylalanine, confirming that the polymorphic (␣- and ␤-) forms
had different growth rates (Kitamura and Ishizu, 1998). Addadi
et al. successfully resolved d- and l-asparagine conglomerations
by adding small amounts of the pure enantiomers to saturated
solutions of d- and l-asparagine mixtures (Addadi et al., 1962).
It is thought that in each of these mechanisms, the analogous
compound adheres to a crystal surface and inhibits crystal growth.
In this study, it is considered that this mechanism also effects the
formation of new polymorphic forms of ST.
0.9 mL/min, and a UV–vis spectrophotometer (250 nm) as detector.
Powder X-ray diffraction was used to determine whether any of the
polymorphic forms had been transformed into another during the
solubility measurement process. The solubilities were determined
in duplicate.
2.12. Solving the crystal structures from the powder X-ray
diffraction data
Evaporating the chloroform from a supersaturated solution of
ST was an effective way of crystallizing the ␥-form, but the other
polymorphic forms were prepared by recrystallization.
The Reflex Plus module in the Materials Studio software (Accel-
rys, Inc, USA) was used for all calculations. The PXRD patterns of
the ␧- and ␩-form crystals of ST were indexed with the X-cell pro-
The DSC curves of the seven polymorphic forms of ST are shown
in Fig. 4. Each curve showed an endothermic peak due to fusion
◦
gram using 30 reflections (2ꢂ < 32 ). The cell and function profile
◦
parameters obtained were refined using the Pawley method. The
space group was determined from within the space group candi-
dates consistent with the systematic absences by a trial-and-error
method.
between 75 and 87 C.
◦
The highest melting points were 86.7 C for the ␣-form and
86.5 C for the ␥-form, showing that these polymorphic forms had
◦
spatially stable molecular arrangements. In contrast, low melting
points were observed for the ␦-, ␧-, and ␨-forms, indicating that
these forms were metastable. Furthermore, the polymorphic forms
prepared by pseudo-seeding with ST-analogous compounds (␦-,
␧-, and ␨-forms) had broad endothermic peaks caused by melting-
point depression. The highest enthalpy of fusion was 39.9 kJ/mol
for the ␩-form, and the lowest was 23.2 kJ/mol for the ␨-form.
The IR spectra are shown in Fig. 5. For the known crystal struc-
tures (␣-, ␤-, and ␦-forms), the carbonyl stretching band was found
Once the initial model molecular conformations of the ␧- and
␩
-form crystals of ST were assigned, the structures were solved
using the Monte Carlo/parallel tempering method in the Powder-
Solve software. Rietveld refinement was then performed, which
involved the following processes: (i) the pseudo Voight func-
tion was used to simulate the peak shape; (ii) the background
was determined by linear interpolation using 20 terms; (iii) the
Finger–Cox–Jephcoat method was used for asymmetric refine-
ment; and (iv) the March–Dollase method was applied to correct
preferred orientation effects. Global isotropic factors were used to
refine the temperature factors. The molecular conformation in each
of the motion groups was refined by varying the torsional angles
−
1
at 1664 cm
for the ␣-form (in which the carbonyl group was
hydrogen bonded to a OH group), whereas this band was shifted to
−
1
1678 and 1684 cm for the ␤- and ␦-forms (in which the carbonyl
groups did not interact with a proton donor group).

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K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
Fig. 2. Microscopy photographs of suplatast tosilate polymorphs: (a) ␣-form, (b) ␤-form, (c) ␥-form, (d) ␦-form, (e) ␧-form, (f) ␨-form, and (g) ␩-form.
the carbonyl group stretching band was found at 1682 cm 1 for
−
It is well known that different hydrogen bonding modes may
cause polymorphism in many compounds, such as barbiturate
derivatives, sulfonamides, and oxalic acid (Mesley and Clements,
the ␧-form, suggesting that there were no hydrogen bonds present.
The ␥- and ␩-forms showed carbonyl stretching bands at 1675 and
1672 cm , respectively. It was not appropriate to assess hydrogen
bond formation in the polymorphs from these data.
