Imipramine affects circadian leaf movements in Oxalis regnellii

The sensitivity of the circadian leaf movement of Oxalis regnellii Mig. to imipramine (a tricyclic dibenzazepine) was investigated. Imipramine, like Li⁺, is used as a therapeutic agent against depressive disorders in man. The therapeutic effects of the two substances might be mediated via effects on basic circadian rhythms and the cellular level. It was indeed possible to influence the circadian movement of Oxalis by imipramine; pulses (10⁻³ M , 4h) phase shifted the rhythms and caused advances. A phase response curve is presented. No period change of the movements was caused by permanent presence of imipramine (5 - 10⁻⁵ or 10⁻⁵ M ). The nature of the imipramine-induced phase shift is discussed and compared with Li⁻¹ effects on the same circadian system.     

Basic properties of the circadian leaf movements of Oxalis regnellii, and period change due to lithium ions

The circadian leaf movement of Oxalis regnellii Mig, has been investigated. The three leaflets of a stalk were normally synchronized, and under the experimental conditions chosen they showed a period of 26.2 ± 0.1 h. Cutting off one or two leaflets led to a successive decrease of the period length (25.7 ± 0.1 and 25.1 ± 0.3 h resp.). It was possible to phase shift the leaf movements by mechanical means (advance of 1.6 ± 0.3 h).
Lithium ions, added permanently to the transpiration stream, increased the period length of the leaf movements by more than one hour (with 10 m M Li⁺). A 24 h pulse of 20 m M LiCl caused a permanent 2–3 h phase delay of the circadian rhythm. Four-h pulses, on the other hand, provoked only transient phase delays, the magnitude being dependent on the phase of application. Lithium concentrations were determined for different regions of leaves and pulvini. It was shown that leaf segments had considerably lower concentrations than pulvini. No significant difference in the lithium concentration was observed between the upper and lower part of pulvini.
In the light leaf position was strongly correlated with water uptake and the consequences for applications of substances to the circadian system via the transpiration stream is discussed. A simple model of the oscillatory system and reactions connected to it is discussed.     

CIRCADIAN GROWTH IN PORPHYRA UMBILICALIS (RHODOPHYTA): SPECTRAL SENSITIVITY OF THE CIRCADIAN SYSTEM

The circadian rhythm in growth of the red macroalga Porphyra umbilicalis (Linnaeus) J. Agardh was investigated under different spectral light conditions in laboratory-grown thalli. A free-running rhythm was observed in constant green or red light at irradiances of 2.5 to 20 μmol photons·m⁻²·s⁻¹, whereas arhythmicity occurred in constant blue light at 6–20 μmol photons·m⁻²·s⁻¹. The circadian oscillator controlling growth rhythmicity in Porphyra uses most of the visible sunlight spectrum and possibly multiple photoreceptors with a high sensitivity for blue light and a lower sensitivity for red light. This was inferred from three experimental results: (1) The free-running period, τ, of the growth rhythm decreased with increasing irradiance, from approximately 25 h at 2.5 μmol photons·m⁻²·s⁻¹ to 22 h at 20 μmol photons·m⁻²·s⁻¹ in red or green light, (2) Dark pulses of 3 h duration, interrupting otherwise continuous green or red light, caused advances during the subjective day and delays during the subjective night; the circadian oscillator in Porphyra can discriminate darkness from green or red light, and (3) Low-irradiance blue light pulses (2.5 μmol photons·m⁻²·s⁻¹) shifted the growth rhythm in red light of higher irradiance (e.g. 10 μmol photons·m⁻²·s⁻¹), and a strong, high amplitude, type 0 phase response curve was obtained that is usually observed with light pulses shifting a circadian rhythm in otherwise continuous darkness.     

Do leaves contribute to the abscisic acid present in the xylem sap of ‘draughted’ sunflower plants?

