UN NOUVEAU MODELE DES CENTRES PHOTOCHIMIQUES DU SYSTEME II*

Abstract— The amount of oxygen evolved by Chlorella cells or by isolated chloroplasts has been measured after illumination by short saturating flashes. In all conditions, the amount of oxygen evolved by one flash is proportional to the fraction of the photochemical centers susceptible to produce oxygen. If dark adapted algae or chloroplasts are illuminated by a sene of flashes, no oxygen is produced by the first flash. This phenomenon is related to the activation process. If y_n is the amount of oxygen evolved by the nth flash of the sequence, it appears that the sene y_n shows large oscillations with a period 4. These oscillations are completely damped after 4–6 periods and the amount of oxygen evolved by a flash reaches a stationary value. For any value of n. the quantities y_n and y_n+2 are linked by a recurrent relation which is the same for Chlorella cells and spinach chloroplasts. No relation can be found between the terms y_n and y_(n+1). The mathematical properties of the series y_n can be understood if one admits that a two memory process is involved in the photochemical reaction. The results have been interpreted in terms of a new model of the System II photochemical centers. The main characteristics of this model are: (1) Each photochemical center includes two electron donors (Z) and one electron acceptor (Q). (2) The formation of one atom of oxygen requires a two quantum process corresponding to the transfer of two electrons from the same electron donor (first memory). (3) The photochemical center acts as a switch which connects alternately each donor to the acceptor (second memory). The switch process occurs after each photoact with an efficiency of about 85 per cent. Other arguments in favor of this model are obtained from studies of the rate of oxygen production at the onset of a weak illumination.     

SPECTROSCOPIE DE LA CHLOROPHYLLE DANS LES CHLOROPLASTES ENTIERS ET DES FRAGMENTS CHLOROPLASTIQUES

Abstract— Chloroplast fragments obtained after treatment with a non-ionic detergent (Triton X-100) are studied spectroscopically. Their absorption and fluorescence spectra show the possibility of obtaining separately, without any visible alteration, the two groups of chlorophyll holochromes corresponding to the two photochemical systems. One of the chloroplastic fractions, enriched in chlorophyll- b shows, like whole chloroplasts, an oxygen burst upon illumination.     

ISOLEMENT ET ACTIVITÉ DE PARTICULES CHLOROPLASTIQUES CORRESPONDANT AUX DEUX SYSTÈMES PHOTOCHIMIQUES

Abstract— Fractionation of the photosynthetic apparatus was performed in detergent-treated chloroplasts. The study in each fraction of: 1–light-induced oxygen exchanges and fluorescence transients, and 2–the pigment content, shows that a separation of the two photochemical systems was obtained. A possible correlation between the modifications of the transient kinetics of system II particles and their isolation is discussed.     

SHORT-TERM ADAPTATION OF HIGHER PLANTS TO CHANGING LIGHT INTENSITIES AND EVIDENCE FOR THE INVOLVEMENT OF PHOSPHORYLATION OF THE LIGHT HARVESTING CHLOROPHYLL alb PROTEIN COMPLEX OF PHOTOSYSTEM II

Abstract The short-term adaptation of intact leaves to an increase in light intensity was studied by an analysis of chlorophyll fluorescence and oxygen evolution monitored by photoacoustics. An increase in light intensity led to an oxygen “gush”. This “gush” was followed by a large (up to 120%) biphasic increase in the yield of oxygen evolution characterized by a fast phase (T = 0.5–2 min) and a slow phase (T = 4–20 min). The fast phase of the increase in oxygen yield was coupled to a decrease of fluorescence, whereas the slow phase was accompanied by a parallel fluorescence increase. A comparison of fluorescence parameters with oxygen yield indicates that the slow phase of the increase in oxygen yield was coupled to an increase in the antenna size of photosystem II. The slow phase was not inhibited by the uncoupler Nigericin but it was absent in chlorophyll-b-less barley mutants dencient in the light harvesting chlorophyll a/b protein complex of photosystem II (LHC II). These experiments indicate that changes in the LHC II mediated energy distribution, which occur in the time-range of several minutes, are involved in the adaptation to changing light intensities. Moreover, electrophoretic analysis of ³²P orthophosphate labeled leaf discs adapted to low and high light intensities suggests that the slow phase of the increase in oxygen evolution involves dephosphorylation of the 25 kDa polypeptide of LHC II, by a small extent of 12%. The trigger for the slow phase of the increase in oxygen yield does not involve the oxidation of the plastoquinone pool. It was found that in response to the increased light intensity, the plastoquinone pool became more reduced as judged by model calculations. Experiments with the uncoupler Nigericin suggest that the control of the slow phase of adaptation to increased light intensity was also not exerted by the pH gradient across the thylakoid membrane. The similarities between the adaptation to increased light intensity and the state II to state I transition suggest that both adaptation phenomena involve LHC II dephosphorylation possibly triggered by the cytochrome b₆/f complex.     

