Na-Ca interactions in barley seedlings: relationship to ion transport and growth

Abstract Salt-stressed plants often show Ca deficiency symptoms. The effects of NaCl salinity (1 to 150 mol m^-3) and supplemental Ca (10 mol m^-3) on Na and Ca transport in barley (Hordeum vulgare L.) and their relationship to growth were investigated. The adjustment of Na and Ca transport was investigated by examining young seedlings exposed to short-term (immediate) and long-term (7 d) exposure to salinity. When the plants were exposed to long-term treatments of salinity, the rate of sodium accumulation in roots was approximately 10 to 15% of short-term treatments. No significant adjustment in the transport to the shoot was observed. Rates of tracer (²²Na) transport were compared to calculated rates based on relative growth rates and tissue element concentrations. Comparisons between measured tracer and calculated rates of transport indicate that ²²Na transport may underestimate transport to the shoot because of dilution of the tracer in the root cytoplasm. Calcium uptake showed only minor adjustment with time. Measured rates of tracer transport to the shoot correlated well with calculated values. The transport and tissue concentrations of Na were significantly affected by supplemental Ca. Calcium transport and tissue concentrations were markedly inhibited by salinity. Supplemental Ca increased Ca transport and accumulation at all NaCl treatments above that of control plants without supplemental Ca. Salinity inhibited plant growth at 150 mol m ^-3NaCl, but not at 75 mol m^-3. Supplemental Ca significantly improved root length but not fresh weight after 7d of salinity, although differences in fresh weight were detected after 9d. There were significant Na-Ca interactions with ion transport, ion accumulation, and growth. The effects of salinity on Na and Ca transport to the shoot do not appear to play a major role in shoot growth of barley.     

Protection of Energy Transfer Process in Isolated Phycobilisome by "White Light" from Ultraviolet-B Induced Damage in the Cyanobacterium Spirulina Platensis

Effect of UV-B (1.9 W m^-2) alone or in combination with supplemental "white light". WL (20 W m^-2) exposure was studied on the energy transfer process of intact phycobilisomes isolated from Spirulina platensis. Exposure of UV-B or supplemental irradiation induced a decrease in room temperature fluorescence intensity and caused a shift towards shorter wavelengths. The low temperature fluorescence measurements showed that UV-B impairs energy transfer from phycocyanin to allophycocyanin B and the extent of damage may be reduced by the exposure to supplemental WL. This revised version was published online in June 2006 with corrections to the Cover Date.     

Inhibition of Vancomycin-Resistant <Emphasis Type="Italic">Enterococcus</Emphasis> by Continuous-Flow Cultures of Human Stool Microflora With and Without Anaerobic Gas Supplementation

A continuous-flow competitive exclusion (CFCE) culture model of human stool microflora was used to examine whether supplemental anaerobic gas is necessary for maintenance of anaerobes and inhibition of vancomycin-resistant Enterococcus (VRE). CFCE cultures of human stool microflora were maintained with supplemental nitrogen, without supplemental nitrogen, or with percolated room air. Cultures with or without supplemental nitrogen maintained >9 log₁₀ CFU mL^–1 of obligate anaerobes and eliminated 10⁶ CFU mL^–1 of VRE. When room air was percolated into the culture, anaerobes were detected at 2 log₁₀ CFU mL^–1, and the same VRE inoculum was not eliminated (P < 0.001). These data demonstrate that human stool CFCE cultures maintain high levels of obligate anaerobes and inhibit VRE without the addition of supplemental anaerobic gas.     

Changes in photosynthesis and antioxidant defenses of Picea asperata seedlings to enhanced ultraviolet-B and to nitrogen supply