Abstract. The root systems of 30-d-old sunflower plants were treated with polyethylene glycol (PEG; osmotic potential - 1.0MPa) for 2h, causing mild and transient wilting. Ten minutes before this treatment was applied, half the plants were defoliated. At varying times after the imposition of the PEG 'drought stimulus', the plant stems were cut and the sap exudate was collected and analysed for abscisic acid (ABA), using an elisa method. When stems were cut 2.25h after the treatments were applied, the ABA concentration in the sap of the controls did not vary with time: the mean concentration was 10.7 ± 1.0μ, mol m⁻³. However, in the treated plants, the first sample contained 78.1 ± 10.1 μmol m⁻³, decreasing to 13.6 ± 2.8 μmol m⁻³ over 8.75h. Defoliation did not affect the ABA concentration in the sap. When stems were cut at varying times (up to 25h) after treatment, the PEG treatment again caused an immediate increase in the ABA concentration in the sap, from 20 ± 1 to 136 ± 21 μmol m⁻³. However, defoliation reduced this increase, but only in plants sampled 4–25h after treatment. We conclude that, after the PEG treatment to the roots, the initial increase of the ABA content of sap, and its attenuation with time, may be ascribed to synthesis in the roots whereas, thereafter, ABA derived from the leaves makes a major contribution to the ABA found in the xylem sap.     

Effect of 2-chloroethylphosphonic acid on capsule formation and alkaloid content in Papaver somniferum

Application of different concentrations of ethephon (2-chloroethylphosphonic acid) to Papaver somniferum L. at the times of stem elongation, bud, and capsule formation produced different effects. Ethephon (10^-2 M ) retarded growth of the plant and inhibited capsule formation during stem elongation, significantly reduced capsule size during the flowering period, but did not alter capsule development during capsule formation. When applied during the period of stem elongation, ethephon (10^-3 M and 10^-4 M ) reduced capsule size; alkaloid accumulation was reduced by ethephon at a concentration of 10^-3 M , but slightly increased by 10^-4 M . Ethephon (10^-3 M and 10^-4 M ) did not alter capsule development or alkaloid content significantly when applied during bud formation, but stimulated capsule size and alkaloid content when applied during capsule formation. Pretreating the plants with Ag⁺ (silver nitrate) did not reverse the ethephon effect. The results suggest that capsule maturation and alkaloid accumulation in P. somniferum are modified by ethylene, which is produced as a result of exogenous ethephon treatment.     

Influence of red irradiation and of removal of integuments on the metabolism of (±) [2-14C]-abscisic acid by Lactuca sativa Grand Rapids achenes

Achenes of Lactuca saliva L. cv. Grand Rapids, imbibed for 6 h in water or in a 10 µ M solution of non-radioactive abscisic acid (ABA), were cultivated on (2-¹⁴C]-ABA (10 µ M ) for 40 to 90 h. Red irradiation (660 ± 2.5 nm, 5 min, 2 W m ^-2) or removal of integuments were carried out before transfer to (2-¹⁴C]-ABA. When both treatments were applied, irradiation preceded removal of integuments. Imbibition and culture took place in darkness at 24°C. Two acidic diethyl ether phases, which contained the free acids (free phase) and the acids released after mild alkaline hydrolysis, respectively, were isolated. They were analyzed by thin layer chromatography (TLC). as well as the remaining aqueous phase.
Both red irradiation and removal of integuments led to increased [2-¹⁴C|-ABA uptake. Application of ABA during imbibition partly limited the stimulating effect of red irradiation on radioactive ABA uptake. Red irradiation stimulated [2-¹⁴C|-ABA metabolism by achenes, favouring the formation of the polar compound found in the remaining aqueous phase. Removal of the integuments stimulated metabolism notably, leading to an increase of the radioactivity in the remaining aqueous phase. This treatment also induced the appearance of new metabolites in the free phase (compound believed to be 7'-hydroxy-ABA) as well as in the remaining aqueous phase. The glucose ester of ABA was the only representative compound of the ester phase. Irrespective of the experimental conditions, there was no classical oxidative metabolism indicating that oxygen was not the limiting factor.     

