**1. Introduction**

Methimazole (1-methyl-1,3-dihydro-1*H*-imidazole-2-thione, **1**), a well-known commercially available thyreostatic drug [1], has an ambidentate heterocyclic anion of the type [N-C-S] and was used as the terminal group in the synthesis of noncyclic crown ethers as scorpionate ligands in diverse aspects of the coordination chemistry [2,3].

Among the bridged bis(methimazole) compounds, 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]methane (**A**) and an analogous ethane derivative 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane (**2a**) were synthesized from **1** and dichloromethane or 1,2-dibromoethane. Synthesis was performed in the presence of a strong base, without or under phase-transfer conditions at an elevated temperature [4,5]. In the context of the pharmaceutical purity profile, these compounds are considered as potential methimazole related substances [6], especially if common solvents, e.g., dichloromethane (DCM) and 1,2-dichloroethane (DCE), are used in the synthetic transformations, isolation, purification and methimazole analytics.

Therefore, it was important to understand their behaviour from a chemical, structural and solid-state point of view. To evaluate these factors, we carried out a preliminary stability study of **1** in DCM and DCE in daylight or dark, at ambient humidity and room temperature. dichloromethane (DCM) and 1,2-dichloroethane (DCE), are used in the synthetic transformations, isolation, purification and methimazole analytics. Therefore, it was important to understand their behaviour from a chemical, structural and solidstate point of view. To evaluate these factors, we carried out a preliminary stability study of **1** in DCM

#### **2. Results and Discussion** and DCE in daylight or dark, at ambient humidity and room temperature.

(Scheme 1).

#### *2.1. Synthesis of Bis Derivative 2* **2. Results and Discussion**

Solutions of **1** in DCM or DCE were left without stirring in daylight or dark, at ambient humidity and at room temperature for 15 days. According to the TLC analysis, no changes of **1** were observed in DCM solutions. However, in both DCE solutions (i.e., in light and dark) spontaneous S-bis alkylation of **1** by 1,2-dichloroethane led to the formation of colourless plate shaped crystals of 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane in the form of dihydrochloride tetrahydrate (**2b**) with a 30% yield; mp. (DSC, onset): 208 ◦C. Purity by HPLC: 98%. Upon its neutralization, extraction, evaporation of the solvent and crystallization of the crude residue from the dry dichloromethane under low humidity conditions of the anhydrous 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane **2a** was obtained with an 84% yield, mp. (DSC, onset): 89 ◦C (lit: 88–90 ◦C, [5]). *2.1. Synthesis of Bis Derivative 2*  Solutions of **1** in DCM or DCE were left without stirring in daylight or dark, at ambient humidity and at room temperature for 15 days. According to the TLC analysis, no changes of **1** were observed in DCM solutions. However, in both DCE solutions (i.e., in light and dark) spontaneous S-bis alkylation of **1** by 1,2-dichloroethane led to the formation of colourless plate shaped crystals of 1,2 bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane in the form of dihydrochloride tetrahydrate (**2b**) with a 30% yield; mp. (DSC, onset): 208 °C. Purity by HPLC: 98%. Upon its neutralization, extraction, evaporation of the solvent and crystallization of the crude residue from the dry dichloromethane under low humidity conditions of the anhydrous 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane **2a** was obtained with an 84% yield, mp. (DSC, onset): 89 °C (lit: 88–90 °C, [5]).

However, if crystallization of the crude residue is attempted from the acetone/water (1/1) mixture, 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane dihydrate (**2c**) is obtained with a 78% yield; mp. (DSC, onset): 65 ◦C. The same product was obtained by the crystallization of pure **2a** from the acetone/water (1/1) mixture. Anhydrous from **2a** was also prepared by drying the dihydrate **2c** (Scheme 1). However, if crystallization of the crude residue is attempted from the acetone/water (1/1) mixture, 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane dihydrate (**2c**) is obtained with a 78% yield; mp. (DSC, onset): 65 °C. The same product was obtained by the crystallization of pure **2a** from the acetone/water (1/1) mixture. Anhydrous from **2a** was also prepared by drying the dihydrate **2c**

**Scheme 1.** Synthesis of 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]—derivatives **A** and **2** (**2a**–**2c**). **Scheme 1.** Synthesis of 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]—derivatives **A** and **2** (**2a**–**2c**).