−
1
1
967; Yang and Guillory, 1972; Pauling, 1960). The occurrence of
hydrogen bonds in the four new polymorphic forms of ST was
assessed by comparing their IR spectra. The carbonyl group stretch-
In general, OH and NH stretching bands that are not hydrogen
−
1
−1
ing band of the ␨-form was found at 1658 cm , indicating the
bonded are found at 3600–3700 and 3400–3500 cm , respectively,
presence of hydrogen bonds with a proton donor group. However,
as intense and sharp peaks. The polymorphic forms of ST other than

K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
87
Fig. 4. Differential scanning calorimetry curves of suplatast tosilate polymorphs:
a) ␣-form, (b) ␤-form, (c) ␥-form, (d) ␦-form, (e) ␧-form, (f) ␨-form, and (g) ␩-form.
(
was assessed by comparing the NMR spectra with the spectra of
the polymorphic forms that were previously characterized. The
chemical shift of the carbonyl carbon was observed at 170.0 ppm
for the ␣-form of ST, in which the carbonyl group is hydrogen
bonded to an OH group, and at 167.9 and 166.7 ppm for the ␤-
and ␦-forms, respectively. In both of the latter forms, the carbonyl
group did not interact with a proton donor group. The chemical
shift for the ␧-form carbonyl was 167.1 ppm, suggesting that the
carbonyl group was not hydrogen bonded. However, the carbonyl
chemical shift for the ␨-form was observed at 170.2 ppm; there-
fore, the carbonyl group in this form was assumed to be hydrogen
bonded. The carbonyl chemical shifts for the ␥- and ␩-forms were
Fig. 3. Powder X-ray diffraction patterns of suplatast tosilate polymorphs: (a) ␣-
form, (b) ␤-form, (c) ␥-form, (d) ␦-form, (e) ␧-form, (f) ␨-form, and (g) ␩-form.
the ␩-form showed broad bands between 3000 and 3500 cm−1
,
revealing that several types of hydrogen bonds were present. The
−
1
␩
-form had a sharp peak at 3495 cm , suggesting that most of the
NH groups were not hydrogen bonded. Since the ␧- and ␨-forms
−
1
of ST had broad bands around 3500 cm , we accessed that sharp
−
1
peaks overlapped in the broad bands around 3500 cm , suggesting
that some of the NH groups were not hydrogen bonded.
The C solid-state NMR results are shown in Fig. 6. The spectra
of all seven polymorphic forms of ST were readily distinguish-
able by the C solid-state NMR. The resonances of methyl carbons
in the dimethylsulfonium group observed around 20 –30 ppm
were different from each other, and the presence of hydrogen
bonds between molecules in the new polymorphic forms of ST
1
3
1
69.1 and 169.9 ppm, respectively, and these intermediate chemi-
1
3
cal shift positions could not be used to assess whether the carbonyl
groups were hydrogen bonded or not in these crystals. The results of
physicochemical properties of the ST polymorphs are summarized
in Table 1.
Fig. 5. Infrared spectra of suplatast tosilate polymorphs: (a) ␣-form, (b) ␤-form, (c) ␥-form, (d) ␦-form, (e) ␧-form, (f) ␨-form, and (g) ␩-form.

8
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K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
Table 1
Summary of physicochemical characteristic for suplatast tosilate polymorphs.
States
Preparation method
DSC
IR
NMR
Crystal structure
Melting
point ( C)
Enthalpy of
fusion (kJ/mol)
Stretching vibration of
Chemical shift of
carbonyl group (ppm)
◦
−1
carbonyl band (cm
)
␣
␤
-form
-form
Known
Known
Recrystallization
Recrystallization
Evaporation
86.7
80.5
35.2
34.8
1664
170.0
167.9
Already determined
Ref: Takahashi et al. (2001)
1678
Already determined
Ref: Ushio et al. (1996c)
␥
-form
New
86.5
74.8
35.0
32.5
1675
1684
169.1
166.7
–
␦
-form
Known
Seeding an analogous
compound
Already determined
Ref: Miura et al. (2003)
␧
-form
New
New
New
Seeding an analogous
compound
75.2
78.1
83.0
29.2
23.2
39.6
1682
1658
1672
167.1
170.2
169.9
Rietveld refinement
␨-form
Seeding an analogous
compound
–
␩
-form
Recrystallization
Rietveld refinement
into the ␣-form (although most of the crystals remained in the orig-
inal ␦-form). The ␧-form was mostly transformed into the ␣-form
during the solubility measurements, and the ␥-form was com-
pletely transformed into the ␣-form.
