This contribution is a practical guide to the measurement of the different chlorophyll (Chl) fluorescence parameters and gives examples of their development under high-irradiance stress. From the Chl fluorescence induction kinetics upon irradiation of dark-adapted leaves, measured with the PAM fluorometer, various Chl fluorescence parameters, ratios, and quenching coefficients can be determined, which provide information on the functionality of the photosystem 2 (PS2) and the photosynthetic apparatus. These are the parameters Fv, Fm, F0, Fm', Fv', NF, and ΔF, the Chl fluorescence ratios Fv/Fm, Fv/F0, ΔF/Fm', as well as the photochemical (qP) and non-photochemical quenching coefficients (qN, qCN, and NPQ). qN consists of three components (qN = qE + qT + qI), the contribution of which can be determined via Chl fluorescence relaxation kinetics measured in the dark period after the induction kinetics. The above Chl fluorescence parameters and ratios, many of which are measured in the dark-adapted state of leaves, primarily provide information on the functionality of PS2. In fully developed green and dark-green leaves these Chl fluorescence parameters, measured at the upper adaxial leaf side, only reflect the Chl fluorescence of a small portion of the leaf chloroplasts of the green palisade parenchyma cells at the upper outer leaf half. Thus, PAM fluorometer measurements have to be performed at both leaf sides to obtain information on all chloroplasts of the whole leaf. Combined high irradiance (HI) and heat stress, applied at the upper leaf side, strongly reduced the quantum yield of the photochemical energy conversion at the upper leaf half to nearly zero, whereas the Chl fluorescence signals measured at the lower leaf side were not or only little affected. During this HL-stress treatment, qN, qCN, and NPQ increased in both leaf sides, but to a much higher extent at the lower compared to the upper leaf side. qN was the best indicator for non-photochemical quenching even during a stronger HL-stress, whereas qCN and NPQ decreased with progressive stress even though non-photochemical quenching still continued. It is strongly recommended to determine, in addition to the classical fluorescence parameters, via the PAM fluorometer also the Chl fluorescence decrease ratio RFd (Fd/Fs), which, when measured at saturation irradiance is directly correlated to the net CO2 assimilation rate (PN) of leaves. This RFd-ratio can be determined from the Chl fluorescence induction kinetics measured with the PAM fluorometer using continuous saturating light (cSL) during 4-5 min. As the RFd-values are fast measurable indicators correlating with the photosynthetic, activity of whole leaves, they should always be determined via the PAM fluorometer parallel to the other Chl fluorescence coefficients and ratios., and H. K. Lichtenthaler, C. Buschmann, M. Knapp.
The photosynthetic activities of three planktonic desmid species (Staurastrum brachiatum, Staurodesmus cuspidatus var. curvatus, and Staurastrum chaetoceras) were compared after adaptation to medium enriched with either a 20 mM Na+-phosphate (P) or HEPES buffer. Incubations up to 2 d were carried out at pH 6 or 8 under normal air or air enriched with 5 % CO2. Gross maximum photosynthetic rate (Pmax) and growth rate were decreased in both S. brachiatum and Std. cuspidatus at higher pH when using the HEPES buffer and this effect was independent of CO2 concentration, indicating that pH had an inhibitory effect on photosynthesis and growth in these species. The P-buffer at pH 8 caused a large decrease in Pmax and quantum yield for charge separation in photosystem 2 (PS2), compared to HEPES-buffered algae. This effect was very large in both S. brachiatum and Std. cuspidatus, two species characteristic of soft water lakes, but also significant in S. chaetoceras, a species dominant in eutrophic, hard water lakes. The decreased Pmax in P-buffer could not be related to a significant increase in cellular P content known to be responsible for inhibition in isolated chloroplasts. Experiments at pH 6 and 8 showed that two conditions, high pH and high Na+ concentration, both contributed to the decreased Pmax and quantum yield in the desmids. Effects of a P-buffer were less pronounced by using K+-P buffer. The use of P-buffer at pH 8 possibly resulted in high irradiance stress in all species, indicated by damage in the PS2 core complex. In the soft water species pH 8 resulted in increased non-photochemical quenching together with a high de-epoxidation state of the xanthophyll cycle pigments. and E. Spijkerman ... [et al.].