SHORT-TERM ADAPTATION OF HIGHER PLANTS TO CHANGING LIGHT INTENSITIES AND EVIDENCE FOR THE INVOLVEMENT OF PHOSPHORYLATION OF THE LIGHT HARVESTING CHLOROPHYLL alb PROTEIN COMPLEX OF PHOTOSYSTEM II

Abstract— The short-term adaptation of intact leaves to an increase in light intensity was studied by an analysis of chlorophyll fluorescence and oxygen evolution monitored by photoacoustics. An increase in light intensity led to an oxygen “gush”. This “gush” was followed by a large (up to 120%) biphasic increase in the yield of oxygen evolution characterized by a fast phase (T = 0.5–2 min) and a slow phase (T = 4–20 min). The fast phase of the increase in oxygen yield was coupled to a decrease of fluorescence, whereas the slow phase was accompanied by a parallel fluorescence increase. A comparison of fluorescence parameters with oxygen yield indicates that the slow phase of the increase in oxygen yield was coupled to an increase in the antenna size of photosystem II. The slow phase was not inhibited by the uncoupler Nigericin but it was absent in chlorophyll-b-less barley mutants deñcient in the light harvesting chlorophyll a/b protein complex of photosystem II (LHC II). These experiments indicate that changes in the LHC II mediated energy distribution, which occur in the time-range of several minutes, are involved in the adaptation to changing light intensities. Moreover, electrophoretic analysis of ³²P orthophosphate labeled leaf discs adapted to low and high light intensities suggests that the slow phase of the increase in oxygen evolution involves dephosphorylation of the 25 kDa polypeptide of LHC II, by a small extent of 12%. The trigger for the slow phase of the increase in oxygen yield does not involve the oxidation of the plastoquinone pool. It was found that in response to the increased light intensity, the plastoquinone pool became more reduced as judged by model calculations. Experiments with the uncoupler Nigericin suggest that the control of the slow phase of adaptation to increased light intensity was also not exerted by the pH gradient across the thylakoid membrane. The similarities between the adaptation to increased light intensity and the state II to state I transition suggest that both adaptation phenomena involve LHC II dephosphorylation possibly triggered by the cytochrome b₆/f complex.     

Light-induced changes in cytochrome b-559 in spinach chloroplasts

Abstract— In isolated spinach chloroplasts, the reduction of cytochrome b-559 by System II and its oxidation by System I can be observed when electron acceptor concentration limits System I activity and when one of a particular class of uncoupling agents is present. This class includes CCCP, desaspidin, Triton X-100 and antimycinA; but not simple amines or atebrin. The effect of the uncoupler is to speed the cytochrome b-559 oxidation rate. In addition, the rate of the off-response of the light-induced P518 absorbance change is increased. To explain these findings, an electron transfer scheme is proposed having two distinct pathways between System II and System I.     

