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A considerable literature has developed around the concept that the complexity of biological organisms, and the development of ever increasing complexity during biological evolution, is driven, in some way, by the maximization of entropy production (MEP). Most of these studies deal in very general terms with the living state and do not examine specific biological models. In the present study, we discuss some of the basic postulates of MEP as they are applied to living organisms. It is concluded that the MEP ideas seem to have little relevance either for the development of biological complexity or biological evolution.

The thermodynamics of light absorption is analysed using, as model system, a two-level molecular system characterised by mirror symmetry between its absorption and emission spectra. This means that the structure of the HOMO-1 and HOMO (highest occupied molecular orbital) ground state and the LUMO, LUMO + 1 (lowest unoccupied molecular orbital) excited states are nearly identical in their vibronic structures. This is a good model for many dyes and, in particular, for chlorophyll molecules, the main chromophore presents in the photosynthetic apparatus of plants. When a photon is absorbed by this two-level system, a transition from the HOMO ground state to the LUMO excited state occurs. Then, the system rapidly relaxes to an equilibrium distribution over the excited state sublevels and this equilibrium distribution is defined by the temperature of the bath that contains the molecule. In the described scenario, taking into account the changes in internal energy and the partition function before and after the photon absorption event, it is demonstrated that no entropy change occurs upon electronic transition from the ground state to the excited state, upon photon absorption. Furthermore, the idea that the photon-induced electronic transition can be described in terms of system heating is considered. We conclude that photon energy cannot be considered as heat. Instead, when thermal excitation is taken into account, the increased system temperature leads inexorably to entropy changes.

It is often suggested that Life may lay outside the normal laws of Physics and particularly of Thermodynamics, though this point of view is refuted by many. As the Living State may be thought of as an open system, often far from equilibrium, most attempts at placing Life under the umbrella of the laws of Physics have been based, particularly in recent years, on non-equilibrium Thermodynamics and particularly the Maximum Entropy Production Principle. In this view it is the dissipation of entropy (heat) which permits the ever increasing complexity of Living Systems in biological evolution and the maintenance of this complexity. However, these studies usually consider such biological entities as whole cells, organs, whole organisms and even Life itself at the entire terrestrial level. This requires making assumptions concerning the Living State, which are often not soundly based on observation and lack a defined model structure. The present study is based on an entirely different approach, in which a classical thermodynamic analysis of a well-defined biological nanoparticle, plant Photosystem I, is performed. This photosynthetic structure, which absorbs light and performs primary and secondary charge separation, operates with a quantum efficiency close to one. It is demonstrated that when monochromatic light is absorbed by the lowest lying electronic transition, the chlorophyll Qy transition, entropy production in the system bath plus entropy changes internal to the system are numerically less than the entropy decrease of the light field. A Second Law violation is therefore suggested for these experimental conditions. This conclusion, while at first sight is supportive of the famous and much discussed statement of Schroedinger, that "Life feeds on negentropy", is analysed and the conditions in which this statement may be considered valid for a Plant Photosystem are defined and delimited. The remarkably high quantum efficiency, leading to minimal entropy production (energy wastage), seems to suggest that evolution of Photosystem I has gone down the road of maximal energy efficiency as distinct from maximal entropy production. Photosystem I cannot be considered a maximum entropy dissipation structure.

It is often suggested that Life may lay outside the normal laws of Physics and particularly of Thermodynamics, though this point of view is refuted by many. As the Living State may be thought of as an open system, often far from equilibrium, most attempts at placing Life under the umbrella of the laws of Physics have been based, particularly in recent years, on non-equilibrium Thermodynamics and particularly the Maximum Entropy Production Principle. In this view it is the dissipation of entropy (heat) which permits the ever increasing complexity of Living Systems in biological evolution and the maintenance of this complexity. However, these studies usually consider such biological entities as whole cells, organs, whole organisms and even Life itself at the entire terrestrial level. This requires making assumptions concerning the Living State, which are often not soundly based on observation and lack a defined model structure. The present study is based on an entirely different approach, in which a classical thermodynamic analysis of a well-defined biological nanoparticle, plant Photosystem I, is performed. This photosynthetic structure, which absorbs light and performs primary and secondary charge separation, operates with a quantum efficiency close to one. It is demonstrated that when monochromatic light is absorbed by the lowest lying electronic transition, the chlorophyll Q

We have investigated the concept of the so-called “antenna entropy” of higher plant photosystems. Several interesting points emerge: 1. In the case of a photosystem which harbours an excited state, the “antenna entropy” is equivalent to the configurational (mixing) entropy of a thermodynamic canonical ensemble. The energy associated with this parameter has been calculated for a hypothetical isoenergetic photosystem, photosystem I and photosystem II, and comes out in the range of 3.5 - 8% of the photon energy considering 680 nm. 2. The “antenna entropy” seems to be a rather unique thermodynamic phenomenon, in as much as it does not modify the free energy available for primary photochemistry, as has been previously suggested. 3. It is underlined that this configurational (mixing) entropy, unlike heat dispersal in a thermal system, does not involve energy dilution. This points out an important difference between thermal and electronic energy dispersal.

The fluorescence emission characteristics of the photosynthetic apparatus under conditions of open ( F 0 ) and closed ( F M ) Photosystem II reaction centres have been investigated under steady state conditions and by monitoring the decay lifetimes of the excited state, in vivo , in the green alga Chlorella sorokiniana . The results indicate a marked wavelength dependence of the ratio of the variable fluorescence, F V = F M − F 0 , over F M , a parameter that is often employed to estimate the maximal quantum efficiency of Photosystem II. The maximal value of the F V / F M ratio is observed between 660 and 680 nm and the minimal in the 690–730 nm region. It is possible to attribute the spectral variation of F V / F M principally to the contribution of Photosystem I fluorescence emission at room temperature. Moreover, the analysis of the excited state lifetime at F 0 and F M indicates only a small wavelength dependence of Photosystem II trapping efficiency in vivo .

State transitions are an important photosynthetic short-term response that allows energy distribution balancing between photosystems I (PSI) and II (PSII). In plants when PSII is preferentially excited compared with PSI (State II), part of the major light-harvesting complex LHCII migrates to PSI to form a PSI-LHCII supercomplex. So far, little is known about this complex, mainly due to purification problems. Here, a stable PSI-LHCII supercomplex is purified from Arabidopsis thaliana and maize (Zea mays) plants. It is demonstrated that LHCIIs loosely bound to PSII in State I are the trimers mainly involved in state transitions and become strongly bound to PSI in State II. Specific Lhcb1-3 isoforms are differently represented in the mobile LHCII compared with S and M trimers. Fluorescence analyses indicate that excitation energy migration from mobile LHCII to PSI is rapid and efficient, and the quantum yield of photochemical conversion of PSI-LHCII is substantially unaffected with respect to PSI, despite a sizable increase of the antenna size. An updated PSI-LHCII structural model suggests that the lowenergy chlorophylls 611 and 612 in LHCII interact with the chlorophyll 11145 at the interface of PSI. In contrast with the common opinion, we suggest that the mobile pool of LHCII may be considered an intimate part of the PSI antenna system that is displaced to PSII in State I.

