New Emitter-Detector-Cuvette Assembly for Measuring Modulated Chlorophyll Fluorescence of Highly Diluted Suspensions in Conjunction with the Standard PAM Fluorometer

A new emitter-detector-cuvette assembly for the standard PAM fluorometer is described which leads to substantial improvement of signal/noise ratio and increased flexibility with respect to the choice of excitation and emission wavelengths. These features are particularly useful for work with very dilute suspensions of unicellular algae and isolated chloroplasts. Instead of fiber optics perspex rods are applied for guiding excitation light to a mirrored 10x10x45 mm cuvette and from there at 90° angle to the photodetector, similarly as recently reported for a PAM fluorometer based on Xe-flash measuring light (Schreiber et al. (1993), Photosynth. Res. 36, 65-72). While the detection limit of the new system does not reach that of the Xe-PAM fluorometer, it is approximately two orders of magnitude higher than that of the standard system. Rapid induction kinetics can be measured at low chlorophyll concen­ trations down to 0.1 (.ig-ml '. Satisfactory quenching analysis for detection of active chloro­ phyll concentration is still possible at 5 |ig chlorophyllT '. The various optical factors con­ tributing to the improved sensitivity are analyzed. An accessory device is described by which the frequency of the measuring light pulses generated by the PAM fluorometer is lowered in order to reduce the actinic effect of the measuring light. The performance of the new system using different excitation and emission wavelengths is demonstrated in measurements with green algae, cyanobacteria and leaves. Applying a newly available blue light-emitting diode with 450 nm peak emission, short wavelength fluorescence enriched in PS II emission can be measured, which is characterized by high values of variable fluorescence relative to maximal fluorescence. Using measuring light covering five different wavelength ranges the fluorescence contributions from cyanobacteria and green algae can be distinguished on the basis of distinct differences in their excitation spectra. This approach should become useful for an estimation of content and activity of different types of phytoplankton in natural surface waters.