#### *2.2. Characterization of 1,2-Bis[(1-methyl-1H-imidazole-2-yl)thio]ethane Forms 2a–2c 2.2. Characterization of 1,2-Bis[(1-methyl-1H-imidazole-2-yl)thio]ethane Forms 2a–2c*

All the studied forms, **2a**–**2c,** were elucidated using spectroscopy, microscopy, thermal and complementary analytical methods. Their structures were confirmed by the single crystal X-ray diffraction analysis. All the studied forms, **2a**–**2c**, were elucidated using spectroscopy, microscopy, thermal and complementary analytical methods. Their structures were confirmed by the single crystal X-ray diffraction analysis.

Proton chemical shifts of dihydrochloride tetrahydrate **2b**, recorded in D2O*,* were different in comparison to the previously reported data for **2a** in CHCl3-C6H6 [5], while relative integrations indicated protonation of the imidazole ring and confirmed the proposed structure (Figure 1). Mass spectra recorded in the positive mode indicated the most abundant ion at 255.0743 m/z, which was attributed to the base peak of **2a,** corresponding to [M+H-2HCl]+. Additionally, two formed fragments at 141.0486 m/z and 114.0249 m/z indicated cleavage of the thioethyl group. However, DSC showed Proton chemical shifts of dihydrochloride tetrahydrate **2b**, recorded in D2O*,* were different in comparison to the previously reported data for **2a** in CHCl3-C6H<sup>6</sup> [5], while relative integrations indicated protonation of the imidazole ring and confirmed the proposed structure (Figure 1). Mass spectra recorded in the positive mode indicated the most abundant ion at 255.0743 m/z, which was attributed to the base peak of **2a,** corresponding to [M+H-2HCl]+. Additionally, two formed fragments at 141.0486 m/z and 114.0249 m/z indicated cleavage of the thioethyl group. However, DSC showed an endothermic loss of volatile solvents between 35 and 70 ◦C. In addition, mass loss by TGA of 3.8% and chloride content of 18%, determined by ionic chromatography, indicated that

diffraction analysis.

C6D6 of **2a** [5].

the product crystallized as a dihydrochloride tetrahydrate. The hypothesis was confirmed by single crystal X-ray diffraction analysis. an endothermic loss of volatile solvents between 35 and 70 °C. In addition, mass loss by TGA of 3.8% and chloride content of 18%, determined by ionic chromatography, indicated that the product crystallized as a dihydrochloride tetrahydrate. The hypothesis was confirmed by single crystal X-ray an endothermic loss of volatile solvents between 35 and 70 °C. In addition, mass loss by TGA of 3.8% and chloride content of 18%, determined by ionic chromatography, indicated that the product crystallized as a dihydrochloride tetrahydrate. The hypothesis was confirmed by single crystal X-ray

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**Figure 1.** 1H (600 MHz) (blue), 13C (151 MHz) (red) NMR in D2O of **2b** and 1H (black) NMR in CDCl3- C6D6 of **2a** [5]. **Figure 1.** <sup>1</sup>H (600 MHz) (blue), <sup>13</sup>C (151 MHz) (red) NMR in D2O of **2b** and <sup>1</sup>H (black) NMR in CDCl<sup>3</sup> -C6D<sup>6</sup> of **2a** [5]. **Figure 1.** 1H (600 MHz) (blue), 13C (151 MHz) (red) NMR in D2O of **2b** and 1H (black) NMR in CDCl3-

The molecular structure of **2b** consisted of two cationic imidazolyl groups bounded to the *S*atoms of the dithioethyl group (Figure 2a). Each chloride anion was a hydrogen bond acceptor from the protonated imidazole nitrogen atom N2 and two water molecules (Figure 2b, Table 1). The two water molecules were also connected by hydrogen bonds. Through these hydrogen bonds, the cations, chloride anions and water molecules were interconnected into a 3D network. The molecular structure of **2b** consisted of two cationic imidazolyl groups bounded to the *S*-atoms of the dithioethyl group (Figure 2a). Each chloride anion was a hydrogen bond acceptor from the protonated imidazole nitrogen atom N2 and two water molecules (Figure 2b, Table 1). The two water molecules were also connected by hydrogen bonds. Through these hydrogen bonds, the cations, chloride anions and water molecules were interconnected into a 3D network. The molecular structure of **2b** consisted of two cationic imidazolyl groups bounded to the *S*atoms of the dithioethyl group (Figure 2a). Each chloride anion was a hydrogen bond acceptor from the protonated imidazole nitrogen atom N2 and two water molecules (Figure 2b, Table 1). The two water molecules were also connected by hydrogen bonds. Through these hydrogen bonds, the cations, chloride anions and water molecules were interconnected into a 3D network.

