Changes In Fluorescence Parameters Research Articles

ФОТОСИНТЕТИЧЕСКАЯ АКТИВНОСТЬ И СОДЕРЖАНИЕ ХЛОРОФИЛЛОВ В ПОБЕГАХ МХА HYLOCOMIUM SPLENDENS HEDW. В УСЛОВИЯХ ТЕМПЕРАТУРНОГО СТРЕССА

Mosses are an attractive model for studying the mechanisms of plant adaptation to unfavorable environmental conditions. There is little information in the literature on the pigment composition and its role in stress resistance of mosses, although this topic is quite widely covered for higher vascular plants. Unfavorable environmental temperature is one stress factors with the strongest effect on plants, since photosynthesis, the main energy-supplying producing process in plants, is sensitive to temperature and often inhibited before other cellular functions are impaired. Chlorophyll is known to play a central role in the process of photosynthesis. In this regard, the aim of the present work was to study the response of the moss Hylocomium splendens Hedw. to temperature stress. The chlorophyll content and chlorophyll a fluorescence parameters in H. splendens shoots were investigated. Results showed that both elevated and negative temperatures decreased the content of chlorophyll a significantly, while, on the contrary, the content of chlorophyll b increased. It is interesting to note that the decrease in chlorophyll a under temperature stress was accompanied by an increase in the content of pheophytin a. The content of chlorophylls in the post-stress period did not recover to the control level. Temperature stress did not affect the maximum photochemical efficiency of photosynthesis, but the rate of electron transfer significantly decreased under the influence of low temperature. Thus, significant changes in the pigment composition of chlorophylls are not always accompanied by changes in fluorescence parameters to the same extent.

Combining fluorescence lifetime with spectral information in fluorescence lifetime imaging ophthalmoscopy (FLIO).

Fluorescence lifetime imaging ophthalmoscopy (FLIO) provides information on fluorescence lifetimes in two spectral channels as well as the peak emission wavelength (PEW) of the fluorescence. Here, we combine these measures in an integral three-dimensional lifetime-PEW metric vector and determine a normal range for this vector from measurements in young healthy subjects. While for these control subjects 97 (±8) % (median (interquartile range)) of all para-macular pixels were covered by this normal vector range, it was 67 (±55) % for the elderly healthy, 38 (±43) % for age-related macular degeneration (AMD)-suspect subjects, and only 6 (±4) % for AMD patients. The vectors were significantly different for retinal pigment epithelium (RPE) lesions in AMD patients from that of non-affected tissue (p < 0.001). Lifetime- PEW plots allowed to identify possibly pathologic fundus areas by fluorescence parameters outside a 95% quantile per subject. In a patient follow-up, changes in fluorescence parameters could be traced in the lifetime-PEW metric, showing their change over disease progression.

Hormonal Regulation in Different Varieties of Chenopodium quinoa Willd. Exposed to Short Acute UV-B Irradiation.

Increased ultraviolet-B (UV-B) due to global change can affect plant development and metabolism. Quinoa tolerates extreme conditions including high UV levels. However, the physiological mechanisms behind its abiotic stress tolerance are unclear, especially those related to UV-B. We previously demonstrated that 9.12 kJ m−2 d−1 may induce UV-B-specific signaling while 18.24 kJ m−2 d−1 promotes a UV-B-independent response. Here, we explored the effects of these UV-B doses on hormonal regulation linked to plant morphology and defense among diverse varieties. Changes in fluorescence parameters of photosystem II, flavonoids and hormones (indoleacetic acid (IAA), jasmonic acid (JA), abscisic acid (ABA) and salicylic acid (SA)) were surveyed under controlled conditions. Here, we showed that the sensitivity to short acute UV-B doses in varieties from different habitats is influenced by their parental lines and breeding time. UV-B sensitivity does not necessarily correlate with quinoa’s geographical distribution. The role of flavonoids in the UV-B response seems to be different depending on varieties. Moreover, we found that the extent of changes in JA and SA correlate with UV-B tolerance, while the increase of ABA was mainly related to UV-B stress.