STUDIES OF SYSTEM II PHOTOCENTERS BY COMPARATIVE MEASUREMENTS OF LUMINESCENCE,FLUORESCENCE, AND OXYGEN EMISSION*

Abstract— New results are presented on the emission of oxygen by algae and chloroplasts illuminated by a sequence of short saturating flashes. These results favor the four-state hypothesis of Kok and co-workers, in which formation of oxygen requires the accumulation of four oxidants produced by four successive photoreactions. Deactivation of the more oxidized precursor states in the dark is studied under different conditions of preillumination. Our results suggest that both a one step and a two step mechanism of deactivation exist. In order to understand the biological significance of Kok's parameter α—the fraction of photochemical centers unable to react on each flash (“misses”)-we study reoxidation of acceptor Q after one flash by fluorescence techniques. It appears that a fraction of Q^- is reoxidized by a back reaction which cancels the effect of the preilluminating flash and is probably responsible for the misses. The results of some luminescence experiments are also reported. These experiments demonstrate that delayed emission of light is associated with the deactivation of states S₂ and S₃. It is possible that excitons produced by deactivation can be reabsorbed by active photochemical centers, which can modify considerably the deactivation process.     

POLYPHASIC CHLOROPHYLL a FLUORESCENCE TRANSIENT IN PLANTS AND CYANOBACTERIA*

Abstract— The variable chlorophyll (Chl) a fluorescence yield is known to be related to the photochemical activity of photosystem II (PSII) of oxygen-evolving organisms. The kinetics of the fluorescence rise from the minimum yield, F₀, to the maximum yield, F_m, is a monitor of the accumulation of net reduced primary bound plastoquinone (Q_A) with time in all the PSII centers. Using a shutter-less system (Plant Efficiency Analyzer, Hansatech, UK), which allows data accumulation over several orders of magnitude of time (40 μs to 120 s), we have measured on a logarithmic time scale, for the first time, the complete polyphasic fluorescence rise for a variety of oxygenic plants and cyanobacteria at different light intensities. With increasing light intensity, the fluorescence rise is changed from a typical O-I-P characteristic to curves with two intermediate levels J and I, both of which show saturation at high light intensity but different intensity dependence. Under physiological conditions, Chl a fluorescence transients of all the organisms examined follow the sequence of O-J-I-P. The characteristics of the kinetics with respect to light intensity and temperature suggest that the O-J phase is the photochemical phase, leading to the reduction of Q_A to Q_A^-. The intermediate level I is suggested to be related to a heterogeneity in the filling up of the plastoquinone pool. The P is reached when all the plastoquinone (PQ) molecules are reduced to PQH₂. The addition of 3-(3–4-dichlorophenyl)-1,1-dimethylurea leads to a transformation of the O-J-I-P rise into an O-J rise. The kinetics of O-J-I-P observed here was found to be similar to that of O-I₁-I₂-P, reported by Neubauer and Schreiber (Z. Naturforsch. 42c , 1246–1254, 1987). The biochemical significance of the fluorescence steps O-J-I-P with respect to the filling up of the plastoquinone pool by PSII reactions is discussed.     

高浓度LaCl3抑制黄瓜(Cucumis sativus Linn)光系统Ⅱ(PS Ⅱ)活性

研究了高浓度LaCl3对黄瓜（Cucumis sativus Linn）光系统Ⅱ（PSⅡ）光诱导荧光动力学参数、低温荧光光谱和放氧活性的影响。结果表明，随着黄瓜体内LaCl3浓度的升高、其荧光量子产率、PSⅡ最大光化学效率、放氧活性和电子传递速率都明显降低。低温荧光分析表明，低浓度LaCl3引起激发能更多的分配给PSⅡ。高浓度LaCl3对黄瓜幼茁的抑制作用表现在对类囊体膜结构的破坏。进而导致PSⅡ光合活性下降，并最终抑制黄瓜生长。     

CORRELATION BETWEEN DELAYED LIGHT EMISSION AND FLUORESCENCE OF CHLOROPHYLL a IN SYSTEM II PARTICLES DERIVED FROM SPINACH CHLOROPLASTS