The paper mainly studied the effects of ultraviolet‐B (UV‐B) radiation, nitrogen, and their combination on photosynthesis and antioxidant defenses of Picea asperata seedlings. The experimental design included two levels of UV‐B treatments (ambient UV‐B, 11.02 KJ m⁻² day⁻¹; enhanced UV‐B, 14.33 KJ m⁻² day⁻¹) and two nitrogen levels (0; 20 g m⁻² a⁻¹ N) – to determine whether the adverse effects of UV‐B are eased by supplemental nitrogen. Enhanced UV‐B significantly inhibited plant growth, net photosynthetic rate (A), stomatal conductance to water vapor (Gs), transpiration rate and photosynthetic pigment, and increased intercellular CO₂ concentration, UV‐B absorbing compounds, proline content, malondialdehyde (MDA) content, and activity of antioxidant enzymes (peroxidase (POD), superoxide dimutase, and glutathione reductase). Enhanced UV‐B also reduced needle DW and increased hydrogen peroxide (H₂O₂) content and the rate of superoxide radical (O₂⁻) production only under supplemental nitrogen. On the other hand, supplemental nitrogen increased plant growth, A, Gs, chlorophyll content and activity of antioxidant enzymes (POD, ascorbate peroxidase, and catalase), and reduced MDA content, H₂O₂ content, and the rate of O₂⁻ production only under ambient UV‐B, whereas supplemental nitrogen reduced activity of antioxidant enzymes under enhanced UV‐B. Carotenoids content, proline content, and UV‐B absorbing compounds increased under supplemental nitrogen. Moreover, significant UV‐B × nitrogen interaction was found on plant height, basal diameter, A, chlorophyll a, activity of antioxidant enzymes, H₂O₂, MDA, and proline content. These results implied that supplemental nitrogen was favorable for photosynthesis and antioxidant defenses of P.asperata seedlings under ambient UV‐B. However, supplemental nitrogen made the plants more sensitive to enhanced UV‐B, although some antioxidant indexes increased.     

The effects of enhanced ultraviolet-B and nitrogen supply on growth,photosynthesis and nutrient status of <Emphasis Type="Italic">Abies faxoniana</Emphasis> seedlings

Abies faxoniana is a key species in reforestation processes in the southeast of the Qinghai-Tibetan Plateau of China. The changes in growth, photosynthesis and nutrient status of A. faxoniana seedlings exposed to enhanced ultraviolet-B (UV-B), nitrogen supply and their combination were investigated. The experimental design included two levels of UV-B treatments (ambient UV-B, 11.02 KJ m⁻² day⁻¹; enhanced UV-B, 14.33 KJ m⁻² day⁻¹) and two nitrogen levels (0; 20 g N m⁻²). The results indicated that: (1) enhanced UV-B significantly caused a marked decline in growth parameters, net photosynthetic rate (Pn), photosynthetic pigments and F _v/F _m, (2) supplemental nitrogen supply increased the accumulation of total biomass, Pn, photosynthetic pigments and F _v/F _m under ambient UV-B, whereas supplemental nitrogen supply reduced Pn, and not affect biomass under enhanced UV-B, (3) enhanced UV-B or nitrogen supply changed the concentration of nutrient elements of various organs.     

Apparent loss of calcium-activated potassium current in internally perfused snail neurons is due to accumulation of free intracellular calcium

Summary Internal perfusion ofHelix neurons with a solution containing potassium aspartate, MgCl₂, ATP, and HEPES causes the calcium-activated potassium current (I _K(Ca)) evoked by depolarizing voltage steps to decrease with time. When internal free Ca⁺⁺ is strongly buffered to 10^–7 m by including 0.5mm EGTA and 0.225mm CaCl₂ in the internal solution,I _K(Ca) remains constant for up to 3 hours of perfusion. In cells whereI _K(Ca) is small at the start of perfusion, perfusion with the strongly buffered 10^–7 m free Ca⁺⁺ solution produces increases inI _K(Ca) which ultimately saturate. In cells perfused with solutions buffered to 10^–6 m free Ca⁺⁺,I _K(Ca) is low and does not change with perfusion. These results lead us to conclude thatI _K(Ca) is stable in perfusedHelix neurons and that the apparent loss ofI _K(Ca) seen initially with perfusion is due to accumulation of cytoplasmic calcium. Since the calcium current (I _Ca) provides the Ca⁺⁺ which activatesI _K(Ca) during a depolarizing pulse,I _Ca is also stable in perfused cells when free intracellular Ca⁺⁺ is buffered.Perfusion with 1 m calmodulin (CaM) produces no effect onI _K(Ca) with either 10^–7 or 10^–6 m free internal calcium. Inhibiting endogenous CaM by including 50 m trifluoperazine (TFP) in both the bath and the internal perfusion solution also produces no effect onI _K(Ca) with 10^–7 m free internal calciu. It is concluded that CaM plays no role inI _K(Ca) activation.     

