ABSTRACT: A high-performance liquid chromatographic (HPLC) method based on a reversed-phase C₈ column and pyridine-containing mobile phases was developed for the simultaneous separation of chlorophylls and carotenoids. The method is selective enough to resolve monovinyl (MV) and divinyl (DV) pairs of polar chlorophylls and DV chlorophyll a (chl a) (the marker pigment for the prokaryote Prochlorococcus marinus) from chl a (the MV analogue). Only the pair DV chl a/chl b was not resolved. This resolution capability for chlorophylls was only previously achieved using polymeric C₁₈ columns in combination with ammonium acetate or pyridine-containing mobile phases. The proposed method also allows the separation of taxon-specific carotenoids belonging to 8 algal classes, including some critical pigment pairs for previous HPLC methods using C₁₈ columns. The method employs a binary gradient, so it can be used with both low-pressure and high-pressure mixing instruments. Method transferability was tested using 3 HPLC systems. Only a slight adjustment of gradient profile was required to obtain similar results with HPLC equipment having different dwell volumes. The selectivity of the method towards some recently discovered chlorophyll and carotenoid pigments makes it especially suitable for studying not only field samples, but also for re-examining the pigment composition of different algal classes.

KEY WORDS: HPLC pigment analysis · Phytoplankton pigments · C₈ column · Pyridine-containing mobile phases · Chemotaxonomy

Published in MEPS Vol. 195 (2000) on March 31
ISSN: 0171-8630. Copyright © Inter-Research, Oldendorf/Luhe, 2000