Dark adapted parameters measured

Dark adaptation inhibits all light-dependent reactions. The resulting absence of photochemistry for a sufficient length of time allows complete re-oxidation of PSII electron acceptor molecules, opening PSII reaction centers and thus maximising the probability that absorbed light can be used for photochemistry. Commonly measured parameters from tissue in this state are used to calculate the maximum quantum efficiency of PSII and are usually used to reference measurements made on light adapted samples.

Fo – The fluorescence origin (Fo) is defined as the chlorophyll fluorescence yield following dark adaptation when all of the PSII reaction centers and electron acceptor molecules are fully oxidised and hence available for photochemistry. As a result, Fo is often measured at the beginning of an experiment when only the modulating beam is illuminated.

Fm – The maximum fluorescence yield (Fm) is attained when the dark adapted sample is exposed to an intense saturating pulse of light from the chlorophyll fluorometer. This temporarily reduces all PSII electron acceptors preventing PSII photochemistry. The temporary absence of competition from photochemistry for absorbed energy ensures maximum chlorophyll fluorescence emission from the sample.

Fv – The difference between the Fo and Fm chlorophyll fluorescence yield relates the maximum capacity for photochemical energy quenching by the sample and is defined as variable fluorescence (Fv).

Fv/Fm – Calculation of the rate constants for competing energy dissipation pathways in tissue under dark-adapted (Fo) and light-saturated (Fm) conditions have shown that the ratio of variable to maximal chlorophyll fluorescence (Fv/Fm) is directly proportional to the quantum efficiency of PSII photochemistry (Butler 1977, 19781). Close correlation with other measures of quantum efficiency of photochemistry in a wide range of species (Björkman and Demmig 19872) has resulted in widespread use of Fv/Fm as a screening parameter for stress response.

Light adapted parameters

In the case of light adapted tissue, a proportion of PSII electron acceptors are reduced, closing some PSII reaction centers. Hence the probability that absorbed energy is used for photochemistry is not maximal as competing non-photochemical processes are operating. The measurement of the light adapted ratio of variable to maximal chlorophyll fluorescence ratio permits the estimation of PS II quantum efficiency (\Phi PSII) using the model of Genty et al. 19893.

Fs, Fm' and \Phi PSII – Several measurements of fluorescence yield from the sample in different defined states are required to estimate \Phi PSII. Initially, the fluorescence yield of the sample under the ambient light regime is required. Such measurements are often made after a sample has adapted to a particular light regime or environment and is operating at steady state. Consequently, the measurement is often referred to as the steady-state fluorescence yield or Fs.

A fully saturating pulse from the chlorophyll fluorimeter is then required to close all of the PSII reaction centers; the temporary inhibition of PSII photochemistry ensures that the maximal fluorescence yield (Fm') is achieved. If a previous dark adapted measurement of Fv/Fm has been made the extent of photochemical and non-photochemical quenching processes can be determined from the equations of Schreiber et al 19864.

Fo' and \Phi PSIIR – Adaptation to high irradiance can involve significant changes in the confirmation of the photosynthetic apparatus which result in non-photochemical energy dissipation in the PSII antennae, before energy reaches the reaction centers (Horton et al. 19915, Horton and Ruban 19946). Failure to account for this effect can lead to inaccuracies in calculation of the relative contributions of photochemical and non-photochemical energy dissipation. This problem can be overcome by transiently shading the sample and using a far-red light source to preferentially excite PSI relative to PSII (electrons are drawn through the electron transport chain effectively opening PSII reaction centers and allowing measurement of a light adapted Fo, usually given the notation Fo'.

Other parameters measured

PAR – Takes a reading of the current incident photosynthetically active radiation value in µmol m-2 s-1 from the on-board PAR sensor of the FMS/PTL if connected to the chlorophyll fluorometer. Please note that the FMS/PTL leafclip is supplied as standard with FMS 2 but must be purchased as an option for FMS 1.

Temp – Takes a reading of the current temperature in °C from the on-board thermocouple of the FMS/PTL if connected to the chlorophyll fluorometer. Please note that the FMS/PTL leafclip is supplied as standard with FMS 2 but must be purchased as an option for FMS 1.

qP – A measure of the photochemical quenching coefficient calculated as:

(Fm'-Fs) / (Fm'- Fo)

qNP – A measure of the non-photochemical quenching coefficient calculated as:

(Fm-Fm') / (Fm-Fo)

NPQ – An alternative definition of non-photochemical quenching calculated as:

(Fm-Fm') / Fm'

ETR – A measure of the electron transport rate calculated as:

PAR \times 0.5 \times \Phi PSII \times  0.84

Measurement of ETR requires the FMS/PTL leafclip which is supplied as standard with FMS 2+ but must be purchased as an option for FMS 1+.

1 Butler, W. L., (1977).
Chlorophyll fluorescence: a probe for electron transfer and energy transfer.
In Encyclopaedia of Plant Physiology, ed. A. Trebst, M. Avron, 5, 149-167. Berlin: Springer-Verlag.

Butler, W. L., (1978).
Energy distribution in the photochemical apparatus of photosynthesis.
Annual Review of Plant Physiology, 29, 345-378.

2 Björkman, O. and B. Demmig (1987).
Photon yield of O2 evolution and chlorophyll fluorescence characteristics at 77K among vascular plants of diverse origins.
Planta, 170, 489-504.

3 Genty, B., Briantais, J-M. and N.R. Baker (1989).
The relationship between the quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence.
Biochimica et Biophysica Acta, 990, 87-92.

4 Schreiber, U., Schliwa, W. and U. Bilger (1986).
Continuous recording of photochemical and non-photochemical chlorophyll fluorescence quenching with a new type of modulation fluorimeter.
Photosynthesis Research, 10, 51-62.

5 Horton, P., Ruban, A.V., Rees, D., Pascal, A. A., Noctor, G. and A. J. Young (1991).
Control of the light-harvesting function of chloroplast membranes by aggregation of the LHCII chlorophyll-protein complexes.
FEBS Lett., 292,1-4.

6 Horton, P. and A. Ruban (1994).
The role of light-harvesting complex II in energy quenching. In Photoinhibition of photosynthesis from molecular mechanisms to the field
ed. N. R. Baker and J. R. Bowyer pp. 111-128. Oxford: BIOS, Scientific Publishers Ltd.