The Functional State of the Photosynthetic Apparatus of Potato Plants upon Destruction of the Tubulin Cytoskeleton

We have investigated the effect of colchicine, a disrupting agent of the tubulin cytoskeleton, on chlorophyll fluorescence parameters, the Hill reaction, noncyclic photophosphorylation, and the lipid peroxidation reaction in Solanum tuberosum grown in soil in a greenhouse environment. The destruction of microtubules caused an increase in the content of lipid fatty acid hydroperoxides, the primary product of lipid peroxidation, and led to changes in fluorescence parameters: F0 increased, Fm and the maximum quantum efficiency of PSII (Fv/Fm) decreased, and the dissipation of electronic excitation energy as heat increased. Simultaneously, it was found that the Hill reaction rate and the intensity of the process of noncyclic photophosphorylation decreased. The results obtained in this work are discussed in view of the destruction of the integrity of the chloroplast membrane and changes in chlorophyll fluorescence parameters due to colchicine application, as well as in light of our previously obtained data on the change in the content of phytohormones and their concentration ratios in potato leaves when microtubules are destroyed.

Effects of heavy metals released from sediment accelerated by artificial sweeteners and humic acid on a green algae Scenedesmus obliquus

Artificial sweeteners (ASs) are emerging polar organic contaminants, which can chelate with heavy metals in the environment. Recently, the ASs have been widely detected in the aquatic environment. However, little is known about the influence of ASs on the mobility and toxicity of heavy metals associated in sediment. In the present study, the release of heavy metals from a lake sediment and the toxicity to a green algae Scenedesmus obliquus were examined in the presence of two most frequently detected ASs, acesulfame and sucralose, and humic acid (HA), respectively. The existence of 1.00–100 mg/L of acesulfame, sucralose, and HA enhanced the release rates (Rrelease) of Cd, Cu, and Pb from the virgin lake sediment significantly by up to 46.7%, 86.0%, and 79.9%, respectively. Further, the results from microcosm experiments revealed that the existence of 1.00–10.0 mg/L of acesulfame, sucralose, and HA in the water phase prompted the release and toxicity of heavy metals associated with Cd- and Cu-spiked sediment, and the promotion capacity followed the order of HA > acesulfame > sucralose. The inhibitions on cell-specific growth rate (μ) of the algae were found increasing significantly with the elevated levels of acesulfame and HA (p < 0.05). Additionally, photosynthesis of the algae was affected by the co-existence of ASs or HA and heavy metals released from sediments, as indicated by the changes in fluorescence parameters. The present study provides valuable insight into the likelihood of the combined effect of ASs and heavy metals on aquatic organisms.

Serum Autofluorescence and Biochemical Markers in Athlete's Response to Strength Effort in Normobaric Hypoxia: A Preliminary Study.

The human organism has the ability to adapt to hypoxia conditions. Training in hypoxia is used in sport to improve the efficiency of athletes; however, type of training affects the direction and scope of this process. Therefore, in this study, the usefulness of serum fluorescence spectroscopy to study the assessment of athlete's response to strength effort in hypoxia is considered in comparison with biochemical assay. Six resistance-trained male subjects took part in a research experiment. They performed barbell squats in simulated normobaric hypoxic conditions with deficiency of oxygen 11.3%, 13% 14.3% compared to 21% in normoxic conditions. Fluorescence intensity of tyrosine revealed high sensitivity on strength effort whereas tryptophan was more dependent on high altitude. Changes in emission in the visible region are associated with altering cell metabolism dependent on high altitude as well as strength training and endurance training. Significant changes in serum fluorescence intensity with relatively weak modifications in biochemical assay at 3000 m above sea level (ASL) were observed. Training at 5000 m ASL caused changes in fluorescence parameters towards the normobaric specific values, and pronounced decreases of lactate level and kinase creatine activity were observed. Such modifications of fluorescence and biochemical assay indicate increased adaptation of the organism to effort in oxygen-deficient conditions at 5000 m ASL, unlike 3000 m ASL. Fluorescence spectroscopy study of serum accompanied by biochemical assay can contribute to the understanding of metabolic regulation and the physiological response to hypoxia. The results of serum autofluorescence during various concepts of altitude training may be a useful method to analyze individual response to acute and chronic hypoxia. An endogenous tryptophan could be exploited as intrinsic biomarker in autofluorescence studies. However, these issues require further research.