**Figure 2.** (**a**) Molecular structure of **2b** with the atomic numbering scheme; symmetry code (i) 1-x, -y, -z, and (**b**) packing diagram of **2b** viewed along the *b-*axis. Hydrogen bounds are marked by dashed blue lines. **Figure 2.** (**a**) Molecular structure of **2b** with the atomic numbering scheme; symmetry code (i) 1-x, -y, -z, and (**b**) packing diagram of **2b** viewed along the *b-*axis. Hydrogen bounds are marked by dashed blue lines. **Figure 2.** (**a**) Molecular structure of **2b** with the atomic numbering scheme; symmetry code (i) 1 − x, −y, −z, and (**b**) packing diagram of **2b** viewed along the *b-*axis. Hydrogen bounds are marked by dashed blue lines.


**Table 1.** Distances and angles of the hydrogen bonds in **2b** and **2c. Table 1.** Distances and angles of the hydrogen bonds in **2b** and **2c. Table 1.** Distances and angles of the hydrogen bonds in **2b** and **2c**.

Transformation of the asymmetric unit: (**a**) 1 − x, −1/2 + y, 1/2 − z; (**b**) −x, 1/2 + y, 1/2 −z; (**c**) x, 3/2 − y, 1/2 + z; (**d**) −x + y, −x, z; (**e**) y, −x + y, −z; (**f**) 1/3 − x + y, 2/3 −x, −1/3 + z. 1/2 + z; (**d**) −x + y, −x, z; (**e**) y, −x + y, −z; (**f**) 1/3 − x + y, 2/3 −x, −1/3 + z. Transformation of the asymmetric unit: (**a**) 1 − x, −1/2 + y, 1/2 − z; (**b**) −x, 1/2 + y, 1/2 −z; (**c**) x, 3/2 − y, 1/2 + z; (**d**) −x + y, −x, z; (**e**) y, −x + y, −z; (**f**) 1/3 − x + y, 2/3 −x, −1/3 + z.

Proton and carbon chemical shifts in the NMR spectra of **2a** in DMSO-d6 exhibited similar values to those in **2b***,* and experimental MS data matched the theoretical data. After several attempts, we obtained good quality, single crystals by crystallisation from dry dichloromethane at dry conditions (see experimental section) that allowed us to confirm the anhydrous structure of **2a**, which crystallized in the monoclinic P21/c space group.

crystallized in the monoclinic P21/c space group.

There are no hydrogen bounds in the structure of **2a**, only van der Waals forces connect the molecules (Figure 3). crystallized in the monoclinic P21/c space group. There are no hydrogen bounds in the structure of **2a**, only van der Waals forces connect the molecules (Figure 3). to those in **2b***,* and experimental MS data matched the theoretical data. After several attempts, we obtained good quality, single crystals by crystallisation from dry dichloromethane at dry conditions (see experimental section) that allowed us to confirm the anhydrous structure of **2a**, which

(see experimental section) that allowed us to confirm the anhydrous structure of **2a**, which

Proton and carbon chemical shifts in the NMR spectra of **2a** in DMSO-d6 exhibited similar values

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Proton and carbon chemical shifts in the NMR spectra of **2a** in DMSO-d6 exhibited similar values

**Figure 3.** (**a**) Molecular structure of **2a** with the atomic numbering scheme; symmetry code (i) 1-x, -y, -z, and (**b**) packing diagram of **2a**. **Figure 3.** (**a**) Molecular structure of **2a** with the atomic numbering scheme; symmetry code (i) 1 − x, −y, −z, and (**b**) packing diagram of **2a**. (**a**) (**b**) **Figure 3.** (**a**) Molecular structure of **2a** with the atomic numbering scheme; symmetry code (i) 1-x, -y,