CG hypomethylation leads to complex changes in DNA methylation and transpositional burst of diverse transposable elements in callus cultures of rice

CG methylation (^mCG) is essential for preserving genome stability in mammals, but this link remains obscure in plants. OsMET1‐2, a major rice DNA methyltransferase, plays critical roles in maintaining ^mCG in rice. Null mutation of OsMET1‐2 causes massive CG hypomethylation, rendering the mutant suitable to address the role of ^mCG in maintaining genome integrity in plants. Here, we analyzed ^mCG dynamics and genome stability in tissue cultures of OsMET1‐2 homozygous (?/?) and heterozygous (+/?) mutants, and isogenic wild‐type (WT). We found ^mCG levels in cultures of ?/? were substantially lower than in those of WT and +/?, as expected. Unexpectedly, ^mCG levels in 1‐ and 3‐year cultures of ?/? were 77.6% and 48.7% higher, respectively, than in shoot, from which the cultures were initiated, suggesting substantial regain of ^mCG in ?/? cultures, which contrasts to the general trend of ^mCG loss in all WT plant tissue cultures hitherto studied. Transpositional burst of diverse transposable elements (TEs) occurred only in ?/? cultures, although no elevation of genome‐wide mutation rate in the form of single nucleotide polymorphisms was detected. Altogether, our results establish an essential role of ^mCG in retaining TE immobility and hence genome stability in rice and likely in plants in general.     

Selection of sulphur sources for the growth of Butyribacterium methylotrophicum and Acetobacterium woodii

Sulphide and cysteine inhibited growth of batch cultures of Butyribacterium methylotrophicum at moderate concentrations (above 0.5 mM) during growth on glucose (10 mM). The ability of several sulphur sources to replace sulphide was tested in cultures of B. methylotrophicum or Acetobacterium woodii. With sulphite (1 mM), thiosulphate (0.5 mM), elemental sulphur, and dithionite (1 mM), but not sulphate (1 mM), cultures of both organisms grew and produced some sulphide. With elemental sulphur as the sulphur source, toxic levels of sulphide accumulated. Optimal levels for the cultivation of B. methylotrophicum with sulphite were 0.5–2.0 mM, but at higher concentrations the growth rate decreased rapidly, while with dithionite up to 4.0 mM the growth rate was relatively unaffected. In chemostat cultures of B. methylotrophicum with dithionite (1 mM) as the sulphur source and glucose as the limiting substrate, dilution rates up to 0.40 h^–1 were obtained. Thiosulphate could only be used in batch cultures in combination with the reductant titanium(III)nitriloacetate, but in continuous cultures the addition of the reductant to the reservoir was not necessary, because once growth had started enough sulphide was produced to keep the fermentor reduced. The maximum growth rate of B. methylotrophicum with thiosulphate in batch and continuous culture was 0.26 h^–1. Both thiosulphate and dithionite are more convenient sulphur sources than sulphide, but dithionite is more versatile because of its reductive properties and the faster growth it allows.Offprint requests to: T. A. Hansen     

Growth and energy production in Bacteroides amylophilus

The molar growth yield (Y _m) of Bacteroides amylophilus strain WP91 on maltose was 68±2 g/mol when determined from batch cultures at the peaks of maximal growth. Continued incubation led to considerable cell lysis. When calculated from batch cultures in exponential phase (specific growth rate, =0.57 h^-1) Y _m was 101 g/mol. The maximum value of Y _m in maltose-limited chemostat cultures at the maximum dilution rate (D) attainable (D==0.39 h^-1) was about 79 g/mol. Ammonia-Fmited chemostat cultures metabolized maltose with a much reduced efficiency and this was associated with a difference in morphology and chemical composition of the cells. The theoretical maximum molar growth yields (Y _m ^max ) were 55 and 114 g/mol for ammonia- and maltose-limited growth respectively. However, if account was taken of extracellular nitrogen-containing material in ammonia-limited cultures, Y _m ^max became 60. The maintenance coefficient (m _s), estimated from the lines relating the specific rate of maltose consumption (q _m) and D (where m _s=q _m at D=0), was 7.4±0.6×10^-4 mol maltose/g x h for both nutrient limitations. A difference in maintenance energy demand, independent of growth-rate, could not account, therefore, for the observed differences in Y _m between ammonia- and maltose-limited growth.     

