Use of eluent modifiers for liquid chromatography particle-beam electron-capture negative-ion mass spectrometry of carotenoids

Liquid chromatography/mass spectrometry (LC/MS) with a particle-beam (PB) interface was investigated as a new method for the analysis of carotenoids. The applicability of this LC/MS technique was evaluated for the determination of carotenes (lycopene, β-carotene) and xanthophylls (astaxanthin, lutein, zeaxanthin, β-cryptoxanthin). Reversed-phase separation of carotenoids was carried out using two in-series narrow-bore columns in gradient-mode; analytes were detected under electron-capture negative-ion chemical ionization (NCI) conditions. The use of additives (ammonium acetate, triethylamine) in the LC mobile phase was investigated and the performance of the PB interface was improved in terms of better sensitivity and linearity. Under selected-ion monitoring NCI-MS conditions the absolute detection limits of carotenes were in the 0.2–3 ng range, whereas for the xanthophylls it was in the 0.02–20 ng range. To study the effect of the addition of the mobile phase modifiers on the PB-MS behaviour of carotenoids, three-way analysis of variance with interactions was applied, which provided evidence for a statistically significant effect of the additives. By applying LC/PB-NCI-MS, characterization of carotenoids in microalgae was successfully performed. NCI mass spectra confirmed the identity of some of the carotenoids investigated; in addition,­α-cryptoxanthin and echinenone were detected in the extract.