Cms l ms

The reagents and solvents were purchased from commercial sources (Fisher and Sigma-Aldrich) and used directly. Analytical thin-layer chromatography was performed on ready-to-use plates with silica gel 60 (Merck, F254). Flash column chromatography was performed over silica gel (grade 60, 230-400 mesh) on Teledyne Isco CombiFlash purification system. Final compounds were purified on preparative reversed phase high-pressure liquid chromatography (RP-HPLC) and was performed on Agilent 1260 Series system with Agilent 1260 Infinity II Variable Wavelength Detector (G7114A, UV = 254 nM) and a Waters BEH C18 (130Å, 5 µm, 10 mm X 250 mm) column. Final compounds were eluted using a gradient 95% water (with 0.1% formic acid) and 5% acetonitrile (with 0.1% formic acid) at a flow rate of 4 mL/min over 30 min.
NMR spectra were acquired on a Bruker AV500 instrument (500 MHz for 1 H-NMR, 126 MHz for 13 C-NMR). TLC-MS were acquired using Advion CMS-L MS. The Agilent 1260 Infinity II Variable Wavelength Detector (G7114A, UV = 254 nM) and an Agilent ZORBAX RR SB-C18 (80Å, 3.5 µm, 4.6 x 150 mm) at a flow rate of 1 mL/min using a solvent system of 100% water with 0.1% TFA to 40 or 60% methanol over 20min were used to assess purity of final compounds.
All the purity of target compounds showed >95% in RP-HPLC. FP was monitored on a BMG CLARIOstar microplate reader.