CRYSTAL GROWTH & DESIGN
Concomitant Polymorphs of the Antihyperlipoproteinemic Bezafibrate Andreas Lemmerer,*,† Nikoletta B. Ba´thori,† Catharine Esterhuysen,‡ Susan A. Bourne,*,† and Mino R. Caira†
2009 VOL. 9, NO. 6 2646–2655
Centre for Supramolecular Chemistry Research, Department of Chemistry, UniVersity of Cape Town, Rondebosch 7701, South Africa, and Department of Chemistry and Polymer Science, UniVersity of Stellenbosch, PriVate Bag X1, Matieland, 7602, South Africa ReceiVed October 10, 2008; ReVised Manuscript ReceiVed March 8, 2009
ABSTRACT: The two polymorphic forms R and β of the active pharmaceutical ingredient 2-[4-[2-(4-chlorobenzamide)ethyl]phenoxy]-2-methylpropanoic acid (bezafibrate) have been reinvestigated and fully characterized by optical microscopy, infrared spectroscopy, differential scanning calorimetry, thermal gravimetric analysis, variable temperature X-ray powder diffraction, and single crystal X-ray diffraction. Form R is obtained from ethanol, whereas form β can be grown from a 2-butanone solution provided that stoichiometric quantities of iso-nicotinamide are present. Without iso-nicotinamide, both forms grow concomitantly from a 2-butanone solution. The two forms are related enantiotropically, with form β converting to form R at 160.7 °C. Form β is the more stable form at room temperature and at temperatures below the transition temperature of 160.7 °C. The two polymorphs arise from conformational differences which result in vastly different hydrogen bonding interactions and subsequent crystal packing in the solid state. The crystal habits of the two forms reflect the different solid-state packing observed by single crystal X-ray diffraction. Computational modelling calculations on the energy of the conformations have been carried out in the solid state and compared to those energy minima calculated in the gas phase. Introduction Co-crystals and polymorphism of active pharmaceutical ingredients (API) are “hot topics” in the ever-expanding crystal engineering field.1 Polymorphic forms of APIs may have different physical properties that can make them desirable or undesirable for formulations. Polymorphs which crystallize under the same growing conditions (same solution vessel and solvent mixture) are an important facet of this phenomenon as they provide vital clues on the complex process of crystallization.2 The compound 2-[4-[2-(4-chlorobenzamide)ethyl]phenoxy]-2-methylpropanoic acid (bezafibrate) is a member of the family of fibrates that help control hyperlipidemia. Bezafibrate is used to regulate the amount of lipid produced by the body, lowering the amount of low density lipoprotein (LDL) and triglycerides and raising the amount of high density lipoprotein (HDL), which reduces the risk of heart disease. The preparation of bezafibrate is reported in the patent literature (DE-PS 21 49 070,3 US-PS 4,749,101,4 and DE-PS 29 20 4135). Crystals of bezafibrate were grown from acetone or water solutions and were reported to have a melting point of 183 °C (US-PS 4,749,101)4 and 186 °C (DE-PS 21 49 070).3 The habit of bezafibrate crystals thus prepared is needle-like. The crystal structure of bezafibrate was reported by Djinovic´ et al.6 in the orthorhombic system with space group P212121. Patent EP 0 625 504 A17 later described the preparation of a new “β-form”,8 which crystallizes as block-shaped crystals. The crystals were grown from a mixture of water and a ketone (examples of ketones used are 2-butanone, 3-pentanone, and 4-methyl-2-oxopentane), together with a 4-10% v/v addition of an alcohol. The ratio of water/ketone may be varied from 10% water to 90% water. However, 2-butanone is the * To whom correspondence should be addressed. (A.L.) E-mail: andreas.
[email protected]; (S.B) E-mail:
[email protected]. Fax: +27-21-6897499. Tel: +27-21-650-2563. † University of Cape Town. ‡ University of Stellenbosch.
preferred ketone. The new form was characterized by powder X-ray diffraction (PXRD), infrared spectroscopy (IR), and differential scanning calorimetry (DSC). The melting point of form β was reported to be 188°C and the DSC curve shows a small endotherm at 173°C. The PXRD trace and IR spectrum confirmed that the blocklike crystals were a new form of bezafibrate.9 The significance of form β is the superior processability and filtration that its block-shaped crystals demonstrate over the needle-like form R crystals. However, the crystal structure of form β is not reported in the patent, and no detailed structural analysis is available, impeding the explanation of the underlying differences of the crystal habits of the two forms of bezafibrate. Here, we report the low temperature crystal structure of form β and compare it to the redetermined, low temperature structure of form R (both determined in this study at 173 K) (Scheme 1). Thermal and spectroscopic characterization of both forms is reported and discussed.10
2. Experimental Procedures 2.1. Reagents. Bezafibrate and iso-nicotinamide were purchased from commercial sources (Aldrich) and used without further purification. All solvents used in the study were of AR (99.9%) quality. 2.2. Differential Scanning Calorimetry (DSC). All DSC measurements were performed using a Perkin Elmer DSC7 at a heating rate of 5 K min-1 under an atmosphere of dry N2 flowing at 40 cm3 min-1. The calorimeter was calibrated with indium and zinc. 2.3. Hot Stage Microscopy (HSM). Thermomicroscopic investigations were performed with a Linkam TH600 hot stage connected to a TP92 temperature controller (Linkam Scientific Instruments Ltd., Tadworth, Surrey, UK). Microscopic images were recorded with a Sony SSC-DC18P (Sony, South Africa) video camera attached to a Nikon SMZ10 microscope (Nikon, South Africa). 2.4. Thermal Gravimetric Analysis (TGA). All TGA measurements were run on a Mettler-Toledo TGA/SDTA 851e at a heating rate of 10 K min-1 under an atmosphere of dry N2 flowing at 30 cm3 min-1.
