Biological fluxes conversion and SXRF experiment with a new active biomonitoring tool for atmospheric metals and trace element deposition.

A new active biomonitoring tool, keeping alive mosses for 2 months, had demonstrated the buffering action of the water presence on the biological activity of three moss species (chlorophyll fluorescence induction method on Pleurozium schreberi, Scleropodium purum, Eurhynchium praelongum). The two more resistant mosses were exposed on four different sites with parallel exposure of monthly bulk collectors during three successive periods of 2 months. The coarse and sedimentable particles of bulk collectors were separated into different size classes (> 1 mm; 1000-200 microm; 200-40 microm; 40-20 microm). Dry depositions and mosses were analysed by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) for a stock (microg g(-1)- fluxes (microg m(-2) t(-1)) conversion. The "moss plate" allowed relative site (Ba, Cs, Sb, Sr, U) and species (Pb, Sb, Ti, V) comparisons. Two months were enough for a significant enrichment (Ba, Cd, Pb, Sb, Ti, U, V). Good explicative models were elaborated for Ba, Pb, Sb and Ti (r2 > 70%) including categorical (sites, moss species) and quantitative variables (fluxes of particles size classes). Entire and intact shoots of Scleropodium purum were analysed by detected synchrotron radiation induced X-ray fluorescence (SXRF). The in vivo distribution of the multi-elemental short term enrichment along the moss stem was mainly localized in the plant older parts (Pb, Ti, Cl, Se).

Chlorophyll high-temperature thermoluminescence emission as an indicator of oxidative stress: perturbating effects of oxygen and leaf water content.

A high-temperature chlorophyll thermoluminescence emission can be observed in plant leaves, without preillumination, following various oxidative stresses. This emission was recorded from 0 degrees C to 160 degrees C on a leaf sample pressed by a teflon ring on a heating plate. A band at 140 degrees C was observed in senescing tree leaves, a band at 130 degrees C with a shoulder at 75 degrees C in tobacco leaves treated with the fungal elicitor cryptogein. Measurements in pure O2 or N2 atmospheres did not affect the 130/140 degrees C band, although oxygen increased the thermoluminescence emission at higher temperatures and should be consequently eliminated during measurements. Preventing desiccation by covering the sample with a glass window suppressed the 130/140 degrees C band, except for its rising edge at about 80/90 degrees C. This broad 130/140 degrees C band results from the chlorophyll-exciting thermolysis of lipid peroxides, but it disappears when water is kept in the sample up to 100 degrees C, due to a non-radiative hydrolysis of the peroxides. Therefore, high-temperature thermoluminescence measurements should be performed on leaf samples allowed to dry under an oxygen-free atmosphere.

Heat stress induces in leaves an increase of the minimum level of chlorophyll fluorescence, Fo: A time-resolved analysis.

A time-resolved study of the effects of heat stress (23 to 50°C) on Fo level of chlorophyll fluorescence of leaves having different antenna content has been performed in order to elucidate the causes of heat induced increase of Fo in vivo. The multi-exponential deconvolution of the decays after a picosecond flash at Fo have shown that the best fit in both wild-type and the mutant chlorina F2 of barley leaves is obtained with three components in the temperature range utilized (100, 400 and 1200 ps at 23°C). In intermittent light greened pea leaves, a fourth long lifetime component (4 ns at 23°C) is needed. The comparison of the three types of leaves at 23°C shows that the content of the LHCII b complex does not affect the lifetimes of the two main components (100 and 400 ps) and affects their preexponential factors. This result suggests that in the PS II unit the exciton transfer from LHC IIb to the rest of the antenna is irreversible. The effects of heat stress on individual lifetime components, Ti, included several changes. Utilizing for PS II unit an extended 'Reversible Radical Pair' model, having three compartments, to interpret the variations of Ti and Ai induced by temperature increases, it can be inferred that heat determines: (i) an irreversible disconnection of a monor antenna complex which is not the LHC IIb complex, this effect is induced by temperatures higher than 40°C; (ii) a decrease of the quantum efficiency of Photosystem II photochemistry which is due to several effects: a decrease of the rate of charge separation, an increase of P(+)I(-) recombination rate constant and a decrease of the stabilization of charges. These effects on Photosystem II photochemistry start to occur above 30°C and are partially reversible.

A Mutation in the D-de Loop of D1 Modifies the Stability of the S2QA- and S2QB- States in Photosystem II.