Regulation of nitrate uptake by amino acids in maize cell suspension culture and intact roots

The effect of amino acids on nitrate transport was studied in Zea mays cell suspension cultures and in Zea mays excised roots. The inclusion of aspartic acid, arginine, glutamine and glycine (15mM total amino acids) in a complete cell-culture media containing 1.0 mM NO₃ ^- strongly inhibited nitrate uptake and the induction of accelerated uptake rates. The nitrate uptake rate increased sharply once solution amino acid levels fell below detection limits. Glutamine alone inhibited induction in the cell suspension culture. Maize seedlings germinated and grown for 7 days in a 15 mM mixture of amino acids also had lower nitrate uptake rates than seedlings grown in 0.5 mM Ca(NO₃)₂ or 1 mM CaCl₂. As amino acids are the end product of nitrate assimilation, the results suggest an end-product feed-back mechanism for the regulation of nitrate uptake.     

Productivity correlated to photobiochemical performance of <Emphasis Type="Italic">Chlorella</Emphasis> mass cultures grown outdoors in thin-layer cascades

This work aims to: (1) correlate photochemical activity and productivity, (2) characterize the flow pattern of culture layers and (3) determine a range of biomass densities for high productivity of the freshwater microalga Chlorella spp., grown outdoors in thin-layer cascade units. Biomass density, irradiance inside culture, pigment content and productivity were measured in the microalgae cultures. Chlorophyll-fluorescence quenching was monitored in situ (using saturation-pulse method) to estimate photochemical activities. Photobiochemical activities and growth parameters were studied in cultures of biomass density between 1 and 47 g L⁻¹. Fluorescence measurements showed that diluted cultures (1–2 g DW L⁻¹) experienced significant photostress due to inhibition of electron transport in the PSII complex. The highest photochemical activities were achieved in cultures of 6.5–12.5 g DW L⁻¹, which gave a maximum daylight productivity of up to 55 g dry biomass m⁻² day⁻¹. A midday depression of maximum PSII photochemical yield (F _v/F _m) of 20–30% compared with morning values in these cultures proved to be compatible with well-performing cultures. Lower or higher depression of F _v/F _m indicated low-light acclimated or photoinhibited cultures, respectively. A hydrodynamic model of the culture demonstrated highly turbulent flow allowing rapid light/dark cycles (with frequency of 0.5 s⁻¹) which possibly match the turnover of the photosynthetic apparatus. These results are important from a biotechnological point of view for optimisation of growth of outdoor microalgae mass cultures under various climatic conditions.     

The effect of ampicillin plus streptomycin on growth and photosynthesis of two halotolerant chlorophyte algae

Streptomycin and ampicillin are antibiotics commonly used to eliminate prokaryotes from the cultures of eukaryotic algae. We studied the effects of 25 mg l⁻¹ streptomycin plus 50 mg l⁻¹ ampicillin on the growth and photosynthesis of two broadly halotolerant algae, Picochlorum oklahomensis and Dunaliella sp. (Chlorophyceae). We measured growth rate, oxygen evolution, chlorophyll fluorescence kinetics, and pigment content in low (150 μmol photons m⁻² s⁻¹) and high (600 μmol photons m⁻² s⁻¹) light grown batch cultures. Our results show only a minor effect of the antibiotics on P. oklahomensis, and none on Dunaliella sp., so this combination of antibiotics is suitable for maintenance of stock cultures for physiological experiments. We also show that these antibiotics can be used in turbidostat cultures of P. oklahomensis, which otherwise tend to succumb to bacteria.     

