II. EXPERIMENTAL

Flumethasone was a commercial reagent, purchased from TargetMol (Batch #T1124), and was used as received. The white powder was packed into a 0.5 mm diameter Kapton capillary and rotated during the measurement at ~2 Hz. The powder pattern was measured at 298(1) K at the Wiggler Low Energy Beamline (Leontowich et al., Reference Leontowich, Gomez, Moreno, Muir, Spasyuk, King, Reid, Kim and Kycia2021) of the Brockhouse X-ray Diffraction and Scattering Sector of the Canadian Light Source using a wavelength of 0.819826(2) Å (15.1 keV) from 1.6 to 75.0° 2θ with a step size of 0.0025° and a collection time of 3 min. The high-resolution powder diffraction data were collected using eight Dectris Mythen2 X series 1 K linear strip detectors. NIST SRM 660b LaB₆ was used to calibrate the instrument and refine the monochromatic wavelength used in the experiment.

The pattern was indexed using JADE Pro (MDI, 2024) on a primitive monoclinic unit cell with a = 6.46144, b = 24.71548, c = 12.15613 Å, β = 90.67°, V = 1941.17 Å³, and Z = 4. The suggested space group was P2₁, which was confirmed by the successful solution and refinement of the structure. A reduced cell search of the Cambridge Structural Database (Groom et al., Reference Groom, Bruno, Lightfoot and Ward2016) with the chemistry C, H, F, and O only yielded no hits.

A structural model of the flumethasone molecule was downloaded from PubChem (Kim et al., Reference Kim, Chen, Cheng, Gindulyte, He, He, Li, Shoemaker, Thiessen, Yu, Zaslavsky, Zhang and Bolton2023) as Conformer3D_COMPOUND_CID_16490.sdf. It was converted to a *.mol2 file using Mercury (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020). The crystal structure was solved using Monte Carlo simulated annealing techniques as implemented in EXPO2014 (Altomare et al., Reference Altomare, Cuocci, Giacovazzo, Moliterni, Rizzi, Corriero and Falcicchio2013), using two molecules as fragments and a bump penalty.

Rietveld refinement was carried out with GSAS-II (Toby and Von Dreele, Reference Toby and Von Dreele2013). Only the 3.5–50.0° portion of the pattern was included in the refinements (d _min = 0.970 Å). The y-coordinate of F1 was fixed to define the origin. All non-H bond distances and angles were subjected to restraints, based on a Mercury/Mogul Geometry Check (Sykes et al., Reference Sykes, McCabe, Allen, Battle, Bruno and Wood2011; Bruno et al., Reference Bruno, Cole, Kessler, Luo, Motherwell, Purkis, Smith, Taylor, Cooper, Harris and Orpen2004). The Mogul average and standard deviation for each quantity were used as the restraint parameters. The restraints contributed 4.8% to the overall χ². The hydrogen atoms were included in calculated positions, which were recalculated during the refinement using Materials Studio (Dassault Systèmes, 2023). The U _iso of the heavy atoms were grouped by chemical similarity. The U _iso for the H atoms was fixed at 1.3× the U _iso of the heavy atoms to which they are attached. The peak profiles were described using the generalized microstrain model (Stephens, Reference Stephens1999). A 2nd-order spherical harmonic model for preferred orientation was included in the refinement. The background was modeled using a 6-term shifted Chebyshev polynomial, with peaks at 10.37 and 40.08° to model the scattering from the Kapton capillary and any amorphous component of the sample.

The final refinement of 207 variables using 18,601 observations and 174 restraints yielded the residual R _wp = 0.05403. The largest peak (1.69 Å from C85) and hole (1.16 Å from C22) in the difference Fourier map were 0.24(6) and −0.24(6) eÅ⁻³, respectively. The final Rietveld plot is shown in Figure 2. The largest features in the normalized error plot are in the shapes of some of the strong low-angle peaks. These misfits probably indicate subtle changes in the specimen during the measurement.