Interaction of triacontanol with gibberellin in control of lettuce seed germination

Triacontanol at concentrations from 2.3 × 10^-9 M to 2.3 × 10^-7 M did not affect the germination of lettuce ( Lactuca sativa L., cv. Grand Rapids) seeds in darkness, stimulated by light at 25°C or by benzyladenine at 31°C. Stimulation of seed germination by gibberellin A₃ (10^-5 M ) was significantly inhibited by triacontanol; the most effective concentration was 4.6 × 10^-8 M. Pulse experiments demonstrated that triacontanol was ineffective when applied later than gibberellin, whereas an inverse sequence of treatment caused an inhibition comparable to that resulting from continuous treatment of seeds with both factors. Possible interaction of triacontanol with gibberellin receptor is discussed.     

2,4-Dichlorophenoxyacetic acid inhibition of potassium transport in barley seedlings

Growth, potassium uptake and translocation as well as transpiration rates were measured in intact low-salt barley seedlings ( Hordeum vulgare L. cv. Union) in the presence of different 2,4-D concentrations at pH 6.5. Growth was only affected at 10^-3 M .
Above 10^-7 M 2,4-D both uptake by the roots and transport to the shoots were inhibited. The inhibition at 10^-5 M remained constant for at least 24 h. Furthermore inhibition of uptake was measurable within 1 h. Excised roots and roots of intact plants showed the same uptake pattern.
It is suggested that the observed effects were caused by 2,4-D-induced changes in uptake and translocation systems in the roots. Pre-treatment with 10^-5 M 2,4-D had no effect upon subsequent potassium uptake. Transpiration was reduced within 1 h in 10^-4 or 10^-3 M 2,4-D, probably due to changes in water transport or root permeability.     

A developmental study of Glycine max cell and protoplast isolation in relation to leaf age and photosynthetic competence

Leaf mesophyll cells were isolated from developing first trifoliate leaves of Glycine max (L.) Merr cv. Fiskeby V using a mechanical isolation procedure combined with low speed centrifugation. Cell yields of 17 ± 1.7% were routinely obtained with 55–75% intactness, as assessed by staining techniques, fluorescence transients and the ability of cells to convert to protoplasts after enzyme treatment. Rates of leaf photosynthesis were maximal in 27-day-old plants [280 μmol O₂ evolved (mg chlorophyll)^-1h^-1], from which isolated cells and protoplasts gave rates of up to 140 μmol O₂ evolved (mg chlorophyll)^-1 h^-1. Results are discussed in relation to leaf development and cell status during the attainment of photosynthetic competence.     

Effect of alcohols on the association of photosynthetic fructose-1,6-bisphosphatase to thylakoid membranes

Aliphatic alcohols have a positive effect on the assoociation of pea ( Pisum sativum L. cv. Lincoln) chloroplast fructose- 1,6-bisphosphatase (FBPase; EC 3.1.3.11) with thylakoid membranes. The alcohol concentration needed to obtain a fixed percentage of enzyme association decreased with increased length of the aliphatic chain of the alcohol; maximum binding was obtained when the lysis medium contained, in molar fractions (or v/v percentages): 48×10^-4(T4 (2.4%), 26×10^-3 (10%), 40×10^-3 (15%), 76×10^-3 (21%), and 13×10^-2 (24%), of 1-butanol, 1-propanol, 2-propanol, ethanol, and methanol, respectively. A good correlation of binding with the octanol/water partition coefficient was observed. Since this coefficient constitutes a measure of hydrophobicity, we suggest that the binding of FBPase to the membranes occurs via hydrophobic clusters of both components.     

