Comp. Biochem. PhysioL 1978. VoL 60B, pp. 51 to 58. Per#an)onPress. Printed in Great Britain A COMPARATIVE STUDY ON THE CAROTENOID PIGMENTATION OF THE ZOOPLANKTON OF LAKE MAARSSEVEEN (NETHERLANDS) AND OF LAC PAVIN (AUVERGNE, FRANCE)---I CHROMATOGRAPHIC CHARACTERIZATION OF CAROTENOID PIGMENTS J. E. PAANAKKER 1 and G. M. HALLEGRAEFF 2 tLaboratory for Analytical Chemistry and 2Institute of Animal Physiology, Dept'. Limnological Laboratory; University of Amsterdam, Amsterdam, The Netherlands (Received 20 June 1977) Abstraet--l. The pigments of the summer zooplankton Of the sandpit Lake Maarsseveen and of the mountain lake Lac Pavin were investigated by means of paper chromatography and high-pressure liquid chromatography (HPLC). Potential possibilities of HPLC as a routine tool in carotenoid analysis are discussed. 2. On a dry weight basis carotenoid content of the vividly-pigmented Lac Pavin zooplankton was about 4.5 times as large as that of the Lake Maarsseveen zooplankton. 3. In contrast to Lac Pavin where carotenoid content of the zooplankton was made up of 80-90% keto-carotenoids, in Lake Maarsseveen carotenoid content of the zooplankton comprised only 40-60To keto-carotenoids. 4. In Lac Pavin the zooplankton, mainly Acanthodiaptomus denticornis (Crustacea, Copepoda), was found to contain hydroxy-echinenone and astaxanthin as major pigments; associated with //-carotene, cryptoxanthin and nine unidentified keto-carotenoids and/or keto-carotenoid esters. In addition trace amounts of zeaxanthin and/or esterified lutein were detected, probably of dietary origin. 5. In Lake Maarsseveen the zooplankton, mainly Daphnia Ionc3ispina (Crustacea, Cladocera), was found to contain //-carotene; echinenone and astaxanthin as major pigments; associated with an un- identified carotene-like pigment and two unidentified keto-carotenoids and/or keto-carotenoid esters. In addition, considerable amounts of (partly esterified) fucoxanthin were detected, most likely derived from the algal diet contained in the animals' digestive tracts. 6. Qualitative and quantitative aspects of carotenoid pigmentation, together with the occurrence of typical algal xanthophylls derived from the digestive tracts of the zooplankton, suggest a more elaborate oxidative metabolism of dietary carotenoids in the Lac Pavin zooplankton. INTRODUCTION In the aquatic environment, carotenoid pigments are synthesized only by the phytoplankton algae. The presence of characteristic animal carotenoids in planktonic crustaceans, which utilize these com- pounds as their major means of pigmentation, should thus reflect the uptake and metabolism of pigments of dietary origin. The hitherto known distribution of carotenoids in Crustacea has been reviewed by Good- win (1971). It is surprising that typical algal xantho- phylls are almost completely absent, and most likely the majority of these compounds are either not absorbed or are rapidly oxidized into characteristic animal carotenoids or into colourless products (Goodwin 1960). The algal pigment//-carotene is most consistently present in the animal tissues in small amounts and astaxanthin most commonly occurs as principal carotenoid pigment. On theoreti- cal grounds it is unlikely that algal xanthophylls can be converted into astaxanthin, and this has led to the supposition of an oxidation sequence starting with //-carotene and ending in astaxanthin as the most probable biosynthetic pathway of carotenoid metabo- lism in Crustacea (Thommen & Wackernagel, 1964; 51 Green, 1965; Herring, 1968b; Katayama et al., 1973). Recently, the observation of diurnal changes in the carotenoid content of natural zooplankton (Ringel- berg & Hallegraeff 1976) has aroused interest in the ecological significance of carotenoid metabolism in planktonic crustaceans. A comparative study was exe- cuted on the carotenoid pigmentation of the summer zooplankton of the sandpit Lake Maarsseveen (Neth- erlands) and of the high-mountain lake Lac Pavin (Auvergne, France). The first paper of this series de- scribes the results of an attempt to investigate the qualitative aspect of carotenoid pigmentation of the zooplankton of the two lakes, using both paper chro- matographic and high-pressure liquid chromato- graphic techniques. The subsequent paper will further investigate concurrent diurnal variations in caro- tenoid content (Hallegraeff et al., 1978). MATERIALS AND METHODS Description of the lakes Lake Maarssevcen is a more or less oligotrophic sandpit (maximal depth 30m; surface area 70ha; 1% light depth 7.5-13 m), situated in the central part of the Netherlands. [For a detailed description see Hallegraeff(1976a).] Investi-