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Predictable Disorder versus Polymorphism in the Rationalization of Structural Diversity: A Multidisciplinary Study of Eniluracil. Royston C. B. Copley...
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Predictable Disorder versus Polymorphism in the Rationalization of Structural Diversity: A Multidisciplinary Study of Eniluracil Royston C. B. Copley,† Sarah A. Barnett,‡,⊥ Panagiotis G. Karamertzanis,‡ Kenneth D. M. Harris,§ Benson M. Kariuki,§ Mingcan Xu,§ E. Anne Nickels,†,# Robert W. Lancaster,‡ and Sarah L. Price*,‡

CRYSTAL GROWTH & DESIGN 2008 VOL. 8, NO. 9 3474–3481

GlaxoSmithKline (GSK) Research & DeVelopment Ltd, New Frontiers Science Park, Third AVenue, Harlow, Essex, CM19 5AW, U.K., Department of Chemistry, UniVersity College London, 20 Gordon Street, London WC1H 0AJ, U.K., and School of Chemistry, Cardiff UniVersity, Cardiff, CF10 3AT, U.K. ReceiVed May 16, 2008

ABSTRACT: Detailed analysis of X-ray diffraction data from four single crystals of eniluracil, prepared under different crystallization conditions, confirms a picture in which the crystals exhibit different degrees of disorder, which is also suggested by the computed low energy crystal structures. Since several of these crystal structures that effectively differ by an interchange of the oxygen and hydrogen atoms on C(4) and C(6) are essentially equi-energetic, growth errors that may be difficult to reverse are practically inevitable. The structural variations observed for the crystals of eniluracil studied are more appropriately described in terms of variable degrees of disorder rather than polymorphism. Analysis of the computed crystal energy landscape for interchangeable hydrogen-bonded (or other strong) motifs is, therefore, shown to be a valuable complement to X-ray diffraction and solid-state NMR for understanding and characterizing disorder in organic solid state systems. In the case of eniluracil, this detailed picture probably accounts for the challenges in devising a robust production process for this anticancer agent in the 1990s. The specific nature of the disorder accounts for different structures being obtained from powder X-ray diffraction data of different samples, and the possibility of publishable single crystal X-ray refinements also being interpreted as polymorphism rather than disorder. Introduction Polymorphism, the ability of a given molecule to adopt different crystal structures1,2 which are chemically identical but may have different physical properties, has been the subject of much research as it is associated with quality control issues for manufacturing crystalline specialty chemicals, such as pharmaceuticals. Understanding the physical and chemical factors that underlie the formation of different polymorphs of a given molecule is fundamentally important3–5 as, in principle, any solid state property may differ between different polymorphic forms of the same molecule. Various definitions of polymorphism1,6 do not specify the degree of difference between the crystalline forms that is required to constitute polymorphism,7 as opposed to variation in the crystalline samples due to growth conditions, such as disorder or through defects, familiar to all crystallographers. A recent study to distinguish8 between genuine polymorphs and redeterminations of the same organic crystal structure in the Cambridge Structural Database9 on the basis of the similarity of their powder X-ray diffraction patterns showed that there was a significant indeterminate range in a carefully constructed similarity measure.10 A proposal for distinguishing between real polymorphs and the consequences of sample dependent disorder, defects and modulations by an energetic fingerprint of the molecular pairs in the coordination environment,11 provides a useful basis for the discussion of recent controversial cases. In this paper we establish, by detailed analysis of the first single crystal X-ray diffraction data of eniluracil in conjunction with computational crystal structure * Author to whom correspondence should be addressed. Tel: +44 (0)20 7679 4622. Fax: +44 (0)20 7679 7463. E-mail: [email protected]. † GlaxoSmithKline (GSK) Research & Development Ltd. ‡ University College London. § Cardiff University. ⊥ Current Address: Diamond Light Source Ltd, Harwell Science and Innovation Campus, Didcot, Oxfordshire, OX11 0DE, U.K. # Current Address: Inorganic Chemistry Laboratory, University of Oxford, South Parks Road, Oxford, OX1 3QR, U.K.

