Effects of oxygen, pH and nitrate concentration on denitrification by Pseudomonas species

Abstract The production of nitrogen-containing gases by denitrification in three organisms was examined using membrane inlet mass spectrometry. The effects of O₂ (during both growth and maintenance) and of pH, nitrate concentration and carbon source were tested in non-proliferating cell suspensions. Two strains of Pseudomonas aeruginosa were capable of co-respiration of NO₃⁻ and O₂ and, under controlled O₂ supply, gave oscillatory denitrification. Variations in culture and assay conditions affected both the rate of denitrification and the ratio of end products (N₂O:N₂). Higher rates were seen following anaerobic growth. Optimum values of pH and nitrate concentration for denitrification are given. Generally, the optimum pH was 7.0–7.5, approximately that of the growth medium. Optimum nitrate concentration was generally 20 mM.     

Compartmentation of triacylglycerol accumulation in plants

Triacylglycerols from plants, familiar to most people as vegetable oils, supply 25% of dietary calories to the developed world and are increasingly a source for renewable biomaterials and fuels. Demand for vegetable oils will double by 2030, which can be met only by increased oil production. Triacylglycerol synthesis is accomplished through the coordinate action of multiple pathways in multiple subcellular compartments. Recent information has revealed an underappreciated complexity in pathways for synthesis and accumulation of this important energy-rich class of molecules.     

Nitrate removal in a combined bioelectrochemical and sulfur autotrophic denitrification system under high nitrate concentration: effects of pH

Bioprocess and Biosystems Engineering - A combined bioelectrochemical and sulfur autotrophic denitrification (CBSAD) system was established to treat high concentration nitrate under different pH...     

Development of accelerated net nitrate uptake : effects of nitrate concentration and exposure time

Upon initial nitrate exposure, net nitrate uptake rates in roots of a wide variety of plants accelerate within 6 to 8 hours to substantially greater rates. Effects of solution nitrate concentrations and short pulses of nitrate (≤1 hour) upon `nitrate-induced' acceleration of nitrate uptake in maize (Zea mays L.) were determined. Root cultures of dark-grown seedlings, grown without nitrate, were exposed to 250 micromolar nitrate for 0.25 to 1 hour or to various solution nitrate concentrations (10-250 micromolar) for 1 hour before returning them to a nitrate-free solution. Net nitrate uptake rates were assayed at various periods following nitrate exposure and compared to rates of roots grown either in the absence of nitrate (CaSO₄-grown) or with continuous nitrate for at least 20 hours. Three hours after initial nitrate exposure, nitrate pulse treatments increased nitrate uptake rates three- to four-fold compared to the rates of CaSO₄-grown roots. When cycloheximide (5 micrograms per milliliter) was included during a 1-hour pulse with 250 micromolar nitrate, development of the accelerated nitrate uptake state was delayed. Otherwise, nitrate uptake rates reached maximum values within 6 hours before declining. Maximum rates, however, were significantly less than those of roots exposed continuously for 20, 32, or 44 hours. Pulsing for only 0.25 hour with 250 micromolar nitrate and for 1 hour with 10 micromolar caused acceleration of nitrate uptake, but the rates attained were either less than or not sustained for a duration comparable to those of roots pulsed for 1 hour with 250 micromolar nitrate. These results indicate that substantial development of the nitrate-induced accelerated nitrate uptake state can be achieved by small endogenous accumulations of nitrate, which appear to moderate the activity or level of root nitrate uptake.     

Combined effect of light and temperature on triacylglycerol accumulation in Dicranum elongatum

In the subarctic moss Dicranum elongatum Schleich & Schwaegr., the level of total lipids and triacylglycerols (TAG) was high in late winter and spring and low in autumn and winter. Four-week exposure of field material to continuous light (135μmol m⁻²s⁻¹) at 1°C resulted in a considerable increase in the amount of TAG in the autumn material acclimated to low temperatures and rhythmic light in the field. In contrast, the same treatment did not cause any increase in TAG in the spring material, acclimated to low temperatures and continuous light in the field. Results from experiments, in which moss cultivated for 4 months at 9°C on 12-h photoperiods (135μmol m⁻²s⁻¹) was kept for 3 weeks at low temperatures (9°C and −3°C) either in continuous light (135 or 70 μmol m⁻²s⁻¹) or with 12-h photoperiods (135 μmol m⁻²s⁻¹), indicated that the TAG level was higher at higher light intensity. At 9°C it was also higher in continuous light of both intensities than in rhythmic light. These results strongly suggest that decreasing irradiance and decreasing daylength limits the accumulation of TAG in D. elongatum during autumn in the subarctic.     