10.1021/cg8011298 CCC: $40.75 2009 American Chemical Society Published on Web 04/22/2009
Polymorphs of Antihyperlipoproteinemic Bezafibrate
Crystal Growth & Design, Vol. 9, No. 6, 2009 2647
Scheme 1. The Molecular Structure of Bezafibrate in Form r and Form βa
a Note the syn conformation of the carboxylic acid hydrogen atom in R and anti conformation in β.
Table 1. Crystallographic Data for Polymorphs r and β of Bezafibrate
formula Mr temperature (K) crystal size (mm3) crystal system space group (No.) a (Å) b (Å) c (Å) R (deg) β (deg) γ (deg) V (Å3) Z F (calcd) (Mg m-3) µ(Mo-KR) (mm-1) theta range for data collection (deg) reflections collected no. unique data [R(int)] no. data with I > 2σ(I) final R (I > 2σ(I)) final wR2 (all data) Flack parameter
form R
form β
C19H20ClNO4 361.81 173 0.70 × 0.124 × 0.034 orthorhombic P212121 (19) 10.3118(4) 17.660(1) 19.713(2) 90 90 90 3589.9(4) 8 1.339 0.236 2.07-25.50
C19H20ClNO4 361.81 173 0.29 × 0.083 × 0.035 monoclinic P21/c (14) 10.7849(5) 15.7886(7) 11.4932(5) 90 115.875(2) 90 1760.9(1) 4 1.365 0.240 2.58-27.99
34303 6678 [0.1190] 4272 0.0440 0.0910 0.04(6)
23368 4241 [0.0915] 2562 0.0458 0.1538
2.5. Fourier Transform Infrared Spectroscopy (FT-IR). FT-IR spectra were recorded on a Perkin Elmer Spectrum One FT-IR spectrometer in KBr pellets at 1 cm-1 resolution from 450 to 4000 cm-1. 2.6. Powder X-ray Diffraction (PXRD). Room temperature X-ray powder diffraction data were collected on a HUBER Guinier 670 Imaging Plate diffractometer with Cu KR1 radiation, (λ ) 1.5405981 Å). The sample was ground, spread on Mylar film, and mounted on a vertical stage. A diffractogram was acquired under ambient conditions at a power setting of 40 kV and 20 mA in transmission mode while the sample oscillated perpendicular to the beam. Variable temperature X-ray powder diffraction data were collected on form β placed in a 1 mm capillary tube. The sample was heated in a HUBER furnace, whose calibrations show temperature control to within 1 °C of the set temperature. 2.7. Single Crystal X-ray Diffraction. Diffraction data for forms R and β were collected on a Nonius Kappa CCD diffractometer11 with graphite-monochromated Mo-KR1 radiation (λ ) 0.71073 Å) at 173 K using an Oxford Cryostream 600. Data reduction and cell refinement were performed using DENZO12 and the space groups of these crystals were determined from systematic absences by XPREP13 and further justified by the refinement results. In both cases, the structures were solved in the WinGX14 Suite by direct methods using SHELXS 9715 and refined using full-matrix least-squares/difference Fourier techniques using SHELXL 97.16 The hydrogen atoms bound to carbon atoms were placed at idealized positions and refined as riding atoms with Uiso (H) ) 1.2 Ueq (ArH, CH2) or 1.5 Ueq(CH3). H atoms bonded to the carboxylic acid and amide group were located in the difference Fourier-
Figure 1. The crystal habits of the two polymorphic forms. (a) Form R grown from ethanol. (b) Form β grown from 2-butanone with stoichiometric amounts of iso-nicotinamide. Panels (c) and (d) show both forms, needle-like and block-shaped, grown concomitantly from 2-butanone with no iso-nicotinamide added. map and their coordinates refined freely but their isotropic displacement parameters were fixed (Uiso (H) ) 1.2 Ueq(O) or Ueq(N)). Diagrams and publication material were generated using ORTEP-3,17 PLATON,18 and DIAMOND.19 Experimental details of the X-ray analyses are provided in Table 1. 2.8. Computational Details. Geometry optimizations were performed with Gaussian 0320 using the B3LYP method,21 a density functional theory (DFT) type of calculation with hybrid functionals, and the 6-311G(d) basis set.22 Hydrogen bond energies were determined by using single point calculations at the DFT level of theory, with the B971 hybrid functional23 and the 6-311G(d) basis set and the Counterpoise correction.24 Pairs of molecules (H-atom positions were optimized at the B3LYP/6-311G(d) level of theory, while the remaining atoms were maintained in the geometries found in the crystal structure) were placed in the same relative orientations as found in the crystal structures in the two forms, and the hydrogen bond stabilization [∆E(Hbond)] calculated by
E(H-bond) ) E(dimer) - E(molecule1) - E(molecule2) (1) where E(molecule1) and E(molecule2) are the energies of the two molecules involved in the hydrogen bond. Similar calculations were performed for clusters of four molecules.
3. Results and Discussion 3.1. Isolation of the Polymorphs. Crystals of form R were grown by slow evaporation of an ethanol solution of commercial bezafibrate as described by Djinovic´ et al.6 Crystals of form β were grown exclusively by slow evaporation from 2-butanone in the presence of a 1:1 stoichiometric ratio of iso-nicotinamide (large crystals of residual iso-nicotinamide were observed in the crystallization vessel and separated out). Crystals of form R and β were obtained concomitantly from a solution of 2-butanone in the absence of iso-nicotinamide. The habits of the two crystal forms are needle (R) and block-like (β) (Figure 1). 3.2. DSC. The DSC curve of form R shows a single melting endotherm at 184.8 °C with a heat of fusion of 53.9 kJ mol-1 (Figure 2a). The DSC curve of form β shows a small endotherm at 160.7 °C with a heat of fusion of 4.3 kJ mol-1 (Figure 2b). This represents the solid-solid phase change to form R, followed by melting of form R (T ) 185.0°C, ∆fusH ) 52.8 kJ
2648
Crystal Growth & Design, Vol. 9, No. 6, 2009
Lemmerer et al.
