Countercurrent chromatography isolation of 11′-γ-tocomonoenol from pumpkin seed oil with detection of novel minor tocochromanols

A non-refined, organic pumpkin seed oil (PSO) was chosen for the isolation and structure verification of the rare vitamin E compound gamma-tocomonoenol (gamma-T1). Initial measurements indicated the presence of similar to 0.4 mg gamma T1 per 100 g PSO. Saponification of similar to 2 L of PSO, followed by repeated countercurrent chromatography (CCC) with the solvent system n-hexane/benzotrifluoride/acetonitrile (v/v/v) and silica gel column chromatography enabled the isolation of 6.8 mg gamma-T1 with a purity of 96.0% according to gas chromatography with mass spectrometry (GC/MS) analysis. Structural analysis by H-1 nuclear magnetic resonance (NMR) spectroscopy and GC/MS of the gamma-T1 isolate confirmed the presence of a double bond in C-11'-position (11'-gamma-tocomonoenol). In addition, CCC fractionation enabled the detection of 18 different tocochromanols, many of which were reported for the first time in PSO. This unmatched variety included among others alpha-/gamma-tocopherol, alpha-/gamma-tocomonoenol, two alpha- and two gamma-tocodienol isomers, alpha-/gamma-tocotrienol as well as the rare 11'-beta-tocomonoenol (beta-T1) and delta-T1. Three uncommon tocochromanols were also detected whose origins and structure remained unclear.

Automated summary

The isolation and structural verification of rare vitamin E compound gamma-tocomonoenol (gamma-T1) was achieved from non-refined, organic pumpkin seed oil (PSO). In addition to gamma-T1, a total of 18 different tocochromanols were detected through CCC fractionation, some of which were reported for the first time in PSO.