Esquire 3000 plus spectrometer

All manipulations were carried out using standard Schlenk techniques and vacuum-line manipulations under a protective atmosphere of nitrogen unless otherwise stated. Compound 1 was prepared according to the published procedure [43 (link)]. All other compounds were purchased from commercial sources unless otherwise specified. The solvents were distilled prior to use and were deaerated according to standard procedures [66 ]. The IR spectra were recorded as KBr disks using a Bruker Tensor 27 FTIR spectrophotometer. UV–vis spectra were recorded on a Jasco V-670 UV/vis/near-IR spectrophotometer. Elemental analyses were carried out on a Vario EL elemental analyzer. NMR spectra were recorded on a Bruker AVANCE DPX-200 spectrometer at 300 K. ¹H and ¹³C chemical shifts refer to solvent signals. ESIMS spectra were recorded on a Bruker Daltronics ESQUIRE3000 PLUS spectrometer. Temperature-dependent magnetic susceptibility measurements on powdered solid samples were carried out using a MPMS 7XL SQUID magnetometer (Quantum Design) over a temperature range 2–330 K at an applied magnetic field of 0.1, 0.5, and 1.0 Tesla. The observed susceptibility data were corrected for underlying diamagnetism.
Safety note! Perchlorate salts of transition metal complexes are hazardous and may explode. Only small quantities should be prepared and great care should be taken.

IR spectra were obtained in a Perkin-Elmer infrared spectrometer model 1750. NMR spectra data were recorded on a Bruker FT 300 (Bruker Biospin, Ettlingen, Germany) or on a Varian Inova 500 (Varian, Palo Alto, CA, USA) spectrometer, operating at 300/500 MHz (¹H) and 75/125 Hz (¹³C), respectively, using CDCl₃, DMSO-d₆, or CD₃OD (Deutero GmbH, Kastellaun, Germany) as solvents and TMS (Sigma-Aldrich, Deisenhofen, Germany) as an internal standard. Chemical shifts (δ) are given in ppm, with coupling constants (J) in Hz. HRMS and LRMS spectra were performed, respectively, on a TIMS-Q-TOF Maxis 3G spectrometer and on an Esquire 3000 Plus spectrometer (Bruker Daltonics, Billerica, Massachusetts, USA), operating with electrospray ionization (ESI) in positive or negative modes. Optical rotation was measured in a digital polarimeter JASCO DIP-370 (Na filter, λ = 588 nm). Column chromatography (CC) was performed with Sephadex LH-20 (Sigma-Aldrich), while thin layer chromatography (TLC) separations were carried out on silica gel 60 PF₂₅₄ (Merck, Darmstadt, Germany). Analytical grade (Labsynth Ltd., São Paulo, Brazil) solvents were used in extraction and chromatographic procedures.

Expression and purification of recombinant human CyPJ protein was carried out as described previously (Chen et al., 2015 (link)). CyPJ protein concentration was determined using the Bradford method. The substrates N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide (Suc-AAPF-pNA), CsA, 2,2,2-trifluoroethanol, and α-chymotrypsin were purchased from Sigma (St. Louis, MO, United States). The libraries of small molecular compounds used for VS were obtained from SPECS (200,000 compounds)¹ and China Natural Products Database (CNPD, 50,000 compounds)². The small organic compounds selected after VS were purchased from SPECS (Zoetermeer, Netherlands). Chemicals and solvents were either purchased from Sigma or purified by standard techniques. Analytical thin-layer chromatography (TLC) was performed on a Merck pre-coated TLC plate (silica gel 60 F₂₅₄). Melting points were recorded on an X₄-Data microscopic melting point apparatus and were uncorrected. Electrospray ionization-mass spectrometry (ESI-MS) was performed on a Bruker Esquire 3000 plus spectrometer. ¹H NMR spectra were recorded on a Bruker Avance 400 spectrometer in CDCl₃ using tetramethylsilane (TMS) as the internal standard. Elemental analysis was performed on Carlo-Erba 1106 (Carlo Erba, Italy).