Figure 2. The Rietveld plot for flumethasone. The blue crosses represent the observed data points, and the green line is the calculated pattern. The cyan curve is the normalized error plot, and the red line is the background curve. The vertical scale has been multiplied by a factor of 20× for 2θ > 25.0°.

The crystal structure of flumethasone was optimized (fixed experimental unit cell) with density functional theory techniques using VASP (Kresse and Furthmüller, Reference Kresse and Furthmüller1996) through the MedeA graphical interface (Materials Design, 2024). The calculation was carried out on 32 cores of a 144-core (768 Gb memory) HPE Superdome Flex 280 Linux server at North Central College. The calculation used the GGA-PBE functional, a plane wave cutoff energy of 400.0 eV, and a k-point spacing of 0.5 Å⁻¹ leading to a 2 × 1 × 2 mesh, and took ~21 h. Single-point density functional calculations (fixed experimental cell) and population analysis were carried out using CRYSTAL23 (Erba et al., Reference Erba, Desmarais, Casassa, Civalleri, Donà, Bush, Searle, Maschio, Daga, Cossard, Ribaldone, Ascrizzi, Marana, Flament and Kirtman2023). The basis sets for the H, C, and O atoms in the calculation were those of Gatti et al. (Reference Gatti, Saunders and Roetti1994), and for F was that of Peintinger et al. (Reference Peintinger, Vilela Oliveira and Bredow2013). The calculations were run on a 3.5 GHz PC using 8 k-points and the B3LYP functional and took ∼7.5 h.

III. RESULTS AND DISCUSSION

There are two molecules in the asymmetric unit of flumethasone. The root-mean-square difference of the non-H atoms in the Rietveld-refined and VASP-optimized structures, calculated using the Mercury CSD-Materials/Search/Crystal Packing Similarity tool, is 0.100 Å. The root-mean-square Cartesian displacement of the non-H atoms in the Rietveld-refined and VASP-optimized structures of molecules 1 and 2, calculated using the Mercury Calculate/Molecule Overlay tool, are 0.064 and 0.083 Å (Figures 3 and 4). The agreements are within the normal range for correct structures (van de Streek and Neumann, Reference van de Streek and Neumann2014). The two molecules have similar conformations (Figure 5); the rms displacement is only 0.091 Å. The asymmetric unit is illustrated in Figure 6. The remaining discussion will emphasize the VASP-optimized structure.

Figure 3. Comparison of the Rietveld-refined (red) and VASP-optimized (blue) structures of molecule 1 of flumethasone. The root-mean-square Cartesian displacement is 0.064 Å. Image generated using Mercury (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020).

Figure 4. Comparison of the Rietveld-refined (red) and VASP-optimized (blue) structures of molecule 2 of flumethasone. The root-mean-square Cartesian displacement is 0.083 Å. Image generated using Mercury (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020).

Figure 5. Comparison of molecule 1 (green) and molecule 2 (orange) of flumethasone. The root-mean-square Cartesian displacement is 0.091 Å. Image generated using Mercury (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020).

Figure 6. The asymmetric unit of flumethasone, with the atom numbering. The atoms are represented by 50% probability spheroids. Image generated using Mercury (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020).

All bond distances, bond angles, and torsion angles fall within the normal ranges indicated by a Mercury Mogul Geometry check (Macrae et al., Reference Macrae, Sovago, Cottrell, Galek, McCabe, Pidcock, Platings, Shields, Stevens, Towler and Wood2020). Quantum chemical geometry optimizations of isolated flumethasone molecules (DFT/B3LYP/6-31G*/water) using Spartan ‘24 (Wavefunction, 2023) indicated that the two molecules converge to the same local minimum (rms difference = 0.012 Å) and are identical in energy. The global minimum-energy conformation (MMFF force field) has the opposite conformation of the side chain but is only 1.7 kcal/mol lower in energy. Intermolecular interactions thus determine the solid-state conformation.

