HPLC DETERMINATION OF LIPOPHILIC PHOTOSYNTHETIC PIGMENTS IN ALGAL CULTURES AND LAKE WATER SAMPLES USING A NON-ENDCAPPED C-18-RP-COLUMN

A method for the determination of lipophilic photosynthetic pigments in algal cultures and lake water samples is described. The extraction of these substances is carried out in a homogenisator with acetone/water (9:1, v/v) and leads to small extraction volumes without further concentration steps during sample preparation. A binary gradient system based on acetonitrile-containing solvents and a non-endcapped C18-RP-column is used for the chromatography of the obtained extracts. Pigments are detected by monitoring the absorbance at 440 nm and identified by their on-line recorded absorption spectra. An evaluation of a variety of C18-RP-columns using one of the most similar pairs of xanthophylls, lutein and zeaxanthin, demonstrates that non-endcapped instead of fully-endcapped C18-RP-phases are able to resolve both substances. Calibration curves for relevant pigments present in lake waters using algal cultures as pigment sources are linear in the tested range from 10 ng to 300 ng for carotenoids and 50 ng to 1200 ng for chlorophylls, respectively. The described method allows the determination of lipophilic algal pigments in lake water samples down to 0.4 mug/l carotenoid or 2 mug/l chlorophyll. The reproducibility of the whole procedure is quite satisfactory and does not exceed 15% for any identified pigment.