Growth and physiological responses of canola (Brassica napus) to three components of global climate change: temperature, carbon dioxide and drought

Elevated CO₂ appears to be a significant factor in global warming, which will likely lead to drought conditions in many areas. Few studies have considered the interactive effects of higher CO₂, temperature and drought on plant growth and physiology. We grew canola ( Brassica napus cv. 45H72) plants under lower (22/18°C) and higher (28/24°C) temperature regimes in controlled-environment chambers at ambient (370 μmol mol⁻¹) and elevated (740 μmol mol⁻¹) CO₂ levels. One half of the plants were watered to field capacity and the other half at wilting point. In three separate experiments, we determined growth, various physiological parameters and content of abscisic acid (ABA), indole-3-acetic acid and ethylene. Drought-stressed plants grown under higher temperature at ambient CO₂ had decreased stem height and diameter, leaf number and area, dry matter, leaf area ratio, shoot/root weight ratio, net CO₂ assimilation and chlorophyll fluorescence. However, these plants had increased specific leaf weight, leaf weight ratio and chlorophyll concentration. Elevated CO₂ generally had the opposite effect, and partially reversed the inhibitory effects of higher temperature and drought on leaf dry weight accumulation. This study showed that higher temperature and drought inhibit many processes but elevated CO₂ partially mitigate some adverse effects. As expected, drought stress increased ABA but higher temperature inhibited the ability of plants to produce ABA in response to drought.     

Long-term effects of abscisic acid on K+ transport in young wheat plants of different K+ status

The effects of abscisic acid (ABA) on growth, uptake and translocation of potassium ions, K⁺,Mg²⁺-ATPase activity and transpiration were investigated in young wheat ( Triticum aestivum L. cv. Martonvásári-8) plants grown at different K⁺ supplies. Long-term treatment with ABA (10 μ M ) reduced growth in high-K⁺ plants, but had less effect under low-K⁺ conditions. K⁺(⁸⁶Rb) uptake was inhibited by about 70 and 40% in low- and high-K⁺ plants, respectively. The stimulation by K⁺ of the Mg²⁺-ATPase activity in the root microsomal fraction was lost with ABA treatment. It is suggested that the inhibitory effect of ABA on K⁺ uptake may be related to this effects on the K⁺,Mg²⁺-ATPase. Translocation of K⁺ to the shoot was inhibited in low-K⁺ plants only, and it was not affected in high-K⁺ plants. In parallel to this, ABA treatment reduced transpiration by about 50% in low-K⁺ plants, whereas a much smaller effect was seen in high-K⁺ plants. These observations suggest that the regulation by ABA of the stomatal movements is strongly counteracted by high-K⁺ status.     

Effect of pH, temperature and culture medium composition on the production of an extracellular cysteine proteinase by Micrococcus sp. INIA 528

Growth and proteinase production by Micrococcus sp. INIA 528 in a batch-operated laboratory fermentor were investigated, with trypticase soy broth as the basal medium for studies on optimum temperature, pH and medium composition. Maximum growth was recorded at 34°C and pH 715, whereas optimum temperature and pH for proteinase production were 31°C and pH 6.25. Maximum rate of enzyme production occurred during the late log and early stationary phases of growth. Addition of 5.0 g 1^-1 yeast extract, 1.0 g 1^-1 glucose, 1.0 g 1^-1 MgSO₄ or 1.0 g 1^-1 K₂HPO₄ to basal medium resulted in a lower enzyme yield, but supplementation of basal medium with 2.5 g 1^-1 (NH₄)₂SO₄ increased enzyme production by 45%. A high initial biomass added to fresh broth supplemented with 2.5 g 1^-1 (NH₄)₂SO₄ only increased enzyme activity by 19%, compared to the maximum enzyme activity achieved with the standard inoculum.     

Identification of 4-chloroindole-3-acetic acid and indole-3-aldehyde in seeds of Pinus sylvestris

4-Chloroindole-3-acetic acid (4-Cl-IAA) and indole-3-aldehyde (IAId) have been characterized as endogenous constituents in seeds of Pinus sylvestris L. by gas chromatography-mass spectrometry. Quantitative estimates indicate that immature seeds contained 640 pg 4-Cl-IAA (g fresh weight)^-1 while mature seeds contained 340 pg (g dry weight)^-1. 4-Cl-IAA could not be detected in seeds five days after germination. The content of IAld increased from 127 pg (g dry weight)^-1 in mature seeds to 315 pg (g dry weight)^-1 after five days of germination.     

