Pregnenolone sulphate

Optical rotations were obtained on a JASCO P200 polarimeter. Melting points were determined using a Büchi^® B-540 melting point apparatus. IR spectra were measured on a Perkin-Elmer FTIR Spectrum 2. NMR spectra were recorded on a Bruker Avance III - 500 MHz NMR spectrometer equipped with a multi-nucleus probe BBFO (5 mm). UV spectra were recorded on the PDA detector of the Varian LC-920 system. LC-HRESIMS data were recorded on an Agilent 6520 Accurate-Mass Q-TOF hyphenated to an Agilent 1200 system equipped with a Zorbax Agilent C18 column (50 mm × 2.1 mm, 1.8 μm). Compounds were purified by semi-preparative HPLC with a Gilson 322 system equipped with an Axia C18 column (21.2 mm × 100 mm, 5 μm). HPLC analyses were performed on a Varian LC-920 system with a Kinetex column C18 100 Å (100 × 3.0 mm, 2.6 μm). Solid-phase extractions were performed on Chromabond^® SPE cartridges (Macherey-Nagel). The reading of the microplates was performed on an ELISA Versamax plus^® (Molecular Devices) plate reader. Solvents for extraction, fractionation and HPLC were supplied by Sigma-Aldrich. The bioassay material was purchased as following: GLP-1 ELISA kit (Active GLP-1, EGLP-35K Millipore), DMEM and PBS (Gibco), BSA (Sigma-Aldrich) and MTS mother solution CellTiter 96 (Promega), compounds standards pregnenolone and pregnenolone sulphate (Sigma-Aldrich).