Figure 2. DSC curves of form R (a) and form β (b).
Figure 3. Phase change of a single crystal of form β converting into form R. The broken area on the right side of the crystal is not due to the phase transition and is present at room temperature already.
Figure 4. FT-IR spectra of form R from commercial (Aldrich, blue trace) and recrystallized (ethanol, red trace) sources, and of form β (green trace). The cream-shaded region and the two peaks indicated by the solid lines are convenient for distinguishing the two forms.
mol-1). Form β is the stable form of bezafibrate below the transition temperature of 160.7 °C and converts enantiotropically to form R. Both forms do not show any crystallization exotherms upon cooling, and when heated again there are no melting exotherms up to 210°C. Both forms form super-cooled melts upon cooling to room temperature. 3.3. HSM. The results obtained using HSM were consistent with the DSC data. The process of phase transformation from form β to form R was observed sequentially as illustrated in Figure 3. The rate of heating at the phase transition temperature was 1 K min-1. The process causes significant fracturing to a single crystal and the crystal becomes opaque.
3.4. TGA. The TGA traces of form R and form β show no mass loss up to the melting point (TGA traces are provided in the supplementary information), consistent with the DSC results. 3.5. FT-IR. Forms R and β were characterized by FT-IR spectroscopy. Comparative FT-IR spectra of bezafibrate from commercial sources (Aldrich) and forms R and β are shown in Figure 4. Clearly, commercial bezafibrate exists in the R form. To distinguish the two forms, peaks in the region 450-1650 cm-1 allow for identification, shown as the shaded region in Figure 4. Peaks at 529, 568, 760, 967, 1013, 1324, 1718, and 3357 cm-1 correspond to form R, and peaks at 606, 618, 821, 847, 1328, 1740, and 3387 cm-1 correspond to form β. The
Polymorphs of Antihyperlipoproteinemic Bezafibrate
Crystal Growth & Design, Vol. 9, No. 6, 2009 2649
Figure 5. Comparative PXRD patterns of bezafibrate obtained from commercial sources (form R, Aldrich, blue trace), recrystallized from ethanol (form R, red trace) and crystallized from 2-butanone (form β, green trace).
Figure 6. Variable temperature PXRD patterns of bezafibrate showing form β transforming into form R.
peak numbers in the FT-IR spectrum of form β are in good agreement with the wavenumbers quoted by Mo¨rsdorf et al. (EP 0 625 504 A17) in their identification of form β. 3.6. PXRD. The powder X-ray diffraction pattern of the commercial product of bezafibrate from Aldrich shows that it corresponds to form R, shown in Figure 5 (characteristic peaks at 2θ angles 13.80°, 15.85°, 16.60°, 17.74°, 18.46°, 19.86° and 20.34°). Needle-like crystals of form R grown from ethanol show the identical PXRD pattern. Block-shaped crystals of form β, grown from 2-butanone, have a different PXRD pattern (characteristic peaks at 2θ angles 10.58°, 10.79°, 16.87°, 17.26°, 17.61°, 17.77°, 19.04°, 21.32°, 22.36°, 23.47° and 25.12°). The measured pattern of form β reported by Mo¨rsdorf et al. (EP 0 625 504 A17) is in good agreement with the pattern reported in this study. Variable temperature PXRD performed on form β of bezafibrate in the range 150-170°C shows form β converting to form
R at the transition temperature indicated by the DSC. The transformation of form β to form R can be monitored by the disappearance of the peaks indicated by the dashed lines in Figure 6. Form β has a peak at 10.50° and the most intense peak at 17.55°. Neither of these peaks is present in the spectrum of form R, which has the most intense peak at 19.53°. The calculated (from single crystal X-ray diffraction) patterns of forms R and β match the experimental patterns.25 3.7. Single Crystal Structures of Forms r and β. The crystal structure of form R was reported in 1989 and determined at room temperature.6 To aid our structural comparison to the newly determined structure of form β, we redetermined the structure of form R of bezafibrate at -100 °C on crystals grown from ethanol, using the same procedure as reported by Djinovic´ et al.6 The asymmetric unit of form R contains two molecules of bezafibrate, molecule A (containing atom Cl1) and molecule B (containing atom Cl2) (Figure 7). The two molecules form a
2650
Crystal Growth & Design, Vol. 9, No. 6, 2009
Lemmerer et al.
Figure 7. The asymmetric unit of form R showing the atomic numbering scheme and 50% thermal displacement ellipsoids. Only the symmetry independent hydrogen bonds are shown.
Figure 8. The 1-D chains of form R showing the O-H · · · O and N-H · · · O hydrogen bonds (dashed red lines). The R44(16) hydrogen bonded ring connecting four bezafibrate molecules is shown in purple outline. Atoms with superscript i and ii are at the symmetry positions (-1 + x, y, z) and (1 + x, y, z) respectively.
pseudo-centrosymmetric hydrogen bonded dimer in space group P212121. The graph set notation26 for the dimer is R22(28). The two molecules are joined by O-H · · · O hydrogen bonds from the carboxylic acid group to the oxygen atom of the amide group. The O · · · O distances are 2.568(3) and 2.643(3) Å. This dimer is connected to adjacent dimers along the [100] direction by N-H · · · O hydrogen bonds from the amide group to the carboxyl O atom, N · · · O distances of 2.896(3) and 2.897(3) Å, to form a tetrameric enclosed ring with graph set notation R44(16) (Figure 8 and Table 2). The tetramers are related by unit cell translations only to form extended one-dimensional (1-D) tubelike chains. Adjacent tubes are generated by the twofold screw axes along the b- and c-axes (Figure 9). In addition, there are four C-H · · · π interactions between each member of the tetrameric unit and one C-H · · · π interaction to an adjacent tube (Figure 9 and Table 2).