A number of spectroscopic characteristics of three almost isoenergetic, red-shifted chlorophylls (chls) in the PS II antenna complex CP29 are investigated with the aim of (i) determining whether their band shapes are substantially identical or not, (ii) addressing the topical problem of whether they are involved in excitonic interactions with other chls, and (iii) establishing whether their binding sites may be defined as "mixed" with respect to their capacity to bind chls a and b. The three chls A2-CHL612, A3-CHL613, and B3-CHL614 were analyzed after in vitro apoprotein-pigment reconstitution using the CP29 coding sequence from Arabidopsis thaliana for both the wild-type and mutant complexes. Difference spectra thermal broadening analyses indicated that the half-bandwidths varied between 12 and 15 nm (at room temperature), due mainly to differences in the optical reorganization energy (25-40 cm(-1)). Moreover, only the A2 chl displayed an intense vibrational band in the 300-600 cm(-1) interval from the 0-0 transition. We conclude that within the red absorbing (approximately 680 nm) antenna chls of a single chl-protein complex a marked spectral band shape heterogeneity exists. By analysis of the absorption and circular dichroism spectra no evidence was found of significantly strong excitonic interactions. The single gene mutation of the A3 and B3 binding sites causes absorption changes in both the long wavelength chl a absorbing region and in the chl b spectral region. This has previously been observed and was attributed to "mixed" chl a/b binding sites [Bassi, R., Croce, R., Cugini, D., and Sandona, D. (1999) Proc. Natl. Acad. Sci. U.S.A. 96,10056-10061]. This interpretation, while in principle not being unreasonable, is shown to be incorrect for these two chls.

The average, corrected attenuance spectra for both spectral forms of phytochrome in a mature leaf were calculated. Optical masking by chlorophyll together with the detour effect (optical path lengthening effect) due to multiple light scattering led to large changes in both the Qy band shape and wavelength position and the effective intensity of the weak vibrational bands increases. The Pfr/Pr oscillator-strength-ratio between 400-750 nm (0.93 in vitro), becomes 1.63 in a leaf. Thus the dominant absorption form is Pfr. These two values permit calculation of the phytochrome photoequilibrium under conditions of "daylight" illumination both in vitro and in folia. These values are 0.6 and 0.38 respectively. Previous literature estimates for the situation in vitro, based on the 660/730 nm absorption ratio, yielded values close to 0.6. It is demonstrated that this large decrease in the phytochrome photoequilibrium in a leaf has the effect of translating this parameter to a position on the dose (red/far-red light ratio)-response (Pfr/Ptot) plot towards greater sensitivity to changes in the environmental red/far-red ratio. The increased sensitivity factor is almost five-fold for the "daylight" environment and is even greater for the various "shade-light" environments. The approximate time taken to attain photoequilibrium (1/e lifetime) has also been calculated for phytochrome in a leaf in different light environments. For the "daylight" environment the photoequilibration time is approximately 5 s, which increases into the 20-80 s interval under different degrees of "shade light". Thus, despite the strong optical masking by chlorophyll in a mature leaf, the phytochrome photoequilibrium is attained quite rapidly on a physiological time scale.

Knox and Parson have objected to our previous conclusion on possible negative entropy production during primary photochemistry, i.e., from photon absorption to primary charge separation, by considering a pigment system in which primary photochemistry is not specifically considered. This approach does not address our proposal. They suggest that when a pigment absorbs light and passes to an excited state, its entropy increases by hν/T. This point is discussed in two ways: (i) from considerations based on the energy gap law for excited state relaxation; (ii) using classical thermodynamics, in which free energy is introduced into the pigment (antenna) system by photon absorption. Both approaches lead us to conclude that the excited state and the ground state are isoentropic, in disagreement with Knox and Parson. A discussion on total entropy changes specifically during the charge separation process itself indicates that this process may be almost isoentropic and thus our conclusions on possible negentropy production associated with the sequence of reactions which go from light absorption to the first primary charge separation event, due to its very high thermodynamic efficiency, remain unchanged.

On the basis of experiments with singlet quenchers and in agreement with previous data, it is suggested that a population of energetically weakly coupled chlorophylls may play a central role in photoinhibition in vivo and in vitro. In the present study, we have used steady state fluorescence techniques to gain direct evidence for these uncoupled chlorophylls. Due to the presence of their emission maxima, near 650 nm and more prominently in the 670–675 nm interval both chlorophylls b and a seem to be involved. A straightforward mathematical model is developed to describe the data which allows us to conclude that the uncoupled/weakly coupled population size is in the range of 1–3 molecules per photosystem.

Visible absorption spectra and circular dichroism (CD) of the red absorption band of isolated photosystem II reaction centers were measured at room temperature during progressive bleaching by electrochemical oxidation, in comparison with aerobic photochemical destruction, and with anaerobic photooxidation in the presence of the artificial electron acceptor silicomolybdate. Initially, selective bleaching of peripheral chlorophylls absorbing at 672 nm was obtained by electrochemical oxidation at +0.9 V, whereas little selectivity was observed at higher potentials. Illumination in the presence of silicomolybdate did not cause a bleaching but a spectral broadening of the 672-nm band was observed, apparently in response to the oxidation of carotene. The 672-nm absorption band is shown to exhibit a positive CD, which accounts for the 674-nm shoulder in CD spectra at low temperature. The origin of this CD is discussed in view of the observation that all CD disappears with the 680-nm absorption band during aerobic photodestruction.

Selective destruction of the strongly dichroic red-shifted chlorophyll form (C709 nm) in photosystem I (PSI) trimers from Spirulina, by either non-selective high intensity illumination (photobleaching) or incubation with low concentrations of Triton X-100 is accompanied by changes in the circular dichroism spectrum of the same amplitude and of opposite sign at 677 nm. The data are interpreted in terms of a dimeric chlorophyll structure with excitonic bands at these two wavelengths. Similar photobleaching experiments with PSI-200 from maize also suggest the presence of bulk antenna/red form excitonic interactions.

