In Vivo Measurements of Light Emission in Plantsa (original) (raw)
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In Vivo Measurements of Light Emission in Plants
Photosynthesis: Open Questions and What We Know Today, Allakhverdiev S. I., Rubin A. B., Shuvalov V.A. (eds.), 2014
There are several types of light emission in plants: prompt fluorescence, delayed fluorescence, thermoluminescence, and phosphorescence. This chapter focuses on two of them: prompt and delayed fluorescence. Chlorophyll a fluorescence measurements have been used for more than 80 years to study photosynthesis; since 1961, it has been used, particularly, for the analysis of PhotosystemII (PS II). Fluorescence is now used routinely in agricultural and biological research where many measured and calculated parameters are used as biomarkers or indicators of plant tolerance to different abiotic and biotic stress. This has been made possible by the rapid development of new fluorometers. Most of these instruments are mainly based on two different operational principles for the measurement of variable chlorophyll a fluorescence: (1) pulse-amplitude-modulated (PAM) excitation followed by measurement of prompt fluorescence and (2) a strong continuous actinic excitation leading to prompt fluorescence. In addition to fluorometers, other instruments have been developed to measure other signals, such as delayed fluorescence, originating mainly from PS II, and light-induced absorbance changes due to the photo-oxidation of the reaction center P700 of PS I, measured as absorption decrease (photobleaching) at about 705 nm, or increase at 820 nm. This chapter includes technical and theoretical basis of newly developed instruments that allow for simultaneous measurement of the prompt fluorescence (PF) and the delayed fluorescence (DF) as well as some other In vivo MEASUREMENTS OF LIGHT EMISSION IN PLANTS 3 parameters. Special emphasis is given here to a description of comparativemeasurements on PF and DF. Since DF is much less used and less known than PF, it is discussed in greater details; it has great potential to provide useful, and qualitatively new information on the back reactions of PS II electron transfer. This chapter, which also deals with the history of fluorometers, is dedicated to David Walker (1928–2012), who was a pioneer in the field of photosynthesis and chlorophyll fluorescence.
Delayed chlorophyll fluorescence as a monitor for physiological state of photosynthetic apparatus
2009
Intact plants emit light quanta called delayed fluorescence (DF). DF is result of radiative deactivation of secondary excited chlorophyll molecules in Photosystem II (PS II) antennae complexes. The excitations are produced by backward electrontransfer reactions both in the donor and acceptor sides of PS II. The poly-exponential dark decay of DF in a time interval of tens of nanoseconds to tens of seconds reflects the kinetics of different forward and backward reactions of the photosynthetic electron transfer. The current work reviews the mechanisms of the DF light quanta generation and the methodical approaches that allow us to obtain quantitative information about the photosynthetic machinery state using the DF signal from native objects. We examine an approach for the simultaneous record of DF and prompt chlorophyll fluorescence during the transition of the photosynthetic machinery from dark-adapted to light-adapted state. A new device (Senior PEA) built by Hansatech (King's Lynn, UK) allows us to measure simultaneously the induction transients of prompt chlorophyll fluorescence, DF decaying in a time range 10 µs-240 ms, and the changes in transmission at 820 nm. The comparative analysis of the three types of signals and the application of a model-based description of the processes and reactions that determine the dynamics of the signals during the light-induced transitions (dynamic models and JIP-test) allow us to obtain, from a single few-seconds-long measurement, quantitative information for: a) energetic fluxes and efficiencies at different steps of energy transformation; b) rate constants of electron transfer in and between the two photosystems; c) energization of the thylakoid membrane. This illustrates that DF in combination with other optical and luminescent measurements is a highly informative method for investigation of the physiological state of the photosynthetic apparatus of plants in vivo and in situ, and is an indispensable tool for the purposes of the biophysical phenomics.
Photosynthesis Research, 2012
This review is dedicated to David Walker , a pioneer in the field of photosynthesis and chlorophyll fluorescence. We begin this review by presenting the history of light emission studies, from the ancient times. Light emission from plants is of several kinds: prompt fluorescence (PF), delayed fluorescence (DF), thermoluminescence, and phosphorescence. In this article, we focus on PF and DF. Chlorophyll a fluorescence measurements have been used for more than 80 years to study photosynthesis, particularly photosystem II (PSII) since 1961. This technique has become a regular trusted probe in agricultural and biological research. Many measured and calculated parameters are good biomarkers or indicators of plant tolerance to different abiotic and biotic stressors. This would never have been possible without the rapid development of new fluorometers. To date, most of these instruments are based mainly on two different operational principles for measuring variable chlorophyll a fluorescence: (1) a PF signal produced following a pulseamplitude-modulated excitation and (2) a PF signal emitted during a strong continuous actinic excitation. In addition to fluorometers, other instruments have been developed to measure additional signals, such as DF, originating from PSII, and light-induced absorbance changes due to the photooxidation of P700, from PSI, measured as the absorption decrease (photobleaching) at about 705 nm, or
Photochemistry and Photobiology, 2003
A method for data acquisition based on recording of light signal from a conventional phophoroscope fluorometer with high-speed digitalization is proposed to extract more information from a delayed chlorophyll a fluorescence (DF) signal. During the signal processing from all points registered by the fluorometer, we obtain simultaneously a large number of induction curves of DF decaying at different time ranges. In addition, it is possible to register a series of dark relaxation kinetics of DF, recorded at different moments during the induction period or at different temperatures. This allows the evaluation of the contribution of DF kinetic components during the induction period or at different temperatures and the comparison between DF signals registered with different phophoroscopes. With the phosphoroscope system used in this study, we have shown that the contribution of the millisecond components (with lifetimes 0.6 and 2-4 ms) predominates during the first second of the induction period. After 1 s of illumination, the amplitudes of the 0.6 ms and 2-4 ms components and of the slower one (with lifetime more than 10 ms) become approximately equal. The change in lifetime of the different components during the induction and during gradual heating is also observed. It is shown that all registered DF kinetic components have different temperature dependences.
