Energieabhängige 63Ni-Aufnahme bei Alcaligenes eutrophus Stamm H 1 und H 16

Zusammenfassung Die Kinetik der Aufnahme von ⁶³Ni wurde an zwei Stämmen von Alcaligenes eutrophus untersucht, die Nickelionen für das chemolithotrophe Wachstum benötigen. Mit Kohlendioxid als einziger Kohlenstoffquelle wird das Wachstum durch niedrige Konzentrationen von Nickel gefördert, wobei das Optimum der Wachstumsförderung bei 0,3 M Nickel lag. Höhere Nickelkonzentrationen wirkten hemmend. Das heterotrophe Wachstum mit Fructose wurde durch Nickelionen nicht gefördert. — Übertragen in Phosphatpuffer, der von Schwermetallionen befreit worden war, zeigten autotroph gewachsene Zellen eine rasche Aufnahme von ⁶³Ni, sofern Wasserstoff, Sauerstoff und Kohlendioxid zugegen waren. Dabei wurde Nickel innerhalb von 60 min aus dem umgebenden Medium bis zur 280 fachen Konzentration in den Zellen angehäuft. Die beobachtete Ni-Aufnahme zeigte ein Temperaturoptimum bei etwa 29° C und wurde durch Hemmstoffe wie Arsenit, Jodacetat, Methylenblau, Natriumazid und Natriumcyanid stark beeinträchtigt. Andere Schwermetallionen (Zn, Co, Mn und Cu) verminderten die Nickelaufnahme nur geringfügig. Durch ⁵⁸NiCl₂ und Toluol wurde der Efflux von ⁶³Ni aus den Zellen gefördert. Die Beobachtungen lassen darauf schließen, daß Nickelionen durch einen energieabhängigen Prozeß in chemolithotroph gewachsenen Zellen dieser Stämme angehäuft werden.

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Energy-dependent ⁶³Ni-uptake by Alcaligenes eutrophus strains H 1 and H 16

Kinetic studies of the uptake of ⁶³Ni were undertaken with two strains of Alcaligenes eutrophus, known to require nickel ions for chemolithotrophic growth. Using carbon dioxide as sole carbon source, growth is stimulated by low concentrations of nickel with optimum concentration for growth stimulation at about 0.3 M nickel. Higher nickel concentrations were inhibitory. Heterotrophic growth on fructose was not stimulated by nickel ions.-Upon transfer into phosphate buffer freed of heavy metal ions, autotrophically grown cells exhibited rapid uptake of ⁶³Ni which was dependent upon th presence of hydrogen, oxygen and carbon dioxide. Within 60 min nickel was accumulated from the medium, reaching 280-fold concentration in the cells. The observed uptake exhibited a temperature optimum at about 29° C and was markedly inhibited by metabolic inhibitors such as arsenite, iodoacetate, methylene-blue, sodium azide and sodium cyanide. Other heavy metal ions (Zn, Co, Mn and Cu) only slightly inhibited ⁶³Ni-uptake. The efflux of ⁶³Ni from the cells was stimulated by ⁵⁸NiCl₂ and by toluene. These data indicate that nickel ions are accumulated by an energy dependent mechanism in chemolithotrophically grown cells of these strains.

     

Uptake and release of63Ni2+ byXenopus embryos during early cleavage stages

Summary Uptake and release of⁶³Ni was studied in dejelliedXenopus laevis embryos exposed to⁶³Ni²⁺ (0.3–30 mol/1) for 0.5-h intervals during the period 1–4.5 h post-fertilization (i.e. from first cleavage to early blastula stage). At first cleavage, the mean uptake of⁶³Ni by embryos was 12-17 times that by non-fertilized eggs, suggesting that conversion of the vitelline envelope to the fertilization envelope enhanced integumental permeability to⁶³Ni²⁺. 63 Ni uptake by embryos at the 1-2-cell stage averaged 1.8–2.5 times that at the early blastula stage. An average of 5% of total⁶³Ni in washed embryos was recovered in isolated fertilization envelopes, indicating that⁶³Ni²⁺ passed through the envelope into internal compartments. Progressive increases of⁶³Ni uptake were seen with increasing exposure levels; after exposure during 1–1.5 h post-fertilization to the highest concentration of⁶³Ni²⁺ (30 mol/1),⁶³Ni uptake averaged 11.4 (SD±5.1) pmol/embryo. Rapid efflux of⁶³Ni was noted after⁶³Ni²⁺-exposed embryos were transferred to nickel-free medium; mean⁶³Ni contents at 0.25 h and 2 h post-exposure diminished to 50% and 15% of the initial values, regardless of the exposure level. The finding thatXenopus embryos are permeable to⁶³Ni²⁺ during early cleavage stages provides a convenient experimental system to investigate the embryotoxicity and teratogenicity of nickel.     