Crystals of **2c** showed the TLC Rf value of 0.59, which was the same as **2a** and **2b**, and a TGA volatile content of 11.31%, indicating the solvated form of the same substance. The structure was solved by the single crystal X-ray diffraction analysis, showing that the dihydrate **2c** crystallizes in the trigonal *R* -3 space group. Six water molecules were mutually interconnected by hydrogen bonds, forming a hexagon in the chair conformation, which is the most common conformation for a cluster of six water molecules. The R6 motif is usually formed by water molecules related by a centre of symmetry [7]. In **2c**, one water molecule formed the R6 pattern by the -3 symmetry element (R66(12) by the graph-set notation). The water molecule was an acceptor of a hydrogen bond from N2 and a weak one from C4, thus forming a 3D network (Figure 4, Table 1). Crystals of **2c** showed the TLC Rf value of 0.59, which was the same as **2a** and **2b**, and a TGA volatile content of 11.31%, indicating the solvated form of the same substance. The structure was solved by the single crystal X-ray diffraction analysis, showing that the dihydrate **2c** crystallizes in the trigonal *R* -3 space group. Six water molecules were mutually interconnected by hydrogen bonds, forming a hexagon in the chair conformation, which is the most common conformation for a cluster of six water molecules. The R6 motif is usually formed by water molecules related by a centre of symmetry [7]. In **2c**, one water molecule formed the R6 pattern by the -3 symmetry element (R<sup>6</sup> 6 (12) by the graph-set notation). The water molecule was an acceptor of a hydrogen bond from N2 and a weak one from C4, thus forming a 3D network (Figure 4, Table 1). -z, and (**b**) packing diagram of **2a**. Crystals of **2c** showed the TLC Rf value of 0.59, which was the same as **2a** and **2b**, and a TGA volatile content of 11.31%, indicating the solvated form of the same substance. The structure was solved by the single crystal X-ray diffraction analysis, showing that the dihydrate **2c** crystallizes in the trigonal *R* -3 space group. Six water molecules were mutually interconnected by hydrogen bonds, forming a hexagon in the chair conformation, which is the most common conformation for a cluster of six water molecules. The R6 motif is usually formed by water molecules related by a centre of symmetry [7]. In **2c**, one water molecule formed the R6 pattern by the -3 symmetry element (R66(12) by the graph-set notation). The water molecule was an acceptor of a hydrogen bond from N2 and a

weak one from C4, thus forming a 3D network (Figure 4, Table 1).

blue dashed lines. **Figure 4.** (**a**) Molecular structure with the atomic numbering scheme; symmetry code (i) 2/3 − x, 1/3 − y, 4/3 − z, and (**b**) packing diagram of **2c** viewed along the *c-*axis. Hydrogen bonds are marked with blue dashed lines. **Figure 4.** (**a**) Molecular structure with the atomic numbering scheme; symmetry code (i) 2/3 − x, 1/3 − y, 4/3 − z, and (**b**) packing diagram of **2c** viewed along the *c-*axis. Hydrogen bonds are marked with blue dashed lines.

The molecular structure of 1,2-bis[(1-methyl-1*H*-imidazole-2-yl)thio]ethane consisted of two imidazole groups bounded to the S-atoms of the dithioethyl group (**2a**, **2c**). The imidazole groups were protonated in **2b**. In all three structures, the ethane moiety lied at the centre of symmetry, and thus only half of the molecule was in the asymmetric unit. Therefore, the imidazole groups were parallel. The difference in the three structures was found in the orientation of the imidazole group, i.e., rotation

about the S-C1 bond resulting in different C5-S-C1-N1 torsion angles (−173.8(2)◦ , −113.4(2)◦ and 151.8(2)◦ , in **2a**, **2b** and **2c,** respectively (Figure 5). thus only half of the molecule was in the asymmetric unit. Therefore, the imidazole groups were parallel. The difference in the three structures was found in the orientation of the imidazole group, i.e., rotation about the S-C1 bond resulting in different C5-S-C1-N1 torsion angles (−173.8(2)°, −113.4(2)° and 151.8(2)°, in **2a**, **2b** and **2c,** respectively (Figure 5).

were protonated in **2b**. In all three structures, the ethane moiety lied at the centre of symmetry, and

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**Figure 5.** Molecule overlay of **2a** (blue), **2b** (magenta) and **2c** (red). Atoms C5 and S and their pairs related by the inversion centre were used for the overlay. Hydrogen atoms are omitted for clarity. **Figure 5.** Molecule overlay of **2a** (blue), **2b** (magenta) and **2c** (red). Atoms C5 and S and their pairs related by the inversion centre were used for the overlay. Hydrogen atoms are omitted for clarity.