Spatial referencing of chlorophyll fluorescence images for quantitative assessment of infection propagation in leaves demonstrated on the ice plant: Botrytis cinerea pathosystem

BackgroundChlorophyll fluorescence analysis is one of the non-invasive techniques widely used to detect and quantify the stress-induced changes in the photosynthetic apparatus. Quantitative information is obtained as a series of images and the specific fluorescence parameters are evaluated inside the regions of interest outlined separately on each leaf image. As the performance of photosynthesis is highly heterogeneous over a leaf surface, the areas of interest selected for generating numeric data are crucial for a reliable analysis. The differences in intact leaf physio-morphological characters and in the structural effects of stress between leaves increase the risk of artefacts.ResultsThe authors propose a new enhanced method for precise assessment of stress-induced spatiotemporal changes in chlorophyll a fluorescence exemplified in the leaves of common ice plants infected with a fungal pathogen. The chl a fluorescence leaf image series obtained with Imaging-PAM fluorometer are aligned both by affine and nonlinear spline transforms based on the set of control points defined interactively. The successive readings were taken on the same leaf and this image sequence registration allows to capture quantitative changes of fluorescence parameters in time and along selected directions on the leaf surface. The time series fluorescence images of attached leaf, aligned according to the proposed method, provide a specific disease signature for an individual leaf. The results for C3 and Crassulacean Acid Metabolism (CAM) plants have been compared with respect to the type of photosynthetic metabolism and the image alignment accuracy has also been discussed.ConclusionsThe image alignment applied to the series of fluorescence images allows to evaluate the dynamics of biotic stress propagation in individual plant leaves with better accuracy than previous methods. An important use of this method is the ability to map the fluorescence signal horizontally in one leaf during disease development and to accurately compare the results between leaves which differ in morphology or in the structural effects of stress. This approach in analysing chlorophyll fluorescence changes can be used to receive spatial and temporal information over a sample area in leaves infected by different pathogenic fungi and bacteria.