Nickel dependence of factor F430 content in Methanobacterium thermoautotrophicum

Factor F₄₃₀ is a yellow compound of unknown structure present in methanogenic bacteria. It has recently been shown to contain nickel. In this communication the influence of the nickel concentration in the growth medium on the factor F₄₃₀ content of Methanobacterium thermoautotrophicum and on the nickel content of factor F₄₃₀ was studied. It was found: (1) The content of factor F₄₃₀ in the cells was strongly dependent on the nickel concentration of the growth medium. Cells grown on media with 2.5 M NiCl₂ contained 28 times as much factor F₄₃₀ per g as those grown on media with 0.075 M NiCl₂; (2) factor F₄₃₀ was synthesized in nickel deprived cells only upon the addition of nickel Nickel uptake paralleled factor F₄₃₀ synthesis; (3) independent of the nickel concentration in the growth medium, the extinction coefficient at 430 nm of factor F₄₃₀ per mol nickel was always near 22,500 cm^-1 (mol Ni)^-1. These findings indicate that nickel is an essential component of factor F₄₃₀.Dedicated to Professor Otto Kandler on the occasion of his 60th birthday     

15N natural abundance studies in Australian commercial sugarcane

The measurement of natural ¹⁵N abundance is a well-established technique for the identification and quantification of biological N₂ fixation in plants. Associative N₂ fixing bacteria have been isolated from sugarcane and reported to contribute potentially significant amounts of N to plant growth and development. It has not been established whether Australian commercial sugarcane receives significant input from biological N₂ fixation, even though high populations of N₂ fixing bacteria have been isolated from Australian commercial sugarcane fields and plants. In this study, ¹⁵N measurements were used as a primary measure to identify whether Australian commercial sugarcane was obtaining significant inputs of N via biological N₂ fixation. Quantification of N input, via biological N₂ fixation, was not possible since suitable non-N₂ fixing reference plants were not present in commercial cane fields. The survey of Australian commercially grown sugarcane crops showed the majority had positive leaf ¹⁵N values (73% >3.00, 63% of which were >5.00), which was not indicative of biological N₂ fixation being the major source of N for these crops. However, a small number of sites had low or negative leaf ¹⁵N values. These crops had received high N fertiliser applications in the weeks prior to sampling. Two possible pathways that could result in low ¹⁵N values for sugarcane leaves (other than N₂ fixation) are proposed; high external N concentrations and foliar uptake of volatilised NH₃. The leaf ¹⁵N value of sugarcane grown in aerated solution culture was shown to decrease by approximately 5 with increasing external N concentration (0.5–8.0 mM), with both NO₃ ^– and NH₄ ⁺ nitrogen forms. Foliar uptake of atmospheric NH₃ has been shown to result in depleted leaf ¹⁵N values in many plant species. Acid traps collected atmospheric N with negative ¹⁵N value (–24.45±0.90) from above a field recently surface fertilised with urea. The ¹⁵N of leaves of sugarcane plants either growing directly in the soil or isolated from soil in pots dropped by 3.00 in the same field after the fertiliser application. Both the high concentration of external N in the root zone (following the application of N-fertilisers) and/or subsequent foliar uptake of volatilised NH₃ could have caused the depleted leaf ¹⁵N values measured in the sugarcane crops at these sites.     

