P.S. Venediktov

Membrane lipid peroxidation, cell viability and Photosystem II activity in the green alga Chlorella pyrenoidosa subjected to various stress conditions

Abstract The unicellular green alga Chlorella pyrenoidosa was subjected to a variety of stress conditions (strong illumination, incubation with Cu 1+ or Zn 2+ , exposure to high temperatures). The amplitude of thermoluminescence (TL) peak at 125°C, accumulation of thiobarbituric acid reactive substances (TBARS), which indicate an accumulation of lipid peroxidation products, efficiency of Photosystem II reactions ( F v / F M ratio) and the percentage of viable cells were measured in stressed culture. Exposure of algae to strong (5000 μmol photons m 2 s 1 ) or to low (60 μmol photons m −2 s −1 ) light combined with the addition of 1.6 μM Cu 2+ or 30 μM Zn 2+ inactivated Photosystem II, decreased the viability of Chlorella cells, and, finally, significantly enhanced TL and the accumulation of TBARS, which was accompanied by chlorophyll bleaching. TL emission started to rise after a lag-period of about 30 min in algae subjected to strong illumination, 2–3 h in copper-treated algae, and 10 h in zinc-treated algae. A vast majority of cells were nonviable to the end of the lag-period. The addition of Cu 2+ or ZN 2+ in darkness caused a slight decrase in the F v / F M ratio without significant changes in TL emission. Incubation of algae at 50°C for 10 min did not affect the F v / F M ratio nd cell viability, whereas no viable cells and Photosystem II activity were detected in the culture incubated at 55°C. Heat stress at temperatures above 55°C significantly enhanced the amplitude of the 125°C TL peak and the accumulation of TBARS when the algae were further incubated at low light at room temperature. We conclude that, under the stress conditions used in this study, (i) lipid peroxides and products of their degradation are not responsible for the cytolethal effect in Chlorella and (ii) lipid peroxidation arises mainly upon illumination of dead cells.

Sublethal concentrations of copper stimulate photosystem II photoinhibition in Chlorella pyrenoidosa

Summary The effect of copper (CuSO 4 , 0.16-1.56 μM) on photosystem II (PS II) was investigated by measuring the chlorophyll fluorescence in Chlorella pyrenoidosa cells exposed to different irradiance levels and different temperatures. The algal culture remained viable only if less than 0.7-0.8 μM Cu 2+ was added. In darkness, PS II inactivation was indicated by a decrease in F v /F m ratio and was detected only at Cu 2+ concentrations higher than 0.7 μM. In the light, Cu 2+ stimulated PS II photoinhibition throughout the whole range of copper concentrations. Low temperature significantly enhanced the PS II photoinhibition in the presence of Cu 2+ . Kinetics of PS II inactivation monitored at algal growth light intensity in the presence of chloramphenicol were only slightly affected by the addition of Cu 2+ . However, Cu 2+ markedly slowed down (0.16 μM) or completely suppressed (0.7 μM) the recovery of PS II activity after the photoinhibitory treatment of the algae with a strong light. The results suggest that sublethal concentrations of copper can nonspecifically retard synthesis of the PS II D 1 protein in Chlorella cells , thus causing PS II inactivation under light.

New experimental approach to the estimation of rate of electron transfer from the primary to secondary acceptors in the photosynthetic electron transport chain of purple bacteria.

A method for calculating the rate constant (KA1A2) for the oxidation of the primary electron acceptor (A1) by the secondary one (A2) in the photosynthetic electron transport chain of purple bacteria is proposed. The method is based on the analysis of the dark recovery kinetics of reaction centre bacteriochlorophyll (P) following its oxidation by a short single laser pulse at a high oxidation-reduction potential of the medium. It is shown that in Ectothiorhodospira shaposhnikovii there is little difference in the value of KA1A2 obtained by this method from that measured by the method of Parson ((1969) Biochim, Biophys. Acta 189, 384-396), namely: (4.5 +/- 1.4)-10(3) s-1 and (6.9 +/- 1.2)-10(3) s-1, respectively. The proposed method has also been used for the estimation of the KA1A2 value in chromatophores of Rhodospirillum rubrum deprived of constitutive electron donors which are capable of reducing P+ at a rate exceeding this for the transfer of electron from A1 to A2. The method of Parson cannot be used in this case. The value of KA1A2 has been found to be (2.7 +/- 0.8)-10(3) s-1. The activation energies for the A1 to A2 electron transfer have also been determined. They are 12.4 kcal/mol and 9.9 kcal/mol for E. shaposhnikovii and R. rubrum, respectively.