The cytotoxic and photodynamic inactivation of micro-organisms by Rose Bengal

Rose Bengal was cytotoxic to the following bacteria at the concentrations given in parentheses (highest concentrations of dye in mol/1 at which growth occurred on nutrient medium): Brochothrix thermosphacta and Deinococcus radiodurans (1 times 10^-6 or less); Streptococcus, Micrococcus, Staphylococcus, Bacillus, Arthrobacter and Kurthia spp. (1 times 10^-5–1 x 10^-4), and Pseudomonas spp. and Enterobacteriaceae (5 times 10^-3–1 x 10^-2 or greater). These organisms were killed rapidly when suspended in illuminated (170 μE/m²/s) solutions of Rose Bengal (1 times 10^-4 mol/1) providing oxygen was present. Singlet oxygen was identified as the lethal agent, because the rate of killing was increased by dissolving the dye in deuterium oxide while the organisms were protected against photoinactivation by L-histidine or crocetin. Yeasts from chilled foods were killed in illuminated solutions of Rose Bengal but a light intensity of 315 μE/m²/s was needed for a death rate comparable with that of bacteria. The yeasts present in a range of chilled meat and dairy products failed to form colonies on Rose Bengal (5 times 10^-5 mol/1) media exposed continuously to modest illumination (55–80 μE/m²/s).     

Endogenous abscisic acid levels are linked to decreased growth of bush bean plants treated with NaCl

This paper studies the relative importance of endogenous ABA and ion toxicity in the leaf growth inhibition caused by NaCl in salt-adapted and unadapted bush beans. Adaptation to salt-stress was achieved by germination of seeds in 75 m M NaCl, while unadapted plants were germinated in tap water. The adaptation process caused a transitory increase in leaf ABA, Na⁺ and Cl⁻ concentrations, while leaf expansion was inhibited. However, when grown for 8 or 13 days in 75 m M NaCl-containing nutrient solution, primary and first trifoliolate leaves of salt-adapted plants had greater areas than those of unadapted plants. Concentrations of ABA, Na⁺ and Cl⁻ in these leaves were lower in adapted plants, and a strong negative correlation between leaf expansion growth and either leaf Na⁺, Cl⁻ or ABA concentrations could be established. However, in the second trifoliolate leaves only the ABA, but not the Na⁺ or Cl⁻, concentrations were significantly correlated with leaf expansion. Our results suggest that salt-induced inhibition of leaf expansion growth in bush beans is mediated by ABA rather than Na⁺ or Cl⁻ toxicity. Moreover, the increase of ABA, induced by the salt-pretreatment, seems to play an important role in limiting the accumulation of Na⁺ and Cl⁻ in the leaves, leading to adaptation of bush beans to salt-stress.     

Involvement of abscisic acid and ethylene in the responses of citrus seedlings to salt shock

The responses of salt‐sensitive citrus rootstocks to 200 m M NaCl were periodically determined on seedlings of citrange Carrizo ( Citrus sinensis [L.] Osbeck × Poncirus trifoliata [L.] Raf) during 30 days. The stressed seedlings adjusted osmotically, reduced stomatal conductance, increased proline content and ethylene production, and showed massive leaf abscission (92%). The salt shock also increased abscisic acid (ABA) and aminocyclopropane‐1‐carboxylic acid (ACC) in roots, xylem fluid and leaves, and in addition promoted Cl⁻ accumulation. The pattern of change of ABA, ACC and proline followed a two‐phase response: an initial transient increase (10‐12 days) overlapping with a gradual and continuous accumulation. This biphasic response appears to be compatible with the proposal that the transitory hormonal rises are induced by the osmotic component of salinity, whereas the Cl⁻ increase determines the subsequent accumulations. During the second phase, Cl⁻ levels correlated with abscission in leaves. Production of leaf ethylene was also concomitant with the increase in the abscission rate. Salt‐induced abscission was either reduced with CoCl₂ (52%) or inhibited with silver thiosulphate (14%). The results suggest that in salt‐stressed citrus, leaf abscission is induced by the chloride build‐up through a mechanism that stimulates leaf ACC synthesis and further conversion to ethylene.     