prediction, that a decade-old industrial problem of solid form diversity arises from variable disorder rather than polymorphism. 5-Ethynyluracil (eniluracil) (Figure 1) is a simple rigid molecule which would have C2V symmetry if C(4)dO(8) and C(6)-H(6) were not different functional groups. Eniluracil was under development by GlaxoWellcome (now GlaxoSmithKline (GSK)) as a potential anticancer agent in the 1990s, and is currently under development by Adherex,12 as it inhibits the enzyme dihydropyrimidine dehydrogenase, thus allowing more effective administration of the anticancer agent 5-fluorouracil.13 The solid form investigations showed that eniluracil formed polycrystalline agglomerates, themselves exhibiting a variety of habits and, since crystals of eniluracil suitable for single crystal X-ray diffraction could not be obtained at that time, two independent studies were carried out to determine the structure from powder X-ray diffraction data. One structure determination14 led to a polar ribbon structure (Figure 1a) in space group P21 with the ethynyl groups interdigitating such that polar sheets are formed and with evidence for disorder in the stacking of these sheets. The second structure determination15 also considered this polar ribbon structure, but finally, although not unequivocally, proposed a structure with nonpolar ribbons in P21/c (Figure 1b) as optimizing the match of the powder pattern under the guidance of a computational search for possible crystal structures. Thus, two independent powder X-ray diffraction studies proposed crystal structures with different relative positions of C(4)dO(8) and C(6)-H(6) and which would clearly be classified as polymorphs.1,11 The two structures were distinguished on the basis of some weak, low-angle peaks, the existence and intensity of which were rather sample dependent. Crystals of eniluracil suitable for single crystal X-ray diffraction studies have now been obtained by different crystallization methods. Careful analysis of the X-ray diffraction data from four crystals, combined with a computational search for the low energy crystal structures employing a realistic model for the intermolecular forces, give a rather different and more

10.1021/cg800517h CCC: $40.75  2008 American Chemical Society Published on Web 08/06/2008

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Figure 1. Alternative variants on the symmetric ribbon hydrogen bonding motif (i.e., the hydrogen bonding arrangement shows mirror symmetry about the uracil C(2)dO(7) axis) for eniluracil (numbering scheme included) showing (a) polar ribbon and (b) nonpolar ribbon.

detailed picture of the solid state of eniluracil. These observations provide a clear example to assist the scientific11 distinction of polymorphism from other possible structural variations found in organic crystal structures, with all the implied legal connotations that this has in the patent protection of solid forms. Experimental Procedures The eniluracil sample used for this study was taken from GlaxoSmithKline batch 0776C 85UAV. Detailed X-ray diffraction studies were carried out on four single crystals, henceforth referred to as 1 to 4, from the following crystallization conditions: crystal 1, slow evaporation of an aqueous THF solution to which a single drop of DMSO had been added; crystal 2, slow evaporation of an aqueous acetone solution to which a single drop of DMSO had been added; crystals 3 and 4, slow cooling (from raised temperature), with seeding, of aqueous acetone solutions. Single Crystal X-ray Diffraction. The single crystal X-ray diffraction data for 1 to 4 were measured at 150 K using a Bruker AXS SMART 6000 diffractometer with graphite monochromated Cu KR radiation (λ ) 1.54178 Å) from a normal focus sealed tube source. The crystal to detector distance was 5.0 cm and between 9 and 15 runs of 970 frames (∆ω ) 0.2°) were collected with the detector set at either a low or high angle position (2θ ) -40.00° and -108.00° respectively). Variations in the number of runs and frame exposure times are detailed in the Supporting Information. The data were integrated and the final unit cell parameters were obtained using SAINT+.16 All data were corrected for absorption using the SADABS program,17 with separate results being considered for point groups 2 and 2/m. The crystal data corresponding to the best models that could be obtained for 1 and 4 are given below (further details on crystals 2 and 3 can be found in the Supporting Information). Crystal information for 1: Colorless truncated octahedron, 0.17 × 0.14 × 0.12 mm, C6H4N2O2, M ) 136.11, monoclinic, space group P21/n (alt. P21/c, #14), a ) 9.0169(3) Å, b ) 6.8086(2) Å, c ) 9.3777(3) Å, β ) 101.2085(17)°, V ) 564.74(3) Å3, Z ) 4, Dcalc ) 1.601 Mg m-3, F(000) ) 280, µ(Cu KR) ) 1.060 mm-1. Crystal information for 4: Colorless block, 0.48 × 0.34 × 0.22 mm, C6H4N2O2, M ) 136.11, monoclinic, space group P21 (#4, origin shifted), a ) 9.0188(2) Å, b ) 6.8105(2) Å, c ) 9.3622(2) Å, β ) 101.1887(14)°, V ) 564.12(2) Å3, Z ) 4, Dcalc ) 1.603 Mg m-3, F(000) ) 280, µ(Cu KR) ) 1.061 mm-1. All the structures were initially solved by direct methods and refined using full-matrix least-squares procedures which minimized the function Σw(Fo2 - Fc2)2. The SHELXTL software package18 was used throughout. Attempts were made to model all four structures in both of the space groups above and the outcome is summarized for each set of data in the Supporting Information. In the P21/n structure, there is one molecule in the asymmetric unit which is disordered between two orientations corresponding to a 180° rotation about the ethynyl axis that is, there is effectively an interchange of the oxygen and hydrogen atoms on C(4) and C(6). A disorder treatment of just these two atoms proved unsatisfactory in terms of both poor geometry and atomic