High-temperature damage and acclimation of the photosynthetic apparatus I. Temperature sensitivity of some photosynthetic parameters of chloroplasts isolated from acclimated and non-acclimated bean leaves

The thermosensitivity of delayed fluorescence, the relative values of variable chlorophyll fluorescence and the degree of quenching of 9-aminoacridine fluorescene were studied in the chloroplasts from heat-acclimated and non-acclimated (treated 6 h at 52,5°C) young bean plants. The temperature sensitivity of each parameter studied was defined by that temperature at which chloroplast activity decreased by 50% (T50) of its maximum value. There was appreciable increase in the thermostability of membrane energization in chloroplasts isolated from acclimated and non-acclimated plants compared with the controls. The photosynthetic parameters differed according to the suspending medium and the preacclimation treatment. When chloroplast were suspended in phosphate buffer with the addition of stabilizing compounds (2 M sucrose or 0.5% human serum albumin) the thermostability of the thylakoid membranes increased, as was evident by the increases in T50 of about 8–10° C (sucrose) and 2–5° C (human serum albumin) for all the parameters investigated. Photoinduced quenching of 9-aminoacridine fluorescence decreased to some extent in the presence of protective compounds, but in chloroplasts from acclimated plants the T50 was practically equal to that for their long-lived luminescence under the same conditions. At the thylakoid membrane level, acclimation was clearly manifested as an increased thermostability of photoinduced proton-gradient formation.

Multiple action sites for photosystem II herbicides as revealed by delayed fluorescence.

DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea) at concentrations higher than 10 μM suppresses the second time range delayed fluorescence (DF) of pea chloroplasts, due to inhibition of the oxidizing side of photosystem II (PS II). The inhibition of the reducing side of PS II resulting in the suppression of millisecond DF takes place at much lower (∼0.01 μM) DCMU concentrations. The variation in the herbicide-affinities of the reducing and oxidizing sides of PS II is not the same for DCMU and phenol-type herbicides. The DCMU-affinity of the oxidizing side considerably increases and approximates that of the reducing side upon mild treatment of chloroplasts with oleic acid. Probably this is a result of some changes in the environment of the binding site at the oxidizing side. At DCMU concentrations higher than 1 mM, the chaotropic action of DCMU leads to the generation of millisecond luminescence which is not related to the functioning of the reaction centres.

Increased toxic effect of methylmercury on Chlorella vulgaris under high light and cold stress conditions

The toxic effect of methylmercury on the photosynthetic activity of Chlorella vulgaris was shown to increase under high illumination and unfavorable low temperature. Increased toxic action of methylmercury resulted from the decreased capacity of photosystem II (PS II) for reparation. It was proposed that mild stress conditions might be used to enhance the detection limit of toxicants by microalgae used as test objects.

The influence of the electric diffusion potential on delayed fluorescence light curves of chloroplasts treated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea.

The potassium salt-induced transient increase of delayed fluorescence yield was studied in pea chloroplasts treated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea. A simple kinetic model is proposed to account for the actinic light intensity dependence of the delayed fluorescence enhancement by the transmembrane diffusion potential induced by sudden salt addition. The electric field dependence of the rate constants for the recombination of primary separated charges with and without subsequent electronic excitation of reaction center chlorophyll was obtained. From the value of enhancement of delayed fluorescence by salt concentration gradients at saturating actinic light intensity, it is concluded that the distance, normal to thylakoid membrane surface, between the primary acceptor and the donor of Photosystem II is smaller than the membrane thickness.

Effects of Deep Dormancy on the Primary Processes of Photosynthesis in Vine (Vitis vinifera) Shoots

Summary The primary processes of photosynthesis in bark phelloderm of one-year-old vine shoots were investigated by means of the delayed fluorescence method. All of the processes investigated (electron transport and oxygen-evolving activity as well as energization of membranes) in the bark phelloderm were shown to take place at a lower level of activity in comparison with the leaves. A further inactivation of the primary photosynthetic processes in the bark phelloderm is observed in the period of deep dormancy whereas after the end of this period a gradual reactivation of the processes takes place. An express method for determination of the beginning and the end of deep dormancy in vine shoots is proposed.

Delayed luminescence yield kinetics in flash illuminated green plants

Abstract Oscillations of a delayed light emission have been studied in Chlorella cells, pea leaves and isolated pea chloroplasts illuminated by series of short saturating flashes. The oscillation is determined by changes of S-state numbers of the O 2 -evolving system and by pH lowering inside the thylakoids after a flash. The rate of deactivation of the S-states drastically decreases upon temperature lowering to + 4°C, and its pH-dependence has a maximum at pH 6. In isolated chloroplasts, transitions between individual S-states are inhibited at some certain temperatures in the range from −17°C to −40°C.

On the interaction of photoactive bacteriochlorophyll with the primary electron acceptor in the reaction centre of Ectothiorhodospira shaposhnikovii

In Ectothiorhodospira shaposhnikovii cells, the half-time of photooxidized bacteriochlorophyll P890+ dark reduction is 60 ms and does not change with increasing actinic illumination period, whereas the half-time of high-potential cytochrome cH+ reduction varies from 80 ms up to 300 ms. Interaction between P890+ and the primary electron acceptor A-1 provides for fast reduction of pigment when cytochrome cH is oxidized or reduced slowly. It is suggested that P890 and A1 form a complex such that A-1 is not able to donate its electron to secondary acceptors until P890+ has accepted an electron from cytochrome.