The asymmetric unit of form β contains only one molecule of bezafibrate and crystallizes in the centrosymmetric space group P21/c (Figure 10). The bezafibrate molecules are joined by the same sequence of alternating N-H · · · O and O-H · · · O hydrogen bond pairs to form tetrameric hydrogen bonded rings (See Table 2). However, the tetramers are not folded towards each other as in form R but are spread out to form a two-dimensional (2-D) hydrogen bonded sheet parallel to the ab-plane where molecules along the b-axis are related by the two-fold screw axis and along the a-axis by unit cell translations (Figure 11). The graph set notation for the ring is R44(48). There is a single C-H · · · π interaction between two bezafibrate molecules (Figure 11 and Table 2). 3.8. Comparison of Form r and Form β. There are three conformational differences at work. As shown in Scheme 1 and Figure 7, the two carboxylic hydrogen atoms in form R, H3
Polymorphs of Antihyperlipoproteinemic Bezafibrate
Crystal Growth & Design, Vol. 9, No. 6, 2009 2651
Table 2. Hydrogen Bonding Details for Form r and Form β D-H · · · A
D-H (Å)
H · · · A (Å)
D · · · A (Å)
∠(D-H · · · A) (deg)
symmetry transformations
C2-H2A · · · Cg
0.95
2.81
3.669(3)
152(1)
x - 1, y, z
Form R N1-H1 · · · O4 O3-H3 · · · O5 N2-H2 · · · O7 O8-H8 · · · O1 C15-H15 · · · Cg C22-H22 · · · Cg C31-H31 · · · Cg C35-H35 · · · Cg
0.78(3) 0.90(4) 0.79(3) 0.91(4) 0.95 0.95 0.95 0.95
2.14(3) 1.69(4) 2.14(3) 1.75(4) 2.69 2.94 2.94 2.66
2.896(3) 2.568(3) 2.897(3) 2.643(3) 3.551(4) 3.697(3) 3.745(4) 3.485(4)
164(3) 163(3) 158(4) 168(3) 150(1) 138(1) 143(1) 146(1)
x - 1, y, z
Form β N1-H1 · · · O4 O3-H3 · · · O5 C17-H17B · · · Cg
0.86(2) 0.83(2) 0.98
2.30(2) 1.79(3) 2.84
2.998(2) 2.583(2) 3.707(2)
139(2) 160(2) 148(1)
-x, y + 1/2, -z + 1/2 x - 1, y, z -x, y - 1/2, -z + 1/2
and H8, are in the syn-conformation, whereas in form β, the same hydrogen atom, H3, is in the anti-conformation (Figure 10). These two conformations of carboxylic acids are well known, and their effect on the hydrogen bonding motif has been explored extensively.27 The syn-conformation favors hydrogen bonding that leads to association of the molecules at right angles to one another, resulting in the observed tube-shaped pattern. In form β, the anti-conformation induces hydrogen bonded molecules to be coplanar, in line with the 2-D sheet network that is observed. The amide hydrogen atom is constrained in a
x + 1, y, z x - 1, y, z x + 1, y, z x - 1/2, -y + 1/2, -z + 1 x + 1, y, z
trigonal planar arrangement and is conformationally the same in both forms. Secondly, the relative orientation of the two aromatic rings in the molecules is significantly different for the two forms. Form R has an interplanar angle of 69.3(1)° between the aromatic ring planes defined by C1-C6 (shown in blue) and C10-C15 (shown in red) in molecule A (Figure 12a) and similarly an angle of 60.6(1)° between the aromatic ring planes defined by C21-C26 (blue) and C30-C35 (red) in molecule B (Figure 12b). The same interplanar angle is 2.0(2)° in the
Figure 9. Packing diagram of the 1-D tube-shaped chains of form R showing the O-H · · · O and N-H · · · O hydrogen bonds and C-H · · · π interactions (dashed blue lines). Adjacent columns are offset along the b-axis.
Figure 10. The asymmetric unit of form β showing the atomic numbering scheme and 50% thermal displacement ellipsoids.
2652
Crystal Growth & Design, Vol. 9, No. 6, 2009
Lemmerer et al.
Figure 11. Packing diagram of form β showing the O-H · · · O and N-H · · · O hydrogen bonds and the 2-D hydrogen bonded network thus formed. The R44(48) hydrogen bonded ring connecting four bezafibrate molecules is shown in purple outline. The C-H · · · π interactions are shown as blue dashed lines. Atoms with superscripts i and ii are at the symmetry positions (-1 + x, y, z) and (-x, 0.5 + y, 0.5 - z), respectively.