The effect of calcium concentration on the toxicity of copper, lead and zinc to yolk-sac fry of brown trout, Salmo trutta L., in soft, acid water

Yolk-sac fry of brown trout were exposed to three levels of single trace metals (Cu, 20,40,80 nmol 1^-1; Pb, 12·5,25,50 nmol 1^-1; Zn, 75,150,300 nmol 1^-1) typical of concentrations reported for acid soft waters, in flowing, artificial, soft water media maintained at pH 4·5 and [Ca] of 20 or 200 μmol 1^-1for 30 days.
Mortalities were high in fry subjected to all levels of the three trace metals at [Ca] 20 μmol 1^-1, with 80% of the total deaths occurring between days 11 and 15 of the experiment. 25% mortality occurred at low [Ca] and pH 4·5 in the absence of trace metals, with only one death recorded at [Ca] 200 μmol1^-1'(Cu, 80 nmol 1^-1). At high [Ca] all three levels of Cu and Pb impaired net Na and K uptake; Cu was the only metal to reduce the uptake of Ca. The Zn treatments had no significant effect on mineral uptake. Calcification of centra was reduced by all three Cu treatments at [Ca] 200 μmol 1^-1. The lowest Zn concentration (75 nmol 1^-1) was the only other treatment to impair skeletal development. In the absence of trace metals, low [Ca] significantly reduced Ca, Na and K uptake, skeletal calcification and dry mass at pH 4·5.
The deleterious effects of low Cu, Pb and Zn concentrations at low pH and low [Ca], and the ameliorative effect of higher ambient [Ca], are discussed in relation to fishery status in soft, acid waters.     

Interaction of photoperiod and gibberellin on growth and photosynthesis of high-latitude Poa pratensis

The relative growth rate of pot-grown plants of Poa pratensis L. cv. Holt, origin 69s°N, was increased by 20–40% by photoperiod extension with low intensity incandescent light from 8 to 24 h at 9–21°C. The main increase occurred over the 14 to 18 h photoperiod range. The true photoperiodic nature of the response was demonstrated by the effectiveness of night interruption in stimulating growth. Fortnightly sprayings with gibberellic acid (GA₃) (3 × 10^-6 to 3 × 10^-5 M ) mimicked all the effects of long days, whereas (2-chloroethyl)-trimethylammonium chloride (CCC) counteracted the effects of long days. Both growth substances exhibited pronounced interactions with photoperiod, GA₃ being most effective in short days and CCC in long days. The growth stimulation, whether caused by long days or GA₃, was exerted mainly through increases in individual and total leaf area. This was associated with a reduction in CO₂, exchange rate and a parallel fall in specific leaf weight. Proportionally, however, the increase in leaf area was greater than the fall in CO₂ exchange rate, resulting in a 38 to 118% increase in photosynthesis per leaf. No evidence was found of any direct and promotive effect of transition to long days on the CO₂ exchange rate of already expanded leaves.     

Effect of some factors on determination of viable microbial cells by peroxyoxalate chemiluminescence method

The effects of physical and chemical factors on the production of H₂O₂ from Escherichia coli cells were studied. When 20 mmol 1^-1 Tris-HCl buffer was used for this purpose the electron transport system (ETS) showed the highest activity at pH 7.6-8.2. KCN promoted the production of H₂O₂ from E. coli cells, and the optimum concentration was changed in different reaction times and pH values. Glucose, 5 mg ml^-1, increased the ETS activity about twofold. The other substrates and surfactants did not increase the chemiluminescence intensity. NaNO₂ and Na₂SO₄ in inorganic salts significantly reduced the ETS activity above 70%. In addition, the optimum temperature for the production of H₂O₂ was 30°C in this study. When glucose (5 mg ml^-1) and KCN (0.2 mmol 1^-1) were added to the reaction buffer containing 0.5 mmol 1^-1 menadione, the detectable minimum cell densities (averages of triplicate assay) of E. coli, Enterobacter cloacae and Serratia marcescens were 5 times 10³ cells ml^-1, 10⁴ cells ml^-1 and 10⁴ cells ml^-1 respectively.