Linomat 5 semi automatic applicator

High performance thin layer chromatography (HPTLC) method was performed on pre-coated HPTLC aluminium sheets silica gel 60F 254 plate (10.0 X10.0 cm) of 250µm thickness (Merck, Darmstadt, Germany). The plates were pre-washed with methanol and then activated at 100°C for 15min before use. The prepared sample solutions of 2µL were spotted as 6.0mm narrow bands using 100µL syringe at a constant dosing speed of 150nL/s with a Linomat V semi-automatic applicator (Camag, Muttenz, Switzerland). The HPTLC plate was developed at a distance of 70mm in a 10.0 X10.0 cm twin trough chamber using a pre saturated mobile phase of hexane: ethyl acetate: formic acid (6:4:0.1 v/v/v). The developed TLC plate was dried using hot air for a minute and visualized under 254nm, 366nm and white light using TLC Visualizer. Further, the plate was scanned at the wavelength of 366nm using a TLC Scanner 180712 (Camag) with winCATS Planar Chromatography Manager software.

TLC analysis was performed using 5 × 10 cm or 7.5 × 10 cm silica gel TLC F 254 aluminum plates (Sigma-Aldrich). Amino acids derivatives solutions were applied using a Linomat V semi-automatic applicator (CAMAG, Muttenz, Switzerland), in a volume of 1 µL, in the form of 4-mm bands, spaced 10 mm apart. The application rate was 250 nL/s. The plates were developed with ethanol-toluene (2:3, V/V) mobile phase, to a distance of 8.0 cm in a horizontal DS chamber (Chromdes, Lublin, Poland). The developed plates were air-dried and then sprayed with spraying solution using SG1 chromatography sprayer (DESAGA, Wiesloch, Germany). Slightly wet plates were exposed to iodine vapors for 20 s in amber glass container containing iodine (iodine chamber), and after being removed outside they were left for 2 min to let the iodine-azide reaction proceed (white spots appear on violet-blue background). The visualized plates were scanned at 300 dpi with an HP ScanJet G4010 office scanner (Hewlett-Packard, Budapest, Hungary). TLSee software was used to proceed the conversion of spots to peaks.