Rapid,nondestructive measurement of chlorophyll content in leaves with nonuniform chlorophyll distribution

A practical microcomputerized video image analysis method is described for quantifying leaf chlorophyll content without extraction. Chlorophyll concentration is estimated from densimetric measurements of whole, intact leaves. Direct comparison with conventional extraction measurements on Epipremnum aureum, a variegated species, verified the image analysis technique's accuracy. The inherent advantages with regard to the nondestructive and convenient nature of the measurement, and suitability for leaves with irregular chlorophyll distribution, are discussed.     

The relationship between chlorophyll and the carotenoids in the algal flagellate,Euglena

1. We have obtained an action spectrum for chlorophyll formation in Euglena gracilis. This action spectrum is similar to the absorption spectrum of protochlorophyll. However, efforts to isolate and identify this pigment have been unsuccessful. 2. Porphyrins have been extracted from both the normal and dark-adapted Euglena and a chlorophyll-free mutant. 3. The "action" spectra for chlorophyll and carotenoid synthesis have been found to almost coincide, indicating that the same porphyrin-like molecule may influence the synthesis of both pigments. 4. It is indicated that two porphyrin-like systems are in operation simultaneously, one concerned with carotenoid "removal" and another involved in carotenoid and chlorophyll synthesis.     

HP1: facts,open questions,and speculation

The demonstration, over a decade ago, that HP1 is a highly conserved constituent of heterochromatin was accompanied by the explicit view that this protein plays a pivotal role in epigenetic regulation (P.B. Singh, J.R. Miller, J. Pearce, R. Kothary, R.D. Burton, R. Paro, T.C. James, and S.J. Gaunt, 1991, Nucleic Acids Res. 19, 789-794). Recent studies have confirmed this view, unveiling specific interactions of HP1 with a variety of histone and nonhistone proteins. We discuss here some of these observations, concentrating on structure-function relationships and intracellular dynamics. Integrating the available information, we also present a hypothetical model describing how HP1, acting as a bifunctional cross-linker, could organize peripheral heterochromatin and contribute in the compartmentalization of the cell nucleus.     

Glycine metabolism and chlorophyll synthesis in barley leaves

α-Hydroxypyridine methane sulphonic acid (HPMS), isonicotinyl hydrazide (INH) and nialamide inhibit chlorophyll synthesis in etiolated barley leaves exposed to light. HPMS lowered the rate of protochlorophyllide regeneration but had little effect on the synthesis of protochlorophyll (P630) from exogenous $\text{δ}$-aminolaevulinic acid (ALA). The addition of glycine to HPMS treated leaves partially overcame the inhibition of chlorophyll synthesis. Glycine-[¹⁴C] was readily incorporated into ALA in dark-grown leaves. HPMS treatment increased the sp. act. of ALA in leaves fed glycine-[¹⁴C]. Glycollate oxidation was lower in extracts from HPMS treated leaves. Plants may therefore have two pathways for ALA production with the glutamate pathway becoming more important in conditions where photorespiration is high.     

Chlorophyll breakdown and chlorophyll catabolites in leaves and fruit

Chlorophyll metabolism probably is the most visible manifestation of life. Total annual turnover of chlorophyll has been estimated to involve more than 1000 million tons. Surprisingly, chlorophyll catabolism has remained an enigma until less than twenty years ago, when a colorless chlorophyll catabolite from senescent plant leaves was identified and its structure was elucidated. In the meantime, chlorophyll breakdown products have been identified in a variety of plant leaves and their structural features have been elucidated. Most recently, chlorophyll breakdown products have also been identified in some ripening fruit. Chlorophyll breakdown in vascular plants only fleetingly involves enzyme-bound colored intermediates. The stage of fluorescent catabolites is also passed rapidly, as these isomerize further to colorless nonfluorescent tetrapyrrolic catabolites. The latter accumulate in the vacuoles of de-greened leaves and are considered the final products of controlled chlorophyll breakdown. The same tetrapyrroles are also found in ripening fruit and are effective antioxidants. Chlorophyll breakdown leads to tetrapyrroles that appear to have physiologically beneficial chemical properties, and it may thus not merely be a detoxification process.     