亚热带人工针叶林叶绿素荧光参数日变化及其与植被生产力的关系

PDF HTML阅读 XML下载 导出引用 引用提醒 亚热带人工针叶林叶绿素荧光参数日变化及其与植被生产力的关系 DOI: 10.5846/stxb201806141327 作者: 作者单位: 中国科学院地理科学与资源研究所,中国科学院地理科学与资源研究所,中国科学院地理科学与资源研究所,中国科学院地理科学与资源研究所,中国科学院地理科学与资源研究所,南京大学,南京大学,中国科学院地理科学与资源研究所 作者简介: 通讯作者: 中图分类号: 基金项目: 国家重点研发计划（2017YFC0503803）；国家自然科学基金项目（41571192）；中国科学院重点部署项目（KFZD-SW-310-01） Diurnal changes in chlorophyll fluorescence parameters and their relationships with vegetation productivity in subtropical coniferous plantations Author: Affiliation: Institute of Geographic Sciences and Natural Resources Research,Institute of Geographic Sciences and Natural Resources Research,,,,,, Fund Project: the National Key Research and Development Program of China (2017YFC0503803), the National Natural Science Foundation of China (41571192), the Key Research Program of the Chinese Academy of Science (KFZD-SW-310-01) 摘要 | 图/表 | 访问统计 | 参考文献 | 相似文献 | 引证文献 | 资源附件 | 文章评论 摘要:叶绿素荧光技术能够快速、灵敏、无损地测定植物光合功能对环境变化的响应，已成为当前植物光合能力测定的重要技术手段之一，但是探讨亚热带森林叶绿素荧光参数在不同季节日变化特征受环境因子影响的研究较少，更缺乏与植被生产力关系的研究。以亚热带人工针叶林为研究对象，利用2016年叶绿素荧光测量和涡度相关通量观测数据，分析了不同季节湿地松和马尾松叶片荧光参数实际光化学效率（Photochemical efficiency of PSⅡ，ΦPSⅡ）和非光化学淬灭（Non-photochemical quenching，NPQ）的日变化特征，并结合增强回归树等统计方法，定量化环境因子对其影响，最后探讨了荧光参数与总初级生产力（Gross primary productivity，GPP）和光能利用率（Light use efficiency，LUE）的关系。结果表明，ΦPSⅡ日变化总体上呈现"U型"，早晚高午间低，而NPQ先升后降，午间达到峰值，与ΦPSⅡ的日变化显著负相关。研究发现，光照是影响亚热带人工针叶林ΦPSⅡ日变化最主要的环境因子，ΦPSⅡ随着光合有效辐射（Photosynthetically active radiation，PAR）的升高而降低，PAR对湿地松和马尾松叶片ΦPSⅡ的相对贡献率分别为83.0%和51.1%。PAR对湿地松叶片NPQ日变化的相对贡献率也最高（77.2%），但马尾松的NPQ主要受叶片温度（Leaf temperature，Tleaf）的影响（59.6%）。此外，GPP与ΦPSⅡ显著负相关，而与NPQ有显著的正相关关系。LUE随着ΦPSⅡ增长，但存在ΦPSⅡ高而LUE低的情况，LUE与NPQ显著负相关，热耗散的比例越高，光能利用效率也相应降低。 Abstract:Chlorophyll fluorescence has become a very powerful technique to estimate the photosynthetic capacities of different plants in a rapid and non-invasive way. However, the effect of environmental factors on the diurnal variations in chlorophyll fluorescence parameters in different seasons has been less studied, and their relation to vegetation productivity has hardly been discussed. In this study, we characterized the diurnal changes in fluorescence parameters (photochemical efficiency of PSⅡ and non-photochemical quenching (NPQ)), and quantified the influence of the three environmental factors (photosynthetically active radiation (PAR), leaf temperature (Tleaf), and relative humidity (RH) using Boosted Regression Trees (BRT) and Pearson correlation analysis). In addition, the relationships between the fluorescence parameters and vegetation productivity (gross primary productivity (GPP) and light use efficiency (LUE)) were explored. We conducted in situ chlorophyll fluorescence measurements on Slash Pine (Pinus elliottii Engelm.) and Masson Pine (Pinus massoniana Lamb.) in the Qianyanzhou (QYZ) subtropical coniferous plantation, Jiangxi province, China, during 2016. Our results showed that ΦPSⅡ decreased to a minimum at midday and subsequently progressively rose, whereas NPQ was highest at midday, significantly contrary to ΦPSⅡ. Among the three environmental factors, PAR played the strongest role in determining the diurnal changes of ΦPSⅡ (83.0% in Slash Pine and 51.1% in Masson Pine). ΦPSⅡ decreased with PAR. PAR also had an important influence on NPQ in Slash Pine (77.2%). However, NPQ in Masson Pine was mainly affected by Tleaf (59.6%). In addition, we obtained the synchronous LUE and GPP from the eddy covariance flux observation in QYZ and found that GPP had a significantly negative correlation with ΦPSⅡ, but a positive correlation with NPQ. LUE increased with ΦPSⅡ, but sometimes high ΦPSⅡ was accompanied by low LUE. LUE decreased with NPQ, indicating that higher proportion of heat dissipation resulting in lower light use efficiency. 参考文献 相似文献 引证文献

High Sensitivity NDD Flim and Pattern Matching Based Fluorophore Identification

Confocal microscopes with multiphoton excitation and non-descanned detection (NDD) can penetrate deep into tissue. The combination with Fluorescence Lifetime Imaging (FLIM) extends the capability of such an instrument for concentration measurements of specific molecules, superior discrimination against autofluorescence as well as time resolved FRET (FLIM FRET).For optimal detection efficiency the fluorescence light is collected directly above the objective and guided to an Hybrid PMA detector combining GaAsP cathode sensitivity with excellent timing performance. The NDD based FLIM adapter replaces the standard objective holder and is easy to mount. A modular detection assembly allows to perform both confocal and NDD FLIM measurements.The fluorescence lifetime of fluorophores is strongly dependent on their photophysical properties and local environment. Together with the emission profile detected in different spectral channels the fluorescence decay can act as a fingerprint for a dye in a specific environment. We present a pattern-matching analysis technique that allows to identify selected patterns consisting of fluorescence decays in different spectral excitation and detection channels. The technique is easy to apply and allows for an excellent separation of different fluorophores, changes of fluorescence parameters due to differences in the environment as well as their discrimination from autofluorescence in biological samples.