single molecule of bezafibrate in form β (Figure 12c). These conformations come about from the free rotation around the -NH-CH2- bond. The torsion angles C7-N1-C8-C9 and C27-N2-C28-C29 are -175.6(3)° and -178.0(3)°, respectively, indicative of the conformation in form R. The same torsion angle C7-N1-C8-C9 is -103.7(2)° in form β. Thirdly, the conformation of the 2-methylpropanoic group relative to the phenoxy group to which it is bonded is variable in all three symmetry independent molecules of bezafibrate. The differences are best seen in the Newman Projections generated by the PLATON17 program. The views are down the C-O ether bond from the central C atom of the propanoic acid group C16 to the linking atom O2. In molecule A of form R the torsion angle C19-C16-O2-C13 is 77.6(3)° (Figure 12d), in molecule B the torsion angle C39-C36-O6-C33 is -86.4(3) (Figure 12e) and in form β, the C19-C16-O2-C13 angle is -66.7(2)° (Figure 12f). The conformational changes might be a consequence of the different hydrogen bonding of the carboxylic acid group as a consequence of the anti- and syn-conformations of the propanoic acid group. Table 3 summarizes the major conformational differences between the two forms. A qualitative and simple visual comparison between the crystal structures of the dimorphs of bezafibrate is available using Hirshfeld Surface Analysis.28 The program CrystalExplorer29 maps the Hirshfeld surface of a molecule and is a tool for visualizing various features of the surface. A Hirshfeld surface is defined as the portion of space where the electron distribution of a sum of spherical atoms for the molecule (promolecule) contributes more than half of the total corresponding sum over the crystal (procrystal) electron density”.28 The Hirshfeld surface as such reflects the intermolecular interactions in the crystal structure. With the Hirshfeld surface,
a 2-D fingerprint plot can be made, which visually shows all the different intermolecular interactions found in the crystal structure. Figure 13 shows three 2-D fingerprint plots of the Hirshfeld surfaces of molecules A and B in form R and the single molecule of form β, where de and di are respectively defined as the distance from the Hirshfeld surface to the nearest atom external and internal to the surface. Molecules A and B of form R have similar intermolecular features. Inspection of the fingerprint plots of form R shows the two molecules in the asymmetric unit to be virtually identical. The N-H · · · O and O-H · · · O hydrogen bonds are spikes (labeled 1). The diffuse region of blue points between the hydrogen bond spikes (2) are of close H · · · H contacts (van der Waals radius of H is 1.2 Å). The five C-H · · · π interactions feature prominently as a chicken-wing shape in the plots of form R (3). From the fingerprint plot of form β, it is clear that the crystal structure is different from that of form R. Visually, there is little to tell the two forms apart in the hydrogen bond spikes, which are the same in all three plots. However, the H · · · H contact region (2) shows that those contacts are further apart in form β. The chicken-wings (3) in form β are less prominent due to the single C-H · · · π interaction in form β, compared to five such interactions in form R. 3.9. Computational Results. The conformational differences were investigated by calculating the energies of the three different conformations (H-atom positions optimized) in the gas phase at the B971/6-311G(d) level of theory (see Table 4). The relative energies of the two conformations of the bezafibrate molecules found in form R are similar, but both are higher in energy than the conformation found in form β. Gas-phase
Polymorphs of Antihyperlipoproteinemic Bezafibrate
Crystal Growth & Design, Vol. 9, No. 6, 2009 2653 Table 3. Summary of Similarities and Differences between Form r and Form β form Ra crystal system melting point crystal habit recrystallization solvent conformation of OdC-O-H
orthorhombic 184.8 needle ethanol, 2-butanone syn
form β monoclinic block 2-butanone + iso-nicotinamide anti
Torsion Angles C3-C4-C7-N1 C23-C24-C27-N2
15.7(5) (A) -19.1(5) (B)
16.7(3)
C11-C10-C9-C8 C31-C30-C29-C28
82.8(4) (A) -95.4(4) (B)
87.1(2)
C7-N1-C8-C9 C27-N2-C28-C29
-175.6(3) (A) -178.0(3) (B)
-103.7(2)
C13-O2-C16-C17 C13-O2-C16-C18 C13-O2-C16-C19 C33-O6-C36-C37 C33-O6-C36-C38 C33-O6-C36-C39
-50.2(4) (A) -169.9(3) (A) 77.6(3) (A) 159.7(3) (B) 40.4(4) (B) -86.4(3) (B)
177.6(2) 58.3(2) -66.7(2)
a
Values quoted separately for molecules A and B in the asymmetric
unit.
Figure 12. The conformations of the two aromatic rings relative to each other in (a) form R (molecule A), (b) form R (molecule B), and (c) form β shown as least-squares planes in blue and red. In molecules A and B, the two aromatic rings are rotated by approximately 60° and 69° relative to each other, whereas in form β the rings are almost coplanar. The conformation of the 2-methylpropanoic acid group in (d) form R (molecule A), (e) form R (molecule B) and (f) form β relative to C13 (or C33) of the phenoxy group. (g) The three molecules overlaid using the same aromatic ring C10-C15 (molecule A: green), C30-C35 (molecule B: purple, symmetry inverted for comparison), and C10-C15 (form β: blue).