The turnover of chlorophyll in greening wheat leaves

A method for the estimation of chlorophyll turnover in wheat leaves is presented. This is based on the inhibition of chlorophyll synthesis by treatment of the cut leaves with laevulinic acid (LA), a competitive inhibitor of δ-aminolaevulinic acid dehydratase. The turnover of chlorophyll in young, greening leaves, given short periods of light was a relatively rapid process. However, in seedlings exposed to light for longer periods the turnover became progressively slower, and was measured in days rather than hours.     

pH dependent chlorophyll fluorescence quenching in spinach thylakoids from light treated or dark adapted leaves

The pH dependence of maximum chlorophyll fluorescence yield (F_m) was examined in spinach thylakoids in the presence of nigericin to dissipate the transthylakoid pH gradient. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) was present to eliminate photochemical quenching. Thylakoids were prepared from dark adapted leaves (dark thylakoids) or preilluminated leaves (light thylakoids). In the latter there had been approximately 50% conversion of the xanthophyll violaxanthin to zeaxanthin, while no conversion had occurred in the former. In the presence of a reductant such as ascorbate, antimycin A sensitive quenching was observed (half maximal quenching at 5 M), whose pH dependence differed between the two types of thylakoid. Preillumination of leaves resulted in more quenching at pH values where very little quenching was observed in dark thylakoids (pH 5–7.6). This was similar to activation of high-energy-state quenching (qE) observed previously (Rees D, Young A, Noctor G, Britton G and Horton P (1989) FEBS Lett 256: 85–90). Thylakoids isolated from preilluminated DTT treated leaves, that contained no zeaxanthin, behaved like dark thylakoids. A second form of quenching was observed in the presence of ferricyanide, that could be reversed by the addition of ascorbate. This was not antimycin A sensitive and showed the same pH dependence in both types of thylakoid. The former type of quenching, but not the latter, showed similar low temperature fluorescence emission spectra to qE, and was considered to occur by the same mechanism.Abbreviations DCMU 3(3,4-dichlorophenyl)-1,1-dimethylurea - DTT dithiothreitol - EDTA Ethylenediaminetetra-acetic acid - F₀ dark level fluorescence yield - F_m maximum fluorescence yield - F_v/F_m ratio of variable to total fluorescence yield - Hepes 4-(2-hydroxyethyl)1-piperazineethanesul-phonic acid - Mes 2-(N-morpholino) ethanesulfonate - pH transthylakoid pH gradient - PS I Photosystem I - PS II Photosystem II - Q_A primary stable electron acceptor of Photosystem II - qE high-energy-state fluorescence quenching     

Differences in photosynthetic activity, chlorophyll and carotenoid levels, and in chlorophyll fluorescence parameters in green sun and shade leaves of Ginkgo and Fagus

The differences in pigment levels and photosynthetic activity of green sun and shade leaves of ginkgo (Ginkgo biloba L.) and beech (Fagus sylvatica L.) are described. Sun leaves of both tree species possessed higher levels in chlorophylls (Chl) and carotenoids on a leaf area basis, higher values for the ratio Chl a/b and lower values for the ratio Chl/carotenoids (a+b)/(x+c) in comparison to shade leaves. The higher photosynthetic rates P(N) of sun leaves (ginkgo 5.4+/-0.9 and beech 8.5+/-2.1 micromol m(-2)s(-1)) were also reflected by higher values for the Chl fluorescence decrease ratios R(F)(d) 690 and R(F)(d) 735. In contrast, the shade leaves had lower P(N) rates (ginkgo 2.4+/-0.3 and beech 1.8+/-1.2 micromol m(-2)s(-1)). In both tree species the stomatal conductance G(s) was significantly higher in sun (range: 70-19 1 mmol m(-2)s(-1)) as compared to shade leaves (range: 5-55 mmol m(-2)s(-1)). In fact, at saturating light conditions there existed a close correlation between G(s) values and P(N) rates. Differences between sun and shade leaves also existed in several other Chl fluorescence ratios (F(v)/F(m), F(v)/F(o), and the stress adaptation index Ap). The results clearly demonstrate that the fan-shaped gymnosperm ginkgo leaves show the same high and low irradiance adaptation response as the angiosperm beech leaves.     

Scale-up of microbial processes: impacts, tools and open questions

Industrial biotechnological production is developing rapidly worldwide. Consequently, more and novel bioprocesses need to perform optimally not only in small lab- but also in large production scales. This article shortly reviews typical impacts found when cells are exposed to micro environmental heterogeneities typically occurring in poorly mixed large scale production reactors. The current state-of-the-art of tool development is presented for analyzing these phenomena. Finally, still open questions are formulated and needs for future research are outlined to further support the expansion of biotech industries by successful research results.     

Haem and chlorophyll formation in etiolated and greening leaves of barley

The amounts of protochlorophyllide (P650) and protohaem were measured in ageing dark-grown barley leaves. Maximum amounts of P650 and protohaem were found in 6- to 8-day-old material after which P650 declined rapidly and protohaem more slowly. In leaves exposed to light maximum chlorophyll was produced in 6-day-old material with progressively less the older the leaves. Haem concentrations increased in seedlings of all ages exposed to light. A lag phase was observed for both chlorophyll and haem formation in leaves given a light treatment. Haem, however, showed a slight yet sig nificant decline as chlorophyll production commenced. The results indicate that chlorophyll and haem synthesis share a common pool of δ-aminolae vulinic acid (ALA). At a certain stage of development, the magnesium porphyrin pathway diverts precursors away from haem synthesis. It is only when the ALA synthesising system is well developed that the production of ALA can satisfy pathways to both haem and chlorophyll. The observed changes in haem under certain conditions suggest that, as in animal systems, haem levels may regulate porphyrin formation (chlorophylls) by controlling the supply of ALA.     