Nickel,a component of factor F 430 from Methanobacterium thermoautotrophicum

Methanobacterium thermoautotrophicum, growing on medium supplemented with 2 mol ⁶³NiCl₂/l, was found to take up 1.2 mol ⁶³Ni per g cells (dry weight). More than 70% of the radioisotope was incorporated into a compound, which dissociated from the protein fraction after heat treatment, was soluble in 70% acetone, and could be purified by chromatography on QAE-Sephadex A-25, Sephadex G-25, and DEAE cellulose. The purified ⁶³Ni labelled compound had an absorption spectrum and properties identical to those of factor F ₄₃₀ and is therefore considered to be identical with factor F ₄₃₀.Factor F ₄₃₀, a compound of molecular weight higher than 1000 with an absorbance maximum at 430 nm, has recently been purified from Methanobacterium thermoautotrophicum (Gunsalus and Wolfe, 1978). The structure and function of this compound are not yet known.     

Kinetic study of a thermostable beta-glycosidase of Thermus thermophilus. Effects of temperature and glucose on hydrolysis and transglycosylation reactions

A -glycosidase of a thermophile, Thermus thermophilus, belonging to the glycoside hydrolase family 1, was cloned and overexpressed in Escherichia coli. The purified enzyme (Ttgly) has a broad substrate specificity towards -D-glucoside, -D-galactoside and -D-fucoside derivatives. The thermostability of Ttgly was exploited to study its kinetic properties within the range 25–80[emsp4 ]°C. Whatever the temperature, except around 60[emsp4 ]°C, the enzyme displayed non-Michaelian kinetic behavior. Ttgly was inhibited by high concentrations of substrate below 60[emsp4 ]°C and was activated by high concentrations of substrate above 60[emsp4 ]°C. The apparent kinetic parameters (k _cat and K _m ) were calculated at different temperatures. Both k _cat and K _m increased with an increase in temperature, but up to 75[emsp4 ]°C the values of k _cat increased much more rapidly than the values of K _m . The observed kinetics might be due to a combination of factors including inhibition by excess substrate and stimulation due to transglycosylation reactions. Our results show that the substrate could act not only as a glycosyl donor but also as a glycosyl acceptor. In addition, when the glucose was added to reaction mixtures, inhibition or activation was observed depending on both substrate concentration and temperature. A reaction model is proposed to explain the kinetic behavior of Ttgly. The scheme integrates the inhibition observed at high concentrations of substrate and the activation due to transglycosylation reactions implicating the existence of a transfer subsite.     

A novel substrate for yeast alcohol dehydrogenase – p-nitroso-N,N-dimethylaniline

Yeast alcohol dehydrogenase (EC 1.1.1.1) catalyzes the novel reduction of p-nitro-so-N,N-dimethylaniline with NADH as a cofactor. Apparent kinetic constants for this enzymatic reaction are: V ₂=2.1 s^–1, K _Q=456 M, K _iQ=119 M, and K _P=1.47 mM, at pH 8.9, 25 °C. This reaction is especially useful for the quantitative determination of NAD⁺ and NADH by enzymatic cycling.     

Inhibitor effects on the accumulation and efflux of nickel ions in plasmid pMOL28-harboring strains of Alcaligenes eutrophus

The accumulation and efflux of ⁶³Ni²⁺ ions were studied using two strains of the strictly respiratory bacterium Alcaligenes eutrophus, the wild type strain N9A and its transconjugant N9A-M243 which harbors plasmid pMOL28.1 encoding constitutive resistance to nickel. When 1 M ⁶³Ni²⁺ is added to respiring cells, strain N9A accumulates high, but M243 only negligibly small amounts of nickel. When the cells were preincubated for about 20 h under anoxic conditions and were then exposed to 1 M ⁶³Ni²⁺ anaerobically, both strains accumulated approximately the same amounts of nickel. Aeration of these preloaded cells resulted in rapid efflux by strain M243 but renewed uptake of nickel by N9A. ⁶³Ni²⁺ uptake and efflux are highly sensitive to protonophores such as CCCP, FCCP and TCS but insensitive to DCCD (each at 20 M concentrations). Measurements on the effects of the inhibitors on biosynthetic processes requiring ATP either from substrate phosphorylation or from electron transport phosphorylation made sure that at the concentrations used the inhibitor effects were specific. Thus the results suggest that in the nickelresistant strain M243 under normal aerobic conditions two constitutive energy-dependent nickel transport systems can function concomitantly, a chromosomally-determined specific uptake system and a plasmid-mediated specific nickel efflux system.     