In photosystem I trimers of Spirulina platensis a major long wavelength transition is irreversibly bleached by illumination with high-intensity white light. The photobleaching hole, identified by both absorption and circular dichroism spectroscopies, is interpreted as the inhomogeneously broadened Q y transition of a chlorophyll form that absorbs maximally near 709nm at room temperature. Analysis of the mean square deviation of the photobleaching hole between 80 and 300K, in the linear electron-phonon frame, indicates that the optical reorganization energy is 52cm −1 , four times greater than that for the bulk, short-wavelength-absorbing chlorophylls, and the inhomogenous site distribution bandwidth is close to 150cm −1 . The room temperature bandwidth, close to 18.5nm, is dominated by thermal (homogeneous) broadening. Photobleaching induces correlated circular dichroism changes, of opposite sign, at 709 and 670nm, which suggests that the long wavelength transition may be a low energy excitonic band, in agreement with its high reorganization energy. Clear identification of the 709-nm spectral form was used in developing a Gaussian description of the long wavelength absorption tail by analyzing the changing band shape during photobleaching using a global decomposition procedure. Additional absorption states near 720, 733, and 743nm were thus identified. The lowest energy state at 743nm is present in substoichiometric levels at room temperature and its presence was confirmed by fluorescence spectroscopy. This state displays an unusual increase in intensity upon lowering the temperature, which is successfully described by assuming the presence of low-lying, thermally populated states.

A photosystem I preparation from maize, containing its full antenna complement (PSI-200) and in which detergent effects on chlorophyll coupling are almost completely absent, has been studied by time-resolved fluorescence techniques with approximately 5 ps resolution at 280 and 170 K in the wavelength interval of 690-780 nm. The data have been analyzed in terms of both the decay-associated spectra (DAS) and the time-resolved emission spectra (TRES). As in a previous room temperature study [Turconi, S., Weber, N., Schweitzer, D., Strotmann, H., and Holzwarth, A. R. (1994) Biochim. Biophys. Acta 1187, 324-334], the 280 K decay is well described by three DAS components in the 11-130 ps time range, the fastest of which displays both positive and negative amplitudes characteristic of excitation transfer from the bulk to the red antenna forms. Both the 57 and 130 ps components have all positive amplitudes and describe complex decay and equilibration processes involving the red forms. At 170 K, four major components in the 10-715 ps time range are required to describe the decay. The fastest represents bulk to red form transfer processes, while the 55, 216, and 715 ps decays, with all positive amplitudes, have maxima near 720, 730, and 740 nm, respectively, in accord with previous steady-state fluorescence measurements. The width and asymmetry of these DAS indicate that they are spectrally complex and represent decay and equilibration processes involving the red forms. Spectral evolution during the fluorescence decay process was analyzed in terms of the TRES. The red shifting of the TRES was analyzed in terms of the first central spectral moment (mean spectral energy) which is biexponential at both temperatures. The slower component, which describes equilibration between the red forms, leads to spectral red shifting during the entire fluorescence decay process, and the mean lifetimes of the spectral moments at 280 and 170 K (86 and 291 ps, respectively) are similar to the mean lifetimes of the fluorescence decays (119 and 384 ps, respectively). Thus, both spectral evolution and the trapping-associated fluorescence decay occur on a similar time scale, and both processes display a very similar temperature sensitivity. On the basis of these data, it is concluded that trapping in PSI-200 is to a large extent rate-limited by excitation diffusion in the antenna and in particular by the slow "uphill" transfer from the low-energy forms to the bulk and/or inner core chlorophyll molecules.

The spectral characteristics of fluorescence quenching by open reaction centres in isolated Photosystem II membranes were determined with very high resolution and analysed. Quenching due to photochemistry is maximal near 687 nm, minimal in the chlorophyll b emission interval and displays a distinctive structure around 670 nm. The amplitude of this `quenching hole' is about 0.03 for normalised spectra. On the basis of the absorption spectra of isolated chlorophyll–protein complexes, it is shown that these quenching structures can be exactly described by assuming that photochemistry lowers the fluorescence yield of the reaction centre complex (D1/D2/cytb 559) plus CP47, with quenching of the former complex being approximately double that of the latter complex. These data, which qualitatively indicate that there are kinetically limiting processes for primary photochemistry in the antenna, have been analysed by means of several different kinetic models. These models are derived from present structural knowledge of the arrangement of the chlorophyll–protein complexes in Photosystem II and incorporate the reversible charge separation characteristic of the exciton/radical pair equilibration model. In this way it is shown that Photosystem II cannot be considered to be purely trap limited and that exciton migration in the antenna imposes a diffusion limitation of about 30%, irrespective of the structural model assumed.

We carried out a kinetic analysis of the light-induced fluorescence quenching (AF) of the light-harvesting chlorophyll a/b pigment-protein complex of photosystem II (LHCII) that was first observed by Jennings et at (Pho-tosynth. Res. 27, 57–64, 1991). We show that during a 2 min light, 2 min dark cycle, both the light and dark phases exhibit biexponential kinetics; this is tentatively explained by the presence of two types of light-induced quenchers in different domains of aggregated LHCII. Quantitative analysis could be carried out on the faster kinetic component; the slower component that was not completed during the measurement was not amenable for quantitative analysis. Our analysis revealed that the rate of the light-induced decrease of the fluorescence yield depended linearly on the light intensity, which shows that the generation of the quencher originates from a reaction that is first order with respect to the concentration of the excited domains. As shown by the estimated rate constant, pho-togeneration of the quencher is a fast reaction that can compete with other excitation-relaxation pathways. Both the decay and the recovery time constants of AF depended strongly on the temperature. Thermodynamic analysis showed that the fast light-induced decline in the fluorescence was determined by a low fraction of the excited states. Recovery was associated with large decrease in the entropy of activation that indicated the involvement of large structural rearrangements. Macroaggregated LHCII exhibited larger ΔF than small aggregates, which is consistent with the proposed role of aggregated LHCII in thy-lakoid membranes in nonphotochemical quenching.

The red tail of the absorption spectrum of the D1-D2-cytb559 complex, defined as the absorption signal not described by the two Gaussian sub-bands associated with the intense electronic transitions at 680 and 683 nm, exhibits anomalous temperature behavior. This tail was analyzed in the temperature interval between 80 and 300 K in terms of the mean square deviation (sigma2) of the total Qy absorption band and by Gaussian sub-band decomposition. The value of the average optical reorganization energy (Snum) obtained from the temperature dependence of sigma2 for the whole absorption band was 32 cm(-1), and changed to 16-20 cm(-1) after subtraction of the sub-bands describing the red tail. This latter value is in agreement with the hole burning literature data for chlorophyll bound to proteins, and indicates that the rather high value for the apparent optical reorganization energy obtained by analysis of the total Qy band of the D1-D2-cytb559 complex is determined by the temperature sensitivity of the red tail. This suggests that the long wavelength absorption tail might be due to vibrational transitions associated with vibrational modes in the range of 80-150 cm(-1) which are thermally accessible and give rise to an absorption signal on the low-energy side of the (0,0) transition. On the basis of this assumption, the electron-phonon coupling strength (S) for these modes is estimated to be in the range 0.028-0.18. This interpretation furthermore supports the idea that the electronic transition near 683 nm is that of a monomer chlorophyll.

