Chromolith semiprep rp 18e

The LC system used for isolation of 2,3-dehydrosilybin and 2,3-dehydrosilychristin consists of an LC-8A preparative HPLC pump, SPD-20A dual-wavelength UV detector, FRC-10A fraction collector, and CBM-20A bus module controlled by the LC solution 1.24 SP1 software (Shimadzu, Kyoto, Japan).
Semi-preparative chromatography of the fraction enriched in 2,3-dehydroflavonolignans (see Section 2.1 and Figure A1 in Appendix B) was conducted using monolithic Chromolith SemiPrep RP-18e (100 × 10 mm, Merck, Darmstadt, Germany) column with monolithic Chromolith RP-18e (5 × 4.6 mm) pre-column in mobile phases containing either 50% methanol, 0.1% formic acid, 5 mL/min flow rate or 45% methanol, 5% acetonitrile, 0.1% formic acid, 5 mL/min flow rate. Both mobile phases produced similar results. Alternatively, ASAHIPAK GS-10G 7B (300 × 25 mm, 20 μm, Shodex, Munich, Germany) column was used with mobile phase 50% methanol, 0.1% formic acid at 5 mL/min flow rate. The injection volume was 300 µL (37 mg/mL) in all cases.

Unless otherwise stated all chemicals were purchased from Sigma-Aldrich Chemie (Schnelldorf, Germany) or Merck (Darmstadt, Germany) at analytical grade and were used without further purification. The Ni catalyst (Shimalilte Ni reduced, 80/100 mesh) was purchased from Shimadzu (Kyoto, Japan). The precursor 6-O-desmethyl-elotinib (OSI-420; GMP grade) was purchased from Syncom B.V. (Groningen, Netherlands) and the reference compound erlotinib (N-(3-ethinylphenyl)-6,7-bis(2-methoxyethoxy)-quinazolin-4-amine) was obtained from Apollo Scientific (Bredbury, UK). Semi-preparative high performance liquid chromatography (HPLC) column (Chromolith® SemiPrep RP-18e, 100–10 mm; guard column: 10–10 mm) and analytical HPLC column (Chromolith Performance RP-18e, 100–4.6 mm; guard column: 10–4.6 mm) were purchased from Merck (Darmstadt, Germany). The analytical HPLC column for the optimized quality control (XBridge® Shield RP-18; 2.5 μm; 3.0–50 mm) and C18plus SepPak® cartridges for solid phase extraction (SPE) were purchased from Waters (Waters® Associates Milford, MA, USA). Low-protein binding Millex GS® 0.22 μm sterile filters were purchased from Millipore® (Bedford, MA, USA). Gas chromatography (GC) capillary column (forte GC Capillary Column ID-BP20; 12 m × 0.22 mm × 0.25 μm) was obtained from SGE Analytical Sciences Pty Ltd. (Victoria, Australia).