Effect of sodium on cellular calcium transport in rat kidney

Summary The influence of extracellular Na (Na _o ) on cellular Ca transport and distribution was studied in rat kidney slices. Calcium efflux from prelabeled slices was depressed when Na _o was completely replaced by choline or tetraethylammonium (TEA) ions and it was markedly stimulated when Na was reintroduced in a Na-free medium. However, reducing Na _o (with choline or TEA as substituting ions) did not increase the total slice⁴⁰Ca, their total exchangeable Ca pool, or the⁴⁰Ca or⁴⁵Ca of mitochondria isolated from these slices. Kinetic analyses of steady-state⁴⁵Ca desaturation curves showed that reducing Na _o depressed the exchange of Ca across the plasma membrane, slightly decreased the cytosolic Ca pool, but did not significantly affect the mitochondrial Ca pool and Ca cycling. Ouabain (10^–3 m) which should reduce the Na gradient across the plasma membrane had no effect on calcium distribution and transport. These results suggest that in kidney cells low Na _o depresses Ca influx as well as Ca efflux; there may be an interaction between Na and Ca at a possible carrier located in the plasma membrane, but there is no Na/Ca exchange as described in several excitable tissues.     

Neurotrophin Protection Against Toxicity Induced by Low Potassium and Nitroprusside in Cultured Cerebellar Granule Neurons

Abstract: Long-term survival of cultured rat cerebellar granule neurons requires depolarizing concentrations of potassium (high potassium; 25 mM KCl). A high-potassium culturing condition has been reported to increase the intracellular calcium concentration ([Ca²⁺]_i) and the expression of brain-derived neurotrophic factor (BDNF), which in turn induces the expression of neurotrophin-3 (NT-3) in these neurons. We therefore examined the neurotrophic effect of these two neurotrophins in low-potassium (5 mM) cultures and their neuroprotective capabilities against sodium nitroprusside-induced neurotoxicity in both low- and high-potassium cultures. Neuronal survival and neurotrophic effects were monitored by [³H]ouabain binding and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. In low-potassium cultures, the neurotrophic effect of BDNF approached that found in high-potassium cultures but was much more robust than that of NT-3. In contrast, undifferentiated neurons cultured in high-potassium medium were much less responsive to BDNF and not responsive at all to NT-3. Induction of nitroprusside neurotoxicity occurred more readily in low- than in high-potassium cultures. BDNF, NT-3, and a high potassium concentration, alone or in combination, were unable to protect neurons treated with nitroprusside at 50 or 100 µM. However, the neurotoxicity of a lower dose of nitroprusside (10 µM) was reversed by the combined actions of these two neurotrophins in low-potassium cultures and by BDNF alone in high-potassium cultures. Because nitroprusside neurotoxicity is less robust in high-potassium cultures, high-potassium-induced BDNF expression and subsequent NT-3 expression may participate in its neuroprotection and neurotrophism in these cultures. Also, we found that toxic doses of nitroprusside antagonized KCl- and NMDA-induced rises in [Ca²⁺]_i, suggesting that this effect is related to nitroprusside-induced neurotoxicity.     

Characterization of the Increase in [Ca2+] i During Hypotonic Shock and the Involvement of Ca2+-activated K+ Channels in the Regulatory Volume Decrease in Human Osteoblast-like Cells

The calcium indicator fura-2 was used to study the effect of hypotonic solutions on the intracellular calcium concentration, [Ca²⁺] _i , in a human osteoblast-like cell line. Decreasing the tonicity of the extracellular solution to 50% leads to an increase in [Ca²⁺] _i from ∼150 nm up to 1.3 μm. This increase in [Ca²⁺] _i was mainly due to an influx of extracellular Ca²⁺ since removing of extracellular Ca²⁺ reduced this increase to ∼250 nm. After cell swelling most of the cells were able to regulate their volume to the initial level within 800 sec. The whole-cell recording mode of the patch-clamp technique was also used to study the effect of an increase in [Ca²⁺] _i on membrane currents in these cells. An increase in [Ca²⁺] _i revealed two types of Ca²⁺-activated K⁺ channels, K(Ca) channels. Current through both channel types could not be observed below voltage of +80 mV with [Ca²⁺] _i buffered to 100 nm or less. With patch-electrodes filled with solutions buffering [Ca²⁺] _i to 10 μm both channels types could be readily observed. The activation of the first type was apparently voltage-independent since current could be observed over the entire voltage range used from −160 to +100 mV. In addition, the current was also blocked by charybdotoxin (CTX). The second type of K(Ca) channels in these cells could be activated with depolarizations more positive than −40 mV from a holding potential of −80 mV. This type was blocked by CTX and paxilline. Adding paxilline to the extracellular solution inhibited regulatory volume decrease (RVD), but could not abolish RVD. We conclude that two K(Ca) channel types exist in human osteoblasts, an intermediate conductance K(Ca) channel and a MaxiK-like K(Ca) channel. MaxiK channels might get activated either directly or by an increase in [Ca²⁺] _i elicited through hypotonic solutions. In combination with the volume-regulated Cl⁻ conductance in the same cells this K⁺ channel seems to play a vital role in volume regulation in human osteoblasts. Received: 8 February 2000/Revised: 13 July 2000     