Causes and prevention of tamoxifen-induced accumulation of triacylglycerol in rat liver

Tamoxifen can induce hepatic steatosis in women. In this study, we wanted to elucidate the mechanism behind the tamoxifen-induced accumulation of triacylglycerol in liver in female rats, and we hoped to prevent this development by combination treatment with the modified fatty acid tetradecylthioacetic acid (TTA). The increased hepatic triacylglycerol level after tamoxifen treatment was accompanied by decreased acetyl-coenzyme A carboxylase (ACC) and FAS activities, increased glycerol-3-phosphate acyltransferase (GPAT) activity, and a tendency to increased diacylglycerol acyltransferase (DGAT) activity. The activities and mRNA levels of enzymes involved in beta-oxidation, ketogenesis, and uptake of lipids from liver were unaffected by tamoxifen, whereas the uptake of lipoproteins was unchanged and the uptake of fatty acids was decreased. Combination treatment with tamoxifen and TTA (Tam+TTA) normalized the hepatic triacylglycerol level and increased the activities of ACC, FAS, GPAT, and DGAT compared with tamoxifen-treated rats. The activities and mRNA levels of enzymes involved in beta-oxidation, ketogenesis, and uptake of lipids were increased after Tam+TTA treatment. In conclusion, tamoxifen increased the hepatic triacylglycerol level, probably as a result of increased triacylglycerol biosynthesis combined with unchanged beta-oxidation. The tamoxifen-induced accumulation of triacylglycerol was prevented by cotreatment with TTA, through mechanisms of increased mitochondrial and peroxisomal beta-oxidation.     

Effect of carbon dioxide on nitrate accumulation and nitrate reductase induction in corn seedlings

Exposure of the leaf canopy of corn seedlings (Zea mays L.) to atmospheric CO₂ levels ranging from 100 to 800 μl/l decreased nitrate accumulation and nitrate reductase activity. Plants pretreated with CO₂ in the dark and maintained in an atmosphere containing 100 μl/l CO₂ accumulated 7-fold more nitrate and had 2-fold more nitrate reductase activity than plants exposed to 600 μl/l CO₂, after 5 hours of illumination. Induction of nitrate reductase activity in leaves of intact corn seedlings was related to nitrate content. Changes in soluble protein were related to in vitro nitrate reductase activity suggesting that in vitro nitrate reductase activity was a measure of in situ nitrate reduction. In longer experiments, levels of nitrate reductase and accumulation of reduced N supported the concept that less nitrate was being absorbed, translocated, and assimilated when CO₂ was high. Plants exposed to increasing CO₂ levels for 3 to 4 hours in the light had increased concentrations of malate and decreased concentrations of nitrate in the leaf tissue. Malate and nitrate concentrations in the leaf tissue of seven of eight corn genotypes grown under comparable and normal (300 μl/l CO₂) environments, were negatively correlated. Exposure of roots to increasing concentrations of potassium carbonate with or without potassium sulfate caused a progressive increase in malate concentrations in the roots. When these roots were subsequently transferred to a nitrate medium, the accumulation of nitrate was inversely related to the initial malate concentrations. These data suggest that the concentration of malate in the tissue seem to be related to the accumulation of nitrate.     

Potential mechanisms and consequences of cardiac triacylglycerol accumulation in insulin-resistant rats