Isolation and characterization of a toxic intracellular protein from Vibrio parahaemolyticus

A toxic factor released from disrupted cells of Vibrio parahaemolyticus was partially purified by gel filtration after precipitation with (NH₄)₂SO₄ at 40% saturation. The factor, which was a thermostable protein of 63 kDa, lysed human erythrocytes at a concentration of 0.15 g ml^-1. Its LD₅₀ by intravenous injection into mice was 6.4 g. Fluid accumulated in suckling mice force-fed with the toxic material (1 to 25 g). Haemolytic activity, which occurred maximall at 37°C and pH 7.0 was enhanced by Ca²⁺, Cu²⁺ and Zn²⁺, each at 1 mm. Anti-toxic-factor serum agglutinated V. parahaemolyticus cells. The factor may play a role in the pathogenesis of V. parahaemolyticus infections and in the host's defence mechanisms against infection by the microorganism.     

Biospeciation, by potentiometry and computer simulation, of Sm-EDTMP, a bone tumor palliative agent

¹⁵³Sm-EDTMP (ethylenediaminetetra(methylenephosphonic) acid) is of considerable interest as a bone therapeutic radiopharmaceutical but its properties in solution are not yet well characterized. The protonation constants of EDTMP and the formation constants of the complexes of Sm-EDTMP have accordingly been measured potentiometrically by glass electrode titrations at 25°C in 0.15 M NaCl. Six protonation constants (log ₀₁₁ = 9.638, log ₀₁₂ = 17.330, log ₀₁₃ = 23.597, log ₀₁₄ = 28.636, log ₀₁₅ = 31.501, log ₀₁₆ = 32.624) and the formation constants of the [Sm(EDTMP)H_-1]^6- (log _11-1 = 4.865), [SmEDTMP]^5- (log ₁₁₀ = 12.018), [Sm(EDTMP)H]^4- (log ₁₁₁ = 17.892) and [Sm(EDTMP)H₂]^3- (log ₁₁₂ = 23.437) complexes were determined. Computer simulations indicate that the [SmEDTMP]^5- and the hydroxy [Sm(EDTMP)H_-1]^6- species are the major Sm(III) complexes formed in blood plasma, which explains the high degree of localization in the kidney and urine observed in biodistribution studies. Calcium ions are probably the maior competitor for EDTMP in blood plasma. As the presence of secondary skeletal metastases results in a high rate of bone turnover, it is possible that the high concentration of calcium at these sites encourages localization of ¹⁵³Sm-EDTMP.     

Nickel-content of urease from Bacillus pasteurii

Urease from Bacillus pasteurii DSM 33 was purified 34-fold to a maximum specific activity of 996.5 mol urea min^-1 mg^-1 at 30°C. Homogeneity was demonstrated by isoelectric focussing which showed a single protein zone corresponding to a pI of about 4.6. The native enzyme was demonstrated to have a molecular mass of 230000 and to consist of identical subunits of 65 500, as measured by SDS electrophoresis. Radioactive ⁶³Ni-nickel co-chromatographed with urease through gel filtration, ion-exchange, and affinity chromatography. Measuring specific radioactivity, the nickel content was found to be 1.00 (±0.1) g-atom Ni per mol of subunit, and 0.82 g-atom Ni per mol as measured by atomic absorption spectrometry. This indicates that 1 atom of nickel is present in each of four subunits of the enzyme.Non-standard abbreviations SDS sodium dodecyl sulfate     

The CO2/O2 specificity of ribulose 1,5-bisphosphate carboxylase/oxygenase

The substrate specificity factor, V _cK_o/V_oK_c, of spinach (Spinacia oleracea L.) ribulose 1,5-bisphosphate carboxylase/oxygenase was determined at ribulosebisphosphate concentrations between 0.63 and 200 M, at pH values between 7.4 and 8.9, and at temperatures in the range of 5° C to 40° C. The CO₂/O₂ specificity was the same at all ribulosebisphosphate concentrations and largely independent of pH. With increasing temperature, the specificity decreased from values of about 160 at 5° C to about 50 at 40° C. The primary effects of temperature were on K _c [Km(CO₂)] and V _c [Vmax (CO₂)], which increased by factors of about 10 and 20, respectively, over the temperature range examined. In contrast, K _o [Ki (O₂)] was unchanged and V _o [Vmax (O₂)] increased by a factor of 5 over these temperatures. The CO₂ compensation concentrations () were calculated from specificity values obtained at temperatures between 5° C and 40° C, and were compared with literature values of . Quantitative agreement was found for the calculated and measured values. The observations reported here indicate that the temperature response of ribulose 1,5-bisphosphate carboxylase/oxygenase kinetic parameters accounts for two-thirds of the temperature dependence of the photorespiration/photosynthesis ratio in C₃ plants, with the remaining one-third the consequence of differential temperature effects on the solubilities of CO₂ and O₂.Abbreviations RuBPC/O(ase) ribulose 1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose 1,5-bisphosphate - CO₂ compensation concentration     