Abstract— The induction transient of delayed light of chlorophyll a, excited by repetitive flashes (0.5 ms in duration) and emitted 0.1 - 1.2 ms after the flashes, was measured in system II particles derived from spinach chloroplasts. An uncoupler, gramicidin S, was always added to the particles in order to eliminate the influence of the phosphorylation system on the delayed light and to isolate a direct relationship between the delayed light emission and the primary photochemical reaction, except for the experiments described in the next paragraph. The yield of delayed light emission from the system II particles was found to be about three–times higher than that of chloroplasts on a chlorophyll content basis. System I particles, on the other hand, emitted much weaker delayed light than chloroplasts. Upon intermittent illumination, induction of delayed light in system II particles showed a decrease from the initial rise level to the steady-state level. The initial rise level was the maximum. The fluorescence induction, on the other hand, exhibited an increase from the initial rise level to the maximum steady-state level. The induction of both delayed light emission and fluorescence arrived at their final steady-state levels after the same period of illumination. Induction of delayed light emission was measured under various conditions that changed the oxidation-reduction state of the primary electron acceptor, X, of photoreaction II: by adding an electron acceptor and an inhibitor of electron transport, and by changing the light intensity. The state of A'was monitored by measuring the fluorescence yield. The yield of delayed light emission excited by each flash was found to depend on the amount of oxidized form of X present before the flash. To examine the role of the primary electron donor Y of photoreaction II in delayed light emission, effects of electron donors of photoreaction II such as Mn²⁺, hydroquinone and p-phenylenediamine were investigated. These agents were found to markedly decrease the yield of delayed light emission without altering the pattern of its induction. They had little effect on the induction of fluorescence. These findings are interpreted by a mechanism in which transformation of the reaction center from the form (X^-Y⁺) into (X Y) produces a singlet excitation of chlorophyll a that is the source of millisecond delayed light emission. This reaction is probably non–physiological and must be very slow if compared to the transformation of (X^-Y⁺) into (X^-Y). Since the form (X^-Y⁺) is produced when the excitation is transferred to the reaction center in the form (XY), it is expected in this scheme that the yield of delayed light emission should depend on the amount of the form (X Y) present before the excitation flashes. Electron donors stimulate transformation of the reaction center from (X^-Y⁺) into (X^-Y). Since this reaction competes with the process of delayed light emission, electron donors are expected to suppress delayed light emission.     

Towards tuning PDT relevant photosensitizer properties: comparative study for the free and Zn2+ coordinated meso-tetrakis[2,6-difluoro-5-(N-methylsulfamylo)phenyl]porphyrin

The spectroscopic, photochemical, and biological studies of 5,10,15,20-tetrakis[2,6-difluoro-5(N-methylsulfamylo)phenyl]porphyrinate Zn(II) (ZnF₂PMet) were carried out including absorption and fluorescence spectra, fluorescence quantum yields, triplet absorption spectra, triplet lifetimes, singlet oxygen quantum yield, and reactive oxygen species (ROS) detection under biological conditions and compared with its free-base analog (F₂PMet). Zinc coordination into the porphyrin ring results in decrease of hydrophobicity and in higher cellular uptake. F₂PMet localized specifically in endoplasmic reticulum and mitochondria while the ZnF₂PMet is more diffused all over the cell, bonded to membrane proteins, as assessed by fluorescence microscopy. Zn-porphyrin exhibits greater singlet oxygen quantum yield than its free-base analog. Studies with fluorescent probes confirm that the ZnF₂PMet produces mostly singlet oxygen, whereas F₂PMet generates more hydroxyl radicals as the ROS. F₂PMet is a more effective photosensitizer in vitro than its zinc complex, thus, the final photodynamic effect depends more on the nature of ROS than on the higher cellular uptake.     