We have investigated the importance of the long wavelength absorbing spectral forms (red forms) of Photosystem I in photosynthetic light harvesting by leaves. To this end leaf spectra were simulated by using a linear combination of absorption (OD) spectra of purified Photosystem I, Photosystem II and LHC II, multiplied by an empirical multiple scattering chloroplast/leaf conversion function. In this way it is demonstrated that while the PS I red forms account for only about 4–5% of light absorption in a normal ‘daylight’ environment, in different ‘shadelight’ environments these long wavelength pigments may be responsible for up to 40% of total photon capture. In the context of maximising the photosynthetic quantum efficiency under the low light conditions of ‘shadelight’, this relative increase in the absorption cross section of PS I can be understood by considering the increased synthesis of the major PS II antenna complex, LHC II, known to occur in plants growing under these light conditions. It is demonstrated that for plants in a moderate to deep ‘shadelight’ regime the PS II cross section needs to increase by 50% to 100% via LHC II synthesis to balance the increased PS I absorption by the red forms. The possibility that under ‘shade light’ conditions the increased PS I cross section may serve in cyclic phosphorylation is also discussed.

Photobleaching of the isolated D1/D2/cytochrome (cyt) b -559 was performed at room temperature and 80K and the absorption spectra analysed. While at room temperature the permanent bleaching of four red absorbing transitions is observed, at 80K the selectivity is greatly increased with bleaching of transitions at 675 nm and 683 nm. The signal associated with the 683 nm structure is sufficiently strong to permit analysis of the bandwidth and band symmetry characteristics, which in the linear electron-phonon coupling assumption indicates an electron-phonon coupling strength ( S ) of about 0.7–1.0 for coupling to bath phonons of frequency ν m =20–30 cm −1 , and an essentially symmetrical Q y (0,0) band shape. Circular dichroism (CD) spectra of the D1/D2/cyt b -559 complex were measured at room temperature and 150K and analysed in terms of gaussian subbands. A minimal description with three subbands at 668 nm, 674 nm and 682 nm was found, though a more convincing description required four subbands with peak positions at 668 nm, 674–675 nm, 680.5 nm and 683 nm. The 680.5 nm transition is thought to be associated with P680 and has bandwidth characteristics in line with the published electron-phonon coupling strength (D. Tang, R. Jankowiak, M. Seibert, C.F. Yocum, G.J. Small, J. Phys. Chem. 94 (1990) 6519–6522), indicating that this transition has not undergone significant excitonic narrowing. The 683 nm CD subband is probably associated with the same transition which gives rise to the major absorption photobleaching signal on the basis of its wavelength position and band shape characteristics. The probable presence of four transitions in the CD spectrum of the D1/D2/cyt b -559 complex is in line with the suggestion that multiple (weak) excitonic interactions may occur between pigments in this chlorophyll-protein complex.

It is argued that the chemical potential analogy does not provide useful information on the thermodynamics of photosystems, as the thermodynamic efficiency of an absorbed quantum is not considered. Instead, the approach based on either entropy balance or entropy flux considerations does provide this information. At high thermodynamic efficiencies, primary photochemistry can, in principle, violate the Second Law of Thermodynamics.

Reaction centres are the sites of primary energy conversion in photosynthesis. The energy of the absorbed photon is trapped photochemically, with high quantum yields, in the form of electrochemical potential in a radical pair, which is then stabilised by a cascade of electron transfer (ET) reactions. These occur through a chain of redox-active cofactors that are coordinated by the reaction centre protein subunits. It is commonly considered that each of the electron transfer steps in the redox chain is associated with a significant driving force and, hence, a relatively large, negative, standard free energy difference (ΔG 0 ≤ − 100 meV). In this scenario, the rate of the reverse reaction which describes the repopulation of the precursor is several orders of magnitude smaller than that of forward electron transfer. Hence, the actual electron transfer rate between each given pair of donor and acceptor molecules is determined almost exclusively by the molecular rate constant of the forward reaction.

The concept that the Carnot efficiency places an upper limit of 0.60-0.75 on the thermodynamic efficiency of photosynthetic primary photochemistry is examined using the PSI-LHCI preparation. The maximal quantum efficiency was determined approximate to 0.99 which yielded a thermodynamic efficiency of 0.96, a value far above that predicted on the basis of the Carnot efficiency. The commonly presented reasoning leading to the Carnot efficiency idea was therefore critically examined. It is concluded that the crucial assumption that the pigment system, under illumination, is in equilibrium with the incident light field, at a black body temperature of T-r, is erroneous, as the temperature of the excited state pigments was experimentally shown to be that of the sample solvent (thermal bath), 280 K in this case. It is concluded that the classical reasoning used to describe the thermodynamics of heat systems is not applicable to "photonic" systems such as plant photosystems.

International audience; Oxygenic photosynthesis is indispensable both for the development and maintenance of life on earth by converting light energy into chemical energy and by producing molecular oxygen and consuming carbon dioxide. This latter process has been responsible for reducing the CO2 from its very high levels in the primitive atmosphere to the present low levels and thus reducing global temperatures to levels conducive to the development of life. Photosystem I and photosystem II are the two multi-protein complexes that contain the pigments necessary to harvest photons and use light energy to catalyse the primary photosynthetic endergonic reactions producing high energy compounds. Both photosystems are highly organised membrane supercomplexes composed of a core complex, containing the reaction centre where electrontransport is initiated, and of a peripheral antenna system, which is important for light harvesting and photosynthetic activity regulation. If on the one hand both the chemical reactions catalysed by the two photosystems and their detailed structure are different, on the other hand they share many similarities. In this review we discuss and compare various aspects of the organisation, functioning and regulation of plant photosystems by comparing them for similarities and differences as obtained by structural, biochemical and spectroscopic investigations.

The fluorescence yield for 694 nm excitation in a Photosystem I-200 particle is significantly lower than that for 665 nm excitation. This supports the previous suggestion, based on a thermodynamic analysis of absorption and emission spectra, that thermal equilibration in the 690–700 nm spectral interval is perturbed, presumably by primary photochemistry [Croce et al. (1996) Biochem 35: 8572–8579]. This equilibrium perturbation was used in the present study as a novel fit parameter in numerical simulations aimed at describing the kinetic/thermodynamic properties of exciton flow and primary photochemistry in PS I. To this end a four energy level scheme was developed which satisfactorily described all the fit parameters, including that of the equilibrium perturbation. An important characteristic which distinguished this model from other model studies is the presence of a number of chlorophyll molecules with absorption maximum near 695 nm, tightly coupled to P700. The main conclusions are: (I) about six chlorophyll molecules absorbing near 695 nm are tightly coupled to P700, in close agreement with the recent crystallographic structure for the Photosystem I core [Kraus et al. (1996); Nature Struct Biol 3: 965–973]; (II) energy transfer from the bulk pigments to the P700 core pigments is slow; (III) analysis of the most physically straightforward model indicates that the primary photochemical charge separation rate is very high (kpc ≥ 2.5 ps-1), though it is possible to simulate the equilibrium perturbation with lower kpc values assuming a large free energy decrease in the excited state of P700; (IV) the red spectral forms slow down reaction centre trapping by a 2–3 fold factor.