Photosystem II electron transfer, charge stabilization, and photoinhibition were studied in three site-specific mutants of the D1 polypeptide of Synechocystis PCC 6803: E243K, E229D, and CA1 (deletion of three glutamates 242-244 and a substitution, glutamine-241 to histidine). The phenotypes of the E229D and E243K mutants were similar to that of the control strain (AR) in all of the studied aspects. The characteristics of CA1 were very different. Formate, which inhibits the QA- to QB- reaction, was severalfold less effective in CA1 than in AR. The S2QA- and S2QB- states were stabilized in CA1. It was previously shown that the electron transfer between QA- and QB was modified in CA1 (P Maenpaa, T. Kallio, P. Mulo, G. Salih, E.-M. Aro, E. Tyystjarvi, C. Jansson [1993] Plant Mol Biol 22: 1-12). A change in the redox potential of the QA/QA- couple, which renders the reoxidation of QA- by back or forward reactions more difficult, could explain the phenotype of CA1. Although the rates of photoinhibition measured as inhibition of oxygen evolution, Chl fluorescence quenching, and decrease of thermoluminescence B and Q bands were similar in AR and CA1, the CA1 strain more quickly reached a state from which the cells were unable to recover their activity. The results described in this paper suggest that a modification in the structure of the D-de loop of D1 could influence the properties of the couple QA/QA- in D2 and the mechanism of recovery from photoinhibition.

Effects of dark- and light-induced proton gradients in thylakoids on the Q and B thermoluminescence bands.

Thermoluminescence experiments have been carried out to study the effect of a transmembrane proton gradient on the recombination properties of the S2 and S3 states of the oxygen evolving complex with QA (-) and QB (-), the reduced electron acceptors of Photosystem II. We first determined the properties of the S2QA (-) (Q band), S2QB (-) and S3QB (-) (B bands) recombinations in the pH range 5.5 to 9.0, using uncoupled thylakoids. The, a proton gradient was created in the dark, using the ATP-hydrolase function of ATPases, in coupled unfrozen thylakoids. A shift towards low temperature of both Q and B bands was observed to increase with the magnitude of the proton gradient measured by the fluorescence quenching of 9-aminoacridine. This downshift was larger for S3QB (-) than for S2QB (-) and it was suppressed by nigericin, but not by valinomycin. Similar results were obtained when a proton gradient was formed by photosystem I photochemistry. When Photosystem II electron transfer was induced by a flash sequence, the reduction of the plastoquinone pool also contributed to the downshift in the absence of an electron acceptor. In leaves submitted to a flash sequence above 0°C, a downshift was also observed, which was supressed by nigericin infiltration. Thus, thermoluminescence provides direct evidence on the enhancing effect of lumen acidification on the S3→S2 and S2→S1 reverse-transitions. Both reduction of the plastoquinone pool and lumen acidification induce a shift of the Q and B bands to lower temperature, with a predominance of lumen acidification in non-freezing, moderate light conditions.

Oxygen-evolving photosystem II preparation from wild type and photosystem II mutants of Synechocystis sp. PCC 6803.

We present here a simple and rapid method which allows relatively large quantities of oxygen-evolving photosystem II- (PS-II-) enriched particles to be obtained from wild-type and mutants of the cyanobacterium Synechocystis 6803. This method is based on that of Burnap et al. [Burnap, R., Koike, H., Sotiropoulou, G., Sherman, L. A., & Inoue, Y. (1989) Photosynth. Res. 22, 123-130] but is modified so that the whole preparation, from cells to PS-II particles, is achieved in 10 h and involves only one purification step. The purified preparation exhibits a 5-6-fold increase of O2-evolution activity on a chlorophyll basis over the thylakoids. The ratio of PS-I to PS-II is about 0.14:1 in the preparation. The secondary quinone electron acceptor, QB, is present in this preparation as demonstrated by thermoluminescence studies. These PS-II particles are well-suited to spectroscopic studies as demonstrated by the range of EPR signals arising from components of PS-II that are easily detectable. Among the EPR signals presented are those from a formal S3-state, attributed to an oxidized amino acid interacting magnetically with the Mn complex in Ca(2+)-deficient PS-II particles, and from S2 modified by the replacement of Ca2+ by Sr2+. Neither of these signals has been previously reported in cyanobacteria. Their detection under these conditions indicates a similar lesion caused by Ca2+ depletion in both plants and cyanobacteria. The protocol has also been applied to mutants which have site-specific changes in PS-II. Data are presented on mutants having changes on the electron donor (Y160F) and electron acceptor (G215W) side of the D2 polypeptide.

The effects of low temperature acclimation and photoinhibitory treatments on Photosystem 2 studied by thermoluminescence and fluorescence decay kinetics.