optimization of the structures shows that the conformations found in the crystal structures are slightly strained, with the greatest differences between the experimental and calculated structures being found for the torsion angles mentioned in section 3.8. In order to investigate the effect of changing the torsion angles, an analysis of the rotation around the O2-C16 (O6-C36), C16-C19 (C36-C39), and C8-N1 (C28-N2) bonds was performed at the B3LYP/6-311G(d) level of theory by changing each of these torsion angles to the values found in the three experimental structures, while keeping the rest of the molecule in the same conformation as found in molecule A of form R. It has been found that the differences in energy at the three different C-N-C-C torsion angles are very low (less than 2.5 kJ mol-1). On the other hand, the energy differences corresponding to changing the O4-C19-C16-O2 and C19C16-O2-C13 torsion angles from those in molecule A of form R to the value in form β are very large (121.8 and 242.2 kJ mol-1, respectively), as a result of repulsive interactions with the rest of the molecule. Hence the propanoic acid group of
bezafibrate must undergo correlated motion, with rotation around the O2-C16 and C16-C19 bonds taking place in a concerted fashion. As mentioned in the crystal structure comparison in section 3.8, the carboxylic acid has the syn conformation in form R, which is the unstable form at room temperature, and the anticonformation in form β, which is the stable form up to 160.7 °C. It is well-known that the syn-conformation of the OH group of COOH is far more stable than the anti-conformation, by 2-4 kcal/mol.30 This means that the syn planar conformation is far more common, with the anti-conformation primarily observed when the OH engages in an intramolecular hydrogen bond (in 88.4% of compounds containing a COOH group the OH is syn).31 Indeed, for bezafibrate the anti-conformer is calculated as being 40.4 kJ mol-1 higher in energy than the syn-conformer at the B3LYP/6-311G(d) level of theory. The barrier to rotation of H3 around the O2-C19 bond is 72.6 kJ mol-1 (see Figure S4, Supporting Information). These are somewhat higher than values calculated for acetic acid, where the anti-conformer has been calculated as 28.9 or 21.4 kJ mol-1 higher in energy than the syn-conformer.32 The fact that the anti-conformer is obtained in the β-form shows that intermolecular interactions play a significant role in stabilizing this conformation, as indicated by the significantly different fingerprint plots (Figure 12) of the two polymorphs. The optimized structures of the two molecules in form R are virtually identical with respect to geometric parameters (with a true centrosymmetric relationship between them) and differ by only 0.3 kJ mol-1 in energy. The optimized β form molecule is more than 5 kJ mol-1 lower in energy than the two optimized R form molecules. These values mean that in the absence of any intermolecular interactions the two conformations found in form R would be expected to be found with almost equal probability (8.3:8.4), while 80% of the bezafibrate molecules would be in the form β conformation. However, the differences between the optimized gas phase and crystal structures show that intermolecular interactions, in particular the O-H · · · O and N-H · · · O hydrogen bonding, play a large role in stabilizing the conformations found in the crystal. The stabilization involved
2654
Crystal Growth & Design, Vol. 9, No. 6, 2009
Lemmerer et al.
Figure 13. Fingerprint plots of the three crystallographically independent bezafibrate molecules. The labels are referred to in the text. Table 4. Relative Energies of Conformations (kJ mol-1) form R (molecule A)
conformation relative energies of conformations (kJ mol-1)a,b relative energies of optimized conformations (kJ mol-1)a,c torsion angles (°) in optimized structures (experimental value)
H-bond energies of O-H · · · O dimers (kJ mol-1)b,d H-bond energies of N-H · · · O dimers (kJ mol-1)b,d relative energies of four molecule clusters (kJ mol-1)a,e
form R (molecule B)
form R (both molecules)
form β
5.2
7.9
0
5.5
5.2
0
O4-C19-C16-O2
-2.7 [33.4(3)]
0.4 [-24.5(3)]
164.3 [135.6(2)]
C7-N1-C8-C9 C19-C16-O2-C13 C16-O2-C13-C12
157.9 [-175.6(3)] 87.4 [77.6(3)] 126.3 [145.8(3)]
-159.4 [-178.0(3)] -87.2 [-86.4(3)] -119.9 [-132.1(3)]
-88.1
-79.9 [-103.7(2)] -93.7 [-66.7(2)] -113.2 [-161.2(2)] -37.1
-15.9
-22.6
-245.33
-124.81
a Relative to energy of form β. b Calculated at the B971/6-311G(d) level of theory with Counterpoise corrections, H-atom positions optimized at the B3LYP/6-311G(d) level of theory. c Calculated at the B971/6-311G(d) level of theory with Counterpoise corrections, all atom positions optimized at the B3LYP/6-311G(d) level of theory. d Calculated according to eq 1. e Calculated according to ∆E ) E(cluster) - 2 × E(molecule1) - 2 × E(molecule2).
Figure 14. Suggested schematic energy vs temperature diagram for form R and β of bezafibrate. Tp,βfR indicates the phase transition from form β to R at constant pressure, mpβ is the melting point of form β and mpR indicates the melting point of form R observed at 185°C.
in these interactions was investigated by calculating the differences in energy between hydrogen bonded dimers or tetramers and the individual component conformations. The conformations of the molecules found in the crystal structures (with H-atom positions optimized) were used as reference for the calculations. The two O-H · · · O hydrogen bonds found within the dimeric asymmetric unit of form R were found to lead to the largest stabilization (88 kJ mol-1, although this value is exaggerated by the fact that the dimer contains two of these interactions,
each contributing 44.6 kJ mol-1), whereas the N-H · · · O interaction was found to contribute the least to the stabilization of form R. The energies of the four molecules forming each of the hydrogen bonded rings in the two forms show that the R44(16) hydrogen-bonding ring found in form R is more stable than the R44(48) found in form β. This is as a result of the greater hydrogen bond energy found for the O-H · · · O interactions in form R. Furthermore, the stabilization is larger than the sum of the O-H · · · O and N-H · · · O hydrogen bonding energies, particularly for form R, suggesting that there is a hydrogen bond cooperativity effect in action in the smaller hydrogen bonded ring of form R. The difference in energies of the tetramers suggests that although in gas phase (and possibly also in solution, although solvent model calculations were not performed) bezafibrate molecules are more stable in the conformation found in form β, in the crystal phase form R is more likely. It should nevertheless be noted that the role of the C-H · · · π interactions has not been investigated in this study, but they are expected to further stabilize both crystalline forms.