#### *2.3. Dehydration Behaviour of Dihydrate 2c and its Transformation to the Anhydrous Form 2a 2.3. Dehydration Behaviour of Dihydrate 2c and Its Transformation to the Anhydrous Form 2a*

Hydrates are the most common type of solvated organic compounds [8], and understanding their dehydration pathways, as a widespread but not properly understood phenomenon, is critical for designing optimal properties for materials, particularly in the case of pharmaceutical solids [9]. Several schemes for the classification of hydrates have been proposed [10–13], but in general, dehydration results in three types of crystallographic behaviour: a) material where the crystal structure changes (different powder pattern) after dehydration*,* i.e., as in the present study, contrary to b) material that undergoes only a slight change in crystal structure (related XRPD pattern) after dehydration, like some azithromycin solvates [14], or c) material that becomes amorphous after dehydration, like some other azithromycin solvates [15]. The dehydration process of **2c** had a markedly different crystal structure of the anhydrous form **2a** and has been studied using different experimental techniques. The dihydrate **2c** was heated below Hydrates are the most common type of solvated organic compounds [8], and understanding their dehydration pathways, as a widespread but not properly understood phenomenon, is critical for designing optimal properties for materials, particularly in the case of pharmaceutical solids [9]. Several schemes for the classification of hydrates have been proposed [10–13], but in general, dehydration results in three types of crystallographic behaviour: a) material where the crystal structure changes (different powder pattern) after dehydration, i.e., as in the present study, contrary to b) material that undergoes only a slight change in crystal structure (related XRPD pattern) after dehydration, like some azithromycin solvates [14], or c) material that becomes amorphous after dehydration, like some other azithromycin solvates [15].

the melting point, at around 55 °C and at reduced pressure of 200 mbar to a constant mass. A significant rate of the dehydration process was detected after 30 min by the formation of opaque crystals. The process was completed after 60 min. DSC measurement of the selected samples showed, after 30 min, endothermic events belonging to the melting of **2c** and **2a**, indicating the coexistence of both forms in the sample (Figure 6, iii), and finally, after 60 min, only a single endothermic event at 90 °C, belonging to the melting of the anhydrous form **2a** (Figure 6, iv). Unfortunately, no sign of recrystallization was detected. The dehydration process of **2c** had a markedly different crystal structure of the anhydrous form **2a** and has been studied using different experimental techniques. The dihydrate **2c** was heated below the melting point, at around 55 ◦C and at reduced pressure of 200 mbar to a constant mass. A significant rate of the dehydration process was detected after 30 min by the formation of opaque crystals. The process was completed after 60 min.

DSC measurement of the selected samples showed, after 30 min, endothermic events belonging to the melting of **2c** and **2a**, indicating the coexistence of both forms in the sample (Figure 6, iii), and finally, after 60 min, only a single endothermic event at 90 ◦C, belonging to the melting of the anhydrous form **2a** (Figure 6, iv). Unfortunately, no sign of recrystallization was detected.

**Figure 6.** Selected DSC thermograms showing solid state transformation of the dihydrate **2c** to the anhydrous form **2a** recorded under a heating rate of 10 ◦C/min and N<sup>2</sup> purge in a pierced lid crucible; (i) anhydrous form **2a**; (ii) dihydrate **2c**; (iii) product after heating **2c** at 55 ◦C/200 mbar for 30 min, and (iv) product after heating **2c** at 55 ◦C/200 mbar for 60 min.

1

Finally, the XRPD powder pattern of the dried sample was different from the powder pattern of dihydrate **2c** and identical to the calculated powder pattern of anhydrous **2a** prepared by crystallization (Figure 7), confirming that **2a** is also the final product of dihydrate **2c** dehydration. and (iv) product after heating **2c** at 55 °C/200 mbar for 60 min. Finally, the XRPD powder pattern of the dried sample was different from the powder pattern of dihydrate **2c** and identical to the calculated powder pattern of anhydrous **2a** prepared by Finally, the XRPD powder pattern of the dried sample was different from the powder pattern of dihydrate **2c** and identical to the calculated powder pattern of anhydrous **2a** prepared by crystallization (Figure 7), confirming that **2a** is also the final product of dihydrate **2c** dehydration.

crystallization (Figure 7), confirming that **2a** is also the final product of dihydrate **2c** dehydration.

