*2.2. Phase Solubility Studies*

Phase-solubility studies of HCT with different cyclodextrins were performed by adding an excess of drug to phosphate buffer solutions (pH 5.5) containing increasing concentrations of CD, i.e., 0 to 12.5 mM for βCD and 0 to 25 mM for the other CD derivatives. The vials were sealed, and the suspensions were electromagnetically stirred (500 rpm) at a constant temperature (25 ◦C) until equilibrium (3 days). Then, an aliquot of solution was withdrawn with a filter-syringe (pore size 0.45 μm), and the drug concentration was spectrometrically determined (UV/Vis 1601 Shimadzu, Tokyo, Japan) at λmax 272.2 nm. The linearity was determined on five concentration levels (from 3 to 20 mg/L). The calibration curve was *y* = 0.0623*x* + 0.003, *r*<sup>2</sup> = 0.999. LOQ = 1 mg/L and LOD = 0.33 mg/L. The presence of CD did not interfere with the drug spectrophotometric assay. Each experiment was performed in triplicate (C.V. < 2.5%). The apparent stability constants of the drug–CD complexes were calculated from the slope of the linear portion of the phase-solubility diagrams and the drug solubility (So) in the dissolution medium according to Equation (1) [18]:

$$\text{Ks} = \text{sloop} \% \text{o}^\* (1 - \text{slope}) \tag{1}$$

The complexation efficiency (CE) was calculated from the slope of the phase-solubility diagrams according to Equation (2) [19]:

$$\text{CE} = \text{slope} / (1 - \text{slope}) \tag{2}$$

The solubilizing efficiency (SE) of CDs towards the drug was calculated as the ratio of drug solubility in the presence of the maximum concentration of CD used with respect to that of the drug alone.

#### *2.3. Preparation of Drug–CD and Drug–NC Binary Systems*

Physical mixtures (PM) were prepared by mixing equimolar amounts of drug and CD with the same granulometric size (75 to 150 μm). By submitting the PM to grinding, using a high-energy vibrational micro-mill for 30 min at 24 Hz (Mixer Mill MM 200, Retsch GmbH, Dusseldorf, Germany), co-ground binary systems were obtained. Kneaded products (KN) were prepared starting from the corresponding PM. Briefly, the PM were placed in a mortar and 0.2 mL of a water/ethanol 50:50 *v*/*v* solution were added until a dough was obtained. The mixture was then manually ground with a pestle until a powder was obtained. A complete drying was achieved putting the powder in an oven at 40 ◦C for 24 h.

HCT binary systems with NCs (PHC, VHS, and SV) were prepared at different w/w ratios (1:1; 1:2; 1:4) by physical mixing (i.e., PM), as described above for HCT-CD systems. Binary systems with the selected NC (SV) at the selected *w*/*w* ratio (1:4 *w*/*w*) were prepared by different techniques. Co-evaporated products (COE) were prepared by dissolving HCT in the minimum amount of ethanol and suspending SV in water, mixing and removing the solvent by rotary evaporation; the solid product was collected and left to dry in an oven for 24 h at 40 ◦C. Cofused products (COF) were realized by heating the PM under magnetic stirring at 300 ◦C for 10 min. Co-ground systems (GR) were obtained by ball-milling the PM in a high-energy vibrational micro-mill (Mixer Mill MM 200, Retsch GmbH, Dusseldorf, Germany) at 24 Hz for 30 min [10,16]. Solvent-heated products (SH) were obtained by adding 5 mL of ethanol to the PM under magnetic stirring for 5 min and heating at 300 ◦C for 5 min until the solvent evaporation [20,21]. Solvent-sonicated products (SS) were prepared by a partial modification of the method of Yendluri et al., 2017 [22,23]. Briefly, 20 mg of drug were dissolved in 1 mL of ethanol to obtain a saturated solution where SV was added. The mixture was sonicated with a Sonopuls HD2200 ultrasound homogenizer (Bandelin Electronic GmbH, Berlin, Germany) equipped with a KE76 probe at 50% of power for 10 min and left to dry overnight to ensure maximum loading. The day after, the sample was washed with fresh ethanol and filtered in order to remove aggregates. The sample was dried in a vacuum desiccator overnight and powdered. The solvent magnetic stirring technique (SMS) is a partial variation of the method used by Lun et al., 2014 [24]. Briefly, the saturated solution of drug in ethanol containing SV was kept under stirring overnight; then, the solid in suspension was separated by centrifugation, filtered and dried overnight at 60 ◦C.