Evidence for carrier-mediated,energy-dependent uptake of urea in some bacteria

Evidence for the existence of an energy-dependent urea permease was found for Alcaligenes eutrophus H16 and Klebsiella pneumoniae M5a1 by studying uptake of ¹⁴C-urea. Since intracellular urea was metabolized immediately, uptake did not result in formation of an urea pool. Evidence is based on observations that the in vivo urea uptake and in vitro urease activity differ significantly with respect to kinetic parameters, temperature optimum, pH optimum, response towards inhibitors and regulation. The K _m for urea uptake was 15–20 times lower (38 M and 13 M urea for A. eutrophus and K. pneumoniae, respectively) than the K _m of urease for urea (650 M and 280 M urea), the activity optimum for A. eutrophus was at pH 6.0 and 35°C for the uptake and pH 9.0 and 65°C for urease. Uptake but not urease activity in both organisms strongly decreased upon addition of inhibitors of energy metabolism, while in K. pneumoniae, potent inhibitors of urease (thiourea and hydroxyurea) did not affect the uptake process. Significant differences in the uptake rates were observed during growth with different nitrogen sources (ammonia, nitrate, urea) or in the absence of a nitrogen source; this suggested that a carrier is involved which is subject to nitrogen control. Some evidence for the presence of an energy-dependent uptake of urea was also obtained in Pseudomonas aeruginosa DSM 50071 and Providencia rettgeri DSM 1131, but not in Proteus vulgaris DSM 30118 and Bacillus pasteurii DSM 33.Non-standard abbreviations CCCP Carbonylcyanide-m-chlorphenylhydrazone - DCCD dicyclohexylcarbodiimide - DNP 2,4-dinitrophenole     

Effect of<Emphasis Type="Italic"> p</Emphasis>CO<Subscript>2</Subscript> and temperature on the boron isotopic composition of the zooxanthellate coral<Emphasis Type="Italic"> Acropora</Emphasis> sp.

Culture experiments were carried out with Acropora sp. (a branching scleractinian coral) in seawater at two pCO₂ conditions (438 and 725 µatm) and two temperatures (25 and 28 °C) in order to establish the pH and temperature dependence of the boron isotopic composition of the skeleton. A clear pCO₂ effect, but no temperature effect, on the coral boron isotope composition is seen. For corals cultured at normal pCO₂ (438 µatm), the ¹¹B of the skeleton was 24.0±0.2 at 25 °C, and 23.9±0.3 at 28 °C. The values of ¹¹B measured for corals cultured at higher pCO₂ (725 µatm) were lower: 22.5±0.1, and 22.8±0.1 at 25 and 28 °C, respectively. The ¹¹B of corals cultivated at both high and normal pCO₂ conditions are consistent with a dominant pH control, and are very close to that calculated from theoretical considerations. Thus, the corals do not seem to significantly alter ambient seawater for calcification with respect to pH. Co-variation between boron and carbon isotope values is explored.Communicated by: Guest Editor A. Grottoli     

