Part of the difficulty in resolving the various carotenoids may
lie in the small differences in polarity between the carotenoids. A
method that increases the number of theoretical plates may be a
strategy to help resolve these analytes (9). Columns packed with
a smaller particle size such as 3 μm provide a higher number of
theoretical plates compared with columns with 5 μm particles,
and could provide a simple way to improve the resolution of the
peaks without the use of themore complex gradient elutions and
column switch devices. Few studies have used C18 stationary
phase columns packed with 3 μm particles to separate
carotenoids (9,28). Steghens, et al. (9) employed a 3 μm, C18
column with a step gradient elution for the separation of seven
carotenoids (lutein zeaxanthin, canthaxanthin β-cryptoxanthin,
lycopenes α-carotene, and β-carotene), retinol, α-tocopherol,
and two internal standards (tocol and echinenone). Because of
the lower sensitivity of UV detection and the baseline fluctuation
caused by gradient elution, this method may not be suitable for
measuring δ- and γ-tocopherol in plasma samples.