Assessment of a microscopic photobleaching technique for measuring the spectral absorption efficiency of individual phytoplankton cells

Abstract

Measurements of the absorptance of photosynthetic pigments in individual phytoplankton cells were made using an epifluorescence microscope equipped with a spectrograph and CCD array detector. Correction for light loss due to scattering was achieved by bleaching the cells with intense light from a mercury arc lamp, and using the bleached cells as a spectrophotometric blank. Absorption efficiency factors were calculated from knowledge of the geometrical cross-section of the cells obtained fromcalibrated video images acquired at the time of measurement. The single-cell efficiency factors were consistent with the average absorption efficiencies of cell suspensions measured using a spectrophotometer over most of the visible spectrum, but they were significantly lower below 420 nm. Cells of the diatom Cyclotella cryptica and the chlorophyte Chlorella salina showed clear spectral differences in spectral shape that could be related to taxonomic differences in pigment content, but absorption efficiency factors of approximately 0.4 at 675 nm were found for both species.