The crystal structure (Figure 7) consists of hydrogen-bonded double layers of flumethasone molecules parallel to the ac-plane. The mean planes of the steroid molecules are approximately −5,14,1 and 5,13,1. Analysis of the contributions to the total crystal energy of the structure using the Forcite module of Materials Studio (Dassault Systèmes, 2023) indicates that the intramolecular energy is dominated by angle distortion terms (as might be expected for a fused ring system), but that bond and torsion distortion terms are also significant. The intermolecular energy is dominated by electrostatic repulsions, which in this force field-based analysis also include hydrogen bonds. The hydrogen bonds are better discussed using the results of the DFT calculation.

Figure 7. The crystal structure of flumethasone is viewed down the a-axis. Image generated using Diamond (Crystal Impact, 2023).

The hydroxyl groups O6 and O63 in the side chains form O–H···O hydrogen bonds to the carbonyl groups O7 and O64 of adjacent molecules (Table I). In addition, they form intramolecular hydrogen bonds to the carbonyl groups O5 and O62. The O63-H114 hydrogen bond forms a pattern with the graph set (Etter, Reference Etter1990; Bernstein et al., Reference Bernstein, Davis, Shimoni and Chang1995; Motherwell et al., Reference Motherwell, Shields and Allen2000) C1,1(14), while the O6-H57 hydrogen bond forms a more complicated pattern with graph set R3,4(34). The hydroxyl groups O4 and O61 act as donors in discrete hydrogen bonds to hydroxyl groups O63 and O6, both with graph set D1,1(2). The hydroxyl groups O3 and O60 form intramolecular hydrogen bonds to the carbonyl groups O7 and O64, both with graph set C1,1(12). The result of these O–H···O hydrogen bonds is a 2-dimensional network parallel to the ac-plane. The energies of the O–H···O hydrogen bonds were calculated using the correlation of Rammohan and Kaduk (Reference Rammohan and Kaduk2018). There are perhaps a surprising number of inter- and intra-molecular C–H···O hydrogen bonds. Molecule 2 acts as a donor in a larger number of these.

Table I. Hydrogen bonds (CRYSTAL23) in flumethasone.

* intramolecular

The volume enclosed by the Hirshfeld surface of flumethasone (Figure 8; Hirshfeld, Reference Hirshfeld1977; Spackman et al., Reference Spackman, Turner, McKinnon, Wolff, Grimwood, Jayatilaka and Spackman2021) is 974.73 Å³, 98.86% of 1/2 of the unit cell volume. The packing density is thus typical. The only significant close contacts (red in Figure 8) involve the hydrogen bonds. The volume/non-hydrogen atom is smaller than normal (Kempster and Lipson, Reference Kempster and Lipson1972), at 17.0 Å³.

Figure 8. The Hirshfeld surface of flumethasone. Intermolecular contacts longer than the sums of the van der Waals radii are colored blue, and contacts shorter than the sums of the radii are colored red. Contacts equal to the sums of radii are white. Image generated using CrystalExplorer (Spackman et al., Reference Spackman, Turner, McKinnon, Wolff, Grimwood, Jayatilaka and Spackman2021).

The Bravais–Friedel–Donnay–Harker (Bravais, Reference Bravais1866; Friedel, Reference Friedel1907; Donnay and Harker, Reference Donnay and Harker1937) algorithm suggests that we might expect needle morphology for flumethasone, with [100] as the long axis. A 2nd-order spherical harmonic model was included in the refinement. The texture index was 1.021(0), indicating that the preferred orientation was slight in this rotated capillary specimen.

IV. DEPOSITED DATA

The powder pattern of flumethasone from this synchrotron data set has been submitted to ICDD for inclusion in the Powder Diffraction File. The Crystallographic Information Framework (CIF) files containing the results of the Rietveld refinement (including the raw data) and the DFT geometry optimization were deposited with the ICDD. The data can be requested at [email protected].