displacement parameters and a much improved model was obtained by having two complete molecules in the two orientations (Figure S1a, Supporting Information). Due to the nature of this disorder, there was concern that either the unit cell and/or the space group choice were incorrect, but exhaustive efforts to improve these were unsuccessful. Common isotropic displacement parameters were used for corresponding atoms in the two components and these were refined with the coordinates of the non-hydrogen atoms and a term establishing the site occupancy factor for each component. This site occupancy factor was then fixed and the coordinates and anisotropic displacement parameters were refined in conjunction with geometric and vibrational restraints. An analogous approach was attempted with the P21 model, in which one independent molecule was similarly disordered while the second was fully ordered (Figure S1b, Supporting Information). In all cases, hydrogen atoms were included in calculated positions and refined using a riding model. Isotropic displacement parameters for the hydrogen atoms were used as appropriate multiples of Ueq for the attached non-hydrogen atom. Crystal structure diagrams were produced using SHELXTL19 for the displacement ellipsoid plots (Figure S1, Supporting Information) and Mercury20 for all packing diagrams. Computational. The molecular geometry was obtained by an ab initio optimization at the MP2/6-31G(d,p) level of theory using GAUSSIAN9821 and then held rigid throughout the generation and lattice energy minimizations of all crystal structures. Nearly four thousand densely packed structures were generated using MOLPAK22 in 38 common coordination environments, with one crystallographically independent molecule, spanning 18 of the most frequently occurring space groups. The lattice energy was subsequently minimized with respect to the unit cell parameters and molecular positions and orientations, using the program DMAREL.23,24 In the evaluation of the lattice energy, the electrostatic contribution was calculated using a set of atomic multipoles calculated from a distributed multipole analysis of the MP2/6-31G(d,p) charge density25 to ensure an accurate description of the electrostatic interactions,26 including the directionality of hydrogen bonding27–29 for the distinct polar hydrogens and hydrogen bond acceptors in the eniluracil molecule. All other terms in the lattice energy were calculated using an empirical atom-atom potential of the form:

U)

∑

i∈1,k∈2

(AιιAκκ)1⁄2exp(-(Bιι + Bκκ)Rik ⁄ 2) -

(CιιCκκ)1⁄2 R6ik

where atom i in molecule 1 is of type ι, and atom k in molecule 2 is of type κ. The parameters for atomic types N, O and HC (nonpolar) were taken from the work of Williams30,31 and HN from the extension of this potential in conjunction with a DMA-based electrostatic model to hydrogen bonded crystals.32 The k ) 0 rigid-body harmonic phonons and elastic constants of the unique low energy structures were also calculated from the second derivative matrix.33 Hence, this determination of the low energy crystal structures of eniluracil is based on a significantly more rigorous modeling of the intermolecular forces than

3476 Crystal Growth & Design, Vol. 8, No. 9, 2008 the Polymorph Predictor search which was used to aid one of the previous structure determinations from powder X-ray diffraction data.15