RELATIONS BETWEEN PHOTOCHEMISTRY AND FLUORESCENCE IN CELLS AND EXTRACTS OF PHOTOSYNTHETIC BACTERIA*

Abstract— Light-induced changes in the yield of bacteriochlorophyll fluorescence have been measured in cells and chromatophores of photosynthetic bacteria, and coordinated with light-induced absorbancy changes. Comparisons were drawn during transitions between dark and light steady states and also between steady states established at different light intensities. Aerobic cell suspensions of Rhodospirillum rubrum, Rhodopseudomonas spheroides, Chromatium and Rhodopseudomonas sp. NHTC 133 showed a strict correspondence between changes in the fluorescence yield and the bleaching of P870 (P985 in Rps. sp. NHTC 133), as reported by Vredenberg and Duysens for R. rubrum cells. The relationship shows that singlet excitation energy in bacteriochlorophyll is quenched by P870 at a rate proportional to the concentration of unbleached P870. This implies that the photosynthetic units are not independent with respect to energy transfer. In anaerobic cell suspensions the change in fluorescence did not follow the bleaching of P870 in the manner described by Vredenberg and Duysens. Here a change in fluorescence may have resulted from the reduction of a primary photochemical electron acceptor as well as from the oxidation (bleaching) of P870. In chromatophore preparations there were further deviations from the Vredenberg and Duysens relationship which could be attributed to changes in the rate constants for quenching of singlet excitation energy. Finally there was a light-induced increase in the fluorescence yield which was related to a band shift of bacteriochlorophyll and not to the bleaching of P870. Aerobic cell suspensions presented a limiting case in which these complications were absent. No change in the fluorescence was associated uniquely with the oxidation of cytochrome or band shifts of carotenoid pigments. These results, when coordinated with earlier findings about the fluorescence of bacteriochlorophyll and P870, indicate that the singlet excitation quantum is the only energy carrier linking the absorption of light with the initiation of photochemistry in bacterial photosynthesis.     

PHOTOCHEMISTRY OF THE REACTION CENTERS OF SYSTEM II UNDER REPETITIVE FLASH GROUP EXCITATION IN ISOLATED CHLOROPLASTS

Abstract— Absorption changes induced in isolated chloroplasts by excitation with repetitive flash groups have been measured at 690 nm, indicating the photochemical turnover of chlorophyll-a_II (Chl-αn), and at 480 nm and 513 nm respectively, reflecting via electrochromic effect the formation of a transmembrane electric field. The data are compared with measurements of oxygen evolution. In chloroplasts with practically fully intact oxygen evolving capacity it was found: 1. The initial amplitude of the 690 nm absorption change induced by the second flash as a function of the time t_v between the first and second flash of a group increases with a half life of about 35 µs. On the other hand, the average oxygen yield due to the second flash as a function of the time t_v rises with a half life of about 600 µs (and a kinetics in the ms-range of minor extent), confirming the data of Vater et al. (1968). 2. Under far red background illumination, where contributions due to PS I in the µs-range can be excluded, the difference spectrum in the red of the absorption changes induced by the first flash corresponds with that of the absorption changes induced by the second flash fired 200 µ after the first flash. 3. The pattern of the absorption changes at 690 nm induced by repetitive double flash groups at t_v= 200 µs does not markedly change in normal chloroplasts by the presence of DBMIB?. Similar 690 nm absorption changes occur in trypsin treated chloroplasts, independent of the presence of DCMU. 4. The fast regeneration in the µs-range of Chl-a_n is also observed in the third flash of a triple flash group at a time t_v= 200 µs between the flashes of the group. 5. The initial amplitudes of the absorption changes with a decay kinetics slower than 100 µs induced by the second flash at 480 nm and 513 nm, respectively, as a function of the time t, between the first and second flash of a group, are characterized by a recovery half-time of about 600 µs, confirming earlier measurements at 520 nm (Witt and Zickler, 1974). On the basis of these results it is inferred that there does exist a photoreaction of Chl-α_n., with an electron acceptor, referred to as X_a, other than the ‘primary’ plastoquinone acceptor X320, if X320 persists in its reduced state. Under conditions of X320 being in the reduced state, this photochemical reaction was shown to be highly dissipative with respect to charging up the watersplitting enzyme system Y. Furthermore, this Chl-a_n-photoreaction with X_a does not lead to a vectorial transmembrane charge separation, which is stable for more than a few microseconds. Different models for the functional and structural organization of PS II are discussed.     