(i) anhydrous form **2a**; (ii) dihydrate **2c**; (iii) product after heating **2c** at 55 °C/200 mbar for 30 min,

and (iv) product after heating **2c** at 55 °C/200 mbar for 60 min.

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**Figure 7.** Photomicrographs of: (**a**) anhydrous **2a** after drying of **2c** during 60 min at 55 °C /200mbar; (**b**) starting dihydrate **2c** and the related PXRD patterns. **Figure 7.** Photomicrographs of: (**a**) anhydrous **2a** after drying of **2c** during 60 min at 55 ◦C /200mbar; (**b**) starting dihydrate **2c** and the related PXRD patterns. **Figure 7.** Photomicrographs of: (**a**) anhydrous **2a** after drying of **2c** during 60 min at 55 °C /200mbar;

Additional studies were performed under atmospheric pressure to determine whether dehydration of **2c** to **2a** proceeds via a metastable monohydrate intermediate, as in lisinopril dihydrate [16], or as a hemihydrate, such as ondansetron hydrochloride dihydrate [17]. Additional studies were performed under atmospheric pressure to determine whether dehydration of **2c** to **2a** proceeds via a metastable monohydrate intermediate, as in lisinopril dihydrate [16], or as a hemihydrate, such as ondansetron hydrochloride dihydrate [17]. (**b**) starting dihydrate **2c** and the related PXRD patterns. Additional studies were performed under atmospheric pressure to determine whether dehydration of **2c** to **2a** proceeds via a metastable monohydrate intermediate, as in lisinopril

The TGA thermogram of **2c** showed a single thermal event in the range 40–110 °C with a mass loss of 11.31%, corresponding to the loss of two water molecules. This was in agreement with the DSC thermogram showing only a sharp endothermic event with an onset at 65 °C, corresponding to the melting of **2c**. Additional exothermic processes of recrystallization and endothermic of melting were not detected (Figure 8). The TGA thermogram of **2c** showed a single thermal event in the range 40–110 ◦C with a mass loss of 11.31%, corresponding to the loss of two water molecules. This was in agreement with the DSC thermogram showing only a sharp endothermic event with an onset at 65 ◦C, corresponding to the melting of **2c**. Additional exothermic processes of recrystallization and endothermic of melting were not detected (Figure 8). dihydrate [16], or as a hemihydrate, such as ondansetron hydrochloride dihydrate [17]. The TGA thermogram of **2c** showed a single thermal event in the range 40–110 °C with a mass loss of 11.31%, corresponding to the loss of two water molecules. This was in agreement with the DSC thermogram showing only a sharp endothermic event with an onset at 65 °C, corresponding to the melting of **2c**. Additional exothermic processes of recrystallization and endothermic of melting were not detected (Figure 8).

**Figure 8.** TGA (i) and DSC (ii) analysis of dihydrate **2c** in perforated crucibles under a heating rate of 10 °C/min and N2 purge. **Figure 8.** TGA (i) and DSC (ii) analysis of dihydrate **2c** in perforated crucibles under a heating rate of <sup>10</sup> ◦C/min and N<sup>2</sup> purge.

10 °C/min and N2 purge.

The changes upon dehydration of **2c** were monitored by the combination of hot stage microscopy (HSM) with DSC at a heating/cooling rate of 10 ◦C/min (Figure 9). Melting started at 40 ◦C until completed at around 65 ◦C (step i). Immediately after the melting point, the melt of **2c** was cooled to room temperature, resulting in a glassy product, and no further crystallisation was observed