Effects of salinity stress on the growth and chlorophyll fluorescence of Phaeodactylum tricornutum and Chaetoceros gracilis (Bacillariophyceae)

Abstract In this study, we investigated changes of fluorescence parameters, specific growth rate, and relative chlorophyll content (the ratio of the chlorophyll content after salinity exposure to that of the same samples at inoculation) in the two diatoms Phaeodactylum tricornutum Bohlin and Chaetoceros gracilis Schuett, during exposure of the cultures to different salinities (1–90 psu for P. tricornutum and 1–70 psu for C. gracilis) for different times (12, 24, and 48 h). The results indicated that the maximal photochemical efficiency of PSII (Fv/Fm), maximal relative electron transport rate (rETRmax), and photochemical quenching (qP) decreased significantly with the degree of salinity stress and increased with increasing exposure times in the two diatoms. For P. tricornutum, the values of Fv/Fm at salinities of 20–40 psu were always higher than those in other salinity treatments, rETRmax reached maximal values at salinities of 20–30 psu, and qP peaked at salinities of 10–20 psu. For C. gracilis, the values of Fv/Fm at salinities of 5–40 psu were always higher than those at 1 psu or at 50–70 psu. The highest values of rETRmax were also observed at salinities of 5–40 psu, whereas qP decreased slowly with increasing salinity from 10 to 60 psu. The changes in non-photochemical quenching (NPQ) were different in the two diatoms. For P. tricornutum, there were no significant differences below 60 psu for all three exposure times. Above 60 psu, however, NPQ in P. tricornutum increased with salinity up to 80 psu and decreased at 90 psu. For C. gracilis, NPQ decreased with increasing salinity from 1 to 20 psu, but increased at salinities of 20–70 psu during the 12- and 24-h periods, and was always low after 48 h. The specific growth rate of the two diatoms was inhibited at both low and high salinities, and decreased with increasing exposure times. The relative chlorophyll content of the two diatoms showed a similar trend to each other, with lower chlorophyll content occurring at extremely low and high salinities. These results indicated that extremely low and high salinity stress negatively influenced the chlorophyll fluorescence parameters and growth of these two diatoms.

A pea mutant (costata) expressing higher activity in thylakoid membrane-bound carbonic anhydrase alters PSII downregulation mechanisms.

The interaction between photosynthetic electron transport and the activities of the thylakoid associated carbonic anhydrase (tCA), estimated as combined tCA activity in pea plants (Pisum sativum L. Borek cv., WT) and mutant form (costata 2/125) that differ in chlorophyll content have been compared. Chlorophyll a fluorescence changes after the inhibition of tCA by ethoxyzolamide (EZ), estimating possible role of tCA in PSII downregulation were investigated. Costata expresses higher tCA activity and higher O2 evolution in comparison to WT. Inhibition of tCA by EZ decreased effective PSII photochemistry that coincided with an enhancement in thermal dissipation, while maximal PSII quantum yield (F(v)/F(m)) did not significantly change. Ethoxyzolamide induced changes in fluorescence parameters that were more strongly expressed in costata 2/125. The results show that tCA is involved in the regulation of the proton gradient across thylakoid membranes and thus limits PSII downregulation.

Fluorescence Investigation of the Binding of Model PDT Drugs to Nonionic and Zwitterionic Surfactants

The binding of two model photodynamic therapy drugs, chlorin p(6) and purpurin 18, with surfactants has been studied using steady-state and time-resolved fluorescence techniques. The surfactants used are amphiphilic nonionic surfactant (Tween 80 and Tween 40) and zwitterionic surfactant (HAPS). These have applications in drug delivery. The studies have been performed at pH 7 and 5 for chlorin p(6) and at pH 7 for purpurin 18. The binding constants have been estimated from the change in fluorescence parameters and have been compared with those for Cremophor EL and human serum albumin. Chlorin p(6) is found to bind to the surfactants to a greater extent at pH 5 than at pH 7. The same ionic species of chlorin p(6) is found to exist at the maximum concentrations of the surfactants.