We examine ergodicity and configurational entropy for a dilute pigment solution and for a suspension of plant photosystem particles in which both ground and excited state pigments are present. It is concluded that the pigment solution, due to the extreme brevity of the excited state lifetime, is non-ergodic and the configurational entropy approaches zero. Conversely, due to the rapid energy transfer among pigments, each photosystem is ergodic and the configurational entropy is positive. This decreases the free energy of the single photosystem pigment array by a small amount. On the other hand, the suspension of photosystems is non-ergodic and the configurational entropy approaches zero. The overall configurational entropy which, in principle, includes contributions from both the single excited photosystems and the suspension which contains excited photosystems, also approaches zero. Thus the configurational entropy upon photon absorption by either a pigment solution or a suspension of photosystem particles is approximately zero. (C) 2014 Elsevier E.V. All rights reserved.

Absorption spectra in the Qy region of the light harvesting complex II (LHCII) have been measured in the temperature range 70-300 K. The spectra were analyzed by evaluating the temperature dependence (a) of the total bandwidth and (b) of the sub-bands obtained by numerical decomposition in terms of double Gaussians. The thermal broadening of the bands are interpreted, in both cases, as a homogeneous component, due to the presence of linear electron-phonon coupling, plus an inhomogeneous component, due to both statistical energy fluctuations at each pigment site and heterogeneity of the sample itself. Sub-bands analysis, in which eight major components are identified, yields a reorganization energy 9 cm(-1)or = Svmor = 14 cm(-1) and an inhomogeneous contribution in the range 120-170 cm(-1). In all cases the bands are substantially symmetrical in the 70-300 K temperature range. This observation gains theoretical support from an analysis of the band moments, when the influence of a low-frequency vibrational mode is considered. Analysis of the total absorption band yields Svm approximately 70 cm(-1); however, this high value is reduced to Svm approximately 11-20 cm(-1) when the red-most sub-band, with maximum at 684 nm, is eliminated at all temperatures. These data are discussed in terms of the underlying transitions, giving strong support to the presence of extreme red absorption bands in LHCII. The presence of another low-frequency mode with vm20-30 cm(-1) is also proposed.

Photosystem I with its full antenna complement (PSI-LHCI) has been prepared by mild detergent solubilization with octyl β-D-glucopyranoside from maize thylakoids. A preliminary polypeptide analysis is presented. At room temperature, the steady-state fluorescence derives from an almost perfectly thermalized state, as demonstrated by a Stepanov analysis, in which about 90% of the excited states are associated with the red chlorophyll spectral forms absorbing above 700 nm. Equilibration is temperature-sensitive and is lost at T < 200 K. A careful analysis of fluorescence between 75 and 280 K clearly demonstrates the presence of at least three red chlorophyll spectral forms with emission maxima at 720, 730, and 742 nm, the absorption origin bands of which have been calculated at 714, 725, and 738 nm. On the basis of a minor deviation from thermal equilibration around 695 nm, it is suggested that at least 3-4 antenna chlorophylls, with an average absorption near 695 nm, are strongly coupled to P700. Thermodynamic analysis of absorption and fluorescence spectra indicates that the equilibrium, absorption-weighted excited state population of the P700 dimer is around 0.013 assuming that the low-energy exciton state possesses all the oscillator strength. The average free energy for excitation transfer from antenna to P700 is thus calculated to be -0.26 kT at room temperature. This indicates that P700 is almost isoenergetic with its antenna at room temperature when the red forms are taken fully into account. From the calculated excited state population of P700, we estimate that the primary charge separation rate in PSI is 1-2 ps-1.

In the present paper the marked changes in photochemical trapping time over the absorption/fluorescence band of isolated PSI–LHCI are studied by means of time resolved fluorescence decay measurements. For emission at 680–690 nm the effective trapping time is close to 17–18 ps, and represents the effective trapping time from the bulk antenna. At wavelengths above 700 nm the effective trapping time increases in a monotonic way, over the entire emission band, to attain values in the range of 70–80 ps near 760 nm. This is argued to be caused by “uphill” energy transfer from the low energy states to the core antenna and reaction centre. These data, together with the steady state emission spectrum, permit calculation of the overall trapping time for maize PSI–LHCI, which is estimated to be approximately 40 ps. The wavelength dependence of the trapping time indicates, that in PSI–LHCI there exists at least one red form which emits at lower energies than the 735 nm state. These data indicate that Photosystem I is about 55% diffusion limited.

The relation between room-temperature absorption and fluorescence spectra of the freshly prepared D1/D2/cytb-559 complex was analysed according to the Stepanov equation (Stepanov, B.I. (1957) Sov. Phys. Dokl. 2, 81–84). It is shown that there is a close correspondence between the emission spectrum calculated from the absorption spectrum and the measured emission spectrum in the wavelength range 665–695 nm. This correspondence was not found for samples which had either been aged at room temperature or subjected to a brief illumination with high intensity light. The data are interpreted to indicate that in freshly prepared D1/D2/cytb-559: (i) excited states are thermally equilibrated between all chromophores including P680, (ii) the fluorescence yield of all chromophores is similar, (iii) the freshly prepared complex does not contain a significant amount of uncoupled pigment. This latter conclusion is supported by straight line Stern-Volmer fluorescence quenching kinetics using the quinone quencher of chlorophyll fluorescence dibromothymoquinone.

The absorption spectrum of the main antenna complex of photosystem II, LHCII, has been modeled using, as starting points, the chlorophyll (chl) atomic coordinates as obtained by the LHCII crystal analysis [Liu, Z., Yan, H., Wang, K., Kuang, T., Zhang, J., Gui, L., An, X., and Chang, W. (2004) Nature 428, 287-292] of three different trimers. The chl site Q(y) transition energies have been obtained in terms of the chl macrocycle deformations influencing the energy level of the chl frontier orbitals. Using these chl site transition energy values and the entire set of interaction energies, calculated in the ideal dipole approximation, the complete Hamiltonians for the three LHCII trimers have been written and the full set of 42 eigenstates of each LHCII trimer have been calculated. With the 42 transition energies and transition dipole strengths, either unperturbed or associated to the eigenstates, the LHCII Q(y) absorption spectrum has been calculated using a chl absorption band shape. These calculations have been performed both in vacuo and in the presence of a medium. Despite the number of approximations, a good correlation with the measured absorption spectrum of a LHCII preparation is obtained. This analysis shows that, although a substantial C3 symmetry of the LHCII trimer in terms of both chl-chl distances and interaction energies is present, a marked variation among monomer subsets of site transition energies is estimated. This leads to a C3 symmetry breaking in the unperturbed chl site transition energies set and, consequently, in the trimer eigenstates. It is also concluded that interactions among chlorophylls do not significantly modify the light absorption role of LHCII in plant leaves.