Photosynthetic formation of inorganic pyrophosphate in phototrophic bacteria

In this paper we report studies on photosynthetic formation of inorganic pyrophosphate (PP_i) in three phototrophic bacteria. Formation of PP_i was found in chromatophores from Rhodopseudomonas viridis but not in chromatophores from Rhodopseudomonas blastica and Rhodobacter capsulatus. The maximal rate of PP_i synthesis in Rps. viridis was 0.15 mol PP_i formed/(min*mol Bacteriochlorophyll) at 23°C. The synthesis of PP_i was inhibited by electron transport inhibitors, uncouplers and fluoride, but was insensitive to oligomycin and venturicidin. The steady state rate of PP_i synthesis under continuous illumination was about 15% of the steady-state rate of ATP synthesis. The synthesis of PP_i after short light flashes was also studied. The yield of PP_i after a single 1 ms flash was equivalent to approximately 1 mol PP_i/500 mol Bacteriochlorophyll. In Rps. viridis chromatophores, PP_i was also found to induce a membrane potential, which was sensitive to carbonyl cyanide p-trifluoromethoxyphenylhydrazone and NaF.Abbreviations BChl Bacteriochlorophyll - F₀F₁-ATPase Membrane bound proton translocating ATP synthase - FCCP Carbonyl cyanide p-trifluoromethoxyphenylhydrazone - H⁺-PPase Membrane bound proton translocating PP_i synthase - TPP⁺ Tetraphenyl phosphonium ion - TPB^- Tetraphenyl boron ion - Transmembrane electrical potential difference     

Thiamin transport by human erythrocytes and ghosts

Summary Thiamin transport in human erythrocytes and resealed pink ghosts was evaluated by incubating both preparations at 37 or 20°C in the presence of [³H]-thiamin of high specific activity. The rate of uptake was consistently higher in erythrocytes than in ghosts. In both preparations, the time course of uptake was independent from the presence of Na⁺ and did not reach equilibrium after 60 min incubation. At concentrations below 0.5 m and at 37°C, thiamin was taken up predominantly by a saturable mechanism in both erythrocytes and ghosts. Apparent kinetic constants were: for erythrocytes,K _m =0.12, 0.11 and 0.10 m andJ _max=0.01, 0.02 and 0.03 pmol·l^–1 intracellular water after 3, 15, and 30 min incubation times, respectively; for ghosts,K _m =0.16 and 0.51 m andJ _max=0.01 and 0.04 pmol·l^–1 intracellular water after 15 and 30 min incubation times, respectively. At 20°C, the saturable component disappeared in both preparations. Erythrocyte thiamin transport was not influenced by the presence ofd-glucose or metabolic inhibitors. In both preparations, thiamin transport was inhibited competitively by unlabeled thiamin, pyrithiamin, amprolium and, to a lesser extent, oxythiamin, the inhibiting effect being always more marked in erythrocytes than in ghosts. Only approximately 20% of the thiamin taken up by erythrocytes was protein-(probably membrane-) bound. A similar proportion was esterified to thiamin pyrophosphate. Separate experiments using valinomycin and SCN^– showed that the transport of thiamin, which is a cation at pH 7.4, is unaffected by changes in membrane potential in both preparations.     

Purification and properties of a hyperthermoactive α-amylase from the archaeobacterium Pyrococcus woesei

The cultivation of the hyperthermophilic archaeobacterium Pyrococcus woesei on starch under continuous gassing (80% H₂:20% CO₂) caused the formation of 250 U/l of an extremely thermoactive and thermostable -amylase. In a complex medium without elemental sulphur under 80% N₂ and 20% CO₂ atmosphere enzyme production could be elevated up to 1000 U/l. Pyrococcus woesei grew preferentially on poly-and oligosaccharides. The amylolytic enzyme formation was constitutive. Enzyme production was also observed in continuous culture at dilution rates from 0.1 to 0.4 h^-1. A 20-fold enrichment of -amylase was achieved after adsorption of the enzyme onto starch and its desorption by preparative gel electrophoresis. The -amylase consisted of a single subunit with a molecular mass of 70 000 and was catalytically active at a temperature range between 40°C and 130°C. Enzymatic activity was detected even after autoclaving at a pressure of 2 bars at 120°C for 5 h. The purified enzyme hydrolyzed exclusively -1,4-glycosidic linkages present in glucose polymers of various sizes. Unlike many -amylases from anaerobes the enzyme from P. woesei was unable to attack short chain oligosaccharides with a chain length between 2 and 6 glucose units.     