Even though the ␨-form was found to be extremely soluble in the
initial dissolution stage, the solubility measurement for this form
was not completed because it rapidly transformed into the ␣-form.
◦
The solubility of the crystal forms at 35 C followed the order
␣
< ␤ < ␥ < ␩ < ␦ < ␧, with the ␣-form being the most stable. The
apparent solubility of the ␧-form was significantly high nearly six
times that of the ␣-form. Since the ␥-form completely transformed
to the ␣-form, it was the second most unstable crystal after the
␨
-form.
3.3. Determination of the crystal structure from the powder X-ray
diffraction data
The crystal structures of the ␣-, ␤-, and ␦-forms were pre-
viously reported (Miura et al., 2003; Ushio et al., 1996c). We
determined the crystal structures of the ␧- and ␩-forms from their
PXRD data using the direct space approach in conjunction with the
Monte Carlo method, followed by Rietveld refinement (Table 3, and
Figs. 7 and 8).
Fig. 6. 13C solid-state nuclear magnetic resonance spectra of suplatast tosilate poly-
morphs: (a) ␣-form, (b) ␤-form, (c) ␥-form, (d) ␦-form, (e) ␧-form, (f) ␨-form, and
(
g) ␩-form.
After two cycles of Monte Carlo calculations (with 266 million
trial structures per cycle) and the subsequent Rietveld refinement,
3.2. Solubility studies
The thermodynamic characteristics of the seven polymorphic
forms were evaluated by determining the solubility of each form
in 2-propanol; the results are shown in Table 2. Once the solubility
measurements had been completed, the remaining crystals were
collected and subjected to PXRD measurements. The ␣-, ␤-, and ␩-
forms did not transform into other polymorphic forms during the
solubility measurements, whereas the ␦-form partly transformed
Table 2
◦
Solubilities of suplatast tosilate polymorphs in 2-propanol (35 C).
Solubility (mg/mL)
Polymorphic form after
solubility determination
␣
␤
-form
-form
20.95
33.63
35.87
42.22
134.28
73.62a
40.17
␣
␤
␣
␣ < ␦
␣ > ␧
␣
␥
-form
␦
␧
-form
-form
␨-form
␩
-form
␩
a
␨-form was rapidly transformed to the ␣-form while measuring solubility.
Fig. 7. Molecular arrangement in ␧-form of suplatast tosilate.

K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
89
Table 3
Crystal data for suplatast tosilate polymorphs.
Cell parameters
␣-form
␤-form
␦-form
␧-form
␩-form
Chemical formula
Formula weight
Sample type
Crystal system
Space group
a (Å)
C23H33NS2O7
499.65
Single crystal
Triclinic
P-1
14.885
15.673
12.435
C23H33NS2O7
499.65
Single crystal
Triclinic
P1
10.748
15.616
8.234
C23H33NS2O7
499.65
Powder
Triclinic
P-1
9.895
15.030
8.909
C23H33NS2O7
499.65
Powder
Monoclinic
P21/c
21.380
12.044
10.756
C23H33NS2O7
499.65
Powder
Triclinic
P-1
16.664
10.357
7.619
b (Å)
c (Å)
˛
(deg)
ˇ (deg)
(deg)
100.17
104.65
108.73
2550.6
100.50
108.47
85.51
1288.0
98.24
90.28
108.15
1242.0
78.18
97.47
115.32
ꢀ
101.59
1255.4
2
0.167; 0.226
–
V (Å)
Z
Rp , Rwp
Ref
2503.5
4
0.146; 0.190
–
4
2
2
a
b
0.063; 0.081
Takahashi et al. (2001)
0.044; 0.047
Ushio et al. (1996c)
0.147; 0.190
Miura et al. (2003)
ꢀ
ꢁ
ꢂ
|
cYsim(2ꢂi)−Iexp(2ꢂi)−Yback(2ꢂi)|
a
ꢁ
x Rp =
.
|
Iexp(2ꢂi)|
ꢀ
ꢁ
ꢂ
1/2
2
sim
(2ꢂi)−Iexp(2ꢂi)−Yback(2ꢂi))
wi(cY
b
ꢁ
y Rwp =
.
wi(Iexp(2ꢂi))2
Fig. 9. Difference plot for the Rietveld simulated and experimental powder X-ray
diffraction patterns for the ␧-form of suplatast tosilate.