LIGHT-INDUCED FORMATION OF O-2˙ OXYGEN RADICALS IN SYSTEMS CONTAINING CHLOROPHYLL

It has been shown recently that photosystem 1 particles, photosystem 1 lipid vesicles and chlorophyll-a lipid vesicles show identical photochemical reactions in the presence of oxygen e.g. H⁺-and O₂-uptake (Van Ginkel, 1979). Therefore, spin-trapping experiments were done to identify the oxygen radicals formed. The spintrap phenyltertiarybutylnitrone (PBN) failed to yield information about oxygen radicals. With the spintrap 5,5-dimethyl-1-pyrroline-1-oxide (DMPO), however, we obtained a mixed spectrum of O^-_2˙ and OH·-adducts generated in chloroplasts, photosystem 1 particles or chlorophyll-a lipid vesicles. These data indicate that chlorophyll-a in an artificial membrane can also catalyze O^-_2˙-formation. Chlorophyll-a lipid vesicles catalyze light-induced formation of the Tiron-semiquinone free radical, which has been proposed as a specific O^-_2˙-probe (Greenstock and Miller, 1975). However, OH· scavengers strongly reduce the formation of this radical, whereas superoxide dismutase does not. Pulse-radiolysis measurements showed that the rate constant for the reaction of Tiron with OH· is 8.2 · 10⁹M^-1 s^-1, which is considerably higher than the published Tiron/O^-_2˙ rate constants. Therefore, Tiron is a better spin probe for OH· than for O^-_2˙. We suggest that light-induced H⁺-and O^-_2˙-uptake in membranes containing chlorophyll-a in the presence of ascorbate is caused mainly by the very rapid reaction of OH· with ascorbate.     

Kinetic Analysis of the Light-induced Fluorescence Quenching in Light-harvesting Chlorophyll a/b Pigment-Protein Complex of Photosystem II

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 COMPLEXITY OF THE FLUORESCENCE OF CHLORELLA PYRENOIDOSA

Abstract— The complexity of the room-temperature emission spectrum of Chlorella was investigated by a matrix analysis method. This approach revealed the presence of two independently fluorescent components in the short-wave region of the spectrum. These components, maximal at about 687 and 695 nm, appeared to correspond to the fluorescence of the bulk pigments of PS II and PS I respectively. The analysis was insensitive to the individual species within the photosystems. As such, other minor fluorescent species, usually observed at low temperatures, which presumably correspond to fluorescence from the trapping centres, did not appreciably complicate the analysis. The absorption spectra of the two photosystems were calculated from the fluorescence data. The results were similar to those that have been obtained by other workers from oxygen evolution and DCMU poisoning data but differed from those obtained by computer analysis of the absorption spectrum. Addition of reduced DCPIP was observed to reverse the increase in fluorescence yield and changes in the spectral distribution of emission taking place on poisoning the algae. The correlation between this and the catalysis of photophos-phorylation in aged or poisoned chloroplasts was noted. This correlation was tentatively interpreted as evidence for a direct interaction between the donor system and the photochemical apparatus associated with PS II, rather than with a member of the electron transport chain as is normally assumed.     

RUTHENIUM RED INHIBITION OF OXYGEN EVOLUTION AND SPECIFIC RELEASE OF THE EXTRINSIC 16 kDa POLYPEPTIDE IN A PHOTOSYSTEM II PREPARATION

The inhibitory effect of the dye ruthenium red was studied in photosystem II-enriched submembrane fractions. A number of distinct types of interaction were found, which differed in their concentration range and required incubation time. Ruthenium red instantaneously quenches the initial chlorophyll a fluorescence level (F₀) and the maximum fluorescence level (F_m) by enhancing radiationless deactivation in the chlorophyll light harvesting complex. Associated with this quenching of fluorescence is an instantaneous decrease in the quantum yield of oxygen evolution. Ruthenium red also inhibited the light saturated rate of oxygen evolution and the variable fluorescence, monitored 80 µs after a saturating excitation-flash. These inhibitions increased with incubation time and became greater than 50% within 5 min. Although ruthenium red was known to affect Ca²⁺ or Cl^? sites specifically, the inhibitory action was more pronounced than simple Ca²⁺ or Cl^? depletion. Incubation with ruthenium red for 5 min blocks the Z P680⁺ → Z⁺ P680 charge transfer reaction. Upon mixing with the photosystem II preparation, ruthenium red induced specific release of the extrinsic 16 kDa polypeptide associated with water-splitting without release of Mn. It is proposed that the inhibitor produces an ionic imbalance which alters the configuration of the donor side of photosystem II.     