Some observations on the ecology,metal uptake and nickel tolerance of Alyssum serpyllifolium subspecies from the Iberian peninsula

Experiments were carried out on the tolerance to, and uptake of, nickel by three iberian subspecies of Alyssum serpylliforium Desf. Two of these subspecies, the serpentine-endemic ssp. lusitanicum from Bragança, Portugal and ssp. malacitanum from Málaga, Spain, are hyperaccumulators (> 1000 g/g in dried material) of nickel. Their possible ancestor, ssp. serpyllifolium (from Granada, Spain) was a non-accumulator of this element. Seeds of the two serpentine-endemics germinated extensively in nickel concentrations up to 12 000 g/g (1.2%) whereas ssp. serpyllifolium only germinated in nickel concentrations below 60 g/ml. Tolerance tests involving measurement of new root lengths of excised seedlings placed in varying nickel concentrations, again showed much greater tolerance of the two serpentinophytes. In both series of experiments, the order of tolerance was: ssp. lusitanicum > ssp. malacitanum > ssp. serpyllifolium. In pot trials involving seedlings of ssp. malacitanum grown in mixtures containing varying amounts of calcium, magnesium, and nickel, the most important finding was that plants will tolerate higher nickel contents in the soil when excess calcium is added. This is achieved by lowering the uptake of nickel. There appeared to be some concomitant reduction in calcium uptake in the presence of nickel, and some increase in uptake of magnesium. The resultant lower calcium/magnesium ratio in the plant, though not symptomatic of a favourable condition for colonization of serpentine soils, probably results from a mechanism which renders nickel innocuous to the plant at the expense of calcium uptake. It is suggested that the physiological characters of ssp. lusitanicum and ssp malacitanum are sufficiently different to support arguments for promoting the latter to full specific rank as has now been done for ssp. lusitanicum.     

Effects of light on nitrate-limited Oscillatoria agardhii in chemostat cultures

The effects of the average light irradiance (I) on growth and nitrate uptake kinetics of the cyanobacterium Oscillatoria agardhii, in nitrate-limited chemostat cultures, were studied. Light was nonsaturating for I <9.4 Wm^–2, for all growth rates () studied. However, was throughout limited by the availability of nitrate. Under light-saturating conditions the kinetics of nitrate-limited growth could be adequately described by both the Monod and Droop equations. Under light-non-saturating conditions the internal nitrogen content (Q) was a function of both and I, for which new formulas were derived. The high uptake capacity (V _max) of nitrate-limited cells was independent of , but was significantly increased for cells growing at I <9.4 Wm^–2. The half-saturation constant for nitrate uptake (K _s ^u ) increased with increasing , but was independent of the prevailing light conditions. The effects of light during nitrate-limited growth were associated with the regulation in the nitrogen-containing pigments.The results reported herein have important consequences for the use of Q, K _s ^u and V _max values as indicators of nutrient-deficiency of natural populations.     

Incorporation of 14CO2 in prenylquinones of Chlorella pyrenoidosa

Incorporation and release of ¹⁴C-label in prenylquinones of Chlorella was investigated under steady state conditions. After one hour of ¹⁴CO₂-photosynthesis all plastid quinones investigated were labeled. The highest label was found in phylloquinone (18%) while -tocopherol exhibits the lowest label (0.38%). Among the plastoquinones, plastohydroquinone-9 shows a higher labeling degree (5.1%) and a faster labeling kinetic than plastoquinone-9 (1.6%). After replacement of ¹⁴CO₂ against ¹²CO₂ the total radioactivity in plastohydroquinone-9, -tocopherol and phylloquinone decreases but in -tocoquinone and plastoquinone-9 proceeds further. From this labeling kinetic we conclude, that newly synthesized [¹⁴C]-tocopherol molecules are converted to [¹⁴C]-tocoquinone and [¹⁴C]plastohydroquinone-9 molecules to [¹⁴C]plastoquinone-9. From their ¹⁴C-incorporation kinetic half-lives could be calculated for all prenylquinones in the same ranges as previously found for the chlorophylls and carotenoids (Grumbach et al., 1978). Half-lives are shorter in plastohydroquinone-9 (30 min) and plastoquinone-9 (40 min) than in phylloquinone (55 min), -tocoquinone (50 min) and -tocopherol (220 min). This means that all prenyl-lipids such as chlorophyll a, -and -carotene, plastohydroquinone-9 and plastoquinone-9 which are more directly involved in the process of photosynthesis are subject to a continuous and higher turnover than the xanthophyll and -tocopherol. From the fast labeling kinetic and short half-lives of -tocoquinone and especially phylloquinone with a labeling degree of 12% after one hour of ¹⁴CO₂ photosynthesis we suppose that perhaps these two prenylquinones are also involved in the photosynthetic activity of chloroplasts.