Visualization of dynamics of plant–pathogen interaction by novel combination of chlorophyll fluorescence imaging and statistical analysis: differential effects of virulent and avirulent strains of P. syringae and of oxylipins on A. thaliana

Pathogen infection leads to defence induction as well as to changes in carbohydrate metabolism of plants. Salicylic acid and oxylipins are involved in the induction of defence, but it is not known if these signalling molecules also mediate changes in carbohydrate metabolism. In this study, the effect of application of salicylic acid and the oxylipins 12-oxo-phytodienoic acid (OPDA) and jasmonic acid on photosynthesis was investigated by kinetic chlorophyll fluorescence imaging and compared with the effects of infection by virulent and avirulent strains of Pseudomonas syringae. Both pathogen strains and OPDA caused a similar change in fluorescence parameters of leaves of Arabidopsis thaliana. The response to OPDA appeared faster compared with that to the pathogens and persisted only for a short time. Infiltration with jasmonic acid or salicylic acid did not lead to a localized and distinct fluorescence response of the plant. To capture the faint early symptoms of the plant response, a novel algorithm was applied identifying the unique fluorescence signature-the set of images that, when combined, yield the highest contrast between control and infected leaf segments. Unlike conventional fluorescence parameters, this non-biased approach indeed detected the infection as early as 6 h after inoculation with bacteria. It was posssible to identify distinct fluorescence signatures characterizing the early and late phases of the infection. Fluorescence signatures of both infection phases were found in leaves infiltrated with OPDA.

Light-Triggered Action Potentials and Changes in Quantum Efficiency of Photosystem II in Anthoceros Cells

Capillary microelectrodes and pulse amplitude-modulated microfluorometry were used to study light-triggered changes in cell membrane potential, chlorophyll fluorescence, and photochemical yield of PSII in chloroplasts of a hornwort Anthoceros sp. The action potential was generated by illuminating the plant sample for a few seconds. It was accompanied by a reversible decrease in quantum efficiency of PSII and by nonphotochemical quenching of fluorescence that continued as long as 10 min after the light stimulus. The presence of ammonium ions (2 mM) enhanced the amplitude and prolonged the duration of dark changes of fluorescence parameters in accordance with the reported increase in duration and amplitude of the light-triggered action potential in the presence of NH 4 + . A rapid retardation of PSII activity within the first seconds of illumination was also evident from absorbance changes at 810 nm reflecting the redox conversions of chlorophyll P700. The PSII-dependent stage of reduction in the induction curves of P700 absorbance was strongly suppressed, and the amplitudes of signals induced by white and far-red light (717 nm) differed insignificantly. It is concluded that a short-term irradiation triggers the generation of ΔpH at the thylakoid membranes, which is accompanied by inhibition of the plasma membrane H+ pump and by reversible inactivation of PSII due to increased thermal dissipation of chlorophyll excitations.

Acrylodan-conjugated Cysteine Side Chains Reveal Conformational State and Ligand Site Locations of the Acetylcholine-binding Protein

We undertook cysteine substitution mutagenesis and fluorophore conjugation at selected residue positions to map sites of ligand binding and changes in solvent exposure of the acetylcholine-binding protein from Lymnaea stagnalis, a nicotinic receptor surrogate. Acrylodan fluorescence emission is highly sensitive to its local environment, and when bound to protein, exhibits changes in both intensity and emission wavelength that are reflected in the degree of solvent exclusion and the effective dielectric constant of the environment of the fluorophore. Hence, cysteine mutants were generated based on the acetylcholine-binding protein crystal structure and predicted ligand binding sites, and fluorescence parameters were assayed on the acrylodan-conjugated proteins. This approach allows one to analyze the environment around the conjugated fluorophore side chain and the changes induced by bound ligand. Introduction of an acrylodan-cysteine conjugate at position 178 yields a large blue shift with alpha-bungarotoxin association, whereas the agonists and alkaloid antagonists induce red shifts reflecting solvent exposure at this position. Such residue-selective changes in fluorescence parameters suggest that certain ligands can induce distinct conformational states of the binding protein, and that mutually exclusive binding results from disparate portals of entry to and orientations of the bound alpha-toxin and smaller acetylcholine congeners at the binding pocket. Labeling at other residue positions around the predicted binding pocket also reveals distinctive spectral changes for alpha-bungarotoxin, agonists, and alkaloid antagonists.