Room temperature and 10 K absorption and linear dichroism spectra of the chlorophyll-protein complexes comprising the outer antenna of PSII (LHCII, CP29, CP26, CP24) have been analyzed in terms of a linear combination of asymmetric Gaussian bands. The results demonstrate the following: (a) The absorption and linear dichroism spectra of each sample can be described by nearly the same set of Gaussian bands at room temperature and 10 K. (b) The relative distributions of the transition moments of the major red-absorbing spectral forms seem to be similar in all four outer antenna chlorophyll-protein complexes at room temperature, with the 684-nm band being oriented closest to the particle plane at room temperature and the 677- and 669-nm bands being tilted at progressively greater angles out of the particle plane. The shorter wavelength transitions seem to be oriented close to the magic angle, but interpretation is complicated in this spectral region due to the low linear dichroism values and by overlap with vibrational bands. (c) The 684-nm band, detected in room temperature absorption and linear dichroism spectra of all complexes, vanishes at 10 K.

The multiexponential fluorescence decay of the CP29 complex in which the apoprotein and pigments were reconstituted in vitro was examined. Of the three decay components observed only the two dominant ones, with about 3 and 5 ns lifetimes, were studied. The main question addressed was whether the multicomponent decay was associated with sample optical heterogeneity. To this end, we examined the optical absorption and fluorescence of the CP29 sample by means of two different and independent experimental strategies. This approach was used as the wavelength positions of the absorption/fluorescence spectral forms has recently been shown to be a sensitive indicator of the binding site-induced porphyrin ring deformation (Zucchelli et al. Biophys J 93:2240-2254, 2007) and hence of apoprotein conformational changes. The data indicate that this CP29 sample is optically homogeneous. It is hypothesised that the different lifetimes are explained in terms of multiple detergent/CP29 interactions leading to different quenching states, a suggestion that allows for optical homogeneity.

Fluorescence emission spectra for the seven chlorophyll-protein complexes comprising the antenna system of Photosystem II (PS II) have been measured. All four outer antenna complexes (LHC II, CP24, CP26, CP29) have relatively greater emission near 648 nm and 680 nm with respect to the inner antenna complexes (CP43, CP47, D1/D2/cyt b-559). The emission spectra for both outer and inner antenna were calculated from the measured emission spectra of the single chlorophyll-protein complexes, using as weighting factors the excited state population at thermodynamic equilibrium in the various chlorophyll-protein complexes suggested by Jennings et al. (Jennings, R.C., Bassi, R., Garlaschi, F.M., Dainese, P. and Zucchelli, G. (1993) Biochemistry 32, 3203–3210). Subsequently, the overall emission spectra for the total PS II antenna (i.e., outer plus inner antenna) were calculated for situations in which varying excited state levels were assumed for the inner and outer antenna. In an attempt to determine the steady-state distribution of excited states between outer and inner antenna these calculated fluorescence spectra were compared with those measured for (a), PS II particles prepared from maize and (b), chloroplasts of wild-type barley and the chlorina F2 mutant. From this comparison it is concluded that at steady-state fluorescence emission, between 28% and 38% of the excited states in PS II are associated with the inner antenna and between 62% and 72% with the outer antenna. These results suggest that the PS II antenna is organised as a very shallow energy funnel. This antenna organisation is discussed in terms of the generation of non-photochemical quenching mechanisms which are designed to protect PS II from high light stress.

The chlorophyll-protein complexes that form the antenna system of photosystem II have been purified and analyzed in terms of the commonly observed chlorophyll spectral forms. With the exception of chlorophyll b, which is known to be associated with the complexes comprising the outer antenna (LHCII, CP24, CP26, CP29), the spectral forms occur with similar absorption maxima and are present in rather similar amounts in each of the antenna complexes. On the basis of the published chlorophyll stoichiometries for the complexes in photosystem II antenna, the distribution of the spectral forms in a "reconstituted" antenna has been determined. These data were used to calculate the equilibrium population of excited states within the various chlorophyll-protein complexes within photosystem II. This was compared with the light absorption capacity of each of the complexes in the "reconstituted" antenna. The ratio of these two parameters (excited-state equilibrium distribution/absorption capacity) was determined to be 1.21 for the inner (core) antenna and 0.88 for LHCII. The standard free energy change for exciton transfer from the outer to the inner antenna was calculated to be -0.17 kcal mol-1. It is concluded that the photosystem II antenna is arranged as a very shallow funnel.

The recent demonstration that the room temperature fluorescence emission spectrum of Photosystem II may be described as a linear combination of Gaussian bands associated with the well-known chlorophyll spectral forms (Zucchelli, G., Jennings, R.C., Garlaschi, F.M. (1992) Biochim. Biophys. Acta, 1099, 163–169) has permitted analysis of a number of fluorescence quenching phenomena in terms of the chlorophyll spectral forms. In this context we have analysed the fluorescence quenching due to open reaction centres and also the light-induced, non-photochemical, non-reversible quenching in isolated thylakoids, a Photosystem II membrane preparation and the major light-harvesting chlorophyll a/b protein complex (LHCII). Whereas open reaction centres quench the emission associated with the spectral form chl688684 with greatest efficiency, thus indicating that it is this spectral form which is the most efficient in transferring energy to reaction centres, all the non-photochemical phenomena investigated show maximal quenching for both chl680678 and chl687684. It is concluded that the light-induced, non-photochemical, non-reversible quenching involves the generation of quenching centres within the Photosystem II antenna, possibly at the level of LHCII.

Room-temperaure steady-state emission and Q1 absorption spectra of light-harvesting chlorophyll ab protein complex II (LHCII) isolated from spinach have been analysed in terms of a linear combination of asymmetric gaussian bands. To investigate a possible correspondence between the absorption and emission bands, the thermal emission spectrum of each absorption band has been calculated. It is demonstrated that the calculated fluorescence bands correspond closely to those obtained by gaussian deconvolution. It is thus possible to associate an emission band with each absorbing chlorophyll spectral species: Chl653648, Chl663660, Chl672670, Chl680678, Chl687684, Chl697695 (Chlmn is the spectral species that absorbs with maximum at wavelength n and emits with maximum at wavelength m) and to interpret the emission spectrum of LHCII as a linear combination of the emission spectra of each absorbing chlorophyll spectral species. In particular, the correspondence between the 684 nm and 695 nm absorption bands and the emission bands with maximum at 687 nm and 697 nm lends support to the presence of long wavelength Chl a spectra forms in LCHII, the external antenna of PS II. A close correlation between the emission and absorption gaussian bands is also found in the analysis of the room temperature absorption and emission spectra of such membrane preparation as BBY-grana from spinach and thylakoids prepared from barley wild type and the chlorina f2 mutant (lacking LHCII). On the basis of these data, the commonly observed 5–7 nm wavelength difference between the absorption and emission spectra maxima of chlorophyll containing biological membranes is interpreted in terms of (a) the Stokes shifts of the single spectral forms together with (b) the increased emission contribution of the longer wavelength forms to the emission spectrum caused by energy transfer.

