**4. Conclusions**

This study demonstrates the first application of HIMO as a useful coformer for the preparation of pharmaceutical cocrystals. As API references, thiobarbituric (TBA) and barbituric (BA) acids were employed. Two different methods for the cocrystals preparation were applied: 1) cocrystalization from MeOH used as a solvent, and 2) the grinding of solid components in a ball mill. In both cases, the desired cocrystals (TBA/HIMO and BA/HIMO) were obtained. An advantage of the ball-mill method is that in a classic, "wet" method, a substantial volume of organic solvents has to be used, and the procedure is relatively long (i.e., 8 days). Another important advantage of the ball milling method is its remarkable reduction of the reaction time required to complete the preparation of the cocrystals. Moreover, the ball milling procedure does not require toxic solvents, which is an important feature in view of economic and ecological concerns. The application of diverse analytical methods such as solid-state NMR spectroscopy, X-ray crystallography of single crystals and powders, and IR spectroscopy allowed us to perform an extended structural analysis of the obtained cocrystals. Solid state NMR techniques were used to monitor the progress of cocrystal formation. In addition, it allowed us to evaluate whether the polymorphic and tautomeric forms of the used components influenced the kinetics of the cocrystal formation and the structures of the final products. It was demonstrated that in each case, identical cocrystals formed, irrespective of the applied method (solvent versus ball milling). However, in the latter procedure, TBA hydrate requires a longer processing time.

The biocompatibility for all investigated compounds was also investigated. Experiments performed with cancer (HeLa, K562) and noncancerous (293T) human cell lines sugges<sup>t</sup> that none of tested compounds is toxic. The obtained results strongly sugges<sup>t</sup> that both TBA/HIMO and BA/HIMO cocrystals can be considered potential candidates for pharmaceutical application at a wide range of concentrations, i.e., varying from 1 μM to at least 50 μM. The solubility of the obtained cocrystals was tested in three media. Although only a small increase of solubility was observed for BA/HIMO in water, it seems obvious that the other characteristics of the studied cocrystals (protection of APIs and providing more than one API) may be useful for the development of new drug delivery systems based on 1-hydroxy imidazole 3-oxide derivatives.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/1999-4923/12/4/359/s1, Figure S1. Calculated powder x-ray diffraction diffractograms (from top to bottom) TBA crystallized from acetonitryl (tautomer A)1; TBA crystallized from ethanol (tautomer B)1; TBA crystallized from water (tautomer B)1; HIMO; TBA/HIMO. Figure S2. 13C CP/MAS spectra recorded with a spinning rate of 8 kHz for TBA/HIMO cocrystals obtained by (a) grinding; (b) cocrystallization from MeOH. Figure S3. Calculated powder x-ray diffraction diffractograms (from top to bottom) for; BA (form II)2, BA dihydrate3; HIMO and BA/HIMO. Figure S4. 13C CP/MAS spectra recorded with a spinning rate of 8 kHz for BA/HIMO cocrystals obtained by (a) grinding; (b) cocrystallization from MeOH.

**Author Contributions:** Data curation, J.S., S.K., E.W. and J.S.; Formal analysis, G.M.; Investigation, A.C.; ´ Methodology, G.D.B.; Project administration, A.W.; Writing—original draft, M.J.P. All authors have read and agreed to the published version of the manuscript.

**Funding:** This project was financially supported by the National Science Center, Poland (Grants: No. UMO-2017/25/B/ST4/02684, M.P., and UMO-2016/23/G/ST5/04115/l, G.M.).

**Conflicts of Interest:** The authors declare no conflict of interest.