Spatial heterogeneity of photosynthesis and the effect of temperature-induced bleaching conditions in three species of corals

Heterogeneity in photosynthetic performance between polyp and coenosarc tissue in corals was shown using a new variable fluorescence imaging system (Imaging-PAM) with three species of coral, Acropora nobilis, Cyphastrea serailia and Pocillopora damicornis. In comparison to earlier studies with fibre-optic microprobes for fluorescence analysis, the Imaging-PAM enables greater accuracy by allowing different tissues to be better defined and by providing many more data points within a given time. Spatial variability of photosynthetic performance from the tip to the distal parts was revealed in one species of branching coral, A. nobilis. The effect of bleaching conditions (33°C vs. 27°C) was studied over a period of 8 h. Marked changes in fluorescence parameters were observed for all three species. Although a decline in ΦPSII (effective quantum yield) and Yi (the first effective quantum yield obtained from a rapid light curve) were observed, P. damicornis showed no visual signs of bleaching on the Imaging-PAM after this time. In A. nobilis and C. serailia, visual signs of bleaching over the 8 h period were accompanied by marked changes in F (light-adapted fluorescence yield), NPQ (non-photochemical quenching) and E k (minimum saturating irradiance), as well as ΦPSII and Yi. These changes were most marked over the first 5 h. The most sensitive species was A. nobilis, which after 8 h at 33°C had reached a ΦPSII value of almost zero across its whole surface. Differential bleaching responses between polyps and coenosarc tissue were found in P. damicornis, but not in A. nobilis and C. serailia. NPQ increased with exposure time to 33°C in both the latter species, accompanied by a decreasing E k, suggesting that the xanthophyll cycle is entrained as a mechanism for reducing the effects of the bleaching conditions.

New Fluorescent Probes for Monitoring Polymerization Reactions: Photocuring of Acrylic Adhesives, 2

The results of following the fluorescence of selected probes during the entire range of curing of acrylic-based adhesives are presented in this work. The change of fluorescence parameters has been continuously measured during the photocrosslinking reactions of acrylic adhesive systems under UV-irradiation. Real-time FT-IR (RT-FT-IR) has been used to measure the conversion reached at different irradiation times. Intensity ratio, maximum emission wavelength and the first moment of fluorescence have been used as parameters to correlate with the double bond conversion degree. Two-slope plots have been obtained during UV-curing, which correspond to the different stages of the crosslinking reaction. Also, the size of fluorescent probe appears to be a parameter that influences the sensing time in which the probe can detect the changes in viscosity/polarity occurring along the UV-curing process. The results are compared with those of commercial probes such as dansylamide and prodan. The use of the fluorescence technique allowed us to understand the mechanism taking place and the role of the binder matrix, depending on the photoinitiator used. This information is not obtained when the reaction is followed by the conventional RT-FT-IR technique.

2-Aminopurine as a real-time probe of enzymatic cleavage and inhibition of hammerhead ribozymes