Photosynthesis Research, 1990
A newly-developed field-portable multi-flash kinetic fluorimeter for measuring the kinetics of the microsecond to millisecond reactions of the oxidizing and reducing sides of photosystem 2 in leaves of intact plants is described and demonstrated. The instrumental technique is a refinement of that employed in the 'double-flash' kinetic fluorimeter (Joliot 1974 Biochim Biophys Acta 357: 439-448) where a low-intensity short-duration light pulse is used to measure the fluorescence yield changes following saturating single-turnover light pulses. The present instrument uses a rapid series of short-duration (2/xs) pulses to resolve a complete microsecond to millisecond time-scale kinetic trace of fluorescence yield changes after each actinic flash. Differential optics, using a matrix of optical fibers, allow very high sensitivity (noise levels about 0.05% Fmax) thus eliminating the need for signal averaging, and greatly reducing the intensity of light required to make a measurement. Consequently, the measuring pulses have much less actinic effect and an entire multi-point trace (seven points) excites less than 1% of the reaction centers in a leaf. In addition, by combining the actinic and measuring pulse light in the optical fiber network, the tail of the actinic flash can be compensated for, allowing measurements of events as rapidly as 20/zs after the actinic flash. This resolution makes practical the routine measurement of the microsecond turnover kinetics of the oxygen evolving complex in leaves of intact plants in the field. The instrument is demonstrated by observing flash number dependency and inhibitor sensitivity of the induction and decay kinetics of flash-induced fluorescence transients in leaves of intact plants. From these traces the period-two oscillations associated with the turnover of the two-electron gate and the period-four oscillations associated with the turnover of the oxygen evolving complex can be observed. Applications of the instrument to extending our knowledge of chloroplast function to the whole plant, the effects on plants of environmental stress, herbicides, etc, and possible applications to screening of mutants are discussed.
Delayed fluorescence in photosynthesis
Photosynthesis Research, 2009
Photosynthesis is a very efficient photochemical process. Nevertheless, plants emit some of the absorbed energy as light quanta. This luminescence is emitted, predominantly, by excited chlorophyll a molecules in the lightharvesting antenna, associated with Photosystem II (PS II) reaction centers. The emission that occurs before the utilization of the excitation energy in the primary photochemical reaction is called prompt fluorescence. Light emission can also be observed from repopulated excited chlorophylls as a result of recombination of the charge pairs. In this case, some time-dependent redox reactions occur before the excitation of the chlorophyll. This delays the light emission and provides the name for this phenomenon-delayed fluorescence (DF), or delayed light emission (DLE). The DF intensity is a decreasing polyphasic function of the time after illumination, which reflects the kinetics of electron transport reactions both on the (electron) donor and the (electron) acceptor sides of PS II. Two main experimental approaches are used for DF measurements: (a) recording of the DF decay in the dark after a single turnover flash or after continuous light excitation and (b) recording of the DF intensity during light adaptation of the photosynthesizing samples (induction curves), following a period of darkness.
Photosynthetica, 2020
An overview is given of several studies on the fast chlorophyll (Chl) a fluorescence (OJIP) transient carried out in the laboratory of Reto Strasser between 2001 and 2009. At the beginning of this period the HandyPEA and PEA-Senior instruments were introduced by Reto Strasser and Hansatech Instruments Ltd. (UK) that gave a lot of experimental flexibility compared to the experiments that were feasible in the preceding years. These technical innovations, including the combination of 820-nm transmission measurements (for the determination of the P700 and PC redox states) and Chl a fluorescence [originating from photosystem II (PSII)], enabled us to establish the effects of electron flow through and at the acceptor side of photosystem I during a dark-to-light transition on fluorescence induction in leaves. These instruments further allowed us to show biological variability between various photosynthetic organisms and how several chemical treatments could modify the Chl a fluorescence kinetics. We also obtained new information on the effect of the inhibitor DCMU [3-(3ꞌ,4ꞌ-dichlorophenyl)-1,1-dimethylurea] on Chl a fluorescence induction. In addition, the effects of heat stress on electron flow through PSII and the entire electron transport chain were investigated in detail. The article also reflects how our perception and interpretation of the OJIP kinetics changed over time.
Photosynthesis Research, 2005
A method of partitioning the energy in a mixed population of active and photoinactivated Photosystem II (PS II) complexes based on chlorophyll fluorescence measurements is presented. There are four energy fluxes, each with its quantum efficiency: a flux associated with photochemical electron flow in active PS II reaction centres (J PS II ), thermal dissipation in photoinactivated, non-functional PS IIs (J NF ), light-regulated thermal dissipation in active PS IIs (J NPQ ) and a combined flux of fluorescence and constitutive, light-independent thermal dissipation (J f,D ). The four quantum efficiencies add up to 1.0, without the need to introduce an 'excess' term E, which in other studies has been claimed to be linearly correlated with the rate coefficient of photoinactivation of PS II (k pi ). We examined the correlation of k pi with various fluxes, and found that the combined flux (J NPQ + J f,D = J pi ) is as well correlated with k pi as is E. This combined flux arises from F s =F 0 m , the ratio of steady-state to maximum fluorescence during illumination, which represents the quantum efficiency of combined non-photochemical dissipation pathways in active PS IIs. Since F s =F 0 m or its equivalent, J pi , is a likely source of events leading to photoinactivation of PS II, we conclude that F s =F 0 m is a simple predictor of k pi .