4. Conclusion This investigation of the dimorphic behaviour of bezafibrate is an ideal example of crystal habit mimicking crystal form with the needle-like (form R) and block-shaped (form β) crystals reflecting major structural differences. The polymorphic behavior of the antihyperlipoproteinemic drug bezafibrate has been extensively investigated and expanded upon from previous
Polymorphs of Antihyperlipoproteinemic Bezafibrate
studies. Using a number of solid-state techniques, we characterized the two concomitant polymorphs R and β using spectral methods (IR), thermal analysis (DSC, TGA and HSM) and diffraction methods (PXRD and single crystal X-ray diffraction). From the information gained, a schematic and qualitative energytemperature diagram can be generated, shown in Figure 14. The system is enantiotropic and converts from form β to form R at 160.7 °C with subsequent melting of form R at 185°C. The effect of solvent and the templating effect of iso-nicotinamide on the form obtained from solutions are significant. Form R can be obtained exclusively using ethanol. Both forms can be grown concomitantly from 2-butanone, while adding a stoichiometric quantity of iso-nicotinamide to the 2-butanone solution produces a 100% yield of form β. Computational modeling of the gas phase conformations confirms that the two molecules in form R are similar in energy, while the conformation adopted in form β is slightly lower in energy than that in form R. Acknowledgment. Financial support was received from the South African National Research Foundation (NRF) (Grant FA2006030100003). A.L. thanks the NRF for a postdoctoral scholarship (SFP2006061500015). N.B.B. thanks the University of Cape Town for a postdoctoral research fellowship. C.E. thanks the University of Stellenbosch and the NRF for support. M.R.C. thanks the University of Cape Town and the NRF for financial assistance. The authors thank Prof. Joel Bernstein for valuable comments on a preliminary draft of this manuscript, Jinjing Li for help with the variable temperature PXRD, and Dr. Gerhard A. Venter for the microscope images of the crystals.
Crystal Growth & Design, Vol. 9, No. 6, 2009 2655 (3) (4) (5) (6) (7) (8)
(9)
(10)
(11) (12) (13) (14) (15) (16) (17) (18) (19) (20)
Supporting Information Available: Crystallographic information files (cif) and thermal gravimetric analysis (TGA) traces are available for forms R and β of bezafibrate, the complete author list for ref 20 and the barrier to rotation calculations. This information is available free of charge via the Internet at http://pubs.acs.org.
(21) (22)
References
(23)
(1) (a) Bernstein, J. Polymorphism in Molecular Crystals; IUCr Monographs on Crystallography; Oxford Science Publications: Oxford, U.K., 2002; Vol. 14. (b) Vishweshar, P.; McMahon, J. A.; Bis, J. A.; Zaworotko, M. J. J. Pharm. Sci. 2006, 95, 499–516. (c) Shan, N.; Zaworotko, M. J. Drug DiscoVery Today 2008, 13, 440–446. (d) Benghiat, V.; Leiserowitz, L. J. Chem. Soc. 1972, 1763–1768. (e) Ahn, S.; Kariuki, B. M.; Harris, K. D. M. Cryst. Growth Des. 2001, 1, 107– 111. (f) Pal, T.; Kar, T.; Bocelli, G.; Rigi, L. Cryst. Growth Des. 2003, 3, 13–16. (g) Timofeeva, T. V.; Kuhn, G. H.; Nesterov, V. V.; Nsterov, V. N.; Frazier, D. O.; Penn, B. G.; Antipin, M. Y. Cryst Growth Des. 2003, 3, 383–391. (h) Munshi, P.; Venugopala, K. N.; Jayashree, B. S.; Row, T. N. G. Cryst. Growth Des. 2004, 4, 1104–1107. (i) Timofeeva, T. V.; Kinnibrugh, T.; Borbulevych, O. Y.; Averkiev, B. B.; Nesterov, V. N.; Sloan, A.; Antipin, M. Y. Cryst. Growth Des. 2004, 4, 1265– 1276. (j) Rafilovich, M.; Bernstein, J.; Harris, R. K.; Apperley, D. C.; Karamertzanis, P. G.; Price, S. L. Cryst. Growth Des. 2005, 5, 2197– 2209. (k) Blagden, N.; Davey, R.; Dent, G.; Song, M.; David, W. I. F.; Pulham, C. R.; Shankland, K. Cryst. Growth Des. 2005, 5, 2218– 2224. (l) Munshi, P.; Row, T. N. G. Cryst. Growth Des. 2006, 6, 708– 718. (m) Wishkerman, S.; Bernstein, J.; Stephens, P. W. Cryst. Growth Des. 2006, 6, 1366–1373. (n) Porter, W. W., III.; Elie, S. C.; Matzger, A. J. Cryst. Growth Des. 2008, 8, 14–16. (o) Griesser, U. J.; Jetti, R. K. R.; Haddow, M. F.; Brehmer, T.; Apperley, D. C.; King, A.; Harris, R. K. Cryst. Growth Des. 2008, 8, 44–56. (p) Barsky, I.; Bernstein, J.; Stephens, P. W.; Stone, K. H.; Cheung, E.; Hickey, M. B.; Henck, J.-O. Cryst. Growth Des. 2008, 8, 63–70. (2) (a) Bernstein, J.; Davey, R. J.; Henck, J.-O. Angew. Chem. Int. Ed. 1999, 38, 3440–3461. (b) Munshi, P.; Venugopala, K. N.; Jayashree, B. S.; Row, T. N. G. Cryst. Growth Des. 2004, 4, 1105–1107. (c) Munshi, P.; Row, T. N. G. Cryst. Growth Des. 2006, 6, 708–718. (d) Rafilovich, M.; Bernstein, J.; Harris, R. K.; Apperley, D. C.; Karamertzanis, P. G.; Price, S. L. Cryst. Growth Des. 2005, 5, 2197– 2209. (e) ter Horst, J. H.; Kramer, H. J. M.; Jansens, P. J. Cryst. Growth Des. 2002, 2, 351–356.
