Recovery of terpenoids from Bixa orellana seeds via high performance- and high speed-countercurrent chromatography guided off-line atmospheric pressure chemical ionization mass spectrometry profiling.

Bixa orellana (Bixaceae) seeds are widely used as food colouring and spice, but also as traditional medicine for treatment of several diseases. Terpenoids from Bixa orellana seeds were extracted by n-hexane and separated by semi-preparative high-performance countercurrent chromatography (HPCCC) and then scaled-up to a high speed countercurrent chromatography apparatus (HSCCC) operated in head-to-tail mode. Evaluation of five biphasic solvent systems led to the suitable system consisting of n-hexane, ethyl acetate, methanol and water (8:2:8:2, v/v/v/v). The CCC-experiments were monitored by off-line atmospheric pressure chemical ionisation mass-spectrometry (APCI-MS/MS) in positive ionisation mode to project single ion traces of target compounds. Off-line mass profiles were converted to column-graphs which showed co-elution effects and areas of pure recoverable substances. Four major compounds bixin (1), alismol (2), geranyl-geraniol (3), δ-tocotrienol (4) were targeted and HPCCC and HSCCC experiment performance was compared by their specific partition ratio values K, the separation factor α, as well as the resolution factor R.