Control of chlorophyll production in rapidly greening bean leaves

The possible involvement of nucleic acid and protein synthesis in light-regulated chlorophyll formation by rapidly greening leaves has been studied.

Removing leaves from illumination during the phase of rapid greening results in a reduction in the rate of pigment synthesis; cessation occurs within 2 to 4 hours. Etiolated leaves which exhibit a lag in pigment synthesis when first placed in the light do not show another lag after a 4 hour interruption of illumination during the phase of rapid greening.

Actinomycin D, chloramphenicol, and puromycin inhibit chlorophyll synthesis when applied before or during the phase of rapid greening. Application of δ-amino-levulinic acid partially relieves the inhibition by chloramphenicol.

It is suggested that light regulates chlorophyll synthesis by controlling the availability of δ-aminolevulinic acid, possibly by mediating the formation of an enzyme of δ-aminolevulinate synthesis. This process may result from gene activation or derepression; the involvement of RNA synthesis of some sort is suggested by the inhibitory effect of actinomycin D on chlorophyll production by rapidly greening leaves.

     

Plant fibers: Initiation,growth, model plants,and open questions

Fibers, which are used as a major raw material in the paper industry, as structural components in timber for building, and in the manufacture of wooden items, are among the most important renewable resources. Billions use wood as a major energy source, and fibers are an energy-rich component of wood. They are used for various textiles and as raw material for composites. In this review, I describe the basic characters of fibers, their structure, development, uses, and some of the current major model plants for fiber formation. I discuss open developmental questions and various aspects of further research. Most of the recent progress in the biology of fiber formation, especially in their cell-wall chemistry, emerged from studies of several model plants: Arabidopsis thaliana, Populus sp., Eucalyptus sp., flax, and hemp. I stress the critical need to combine the use of modern methods of research with classical botany. Approaching the issue of fiber formation only by molecular or only by classical methods will not only limit the progress, but may result in critical mistakes. Considering the importance of fibers to humanity, it is surprising how little we know about the biology of fiber formation and how little it is studied as compared, for instance, to the effort to study the genetics and cell biology of flower organ identity.     

Interactions between riparian leaves and algal/microbial activity in streams

The effects of leaf leachates derived from six forest trees on algal biomass development was examined in a 3-week experiment in a New Zealand spring. The trees all occur in riparian zones and were Fagus sylvatica and Alnus glutinosa from Europe, Eucalyptus globulus from Australia, Pinus radiata from western North America, and two New Zealand species, Fuchsia excorticata and Nothofagus solandri. Leachates from five species inhibited algal growth relative to controls, whereas leachate from a sixth species (F. excorticata) was stimulatory. In contrast, cellulose breakdown was unaffected by five leachates and inhibited by the other (F. sylvatica). In a second experiment, colonization of leaves and inert substrata (polythene strips) by algae was examined with SEM after 16 days. Algal abundance was greatest on polythene strips and leaves of F. sylvatica, and substantially lower on the other species. Our results indicate that both leaf texture and chemical composition can affect algal colonization and growth, and that chemicals derived from leaves have the potential to both inhibit and stimulate ecosystem processes in streams.     

Energy-dependent quenching of dark-level chlorophyll fluorescence in intact leaves

A new type of modulation fluorometer was used in the study of energy-dependent chlorophyll fluorescence quenching (q_E) in intact leaves. Under conditions of strong energization of the thylakoid membrane (high light intensity, absence of CO₂) not only variable fluorescence, F_V, but also dark-level fluorescence, F_O, was quenched, leading to definition of a quenching coefficient, q_O. Information on q_O was shown to be essential for correct determination of photochemical (q_Q) and energy dependent quenching (q_E) by the saturation pulse method. The relationship between q_E and q_O was analysed over a range of light intensities at steady state conditions. q_E was found to consist of two components, the second of which is linearly correlated with q_O. q_O and the second component of q_E are interpreted to reflect the state 1 — state 2 shift caused by LHC II phosphorylation.     

Photosynthesis and chlorophyll content in different areas of fodder cabbage leaves

Byla stanovena intensita fotosynthesy a obsah chlorofylu v terěících z r?znych okrsk? ?epele listu krmné kapusty odr. Coulet de Flandre. Obsah chlorofylu na jednotku plochy byl vy??í v apikální ?ásti list? ne? v ?ásti basální, vy??í ve st?edové ne? v okrajové ?ásti listu. Nebyly nalezeny podstatné rozdily v intensitě fotosynthesy ve vzorcích z r?znych ?ástí listu. Intensita fotosynthesy není p?ímo úměrná váze su?iny daného vzorku, a?koliv vàha su?iny na jednotku plochy je podstatně vy??í v apikálni ne? v basální ?ásti listové plochy.     