Using CO2 to enhance carbon capture and biomass applications of freshwater macroalgae

A major limiting factor in the development of algae as a feedstock for the bioenergy industry is the consistent production and supply of biomass. This study is the first to access the suitability of the freshwater macroalgal genus Oedogonium to supply biomass for bioenergy applications. Specifically, we quantified the effect of CO₂ supplementation on the rate of biomass production, carbon capture, and feedstock quality of Oedogonium when cultured in large‐scale outdoor tanks. Oedogonium cultures maintained at a pH of 7.5 through the addition of CO₂ resulted in biomass productivities of 8.33 (±0.51) g DW m^?2 day^?1, which was 2.5 times higher than controls which had an average productivity of 3.37 (±0.75) g DW m^?2 day^?1. Under these productivities, Oedogonium had a carbon content of 41–45% and a higher heating value of 18.5 MJ kg^?1, making it an ideal biomass energy feedstock. The rate of carbon fixation was 1380 g C m^?2 yr^?1 and 1073.1 g C m^?2 yr^?1 for cultures maintained at a pH of 7.5 and 8.5, and 481 g C m^?2 yr^?1 for cultures not supplemented with CO₂. This study highlights the potential of integrating the large‐scale culture of freshwater macroalgae with existing carbon waste streams, for example coal‐fired power stations, both as a tool for carbon sequestration and as an enhanced and sustainable source of bioenergy.     

Distribution of 45Ca in the superior cervical ganglion of the cat

Superior cervical ganglia isolated from immature cats accumulated 0.9 ng atoms of ⁴⁵Ca per mg wet weight during 10-min incubations at 37°C; when expressed as an equivalent volume of medium the accumulation was four times the uptake of ³H-inulin. Orthodromic stimulation of the ganglia doubled ⁴⁵Ca accumulation, whereas excitation with 50 mM KCl, 5 mM glutamate, or antidromic stimulation increased accumulation by one-half. Hexamethonium reduced the increment in ⁴⁵Ca accumulation due to orthodromic stimulation only, but another ganglionic blocking agent, tetraethylammonium, did not reduce accumulation in any case. Both agents blocked ganglionic transmission monitored electrophysiologically. To resolve this discrepancy, and to approach the localization of ⁴⁵Ca within the ganglia, the efflux of previously accumulated ⁴⁵Ca was examined. The data could be fitted by an equation incorporating the sum of three exponentials, representing a rapidly exchanging compartment plus two more slowly exchanging ones. The latter two appeared to reflect the pre- and postganglionic elements in the ganglia: ⁴⁵Ca content of the “preganglionic” compartment was increased by orthodromic but not by antidromic stimulation, and was not decreased by either blocking agent; conversely, ⁴⁵Ca content of the “postganglionic” compartment was increased by both orthodromic and antidromic stimulation, and was decreased by both blocking agents after orthodromic stimulation. The lack of effect of tetraethylammonium on the whole ganglion resulted from an increase in “preganglionic” accumulation that offset the “postganglionic” decrease. After preganglionic denervation, the ⁴⁵Ca content of the “preganglionic” compartment was reduced by two-thirds, while the ⁴⁵Ca content of the “postganglionic” compartment was unchanged. Chemical stimulation increased ⁴⁵Ca accumulation in both compartments. Diphenylhydantoin, 0.1 mM, decreased the increment in ⁴⁵Ca accumulation due to electrical stimulation and to 50 m^M KCl; this inhibition occurred in the “preganglionic” compartment (and perhaps also in the “postganglionic”), and was accompanied by an increased efflux of ⁴⁵Ca.     