(24)
(25)
(26) (27)
(28)
(29) (30)
(31)
(32)
Witte, E. C.; Stach, K.; Thiel, M.; Schmidt, F.; Stork, H. 1973. Bourgogne, J.-P.; Sornay, R. 1988. Beyer, P. 1979. Djinovic´, K.; Globokar, M.; Zupet, P. Acta Crystallogr. 1989, C45, 772–775. Mo¨rsdorf, P.; Maier, P.; Schmitt, J.; Ahrens, K.-H. 1994. No labelling of the previously known needle-like form is given. This is often the case for the first reported structure of a system not known to be polymorphic. This is thus assumed to be the R form of bezafibrate. The measured PXRD trace of form β is compared to the calculated (from the single crystal structure reported by Djinovic´ et al.) trace of form R. However, no measured IR spectrum of form R was reported or referred to in the patent. Form β of bezafibrate was obtained serendipitously as part of our program to obtain co-crystals of a number of APIs (containing carboxylic acid functional groups) with nicotinamide and iso-nicotinamide. Form β of bezafibrate was crystallized from an unsuccessful attempt to co-crystallize bezafibrate with iso-nicotinamide from a solution of 2-butanone. COLLECT; Nonius BV: Delft, The Netherlands, 1998. Otwinowski, Z.; Minor, W. In International Tables for Crystallography; Rossman, M. G., Arnold, E.; Kluwer: Dordrecht, 2000; Vol. F. XPREP2, Version 6.14; Bruker AXS Inc.: Madison, Wisconsin, USA, 2003. Farrugia, L. J. J. Appl. Crystallogr. 1997, 30, 565; WinGX. Sheldrick, G. M. SHELXS-97, Program for the Solution of Crystal Structure; University of Gottingen: Germany, 1997. Sheldrick, G. M. SHELXL-97, Program for the Refinement of Crystal Structure; University of Gottingen: Germany, 1997. Spek, A. L. J. Appl. Crystallogr. 2003, 36, 7–13. Farrugia, L. J. J. Appl. Crystallogr. 1999, 32, 837–838; WinGX. Brandenburg, K. Diamond, Version 2.1e.; Crystal Impact GbR: Bonn, Germany. Frisch, M. J. Gaussian 03, Revision B.05; Gaussian, Inc.: Pittsburgh PA, 2003. (a) Becke, A. D. J. Chem. Phys. 1993, 98, 5648–5652. (b) Lee, C.; Yang, W.; Parr, R. G. Phys. ReV. 1988, B37, 785–789. (a) Hehre, W. J.; Ditchfield, R.; Pople, J. A. J. Chem. Phys. 1972, 56, 2257–2261. (b) Hariharan, P. C.; Pople, J. A. Theor. Chim. Acta 1973, 28, 213–222. Hamprecht, F. A.; Cohen, A. J.; Tozer, D. J.; Handy, N. C. J. Chem. Phys. 1998, 109, 6264–6271. (a) Simon, S.; M. Duran, M.; Dannenberg, J. J. J. Chem. Phys. 1996, 105, 11024–11031. (b) Boys, S. F.; Bernardi, F. Mol. Phys. 1970, 19, 553–566. The peaks in the calculated trace are shifted to higher 2θ values due to the temperature difference of the calculated (-100 °C) and measured patterns for form R (163 °C) and form β (138°C). Bernstein, J.; Davis, R. E.; Shimoni, L.; Chang, N.-L. Angew. Chem. Intl. Ed. Engl. 1995, 34, 1555–1573. (a) Leiserowitz, L. Acta Crystallogr. 1976, B32, 775–802. (b) Dinabandhu, D.; Desiraju, G. R. Chem. Asian J. 2006, 1-2, 231– 244. (c) Dinabandhu, D.; Desiraju, G. R. CrystEngComm 2006, 8, 674–679. (a) McKinnon, J. J.; Spackman, M. A.; Mitchell, A. S. Chem.sEur. J. 1998, 4, 2136–2141. (b) McKinnon, J. J.; Mitchell, A. S.; Spackman, M. A. Chem. Commun. 1998, 2071–2072. (c) Spackman, M. A.; McKinnon, J. J. CrystEngComm 2002, 4, 378–392. (d) McKinnon, J. J.; Spackman, M. A.; Mitchell, A. S. Acta Crystallogr., Sect. B 2004, 60, 627–668. (e) McKinnon, J. J.; Fabbiani, F. P. A.; Spackman, M. A. Cryst. Growth Des. 2007, 7, 755–769. Wolff, S. K.; Grimwood, D. J.; McKinnon, J. J.; Jayatilaka, D.; Grimwood, D. J. Crystal Explorer, Version 2.1 (381); University of Western Australia, Australia. (a) Leiserowitz, L. Acta Crystallogr. Sect. B 1976, 32, 775–802. (b) Li, Y.; Houk, K. N. J. Am. Chem. Soc. 1989, 111, 405–4507. (c) Montzka, T. A.; Swaminathan, S.; Firestone, R. A. J. Phys. Chem. 1994, 98, 13171–13176. Cambridge Structural Database, Feb. 2009 update. Search criteria: Fragments containing a COOH group where the OdC-O-H torsion angle had a value between-40 and 40 deg, corresponding to a syn conformation . Allen, F. H. Acta Crystallogr., Sect. B 2002, 58, 380– 388. Sato, H.; Hirata, F. J. Mol. Struct.: Theochem 1999, 461-462, 113–120.
CG8011298
