717 plus auto sampler

Kinetic water solubility was experimentally determined utilising analytical HPLC [Waters HPLC system (1525 Binary pump, 2487 dual wavelength absorbance detector, and 717 plus Autosampler) with a Phenomenex Luna^® 5μ C18(2) 100 Å (150 × 4.6 mm) column]. The following gradient was operated: 20–100% CH₃CN/water over 15 min, maintained at 100% CH₃CN for 10 min, and finally returned to 20% CH₃CN in water over 5 min. Both CH₃CN and Milli-Q water solvents used were containing 0.05% TFA and the flow rate was set at 1.0 mL/min. First, 4000 mg/L stock solutions in DMSO were created for compounds 8b, 8c, 8k, and 8l, followed by serial dilutions using CH₃CN/H₂O (9:1) to create a calibration curve for each compound by plotting absorbance versus concentrations of 200 mg/L, 400 mg/L, 600 mg/L, 800 mg/L and 1000 mg/L. The absorbance values of each concentration are an average of two individual measurements. Next, from the 4000 mg/L stock solution in DMSO, three samples of each compound were diluted 1:20 (200 mg/L) in deionised water and centrifuged. A 100 μL sample from the supernatant was diluted 1:1 with 100 μL neat CH₃CN, mixed, filtered using a 0.45 μM filter, and 100 μL sample was taken for analytical HPLC testing. Two absorbance values of each sample were measured and averaged, followed by appropriate calculation and extrapolation to the established calibration curve.