Light scattering,chlorophyll fluorescence and state of the adenylate system in illuminated spinach leaves

Scattering of green light and chlorophyll fluorescence by spinach leaves kept in a stream of air or nitrogen were compared with leaf adenylate levels during illumination with blue, red or far-red light. Energy charge and ATP-ADP ratios exhibited considerable variability in different leaves both in the dark and in the light. Variability is explained by different possible states of the reaction oxidizing triose phosphate or reducing 3-phosphoglycerate. Except when oxygen levels were low, there was an inverse relationship between light scattering and chlorophyll fluorescence during illumination with blue or red light. When CO₂ was added to a stream of CO₂-free air, chlorophyll fluorescence increased, sometimes after a transient decrease, and both light scattering and leaf $\text{ATP}\text{ADP}$ ratios decreased. Similar observations were made when air was replaced by nitrogen under blue or high-intensity red light. Under these conditions, over-reduction caused inhibition of electron transport and phosphorylation in chloroplasts. However, when air was replaced by nitrogen during illumination with low-intensity red light or far-red light, light scattering increased instead of decreasing. Under these light conditions, $\text{ATP}\text{ADP}$ ratios were maintained in the light. They decreased drastically only after darkening. Although $\text{ATP}\text{ADP}$ ratios responded faster than light scattering or the slow secondary decline of chlorophyll fluorescence due to illumination, it appeared that in the steady state, light scattering and chlorophyll fluorescence are useful indicators of the phosphorylation state of the leaf adenylate system at least under aerobic conditions, when chloroplast and extrachloroplast adenylate systems can effectively communicate.     

Effects of drought on photosynthesis,chlorophyll fluorescence and photoinhibition susceptibility in intact willow leaves

Plants from clonal cuttings of Salix sp. were subjected to a drying cycle of 10 d in a controlled environment. Gas exchange and fluorescence emission were measured on attached leaves. The light-saturated photosynthetic CO₂ uptake became progressively inhibited with decreased leaf water potential both at high, and especially, at low intercellular CO₂ pressure. The maximal quantum yield of CO₂ uptake was more resistant. The inhibition of light-saturated CO₂ uptake at leaf water potentials around-10 bar, measured at a natural ambient CO₂ concentration, was equally attributable to stomatal and non-stomatal factors, but the further inhibition below this water-stress level was caused solely by non-stomatal factors. The kinetics of fluorescence emission was changed at severe water stress; the slow secondary oscillations of the induction curve were attenuated, and this probably indicates perturbations in the carbon reduction cycle. The influence of light level during the drought period was also studied. Provided the leaves were properly light-acclimated, drought at high and low light levels produced essentially the same effects on photosynthesis. However, low-light-acclimated leaves became more susceptible to photoinhibitory treatment under severe water stress, as compared with well-watered conditions.     

The mechanism of the ozone-induced changes in thermoluminescence glow curves of barley leaves

The changes in thermoluminescence (TL) signals induced by short-term ozone exposure of leaves are characterized by a down-shift of the peak-temperature of the TLB-band and an increase of a TL band at 55°C. We investigated the relationship of these changes to photosystem 2 (PS2) photochemistry. The changes were not only detectable in the presence of ozone, but also after irradiation of dark-adapted leaves and after aging of irradiated detached leaf segments. The opposite effect on TL, an up-shift of the peak-temperature of the B-band and the decrease of the intensity of the band at 55°C were found after infiltration of leaves with nigericin, antimycin A, and diphenyleneiodonium chloride (DPI). Propyl gallate down-shifted the peak-temperature of the B-band. 2,5-dimethyl-1,4-benzoquinone up-shifted the peak-temperature of the B-band and decreased the intensity of the 55°C band. The intensity of the 55°C band did not change significantly in the presence of oxygen in comparison to that in nitrogen atmosphere. It decreased with time of dark adaptation (50% intensity was observed after 3 h of dark adaptation at room temperature), however, it was reactivated to its initial value (at 5 min of dark adaptation) after 1 single-turnover flash. The 55°C band was not significantly changed in the presence of DCMU. Thus the ozone-induced band at 55°C is assigned to charge recombination in PS2. Changes in the electron transport chain at the acceptor side of PS2, probably related to the cyclic electron transport around photosystem 1 and/or chlororespiration, could play an important role in the increase of the 55°C band and the down-shift of the B-band. The changes at the acceptor side indicated by TL can be an ex pression of a physiological regulatory mechanism functional under stress conditions.