Results Variability of Single Crystal X-ray Diffraction Results. On initial examination of the single crystal X-ray diffraction data, it was believed that two polymorphs with different lattice parameters had been found since crystal 2 indexed to a unit cell with approximately half the volume of crystal 1. This smaller unit cell corresponds to that reported for the determination by powder diffraction, PXRD_1.14 However, careful examination of the frame data for crystal 2 revealed the presence of additional weak reflections that were not explained by the smaller unit cell and reintegration with a unit cell analogous to that obtained for crystal 1 gave a superior result. Analysis of the intensity data showed this new unit cell to be pseudo B-centered, with the smaller unit cell corresponding to a true centring. The structures of both 1 and 2 were modeled in space group P21/n, as reported previously for the isomorphous disordered structures of bromouracil34 and chlorouracil.34 In this space group, there is disorder of the molecule corresponding to an approximate 2-fold rotation about the ethynyl axis. Further data collections on crystals 3 and 4, grown from slow cooling experiments as opposed to evaporation, were carried out to observe how the site occupancy factors of the two components vary in crystals prepared by different crystallization procedures. Both data sets gave satisfactory results in space group P21/n, with all four crystals having comparable unit cell dimensions (Table S2, Supporting Information). A relatively small variation was noted in the site occupancy factors of the major components [1, 0.742(3); 2, 0.705(3); 3, 0.738(3); 4, 0.841(3)], but these were clearly not 0.50 which would have allowed modeling in P21/m. It should be stressed that, at this stage, all four crystals appeared to represent the same solidstate form and all would probably have passed the normal crystallographic tests applied by most referees and journals. At this point, a reduction in symmetry to space group P21 was tried (corresponding to two independent molecules in the asymmetric unit). Surprisingly, this was found to lead to ordering of one of the two independent molecules for crystals 3 and 4. This would imply that the structures for crystals 3 and 4 are no longer equivalent to those for 1 and 2. The improvement for 3 was debatable as it could be argued that the additional refined parameters required in the noncentrosymmetric space group were responsible for the drop in the R1 value from 0.0563 to 0.0535. For crystal 4, however, the improved refinement in space group P21 was clear, with the R1 value decreasing from 0.0481 to 0.0367, no unexplained residual electron density and a very stable refinement. Using the P21 model for 1 and 2 was unsuccessful, even when applying strict restraints. It was, therefore, concluded that the best structural model for crystals 1 and 2 was P21/n, whereas for crystal 4 it was P21. For crystal 3, the structure could be described adequately using either structural model. The modeling possibilities for this molecule are extensive and, while the utmost care was taken to achieve the best result, it is feasible that an alternative treatment could lead to further ordering. The point of interest here is that the diffraction data appear to suggest that a different model is needed for crystal 4 in contrast with the other crystals studied. The crystal structures of 1 and 4 are characterized (Figure 1) by classical hydrogen-bonded ribbons that utilize both N-H groups but only one of the carbonyl (C2dO7) functionalities; we term this structure the symmetric ribbon due the symmetry of the hydrogen bonding motif about the carbonyl in the 2

Copley et al.