Steady-state room-temperature emission spectra have been determined for spinach and barley (both wild-type and the chlorina mutant lacking Chl b and LHCII) thylakoids, a spinach Photosystem II membrane preparation and Chlorella cells, under conditions in which the Photosystem II reaction centres were either open or closed. The data show that the fluorescence quenching efficiency of open reaction centres is greatest for those chlorophyll species emitting in the 680–690 nm wavelength range. The reaction centre quenching efficiency decreases at both shorter and longer wavelengths. Calculation of the fluorescence emission ratio spectra for membrane preparations containing LHCII (wild-type barley thylakoids and isolated LHCII) with respect to those without LHCII (chlorina barley mutant thylakoids) indicates that this complex has characteristic emissions around 650 nm (Chl b) and 680 nm. The Chl b emission is about 2% that of the main Chl a emission at 683 nm (open reaction centres). As the characteristic LHCII emission bands near 650 nm and 680 nm are almost absent in the ratio between LHCII emission spectrum and both spinach and wild-type barley thylakoid emission spectra at open reaction centres, it is concluded that most Photosystem II fluorescence is from LHCII when reaction centres are open. Thus, excitation energy does not seem to be preferentially localised in the core antenna complexes prior to fluorescence emission. The data are interpreted in terms of a model in which energy transfer from antenna to reaction centres is basically diffusion limited.

The photosystem II kinetic model (diffusion or trap-limited) is still much debated. There is discussion about whether energy transfer from the core antenna (CP47 and CP43) to the reaction center complex (D1-D2-cyt b 559) is rate-limiting (transfer to trap-limited). This study investigates this problem in isolated core particles by exploiting the different optical properties of the core antenna and the reaction center complex near 680 nm, due to P680 and an isoenergetic pheophytin. This was used as a marker feature for the reaction center complex. If the transfer to the trap-limited model were correct, assuming excited-state thermalization, the specific reaction center fluorescence decay lifetime should be shorter near 680 nm, where there is reaction center complex specificity, than at the other emission wavelengths. Such a selective reaction center feature was not observed in fluorescence decay measurements. At the experimental resolution used here, we conclude that the trap-limited energy transfer to the reaction center could, at the most, be 20% limiting. Thus, the transfer to the trap-limited model is not supported. A kinetic, compartmental analysis was also performed on the data, taking into account a large number of separate measurements and the associated errors. Target analysis, considering these intermeasurement errors, yielded two minima which adequately describe the fluorescence lifetime data. The nonunique nature of the description is due to the fact that we have taken into consideration these intermeasurement errors. In our case, due to these errors, a correct kinetic model interpretation required additional experimental information.

Room temperature absorption spectra (in the wavelength range 600–740 nm) of light-harvesting chlorophyll a/b protein complex II (LHCPII) isolated from spinach and pea have been analysed in terms of a linear combination of asymmetric gaussian bands. All the commonly observed chlorophyll spectra bands are found, including significant levels of the components which peak around 684 and 693 nm. The presence of these long-wavelength absorbing forms in LHCPII is also suggested by analysis of the room temperature absorption spectra of thylakoids of the chlorina barley mutant, which lacks LHCPII, and pea grown in intermittent light. From a comparative analysis of room temperature and liquid nitrogen temperature absorption spectra of LHCPII, it is concluded that temperature modifies the spectral bands, narrowing the bands up to 677 nm and markedly depressing the 684 nm band. These observations are not in close agreement with the commonly accepted ideas on the distribution of the chlorophyll spectral species with respect to reaction centres in the photosystem II (PSII) antenna matrix.

Excitation energy transfer from Photosystem II antenna to reaction centres as a function of excitation wavelength, between 646–701 nm, was studied by analysis of the fluorescence quenching efficiency of RCs in both a BBY-grana preparation and thylakoids of the chlorina barley mutant lacking LHCPII. For BBY-grana, maximum transfer was observed with an excitation wavelength slightly greater than 680 nm, with significant decreases occurring for both longer and shorter wavelengths. The data are analysed in terms of the chlorophyll spectral forms as determined by asymmetric gaussian deconvolution of room temperature absorption spectra. For the short-wavelength side of the transfer maximum (646–682 nm) the following relative transfer efficiencies have been determined: 684 > 670 > 650; 678 > 661; 684 > 678 nm. Energy transfer between Photosystem II units is shown to increase with decreasing absorption wavelength below 683 nm. Thus, the above relative transfer efficiencies are associated with transfer to reaction centres within Photosystem II units and not between PS II units. The data are discussed in terms of alternative antenna models. It is argued that they are best accommodated by a model in which the antenna spectral forms have no particular macroscopic distribution with respect to RCs and in which the lower-wavelength-absorbing spectral forms may function as ‘antitraps’ for the longer-wavelength forms as determined by the transfer microparameters. Analysis of the gaussian band associated with Chl b in BBY-grana and LHCII suggest that this antenna component has a low-transition dipole strength within Photosystem II antenna complexes. It is suggested that this will enhance the role of Chl b as an ‘antitrap’ for excited states associated with the longer-wavelength-absorbing chlorophyll species.

The analysis of FDMR spectra, recorded at multiple emission wavelengths, by a global decomposition technique, has allowed us to characterise the triplet populations associated with Photosystem I and Photosystem II of thylakoids in the green alga Chlamydomonas reinhardtii. Three triplet populations are observed at fluorescence emissions characteristic of Photosystem II, and their zero field splitting parameters have been determined. These are similar to the zero field parameters for the three Photosystem II triplets previously reported for spinach thylakoids, suggesting that they have a widespread occurrence in nature. None of these triplets have the zero field splitting parameters characteristic of the Photosystem II recombination triplet observed only under reducing conditions. Because these triplets are generated under non-reducing redox conditions, when the recombination triplet is undetectable, it is suggested that they may be involved in the photoinhibition of Photosystem II. At emission wavelengths characteristic of Photosystem I, three triplet populations are observed, two of which are attributed to the P700 recombination triplet frozen in two different conformations, based on the microwave-induced fluorescence emission spectra and the triplet minus singlet difference spectra. The third triplet population detected at Photosystem I emission wavelengths, which was previously unresolved, is proposed to originate from the antenna chlorophyll of the core or the unusually blue-shifted outer antenna complexes of this organism.