The effects of low temperature acclimation and photoinhibitory treatment on Photosystem 2 (PS 2) have been studied by thermoluminescence and chlorophyll fluorescence decay kinetics after a single turnover saturating flash. A comparison of unhardened and hardened leaves showed that, in the hardened case, a decrease in overall and B-band thermoluminescence emissions occurred, indicating the presence of fewer active PS 2 reaction centers. A modification in the form of the B-band emission was also observed and is attributed to a decrease in the apparent activation energy of recombination in the hardened leaves. The acclimated leaves also produced slower QA (-) reoxidation kinetics as judged from the chlorophyll fluorescence decay kinetics. This change was mainly seen in an increased lifetime of the slow reoxidation component with only a small increase in its amplitude. Similar changes in both thermoluminescence and fluorescence decay kinetics were observed when unhardened leaves were given a high light photoinhibitory treatment at 4°C, whereas the hardened leaves were affected to a much lesser extent by a similar treatment. These results suggest that the acclimated plants undergo photoinhibition at 4°C even at low light intensities and that a subsequent high light treatment produces only a small additive photoinhibitory effect. Furthermore, it can be seen that photoinhibition eventually gives rise to PS 2 reaction centers which are no longer functional and which do not produce thermoluminescence or variable chlorophyll fluorescence.

Graphical and numerical analysis of thermoluminescence and fluorescence F0 emission in photosynthetic material.

A set-up for recording thermoluminescence emission together with the constant F0 fluorescence yield is described briefly. It is driven by a microcomputer through plugged-in cards.Practical aspects of the simulation of TL bands and of decomposition of complex TL signals are examined. A reproducible and linear temperature gradient and the use of photon counting for luminescence detection are important features for further analyzing the recorded signal. The simulation procedure used is a step-by-step calculation of the number of charge recombinations, which is then substracted from the number of remaining charge pairs able to produce luminescence. This procedure consists first of a graphical fitting, followed by a numerical minimization, with a maximum of five simulated components. The quality of the simulation is evaluated by the sum of squares of differences (signal-simulation), related to the signal area. Equivalent decomposition patterns may be found for the same recording and additional information is needed for interpretation of TL data. Averaging signals is feasible, provided that maximum temperatures Tm of averaged bands are sufficiently similar (±3°C). Simultaneous measurement of the antenna fluorescence yield F0, using an ultra-weak pulsed blue LED, gives an estimate of the luminescence yield. This has to be taken into account in the analysis of the Q band and of high temperature (>40°C) bands.The simulation parameters appear to be dependent on plant growth conditions. Quantitative analysis of thermoluminescence emission could be useful in the study of the effects of climatic factors on the photosynthetic apparatus in plants.

[Influence of linoleic acid (18:2 n-6) and alpha-linolenic acid (18:3 n-3) on the composition, permeability and fluidity of cardiac phospholipids in the rat: study using membrane models (liposomes)]. / Influence des acides linoléique (18:2 n-6) et alpha-linolénique (18:3 n-3) sur la composition, la perméabilité et la fluidité des phospholipides cardiaques du rat: étude à l'aide de modèles membranaires (liposomes).

For 9 weeks, 80 male SPF Wistar rats were fed purified diets containing mixtures of vegetable oils (15% by weight) with different linoleic and linolenic contents. Diet (L+) contained large amounts of linoleic acid (53% of the total fatty acids), diet (L+Ln) contained the same amount of linoleic acid but also 10% of alpha-linolenic acid, diet (L-) supplied a low level of linoleic acid (12% of the total fatty acids) and so did diet (L-Ln) which also contained 10% of alpha-linolenic acid. The levels and fatty acid composition of heart phospholipids were determined. Liposomes, prepared from the total phospholipids extracted from rat hearts, were tested at different temperatures (15 to 50 degrees C) for their permeability to urea and fluidity; fluidity was monitored by fluorescence depolarization of 1,6-diphenyl-1,3,5-hexatriene (DPH). As already demonstrated, dietary alpha-linolenic acid substituted C22 polyunsaturated fatty acids of the linoleic family (n-6) for those of the linolenic acid family (n-3). This substitution remained high, even though linoleic acid represented more than 50% of the total dietary fatty acids. Diphosphatidylglycerol markedly increased in heart phospholipids of rats fed diets (L+) and (L+Ln). The changes in liposome permeability observed were rather well correlated with the unsaturation index of the phospholipid fatty acids, whereas fluidity changes were not. Fluidity decreased in liposomes of rats fed high levels of linoleic acid. Factors such as diphosphatidylglycerol or (n-3) fatty acid concentration in heart phospholipids could explain these results.