position. The ribbon is formed such that the ethynyl groups protrude perpendicular to the direction of the hydrogen bonded core, as is commonly found in other 5-substituted uracils.34–39 Since the second carbonyl plays no role in this hydrogen bonding motif, it can be seen that the eniluracil molecules can add to this ribbon either way around such that the R22(8) rings either all involve both N1 and N3 leading to polar ribbons (Figure 1a) or alternatively involve only N1 or N3 resulting in nonpolar ribbons (Figure 1b). For both components of structure 1 and the major component of structure 4, the ribbons formed are nonpolar (Figure 2a), that is, the R22(8) rings involve either only N1-H1 · · · O7 hydrogen bonds or only N3-H3 · · · O7 hydrogen bonds. The minor component of structure 4, however, has R22(8) rings which include both N1 and N3 type nitrogens, thus leading to polar ribbon formation. The disorder model for structure 1 is shown in Figure 2b and that for structure 4 in Figure 2c. Because the two disorder components in crystal 4 are different, the resulting crystallographic structure combining these components (Figure 2c) shows that that one of the two molecules appears ordered as opposed to crystal 1 which appears fully disordered (Figure 2b). Additionally, it can be seen that there is a slight deformation from linearity in the ethynyl group (C5-C9-C10 angle range is 176.9(17)-178.0(3)°) in order to improve the C≡C-H · · · O8 interaction. In all cases, however, these hydrogen bonded ribbons interdigitate to form sheets (parallel to [1j 0 3]) with a small vertical gap of approximately 0.4 Å between the ribbon layers and a 3.2 Å gap between sheets. Although the stacking is identical in both 1 and 4, consideration of the relative positions of C(4) and C(6) means that the sheets of nonpolar ribbons stack such that only alternate layers can be overlaid by translation (Figure S3f, Supporting Information), whereas for the sheets composed of polar ribbons all of the layers can be overlaid (Figure S3a, Supporting Information). Computational Search for Low Energy Crystal Structures. A large number of distinct eniluracil crystal structures were found in the computational search with 20 unique structures within 5 kJ mol-1 of the global minimum (Figure S2 and Table S5, Supporting Information). Six distinct structures based on the ribbon motifs in Figure 1 were up to 2 kJ mol-1 more stable than the lowest energy structure (am40, Table S5 and Figure S4c, Supporting Information) that utilizes both carbonyl groups as hydrogen bond acceptors. Thus, eniluracil is another example of the subtle balance between hydrogen bonding and close packing which can lead to unused acceptors in the crystal structure.40,41 The packing arrangements of the four, distinct, near-planar sheets of interdigitating ribbons, all found within the five most stable structures (Table S5, Supporting Information), are shown in Figure 3. The ribbons are all based on the symmetric ribbon hydrogen bonded motif, and it is notable that these structures look identical if the difference between C(4)dO(8) and C(6)-H(6) is not considered. If the distinction is made, however, then the structures in Figure 3 divide into those with polar and nonpolar ribbons, as illustrated in Figure 1. These ribbons can pack in two ways to give four distinct sheets. The global minimum (fc85, Figure 3b) consists of polar ribbons that interdigitate in an antiparallel fashion to form nonpolar sheets. The second most stable structure (ak56, Figure 3d) has nonpolar ribbons, interdigitated in an antiparallel manner. However, reversing the direction of the second nonpolar ribbon, such that both ribbons now run in the same direction, gives a very similar structure with hardly any loss in stability (ab20, Figure 3c). The fourth alternative, with parallel polar ribbons, results in a polar

Multidisciplinary Study of Eniluracil

Crystal Growth & Design, Vol. 8, No. 9, 2008 3477

Figure 2. The sheet arrangement (a) adopted by the major component of crystal 1 of eniluracil with displacement ellipsoids drawn at the 50% probability level (C - dark gray, H - light gray, N - blue, O - red, hydrogen bonds - green). This arrangement also corresponds to the minor component of crystal 1 and the major component of disorder for crystal 4. The disordered packing of eniluracil in (b) crystal 1 in P21/n (the major component of disorder is shown in red and the minor component in pink) and (c) crystal 4 in P21 (the major component of disorder is shown in red and the minor component in pink while the fully ordered molecules are colored blue). An origin shift was applied to the P21 model of 4 to allow direct comparison of the coordinates.

sheet (ah27, Figure 3a) which is only 0.91 kJ mol-1 less stable than the global minimum. These four alternative sheet structures can stack in a variety of ways that, again, only differ when the distinction between C(4)dO(8) and C(6)-H(6) is made, still with very little energy difference (Figure S3a-g, Supporting Information). The nonpolar antiparallel sheets can stack in the same way (fc83, Figure S3e, Supporting Information) rather than the opposite, alternating stacking of ak56 (Figure S3f, Supporting Information) and the components of 1, to give a crystal structure that is only 2.5 kJ mol-1 less stable. Similarly, the fourth most stable structure (ak75, Figure S3b, Supporting Information) has the same polar sheets as ah27 (Figure S3a, Supporting Information), but stacked in a nonpolar fashion that is, at most, only 0.4 kJmol-1 less stable. (For polar crystal structures, such as ah27, there may be a surface destabilizing contribution,42 which has been neglected in this work.) Thus, there are a number of variations on the symmetric ribbon crystal structure within a small energy range of the global minimum, even within a relatively limited search (Z′ ) 1 in common space groups). The equivalence of the interaction energies for a range of coordination environments (distinguished by CdO and C-H) implies that there are a large number of possible structures consisting of mixtures of the symmetric