**2c** and the anhydrous form **2a**.

after prolonged standing. However, if **2c** was heated to 80 ◦C and immersed in oil during the HSM experiment, followed by cooling, recrystallization of the dihydrate **2c** was observed due to the conditions that inhibited efficient water removal. However, if the sample was heated over melting, up to 95 ◦C, in an open pan followed by cooling to room temperature, spontaneous crystallisation occurred, and according to XRPD, the obtained product was a mixture of the dihydrate **2c** and the anhydrous form **2a**. formation of potential intermediary monohydrate or hemihydrate phases, were observed. Dehydration of **2c** in vacuo favours rapid water removal below the melting point, without the appearance of a liquid or melt phase. Removal of water molecules from **2c** under these conditions resulted in the solid-solid rearrangement of the molecules in the crystal lattice, leading to anhydrous **2a**. On the contrary, crystallization of **2a** from the melt of dihydrate **2c** under atmospheric pressure only occurred when all the residual water was removed from the sample (at 120 °C). When a sample of melted **2c** was only heated to 95 °C, concomitant crystallization of **2a** and **2c** was observed, and

this was identical to the partial dehydration of **2c** after 30 min under reduced pressure.

no additional exothermic and endothermic events or changes in XRPD, which could indicate the

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The changes upon dehydration of **2c** were monitored by the combination of hot stage microscopy (HSM) with DSC at a heating/cooling rate of 10 °C/min (Figure 9). Melting started at 40 °C until completed at around 65 °C (step *i*). Immediately after the melting point, the melt of **2c** was cooled to room temperature, resulting in a glassy product, and no further crystallisation was observed after prolonged standing. However, if **2c** was heated to 80 °C and immersed in oil during the HSM experiment, followed by cooling, recrystallization of the dihydrate **2c** was observed due to the conditions that inhibited efficient water removal. However, if the sample was heated over melting, up to 95 °C, in an open pan followed by cooling to room temperature, spontaneous crystallisation occurred, and according to XRPD, the obtained product was a mixture of the dihydrate

In contrast, when the melt of **2c** was heated to 120 °C and cooled to room temperature, according to the XRPD slow nucleation and growth process of the anhydrous form **2a** occurred after prolonged

**Figure 9.** Selected DSC thermograms and HSM micrographs of dihydrate **2c,** recorded using a heating/cooling rate of 10 °C/min with perforated DSC crucibles: (i) dihydrate **2c** heating step up to 120 °C; (ii) cooling step to room temperature, and (iii) heating step of the recrystallized anhydrous form **2a**, i.e., re-run (re-heating) of the sample in the bottom thermogram. **Figure 9.** Selected DSC thermograms and HSM micrographs of dihydrate **2c,** recorded using a heating/cooling rate of 10 ◦C/min with perforated DSC crucibles: (i) dihydrate **2c** heating step up to 120 ◦C; (ii) cooling step to room temperature, and (iii) heating step of the recrystallized anhydrous form **2a**, i.e., re-run (re-heating) of the sample in the bottom thermogram.

Finally, we can conclude that the dehydration at both conditions, i.e., under reduced and atmospheric pressure, proceeds to the anhydrous form **2a** and it is in accordance with the well-known fact that dehydration processes are greatly dependent on the atmospheric environment [11,18]. **3. Materials and Methods**  *3.1. General*  In contrast, when the melt of **2c** was heated to 120 ◦C and cooled to room temperature, according to the XRPD slow nucleation and growth process of the anhydrous form **2a** occurred after prolonged standing indicating complete dehydration (step ii). In the re-run the sample melting started at 89 ◦C until completed at around 90 ◦C (step iii), i.e., at the melting point of the anhydrous form **2a**. However, no additional exothermic and endothermic events or changes in XRPD, which could indicate the formation of potential intermediary monohydrate or hemihydrate phases, were observed.

Dehydration of **2c** in vacuo favours rapid water removal below the melting point, without the appearance of a liquid or melt phase. Removal of water molecules from **2c** under these conditions resulted in the solid-solid rearrangement of the molecules in the crystal lattice, leading to anhydrous **2a**. On the contrary, crystallization of **2a** from the melt of dihydrate **2c** under atmospheric pressure only occurred when all the residual water was removed from the sample (at 120 ◦C). When a sample of melted **2c** was only heated to 95 ◦C, concomitant crystallization of **2a** and **2c** was observed, and this was identical to the partial dehydration of **2c** after 30 min under reduced pressure.

Finally, we can conclude that the dehydration at both conditions, i.e., under reduced and atmospheric pressure, proceeds to the anhydrous form **2a** and it is in accordance with the well-known fact that dehydration processes are greatly dependent on the atmospheric environment [11,18].