Fig. 8. Molecular arrangement in ␩-form of suplatast tosilate.
the weighted- profile R value (Rwp) converged at below 23%. The
experimental and calculated diffraction patterns and the corre-
sponding difference profiles are shown in Figs. 9 and 10.
The ␧-form crystal was found to be in a monoclinic system, and
it was very different from the polymorphic forms that were previ-
ously reported. This crystal was composed of equal amounts of the
two ST enantiomers (P21/c, Z = 4) and had a center of symmetry like
the ␣- and ␦-forms.
The R and S ST enantiomers in the ␧-form were hydrogen bonded
between the hydroxyl groups and ethoxy oxygen atoms (with an
O· · ·O distance of 2.873 A˚ ), as in the ␦-form. However, unlike the
Fig. 10. Difference plot for the Rietveld simulated and experimental powder X-ray
diffraction patterns for the ␩-form of suplatast tosilate.
other ST crystals, the ␧-form was found to have a unique spatial

9
0
K. Nagai et al. / International Journal of Pharmaceutics 460 (2014) 83–91
molecular arrangement, in which the amide NH hydrogen was not
hydrogen bonded to the sulfonate ion oxygen, which was consistent
with the IR result.
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The ␩-form crystal structure was similar to that of the ␣-form,
consisting of a racemic mixture of R and S enantiomers in the unit
cell (Z = 2) of the centrosymmetric space group P-1. The R and S
enantiomers were found to be hydrogen bonded between some of
a hydroxyl group and carbonyl oxygen (with an O· · ·O distance of
3
.036 A˚ ), similarly to the ␣-form but unlike the ␤- and ␦-forms.
Concurrently, some of the hydroxyl groups in the ␩-form crys-
tal structure were also found to be hydrogen bonded to sulfonate
group oxygen atoms (with an O· · ·O distance of 2.547 A˚ ), as in the ␤-
form. From these conflicting results, we estimated that some of the
hydroxyl groups should therefore be disordered, and free carbonyl
groups did seem to exist.
The ␧- and ␩-form crystal structures supported the conclusions
13
of intermolecular hydrogen bonding drawn from the IR and
solid-state NMR spectra.
C
The determination of crystal structures of the ␥- and ␨-
forms were also examined, however, they transformed into
the ␣-form during the measurements and they are still being
studied.
Kuroda, K., Yokoyama, T., Uneda, T., Kira, Y., Konishi, A., Kuroda, T., 1979. Studies on
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Mesley, R.J., Clements, R.L., 1967. Infrared identification of barbiturates with
particular reference to the occurrence of polymorphism. J. Pharm. Sci. 20,
4
. Conclusions
We identified four new polymorphic forms (␥-, ␧-, ␨-, and ␩-
341–347.
forms) of ST by recrystallization and seeding with ST-analogous
compounds. The physicochemical properties of the new ST poly-
Miura, H., Ushio, T., Nagai, K., Fujimoto, D., Lepp, Z., Takahashi, H., Tamura, R., 2003.
Crystallization of a desired metastable polymorph by pseudoseeding crystal
structure solution from its powder X-ray diffraction data, and confirmation of
polymorphic transition. Cryst. Growth Des. 3, 959–965.
Neumann, M.A., 2003. X-Cell: a novel indexing algorithm for routine tasks and dif-
ficult cases. J. Appl. Crystallogr. 36, 356–365.
Pauling, L., 1960. The Nature of the Chemical Bond, 3rd ed. Cornell University Press,
New York, U.S.A., pp. 479–480.
Rietveld, H.M., 1969. A profile refinement method for nuclear and magnetic struc-
tures. J. Appl. Crystallogr. 2, 65–71.
Stephenson, G.A., 2000. Structure determination from conventional powder
diffraction data: application to hydrates, hydrochloride salts, and metastable
polymorphs. J. Pharm.Sci. 89, 958–966.
Takahashi, H., Tamura, R., Fujimoto, D., Lepp, Z., Kobayashi, K., Ushio, T., 2002. Pref-
erential enrichment: full crystallographic analysis of the unusual phenomenon
in the mixed crystals’ version. Chirality 14, 541–547.