ELIPs (early light-induced proteins) are thylakoid proteins transiently induced during greening of etiolated seedlings and during exposure to high light stress conditions. This expression pattern suggests that these proteins may be involved in the protection of the photosynthetic apparatus against photooxidative damage. To test this hypothesis, we have generated Arabidopsis (Arabidopsis thaliana) mutant plants null for both elip genes (Elip1 and Elip2) and have analyzed their sensitivity to light during greening of seedlings and to high light and cold in mature plants. In particular, we have evaluated the extent of damage to photosystem II, the level of lipid peroxidation, the presence of uncoupled chlorophyll molecules, and the nonphotochemical quenching of excitation energy. The absence of ELIPs during greening at moderate light intensities slightly reduced the rate of chlorophyll accumulation but did not modify the extent of photoinhibition. In mature plants, the absence of ELIP1 and ELIP2 did not modify the sensitivity to photoinhibition and photooxidation or the ability to recover from light stress. This raises questions about the photoprotective function of these proteins. Moreover, no compensatory accumulation of other ELIP-like proteins (SEPs, OHPs) was found in the elip1/elip2 double mutant during high light stress. elip1/elip2 mutant plants show only a slight reduction in the chlorophyll content in mature leaves and greening seedlings and a lower zeaxanthin accumulation in high light conditions, suggesting that ELIPs could somehow affect the stability or synthesis of these pigments. On the basis of these results, we make a number of suggestions concerning the biological function of ELIPs.

The time-resolved fluorescence decay of plant PSI-LHCI has been analyzed and compared with its component parts, the PSI core and the peripheral antenna LHCI, in an attempt to (i) define the physical domains associated with the multicomponent decay-associated spectra (DAS) and determine the origin of the kinetically slow steps responsible for them, (ii) formulate a clear working hypothesis for the positive decay-associated spectral amplitudes of the two slowest decay components, and (iii) determine the impact of the peripheral antenna complexes (LHCI) on the effective trapping rate for the photosystem. The results for PSI-LHCI indicate that the three exponential component DAS description, previously reported in the literature, is not numerically unique. The fit minimum is rather broad, which necessitated the introduction of other fit criteria in addition to the purely numerical one. The analysis demonstrates that (i) the physical domains associated with the multicomponent decay are associated with the antenna and particularly with the low-energy spectral forms, (ii) the positive DAS amplitudes of the two slowest decay components are suggested to be due to energy transfer kinetic heterogeneity to different F735 low-energy forms, and (iii) the peripheral antenna slows down the effective photosystem photochemical rate by about 3 times, and this is approximately half due to antenna degeneracy and half due to the low-energy forms.

Thermo-optically induced structural reorganizations have earlier been identified in isolated LHCII, the main chlorophyll a/b light harvesting complexes of Photosystem II, and in granal thylakoid membranes [Cseh et al. (2000) Biochemistry 39: 15250-15257; Garab et al. (2002) Biochemistry 41: 15121-15129]. According to the thermo-optic mechanism, structural changes can be induced by fast, local thermal transients due to the dissipation of excess excitation energy. In this paper, we analyze the temperature and light-intensity dependencies of thermo-optically induced reversible and irreversible reorganizations in the chiral macrodomains of lamellar aggregates of isolated LHCII and of granal thylakoid membranes. We show that these structural changes exhibit non-Arrhenius type of temperature dependencies, which originate from the 'combination' of the ambient temperature and the local thermal transient. The experimental data can satisfactorily be simulated with the aid of a simple mathematical model based on the thermo-optic effect. The model also predicts, in good accordance with experimental data published earlier and presented in this paper, that the reorganizations depend linearly on the intensity of the excess light, a unique property that is probably important in light adaptation and photoprotection of plants.

We have investigated the previous suggestions in the literature that the outer antenna of Photosystem II of barley does not influence the effective photosystem primary photochemical trapping rate. It is shown by steady state fluorescence measurements at the F(0) fluorescence level of wild type and the chlorina f2 mutant, using the chlorophyll b fluorescence as a marker, that the outer antenna is thermally equilibrated with the core pigments, at room temperature, under conditions of photochemical trapping. This is in contrast with the conclusions of the earlier studies in which it was suggested that energy was transferred rapidly and irreversibly from the outer antenna to the Photosystem II core. Furthermore, the effective trapping time, determined by single photon counting, time-resolved measurements, was shown to increase from 0.17+/-0.017 ns in the chlorina Photosystem II core to a value within the range 0.42+/-0.036-0.47+/-0.044 ns for the wild-type Photosystem II with the outer antenna system. This 2.5-2.8-fold increase in the effective trapping time is, however, significantly less than that expected for a thermalized system. The data can be explained in terms of the outer antenna increasing the primary charge separation rate by about 50%.

The average fluorescence decay lifetimes, due to reaction centre photochemical trapping, were calculated for wavelengths in the 690- to 770-nm interval from the published fluorescence decay-associated emission spectra for Photosystem I (PSI)-light-harvesting complex of Photosystem I (LHCI) [Biochemistry 39 (2000) 6341] at 280 and 170 K. For 280 K, the overall trapping time at 690 nm is 81 ps and increases with wavelength to reach 103 ps at 770 nm. For 170 K, the 690-nm value is 115 ps, increasing to 458 ps at 770 nm. This underlines the presence of kinetically limiting processes in the PSI antenna (diffusion limited). The explanation of these nonconstant values for the overall trapping time band is sought in terms of thermally activated transfer from the red absorbing states to the "bulk" acceptor chlorophyll (chl) states in the framework of the Arrhenius-Eyring theory. It is shown that the wavelength-dependent "activation energies" come out in the range between 1.35 and 2.7 kcal mol-1, increasing with the emission wavelength within the interval 710-770 nm. These values are in good agreement with the Arrhenius activation energy determined for the steady-state fluorescence yield over the range 130-280 K for PSI-LHCI. We conclude that the variable trapping time in PSI-LHCI can be accounted for entirely by thermally activated transfer from the low-energy chl states to the bulk acceptor states and therefore that the position of the various red states in the PSI antenna seems not to be of significant importance. The analysis shows that the bulk antenna acceptor states are on the low-energy side of the bulk antenna absorption band.

Selective excitation, at room temperature, in the long wavelength absorption tail of the photosystem I antenna complexes, known as light harvesting complex I, induces pronounced pre-equilibration fluorescence from the directly excited pigment state. This has allowed determination of the fluorescence band shape of this low energy photosystem I chlorophyll antenna state, at room temperature, for the first time. The emission maximum is near 735 nm. The remarkable band width (55 nm) and asymmetry have never been previously reported for chlorophyll a states.

The dependence of PSII triplet populations as a function of the primary quinone acceptor QA redox state has been investigated by means of the fluorescence detected magnetic resonance (FDMR) technique in isolated thylakoid membranes. The results show that the population level of a microsecond decaying component having zero field splitting parameters |D|=0.0292 cm−1 and |E|=0.0037 cm−1 is inversely correlated to that of the recombination triplet (|D|=0.0285 cm−1 |E|=0.045 cm−1). The results could be interpreted in terms of the fast (50–150 μs) decaying population being either the recombination triplet when the primary quinone QA is singly reduced or a triplet located on a chlorophyll which is in tight singlet transfer coupling with the primary donor.