Ion accumulation in lucerne plants regenerated from salt-adapted suspension cultures compared with recultured cells from these plants

Lucerne (Medicago media cv. Rambler) plants, regenerated from cell suspension cultures adapted to 200 mm NaCl, and cell cultures derived from the leaflets of regenerated plants showed enhanced Na⁺ exclusion compared with the unselected plants and cell cultures. There was a decrease in the K⁺ concentration in response to NaCl treatment but the selected plants and cell cultures maintained a significantly higher K⁺ concentration and showed much lower Na⁺/K⁺ ratios than unselected plants and cell cultures. The proline concentration increased in the shoots and cell cultures in response to NaCl stress but the selected plants and cell cultures showed a significantly higher proline concentration than unselected plants and cell cultures. Received: 10 December 1996 / Revision received: 28 February 1997 / Accepted: 25 August 1997     

Safrole Analysis by GC-MS of Prototrophic (Ocotea odorifera (Vell.) Rohwer) Cell Cultures

Ocotea odorifera, a tree native to the Atlantic rainforest in south Brazil, has been used as a source of sassafras oil rich in safrole [5-(2-propenyl)-1,3-benzodioxole], an aromatic ether used as a flavoring agent and also in the manufacture of insecticides. The intensive deforestation process of the Brazilian Atlantic rainforest has threatened with extinction some species including O. odorifera. In this context, O. odorifera cell cultures might be an interesting alternative for the production of secondary metabolites of value (i.e., safrole), without risk of damage to the native germplasm. Insights into the secondary metabolites of organosolvent extracts of prototrophic cell cultures were performed by GC-MS and MALDI-TOF MS. GC-MS analysis revealed the occurrence of safrole in concentration of ca. 62.6 μg ml⁻¹ and allowed the identification of other compounds besides safrole in the crude extract of these cultures. Thus, alkyl phenol (C₈) [m/z= 206], C₁₄ myristic acid [m/z= 228], long chain olefin or alcohol and essential oil were detected. However, safrole [m/z= 162] was not detected by MALDI-TOF MS, indicating that it is not easily protonated (M + H)⁺.     

Influence of ultraviolet-B (UV-B) radiation on photosynthetic and growth characteristics in field-grown cassava (Manihot esculentum Crantz)

The effects of ultraviolet-B (UV-B between 290 and 320 nm) on photosynthesis and growth characteristics were investigated in field grown cassava (Manihot esculentum Crantz). Plants were grown at ambient and ambient plus a 5.5kJ m^?2 d^?1 supplementation of UV-B radiation for 95 d. The supplemental UV-B fluence used in this experiment simulated a 15% depletion in stratospheric ozone at the equator (0°N). Carbon dioxide exchange, oxygen evolution, and the ratio of variable to maximum fluorescence (F_v/F_m) were determined for fully expanded leaves after 64–76 d of UV-B exposure. AH plants were harvested after 95 d of UV-B exposure, assayed for chlorophyll and UV-B absorbing compounds, and separated into leaves, petioles, stems and roots. Exposure to UV-B radiation had no effect on in situ rates of photosynthesis or dark respiration. No difference in the concentration of UV-B absorbing compounds was observed between treatments. A 2-d daytime diurnal comparison of F_v to F_m ratios indicated a significant decline in F_v/F_m ratios and a subsequent increase in photoinhibition under enhanced UV-B radiation if temperature or PPF exceeded 35°C or 1800μmol m^?2 s^?1, respectively. However, UV-B effects on fluorescence kinetics appeared to be temporal since maximal photosynthetic rates as determined by oxygen evolution at saturated CO₂ and PPF remained unchanged. Although total biomass was unaltered with UV-B exposure, alterations in the growth characteristics of cassava grown with supplemental UV-B radiation are consistent with auxin destruction and reduced apical dominance. Changes in growth included an alteration of biomass partitioning with a significant increase in shoot/root ratio noted for plants receiving supplemental UV-B radiation. The increase in shoot/root ratio was due primarily to a significant decrease in root weight (–32%) with UV-B exposure. Because root production determines the harvest-able portion of cassava, UV-B radiation may still influence the yield of an important tropical agronomic species, even though photosynthesis and total dry biomass may not be directly affected.