Ultrafast spectroscopy studies on the mechanism of electron transfer and energy conversion in the isolated pseudo ginseng, water hyacinth and spinach chloroplasts

The spectroscopy characteristics and the fluorescence lifetime for the chloroplasts isolated from the pseudo ginseng, water hyacinth and spinach plant leaves have been studied by absorption spectra, low temperature steady-state fluorescence spectroscopy and single photon counting measurement under the same conditions and by the same methods. The similarity of the absorption spectra for the chloroplasts at room temperature suggests that different plants can efficiently absorb light of the same wavelength. The fluorescence decays in PS II measured at the natural QA state for the chloroplasts have been fitted by a three-exponential kinetic model. The three fluorescence lifetimes are 30, 274 and 805 ps for the pseudo ginseng chloroplast; 138, 521 and 1494 ps for the water hyacinth chloroplast; 197, 465 and 1459 ps for the spinach chloroplast, respectively. The slow lifetime fluorescence component is assigned to a collection of associated light harvesting Chl a/b proteins, the fast lifetime component to the react     

Kinetics of chloroplast-mediated photoxidation of diketogluonate

Abstract— Illuminated chloroplasts can mediate a photoxidation of diketogulonic acid (DKGA) with rates of oxygen uptake equivalent to rates of Hill reactions with ferricyanide or quinone. The photoxidation of DKGA is sensitive to dichlorophenyl dimethylurea (DCMU) and exhibits the drop in quantum yield at long wavelengths characteristic of photosystem II. Still, the reaction is only partially inactivated by heating chloroplasts at 50° for 10 min (which destroys oxygen evolution). The photoxidation is inhibited by copper and detergents; and is stimulated by added flavin (or methyl viologen) and manganous ions. A model system containing Mn³⁺ (as manganipyrophosphate) and DKGA, mimics the chloroplast system. Pre-illuminated chloroplast suspensions can be substituted for Mn³⁺ in the model dark reaction. It seems that a light-dependent oxidation of Mn²⁺ to Mn³⁺ by photosystem II is the essential contribution of the chloroplasts. Electrons from Mn²⁺ move through the electron transport system to ferricyanide or to photosystem I where, via flavin (or methyl viologen), oxygen is reduced to H₂O₂.     

ON THE ORIGIN OF GLOW PEAKS IN EUGLENA CELLS, SPINACH CHLOROPLASTS AND SUBCHLOROPLAST FRAGMENTS ENRICHED IN SYSTEM I OR II

Abstract— The origin of glow peaks (thermoluminescence) was investigated in isolated spinach chloroplasts and Euglena cells by pretreatment with various concentrations of 3-(3,4 dichlorophenyl)-1,1-dimethylurea (DCMU)?, different light intensities, and after mild heating at various temperatures. Experiments are also reported on subchloroplast fractions enriched in pigment systems I (PSI) or II (PSII) (prepared under conditions to reduce destruction of membranes by excessive detergent contact). These results provide the following, most likely, suggestion for the origin of glow peaks: (1) Z peak originates in metastable states; it is insensitive to DCMU, temperature (320–328 K), and appears only when other peaks are saturated (10 Wm^-2). (2) Peak I involves the use of a reducing entity A (plastoquinone) beyond Q (the primary electron acceptor of pigment system II, PSII), or, of a high “S” state (charge accumulator) of oxygen evolving system; its intensity is dramatically reduced by low concentrations (1 μM) of DCMU, and, there is more of it in PSII than in PSI particles. (3) Peak II is due to reaction of Q^- with the “S” states of the oxygen evolving system; its intensity increases upon the addition of low concentrations of DCMU, at the expense of peak I; it is most sensitive to mild heating, and there is more of it in PSII than in PSI particles. (4) Peak III was not studied here as it was not resolved in most of our preparations. (5) Peak IV is from both pigment system I and II; it is sensitive to heating (>50°C), is somewhat sensitive to DCMU, and is present in both PSI and PSII particles. (6) Peak V is from PSI; it is least sensitive to mild heating, and it is enriched in PSI particles. The present studies have extended our knowledge regarding the origin of glow peaks in spinach chloroplasts and Euglena cells; in particular, the involvement of the charge accumulating “S” states of oxygen evolution (for peaks I and II) and of system I (for peak V) are emphasized in this paper.