ribbon motifs, sheet structures and polytypic variations in stacking that will be approximately equi-energetic. Comparison of Computational and Experimental Results. The second lowest energy predicted structure (ak56) is an excellent approximation to both components of the P21/n structure of 1, determined by single crystal X-ray diffraction, with a root mean squared difference in the non-hydrogen atoms of the 15 molecule coordination sphere (rms15), calculated using COMPAK,43 of 0.144 Å for the structure corresponding to the major component of disorder and 0.145 Å for the minor component. Some of this discrepancy arises from the differences in the molecular structures, mainly due to the deformation of the ethynyl group which is slightly distorted from linearity to improve the C(9)tC(10)-H(10) · · · O(8)dC(4) interaction, as the ab initio molecular model has an rms difference in the position of the non-hydrogen atoms of 0.031 Å with the molecule of the major component, and 0.038 Å for the minor component.20 The P21 structure found for crystal 4 can be considered as a combination of two of the predicted structures: ak56 (c.f. 1) which corresponds to the major disorder component (rms15 of 0.146 Å), and a polar structure very similar to the third lowest energy structure, ah27, when the minor component is considered (rms15 of 0.141 Å). We note that the predicted structure ah27

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Figure 3. The packing arrangement in the four distinct sheet structures in lowest energy crystal structures found in the search (a) parallel polar ribbons in ah27 (P21; rank #3, -115.51 kJ/mol) corresponding to minor component of crystal 4, (b) antiparallel polar ribbons in fc85, the global minimum energy structure (P21/c; rank #1, -116.42 kJ/mol), (c) parallel nonpolar ribbons in ab20 (P1j; rank #5, -114.87 kJ/mol) and (d) antiparallel nonpolar ribbons in ak56 (P21/c; rank #2, -116.12 kJ/mol) corresponding to the ordered experimental structure 1. All structures are indistinguishable if the relative positions of the C(4)dO(8) and C(6)-H(6) groups are ignored, but the distinction gives the polarity indicated by the pink and green arrows.

matches that of PXRD_1,14 and the unit cell of this structure corresponds to the B-centered version of the experimental singlecrystal unit cells. Thus, the experimental structures are comprised of the ordered low energy computed structures. Discussion The experimental crystal structures of eniluracil appear intrinsically disordered; detailed analysis of single crystal X-ray diffraction data shows that the degree of disorder varies between crystals prepared under different crystallization conditions and the production of a fully ordered structure based on exhaustive experimental work seems a practically unobtainable ideal. It is apparent44 (Figure 1) that the disorder cannot be local disorder of the orientation of molecules about the ethynyl axis, as this would bring two carbonyl oxygen atoms into close contact45 and, if dynamic disorder, would break four hydrogen bonds during a sterically implausible transition. A more plausible model for the disorder is suggested by the calculations. The single crystal predominantly grows as nonpolar ribbons which interdigitate such that the ribbons

are antiparallel but, occasionally, they interdigitate in the “wrong” direction, that is, the predominant structure is as illustrated in Figure 3d (ak56) but with some interdigitation occurring as in Figure 3c (ab20). While these variations in the sheets of nonpolar ribbons are more consistent with the single crystal X-ray diffraction results, it is also plausible that, under certain growth conditions, polar ribbons, or ribbons whose character changes at a vacancy defect, could form. The similar energies of the polar ribbon variants of essentially the same crystal structure suggest that these could readily be incorporated in the structure. Once a ribbon had formed and interdigitated within a sheet, this growth error could only be corrected by the removal of the ribbon from the growing crystallite requiring a considerable rearrangement. Since the various sheets can stack in the same way, with the C(4)dO(8) and C(6)-H(6) distinction barely affecting the energy, alternative stacking arrangements of the same sheet can also lead to disorder, and such changes, as well as differences in ribbon type and interdigitation, may also be propagated through the stacking.

Multidisciplinary Study of Eniluracil

Figure 4. The simulated powder X-ray diffraction patterns for the five lowest energy predicted structures based on the symmetric ribbon packing and the disordered single crystal X-ray structures 1 (P21/n) and 4 (P21). All powder patterns were calculated using PLATON57 with λ ) 1.54056 Å.

