Sources of Variability in Chlorophyll Analysis by Fluorometry and by High Performance Liquid Chromatography

The need for accurate determination of chlorophyll a (chl a) is of interest for numerous reasons. From the need for ground-truth data for remote sensing to pigment detection for laboratory experimentation, it is essential to know the accuracy of the analyses and the factors potentially contributing to variability and error. Numerous methods and instrument techniques are currently employed in the analyses of chl a. These methods range from spectrophotometric quantification, to fluorometric analysis and determination by high performance liquid chromatography. Even within the application of HPLC techniques, methods vary. Here we provide the results of a comparison among methods and provide some guidance for improving the accuracy of these analyses. These results are based on a round-robin conducted among numerous investigators, including several in the Sensor Intercomparison and Merger for Biological and Interdisciplinary Oceanic Studies (SIMBIOS) and HyCODE Programs. Our purpose here is not to present the full results of the laboratory intercalibration; those results will be presented elsewhere. Rather, here we highlight some of the major factors that may contribute to the variability observed. Specifically, we aim to assess the comparability of chl a analyses performed by fluorometry and HPLC, and we identify several factors in the analyses which may contribute disproportionately to this variability.