Takahashi, H., Tamura, R., Lepp, Z., Kobayashi, K., Ushio, T., 2001. Preferential enrich-
ment: an essential crystal structure. Enantiomer 6, 57–66.
Takahashi, H., Tamura, R., Ushio, T.z, Nakajima, Y., Hirotsu, K., 1998a. Ideal enan-
tiomeric resolution (preferential enrichment) by recrystallization of a racemic
compound. V: relationship between preferential enrichment and crystal struc-
tures. Chirality 10, 705–710.
13
morphs were investigated using DSC, IR, and C solid-state NMR
spectroscopies, and PXRD The PXRD patterns for the new crystal
forms were completely different from the patterns for the three
previously characterized polymorphs, as well as being different
from each other.
The highest melting points observed from DSC measurements
◦
◦
were 86.7 C for the ␣-form and 86.5 C for the ␥-form, showing
that both these forms were relatively stable. Relatively low melting
points were found for the ␦-, ␧-, and ␨-forms (which were pre-
pared by pseudo-seeding with ST-analogous compounds); these
forms had broad endothermic peaks associated with melting-point
depression.
We assessed the likelihood of the presence of hydrogen bonds in
13
the four new polymorphic forms of ST from their IR and C solid-
state NMR spectra. The ST carbonyl group in the ␧-form did not
appear to be hydrogen bonded, in contrast to the ␨-form carbonyl
group.
The crystal structures of the ␧- and ␩-forms were determined
from their PXRD data using the direct space approach in con-
junction with the Monte Carlo method, followed by Rietveld
refinement. These crystal structures supported the conclusions
drawn from the IR and solid-state NMR spectra, that there
were hydrogen bonds between the molecules in these crystal
forms.
Takahashi, H., Tamura, R., Ushio, T., Nakai, Y., Hirotsu, K., Toda, F., 1998b. Preferen-
tial enrichment: mode of polymorphic transformation of a mixed crystal into a
racemic compound crystal. Mol. Cryst. Liq. Cryst. 313, 211–216.
Tamura, R., Iwama, S., Gonnade, R.G., 2011. Control of polymorphic transition induc-
ing preferential enrichment. Cryst. Eng. Commun. 13, 5269–5280.
Tamura, R., Takahashi, H., Hirotsu, K., Nakajima, Y., Ushio, T., 2001a. Preferential
enrichment and crystal structure. Mol. Cryst. Liq. Cryst. 356, 185–194.
Tamura, R., Takahashi, H., Hirotsu, K., Nakajima, Y., Ushio, T., Toda, F., 1998. Unusual
disordered crystal structure of a racemate exhibiting a novel enantiomeric res-
olution: preferential enrichment. Angew. Chem. Int. Ed. 37, 2876–2878.
Tamura, R., Takahashi, H., Miura, H., Lepp, Z., Nakajima, Y., Hirotsu, K., Ushio,
T., 2001b. Comparison of crystal structures of new racemic chiral com-
pounds showing and not showing the phenomenon of preferential enrichment.
Supramol. Chem. 13, 71–78.
Acknowledgments
Tamura, R., Ushio, T., Takahashi, H., Nakamura, K., Azuma, N., Toda, F., Endo, K.,
1
997. Ideal enantiomeric resolution by recrystallization of a racemic compound
The authors would like to thank Dr. R. Tamura and Dr. H. Tsue of
the Graduate School of Human and Environmental Studies, Kyoto
University, for conformation analyses, and Dr. H. Takahashi of the
Graduate School of Human and Environmental Studies, Kyoto Uni-
versity, for performing NMR measurements.
(part 4): relationship between enantiomeric resolution phenomenon and crystal
properties. Chirality 9, 220–224.
Thirunahari, S., Atipamula, S., Chow, P.S., Tan, R.B.H., 2010. Conformational poly-
morphism of Tolbutamide: a structural, spectroscopic, and thermodynamic
characterization of Burger’s forms I–IV. J. Pharm. Sci. 99, 2975–2990.
Tozuka, Y., Ito, A., Seki, H., Oguchi, T., Yamamoto, K., 2002. Characterization and
quantitation of clarithromycin polymorphs by powder X-ray diffractometry and
solid-state NMR spectroscopy. Chem. Pharm. Bull. 50, 1128–1130.
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