The occurrence of growth mistakes would be extremely dependent on the growth conditions, and could be of sufficient extent and frequency to hinder the growth of the crystals. Since it has been noted that disorder is more prevalent in cases where problems of crystal growth have required the use of synchrotron radiation,46 it seems likely that the disorder would be greater in microcrystalline than in our single crystal samples. In retrospect, it seems likely that the problems associated with obtaining a robust process in the 1990s for producing solid eniluracil of a consistent size and shape during the drug development phase, as well as attaining single crystals, arose because of this underlying tendency to disorder. Disorder or Polymorphism in Microcrystalline Samples? The disorder models established from the single crystal X-ray diffraction data are particularly significant given the results of the powder X-ray diffraction work undertaken previously. The two structures reported from powder X-ray diffraction unequivocally represent different polymorphs since PXRD_215 is an ordered variant of the nonpolar ribbon structure 1 and ak56, while PXRD_114 is the polar ribbon structure (ah27). However, one determination15 noted the sample dependence of some weak reflections which are critical for differentiating between these structures while the other14 suggested evidence for stacking disorder. It is unlikely that the two microcrystalline samples were identical as many crystallization batches, differing in agglomeration and habit as observed by scanning electron microscopy, were produced during the 1990s development phase. However, it seems probable that the two structures derived from powder X-ray data represent just two of the ordered extremes of a range of disordered structures produced under various crystallization conditions. We have attempted to detect the disorder using highresolution solid state 13C NMR, in order to obtain independent evidence for variation in the degree of disorder between different microcrystalline samples. However, no disorder could be detected, undoubtedly because the differences in the local environment of 13C nuclei in even the ethynyl carbons do not give rise to differences in isotropic 13C chemical shifts that are sufficiently large to be detected (see Figure 3). Figure 4 shows that the simulated X-ray diffraction powder patterns for the single crystal structures of 1 and 4 are virtually

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indistinguishable, and only differ subtly from the predicted patterns for many of the computed symmetric ribbon structures. A normalized similarity measure8,47 of powder patterns gives a similarity coefficient between 1 and 4 of 0.997. This would clearly classify the two structures in different space groups as not being polymorphs as a coefficient of 0.97 appeared to be the approximate cutoff8 for distinguishing between polymorphs and redeterminations of structures in the Cambridge Structural Database after volume normalization to minimize the effect of temperature differences. Further, simulated powder pattern similarity coefficients (Tables S6 and S7, Supporting Information) for ordered variants of the experimental structures with C(4)dO(8) and C(6)-H(6) interchanges (Figures 3 and S3, Supporting Information) are around this cutoff of 0.97, whereas the other low energy packings are clearly dissimilar with coefficients of less than 0.8. In particular ah27 and ak56, which correspond to PXRD_1 and PXRD_2, respectively, have a similarity coefficient8 of 0.98. Thus, the simulated powder patterns are fully consistent with there being a range of disordered structures for eniluracil rather than polymorphism, and demonstrate the challenges of using powder X-ray diffraction to distinguish between structures in which disorder involves only a fraction of the atoms present in the structure. Indeed, we note a previous case48 in which a completely satisfactory Rietveld refinement of powder X-ray diffraction data was obtained for an ordered structural model, but (guided by evidence of disorder from solid state NMR) an improved Rietveld refinement was obtained for a disordered structural model. Computational Prediction of Disorder. The two disordered single crystal structures of eniluracil can be produced by mixing motifs from approximately equi-energetic computed structures by plausible errors in crystal growth. There are other examples where computed crystal landscapes, which show structures with interchangeable motifs within a small energy range, rationalize a disordered solid-state structure. For example, predicted equienergetic distinct stackings of identical sheets helped characterize the disordered R3jm form 2 of chlorothalonil49 and explain why aspirin50 can either be found as having two polymorphs51 or as an intergrowth of the corresponding polymorphic domains.52 The occurrence of almost equi-energetic stackings of the same sheet is highly likely to produce stacking errors or polytypism. Different packings of antiparallel stacked dimers of caffeine are so close in energy53 that the inherently disordered structure of anhydrous caffeine is plausible. Thus, the analysis of the low energy computed structures can readily show whether there are possibilities for interchanging the dominant strongly bound motifs such that disordered structures are highly probable. The nature of the disorder is important in determining whether the rearrangement to the most stable ordered form is facile or so difficult that ordered crystals are practically unobtainable. The latter appears to be the case for eniluracil and it is not unique; the published structures of 5-chloro- and 5-bromouracil are34 isomorphic with the P21/n structure 1 of eniluracil, displaying the same disorder. The computed low energy structures have been contrasted with the experimental structures for other 5-substituted uracils54 which demonstrates that the nature of the substituent influences the relative energies of the hydrogen bonded motif and that less symmetric substituents like hydroxyl and ethyl adopt an ordered symmetric ribbon motif. Relating computed low energy crystal structures to either distinct polymorphs or various types of disorder and modulated structures requires further investigation. Other correlations between the nature of a multitude of computed low energy

