*3.3. Chloroform Solvates*

During the preparation of the anhydrates, new forms were detected when chloroform was used. Therefore, a solid from screening starting from anhydrous form I and form III has been performed in chloroform as solvent. Different thermodynamic and kinetic conditions have been applied, and several antisolvents have been used. Only one chloroform solvate was identified by PXRD, and it has been fully characterized (SCHCl3-H2O). Its DSC analysis (Figure 5) shows a first wide endothermic phenomenon overlapped with an endothermic phenomenon at 77 ◦C, which could be attributed to the evaporation of the solvent and the melting of the solvate taking place simultaneously. Thermal desolvation is a complex process in which the melting, release of solvent vapor and crystallization of a new phase may occur simultaneously. Next, a low intensity phenomenon is observed at 198 ◦C which could be attributed to the melting of form III, followed by the melting of form II at 203 ◦C. Its TGA analysis shows a weight loss of 19.4% from 32 to 100 ◦C which could be attributed to 1 chloroform molecule and 1 H2O molecule (theoretical weight loss of 22.9%).

**Figure 5.** DSC (blue) and TGA (red) of form SCHCl3-H2O.

Temperature variable powder X-ray diffraction of form SCHCl3-H2O has been performed in synchrotron Alba. In Figure 6, we can appreciate the diffractogram of SCHCl3-H2O which is stable until 74 ◦C and from this temperature, some new peaks corresponding to form

III begin to appear, while characteristic peaks of SCHCl3-H2O disappear. Form SCHCl3-H2O progressively transforms into form III while increasing temperature until 85 ◦C.

**Figure 6.** Temperature variable PXRD of SCHCl3-H2O showing its transformation into form III.

In addition, different experiments in chloroform (vapor diffusion, slow cool crystallizations) were performed in order to get a single crystal of SCHCl3-H2O. Curiously, we always obtained under different conditions good quality crystals of a new solid form, as the PXRD calculated from its crystal structure solved did not match with any of the previously known forms. As shown by its crystal structure determination, this form consisted of one molecule of BL, one molecule of water and three chloroform molecules, so it was identified as the heterosolvate or mixed solvate S3CHCl3-H2O. Its structure was solved at low temperature, and when leaving the crystal to reach r.t., the crystal collapsed, and it was impossible to measure it at ambient conditions. The presence of opaque crystals indicated that the product had transformed opaque due to pseudomorphosis. Therefore, the single crystals (which darkened immediately at r.t.) were analyzed by PXRD at r.t., and their diffractogram matched with the one of form SCHCl3-H2O. Therefore, the new form obtained at low temperature in SCXRD is a precursor of SCHCl3-H2O by losing 2 chloroform molecules from its crystal structure. Indeed, form S3CHCl3-H2O is the one which is always obtained in chloroform solutions, and it immediately transforms into SCHCl3-H2O when removing it from the mother liquor, preventing its detailed characterization by other methods. The instability of this new identified solvate of BL means that we cannot exclude the discovery of other labile solvate forms. Highly unstable solvates are more common among small organic molecules than is generally believed [25].

The discovery of those mixed chloroform-water solvates led us to conduct a screening in anhydrous chloroform too. A new chloroform solvate SCHCl3 was discovered; however, it was only obtained as single crystal. Again, this solvate darkened when removing it from the mother liquor at r.t., and it was only possible to solve its structure at low temperature. The analysis of the corresponding PXRD at r.t. resulted in SCHCl3-H2O, revealing that SCHCl3 transformed also into SCHCl3-H2O, but in this case by incorporating one water molecule in its crystal structure. Figure 7 shows the PXRD diffractograms of the three chloroform solvates of BL.

**Figure 7.** PXRD comparison among the different chloroform solvates.

Thus, we can conclude that both chloroform solvates S3CHCl3-H2O and SCHCl3 are only stable at low temperature or inside chloroform solutions (anhydrous in the case of SCHCl3) and that they transform immediately into SCHCl3-H2O at r.t. Both forms can be best described as transient or elusive phases as they become unstable after removal from the mother liquor. There are other examples in the literature of these labile solvates, and their existence needs to be known and considered [26]. Solvates that desolvate readily as soon as they are removed from the mother liquor can easily be overlooked in solid form screenings and wrong conclusions concerning the solvent effects on the nucleation of individual phases may result [27].
