*3.2. PXRD Analysis 3.2. PXRD Analysis*

the new crystalline phases.

PXRD enables the study and characterization of novel crystalline materials [30].PXRD patterns of the four crystal samples are shown in Figure 2. The patterns for **1** were different from those of the starting material and from the previously published PXRD patterns for the polymorphic form [31]. The purity of all products was confirmed by comparing the PXRD patterns with the simulated patterns obtained from the single crystal data (see the Supplementary Materials, Figures S1−S4). No apparent peaks corresponding to impurities were observed, so the obtained powder compounds were considered to be of high chemical purity. Further, these PXRD patterns of enrofloxacin compounds **2**–**4** differ significantly from the patterns of individual APIs, proving the formation of PXRD enables the study and characterization of novel crystalline materials [30]. PXRD patterns of the four crystal samples are shown in Figure 2. The patterns for **1** were different from those of the starting material and from the previously published PXRD patterns for the polymorphic form [31]. The purity of all products was confirmed by comparing the PXRD patterns with the simulated patterns obtained from the single crystal data (see the Supplementary Materials, Figures S1−S4). No apparent peaks corresponding to impurities were observed, so the obtained powder compounds were considered to be of high chemical purity. Further, these PXRD patterns of enrofloxacin compounds **2**–**4** differ significantly from the patterns of individual APIs, proving the formation of the new crystalline phases.

**Figure 2.** Powder X-ray diffraction (PXRD) patterns of compounds **1**–**4**. **Figure 2.** Powder X-ray diffraction (PXRD) patterns of compounds **1**–**4**.

#### *3.3. Spectroscopic Characterization 3.3. Spectroscopic Characterization*

The FTIR analysis results for the products are given in Figure 3. The absorption band of crystalline enrofloxacin was located at 1737 cm−1 because of the carbonyl stretching of its unionized carboxylic acid C=O group [32]. The C=O band in **1** was at 1736.4 cm−1, which indicates that the enrofloxacin exists as a neutral molecule. Slight peak shifts were also seen in compounds **2**–**4**, in particular, for the carboxylic acid C=O stretching, which shifted to 1728.8 cm−1, 1728.5 cm−1 and 1729.3 cm−1, respectively. This can be attributed to intermolecular interactions such as the formation of hydrogen bonds [33]. The terminal amino group of the piperazine ring was protonated in the process of crystallization, and this is proved by the presence of broad bands from 2600 cm−1 to 3000 cm−1. However, confirming this is difficult because the broad IR absorption bands possibly overlap with others, e.g., in the case of the C−H stretching [34]. However, Karanam et al. reported FTIR analysis results that showed that enrofloxacin salts were protonated in the process of crystallization [31]. The medium-intensity broad band in the region of 3300–3500 cm−1 is attributed to the O−H stretching of the water molecule **2** and ethanol molecule **3–4**. The existence of enrofloxacin as a neutral molecular and in the ionic state in the crystal structures was confirmed by SCXRD analysis. The FTIR analysis results for the products are given in Figure 3. The absorption band of crystalline enrofloxacin was located at 1737 cm−<sup>1</sup> because of the carbonyl stretching of its unionized carboxylic acid C=O group [31]. The C=O band in **1** was at 1736.4 cm−<sup>1</sup> , which indicates that the enrofloxacin exists as a neutral molecule. Slight peak shifts were also seen in compounds **2**–**4**, in particular, for the carboxylic acid C=O stretching, which shifted to 1728.8 cm−<sup>1</sup> , 1728.5 cm−<sup>1</sup> and 1729.3 cm−<sup>1</sup> , respectively. This can be attributed to intermolecular interactions such as the formation of hydrogen bonds [32]. The terminal amino group of the piperazine ring was protonated in the process of crystallization, and this is proved by the presence of broad bands from 2600 cm−<sup>1</sup> to 3000 cm−<sup>1</sup> . However, confirming this is difficult because the broad IR absorption bands possibly overlap with others, e.g., in the case of the C−H stretching [33]. However, Karanam et al. reported FTIR analysis results that showed that enrofloxacin salts were protonated in the process of crystallization [34]. The medium-intensity broad band in the region of 3300–3500 cm−<sup>1</sup> is attributed to the O−H stretching of the water molecule **2** and ethanol molecule **3–4**. The existence of enrofloxacin as a neutral molecular and in the ionic state in the crystal structures was confirmed by SCXRD analysis.

**Figure 3.** Fourier transform infrared (FTIR) spectrum of compounds **1**–**4**. **Figure 3.** Fourier transform infrared (FTIR) spectrum of compounds **1**–**4**.
