High performance liquid chromatography system

Bisoprolol base (98.5%) was acquired from Mikromol, while other reagents such as acetic anhydride (98.5%), formic acid (98–100%) and ZnCl₂ (98–100.5%) were obtained from Merck. All common solvents used in the experiments were of chemically pure and analytical research (AR) grade, sourced from Merck and Fluka and used without further purification. The ¹H-NMR signals were measured using an Agilent NMR instrument with a frequency of 500 MHz, while the ¹³C-NMR signals were measured using an Agilent NMR instrument with a frequency of 125 MHz. Fourier transform infrared (FTIR) spectroscopy was performed using a Prestige 21 Shimadzu instrument to obtain absorption spectra of the functional groups present in the products. Pure compounds were obtained using a Shimadzu high-performance liquid chromatography (HPLC) system equipped with a photodiode array (PDA) detector.

Pharmacokinetics of CPT was performed as described [71 (link)] with minor modifications. One cohort of male C57BL/6J mice (weighing 33 to 35 g) orally received CPT (1 mg kg⁻¹). Plasma samples were collected at indicated points (0, 1, 4, 6, and 12 h) after oral administration and stored at −20°C for further processing. CPT concentrations in the mice plasma were quantified by a Shimadzu high-performance liquid chromatography (HPLC) system.

A Shimadzu high-performance liquid chromatography (HPLC) system (Shimadzu, Tokyo, Japan), equipped with an LC-20A UV detector (Shimadzu), was used with a COCOSMOSIL 5C18-PAQ column (5 μm, 4.6 mm × 250 mm) for the separation of compounds. The temperature of the column compartment was maintained at 30°C throughout the analysis. The wavelength of detection was 254 nm, and the injection volume was 20 μL. All chromatographic assays were performed with a flow rate of 1 mL/min, and 25 mM potassium dihydrogen phosphate buffer was used as the mobile phase. The method was validated by checking the accuracy, precision, linearity, limit of detection, limit of quantitation, and specificity prior to sample analysis.

A Shimadzu high-performance liquid chromatography (HPLC) system, combined with a Diode-Array Detection (DAD) detector and an RF-20A XS fluorescence detector (Shimadzu, Kyoto, Japan) were used.

A Shimadzu^® High-Performance Liquid Chromatography (HPLC) system (Kyoto, Japan) equipped with a photodiode array (PDA) detector was used for separation and absorbance analysis, respectively. Data acquisition and system control were performed by analytical software (LC Solution, Release 1.22 SP1). The mobile phase comprised acetonitrile:water:triethylamine (90:10:0.3 v/v/v). The aqueous phase was adjusted to pH 7.0 with phosphoric acid. The TQ was separated using a chromatographic column at 25 ± 1 °C (5 μm, 4.6 × 150 mm; Nano Separation Technologies RP-18). The flow rate was defined as 1.0 mL/min, and TQ was detected at 325 nm after injecting 20 μL.

A Shimadzu High Performance Liquid Chromatography (HPLC) system coupled to UV‐VIS detection (Shimadzu Corp., Kyoto, Japan) was employed to analyze melatonin receptor ligands in metabolic stability and physicochemical characterization assays. It consisted of a LC‐10ADvp solvent delivery module, a 20 μL Rheodyne sample injector (Rheodyne LLC, Rohnert Park, CA, USA) and a SPD‐10Avp UV‐VIS detector. PeakSimple 2.83 software was employed for data acquisition and HPLC peak integration. HPLC columns were a RP‐C18 Supelco Discovery (Supelco, Bellefonte, PA, USA), 5 μm, 150×4.6 mm i.d. for all compounds except 13, and a Phenomenex Gemini, 5 μm, 150×4.6 mm i.d. for compound 13 (Phenomenex). Elution conditions were optimized for each compound at a flow rate of 1 mL min⁻1 employing mobile phases consisting of water and acetonitrile at different percentages, while UV detection was set at λ = 254 nm. For compound 13, the mobile phase consisted of a 10 mM ammonium acetate buffer pH 7.0 and methanol and the UV detection was set at λ = 281 nm.