The design, synthesis and study of internally fluorescent hammerhead (HH) ribozymes, where changes in fluorescence parameters directly reflect the progress of the ribozyme's cleavage chemistry, are described. The approach relies on a HH substrate modified at position 1.1, proximal to the cleavage site, with 2-aminopurine (2AP), an intensely fluorescent adenosine isoster. The incorporation of 2AP, an unnatural nucleoside, does not interfere with the ribozyme folding and catalysis. Since 2AP is highly sensitive to environmental changes, its fluorescence is dramatically altered upon ribozyme-mediated cleavage of the substrate. This generates a measurable signal that directly reflects the progress of the ribozyme's reaction in real time. Identical pseudo first order rate constants are obtained for HH constructs using both continuous fluorescence monitoring and radioactive labeling. This rapid and real-time monitoring facilitates the study of ribozyme activity under different conditions (e.g., ionic strength, pH, etc.), and provides a useful assay to rapidly screen potential inhibitors. Three hitherto unknown HH inhibitors are presented and compared to neomycin B and chlortetracycline, two previously studied HH inhibitors. All three new small molecules, neo-acridine, guanidino-neomycin B, and [Δ-(Eilatin)Ru(bpy) 2] 2+, prove to be better inhibitors than neomycin B or chlortetracycline. Investigating HH inhibition under different ionic strengths reveals that the binding of neo-acridine, [Δ-(Eilatin)Ru(bpy) 2] 2+, and chlortetracycline to the HH involves hydrophobic interactions as their RNA affinities are largely unaffected by increasing salt concentrations. In contrast, neomycin B loses more than 50-fold of its inhibitory ability as the NaCl concentration is increased from 50 to 500 mM.

Are Calcium Ions and Calcium Channels Involved in the Mechanisms of Cu&lt;sup&gt;2+&lt;/sup&gt; Toxicity in Bean Plants? The Influence of Leaf Age

The influence of calcium channel blockers and ionophore on Cu2+-induced changes of the photosynthetic activity of runner bean plants (Phaseolus coccineus L.) was investigated. Excess Cu2+ was applied to leaves by injection or via the roots to examine a short/local or a long time/systemic effect of this metal, respectively. The changes in fluorescence parameters indicated that the mechanism of toxic action of Cu2+ ions on the photosynthetic apparatus was only partially connected with Ca2+ or Ca2+ channels. In young plants Ca2+ diminished especially photochemical and nonphotochemical dissipative processes induced by short- and long-term influence of excess Cu2+. Blocking of Ca2+ channels did not change direct Cu2+ action on the photosynthetic activity, however, their opening distinctly intensified the inhibitory effect of the metal. After a longer accumulation peri od the effect of Cu2+ ions did not change significantly due to modified Ca2+ penetration through membranes (except that caused by La3+). Copper directly introduced into older leaves diminished only at its highest concentration the activity both of the donor and acceptor sides of photosystem 2 (PS2) connected with Rfd decrease and increase of LNU. A similar effect was observed also after a long-term Cu2+ action, but disturbances on the acceptor side of PS2 were observed only at a higher Ca2+ content in the nutrient solution. Ca2+ ions, particularly after openning of channels, intensified direct inhibitory Cu2+ action on the photosynthetic activity expressed by decreased values of Fv/F0 and Rfd. Lanthanum and verapamil, at a lower Ca2+ content in the medium, decreased the photosynthetic activity of Cu2+-treated plants. This effect was also seen after additional Ca2+ supply to the leaves.

Leaf Age Dependence of Chlorophill Fluorescence Parameters in Water-Stressed Leaves of Phaseolus vulgaris L.

To investigate leaf age dependence in the responses of photosynthetic functions to water stress, the changes in fluorescence parameters of bean (Phaseolus vulgaris L.) leaves were examined while the leaf water potential (ΨLW) was gradually decreased by withholding water supply to the soil; leaves of three different ages were used from each plant. No significant leaf age dependence was detected in any fluorescence parameters when the ΨLW was higher than -0.9MPa, the point at which the net CO2 uptake rate approached zero. However, at ΨLW lower than -0.9MPa, both the maximum PS II photochemical efficiency (FV/FM) and the quantum yield of non-cyclic electron transport in PS II (φe) decreased with decreasing ΨLW. The decreases in both FV/FM and φe were larger in younger leaves. More conspicuous leaf age dependence was detected in the response of the steady-state nonphotochemical quenching coefficient (qN). Water stress-induced changes in all these fluorescence parameters recovered to pre-stress levels within a day after rewatering. The results showed that the changes in the fluorescence parameters did not result from damage to the photosynthetic systems, but from changes in physiological conditions caused by the cessation of CO2 supply to the stroma or changes in stromal water status. It was concluded that leaf age is an important factor in determining the responses of fluorescence parameters to water stress.