3480 Crystal Growth & Design, Vol. 8, No. 9, 2008

crystal structures and solid state transformations include the observation of an intermediate low-symmetry disordered phase of cyclopentane,55 and the polyamorphism to polymorphism conversion of carbonic acid.56 The significance of the present study of eniluracil is that the exceptionally detailed single crystal X-ray diffraction work, following a large solid form development program spanning many years, provides strong support for the particular static disorder model that can be derived by analyzing the computed structures. Forewarning of the possibility of this type of disorder could have aided the experimental developmental studies, as well as providing the molecular picture that has helped explain the ambiguities in the diffraction results. Thus, computing the energetically feasible crystal structures has considerable potential for unravelling the solid state behavior in systems where growing single crystals proves highly problematic. Concluding Remarks Structures determined from single crystal X-ray diffraction data for four separate crystals of eniluracil show a variable degree of disorder, which can be explained as errors in the interdigitation and stacking of the hydrogen bonded ribbons. This disorder is rationalized by the computed crystal energy landscape, which shows that both nonpolar and polar hydrogen bonded ribbons can interdigitate to form a variety of sheets and stackings resulting in a range of crystal structures which differ little in their lattice energies and simulated powder X-ray diffraction patterns, but have very distinct relative positions of the C(4)dO(8) and C(6)-H(6) groups. The range of models that can fit the X-ray diffraction data raises the question as to whether structures with a different space group but the same unit cell parameters automatically constitute different polymorphs, without consideration of the packing arrangement. Certainly, there are no practical differences between single crystals 1 and 4 although they are better described by the P21/n or P21 crystallographic models. The differences between their simulated powder patterns are subtle, and even those between the various idealized extremes in the computed structures are below the level usually reported in patents. The solid state chemistry of eniluracil appears to be one of variable disordered structures which should be carefully distinguished from polymorphism.7,11 However, variability in physical properties arising from differing degrees of disorder, or other forms of structural modulation, has important implications for process development and the generation of robust manufacturing processes. The ability of the computed crystal energy landscapes to warn of the possibility and nature of a variable disordered solid state has been shown to be a valuable complementary technique for characterizing the organic solid state. Sample dependent disorder can be readily confused with polymorphism, as demonstrated by the earlier powder X-ray diffraction work on eniluracil, arriving at two solutions with slightly different hydrogen bonded ribbons in different unit cells, and the single crystal structures being acceptably modeled in different space groups. A multidisciplinary approach of the type applied here may often be required to develop a detailed understanding of the distinction between polymorphism and disorder for specific cases. Acknowledgment. We would like to thank the members of GSK who have assisted with this reappraisal of the solid state of eniluracil, particularly Mark Coleman. In addition we thank Lucie Deprez for crystallization work; Prof. Derek Tocher, Dr. Maryjane Tremayne, and other members of the “Control and

Copley et al.

Prediction of the Organic Solid State” (CPOSS project www. cposs.org.uk) group for useful discussions. CPOSS is funded by the Basic Technology program of the Research Councils UK. Supporting Information Available: CIFs for crystals 1 and 4; S1 crystal structure information for crystals 1-4, including thermal ellipsoid plots for crystals 1 and 4; S2 further details of the computed crystal structures, including structure diagrams and similarity calculations. Computed crystal structures are stored on the STFC e-Science center dataportal and are available from the authors on request. Full ref 21. This material is available free of charge via the Internet at http:// pubs.acs.org.

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Multidisciplinary Study of Eniluracil

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