The accumulation of intracellular triacylglycerol (TG) is highly correlated with muscle insulin resistance. However, it is controversial whether the accumulation of TG is the result of increased fatty acid supply, decreased fatty acid oxidation, or both. Because abnormal fatty acid metabolism is a key contributor to the pathogenesis of diabetes-related cardiovascular dysfunction, we examined fatty acid and glucose metabolism in hearts of insulin-resistant JCR:LA-cp rats. Isolated working hearts from insulin-resistant rats had glycolytic rates that were reduced to 50% of lean control levels (P < 0.05). Cardiac TG content was increased by 50% (P < 0.05) in the insulin-resistant rats, but palmitate oxidation rates remained similar between the insulin-resistant and lean control rats. However, plasma fatty acids and TG levels, as well as cardiac fatty acid-binding protein (FABP) expression, were significantly increased in the insulin-resistant rats. AMP-activated protein kinase (AMPK) plays a major role in the regulation of cardiac fatty acid and glucose metabolism. When activated, AMPK increases fatty acid oxidation by inhibiting acetyl-CoA carboxylase (ACC) and reducing malonyl-CoA levels, and it decreases TG content by inhibiting glycerol-3-phosphate acyltransferase (GPAT), the rate-limiting step in TG synthesis. The activation of AMPK also stimulates cardiac glucose uptake and glycolysis. We thus investigated whether a decrease in AMPK activity was responsible for the reduced cardiac glycolysis and increased TG content in the insulin-resistant rats. However, we found no significant difference in AMPK activity. We also found no significant difference in various established downstream targets of AMPK: ACC activity, malonyl-CoA levels, carnitine palmitoyltransferase I activity, or GPAT activity. We conclude that hearts from insulin-resistant JCR:LA-cp rats accumulate substantial TG as a result of increased fatty acid supply rather than from reduced fatty acid oxidation. Furthermore, the accumulation of cardiac TG is associated with a reduction in insulin-stimulated glucose metabolism.     

pH effects on growth and lipid accumulation of the biofuel microalgae Nannochloropsis salina and invading organisms

Biofuels derived from non-crop sources, such as microalgae, offer their own advantages and limitations. Despite high growth rates and lipid accumulation, microalgae cultivation still requires more energy than it produces. Furthermore, invading organisms can lower efficiency of algae production. Simple environmental changes might be able to increase algae productivity while minimizing undesired organisms like competitive algae or predatory algae grazers. Microalgae are susceptible to pH changes. In many production systems, pH is kept below 8 by CO₂ addition. Here, we uncouple the effects of pH and CO₂ input, by using chemical pH buffers and investigate how pH influences Nannochloropsis salina growth and lipid accumulation as well as invading organisms. We used a wide range of pH levels (5, 6, 7, 8, 9, and 10). N. salina showed highest growth rates at pH 8 and 9 (0.19?±?0.008 and 0.19?±?0.011, respectively; mean ± SD). Maximum cell densities in these treatments were reached around 21 days into the experiment (95.6?×?10⁶?±?9?×?10⁶ cells mL^?1 for pH 8 and 92.8?×?10⁶?±?24?×?10⁶ cells mL^?1 for pH 9). Lipid accumulation of unbuffered controls were 21.8?±?5.8 % fatty acid methyl esters content by mass, and we were unable to trigger additional significant lipid accumulation by manipulating pH levels at the beginning of stationary phase. Ciliates (grazing predators) occurred in significant higher densities at pH 6 (56.9?±?39.6?×?10⁴ organisms mL^?1) than higher pH treatments (0.1–6.8?×?10⁴ organisms mL^?1). Furthermore, the addition of buffers themselves seemed to negatively impact diatoms (algal competitors). They were more abundant in an unbuffered control (12.7?±?5.1?×?10⁴ organisms mL^?1) than any of the pH treatments (3.6–4.7?×?10⁴ organisms mL^?1). In general, pH values of 8 to 9 might be most conducive to increasing algae production and minimizing invading organisms. CO₂ addition seems more valuable to algae as an inorganic carbon source and not as an essential mechanism to reduce pH.     

Apomyoglobin stability as dependent on urea concentration and temperature at two pH values

Equilibrium unfolding of apomyoglobin (ApoMb) in the presence of urea was studied as dependent on the temperature (5–2°C) at two pH values (5.7 and 6.2). Thermodynamic parameters of ApoMb transition from the native to the unfolded state were estimated under various conditions. Conformational changes in ApoMb were detected by tryptophan fluorescence and far-UV circular dichroism. The ApoMb stability and the cooperativity of its unfolding at 5°C were considerably lower than at other temperatures at both pH values, where ApoMb is in the native conformation.__________Translated from Molekulyarnaya Biologiya, Vol. 39, No. 2, 2005, pp. 330–335.Original Russian Text Copyright © 2005 by Baryshnikova, Sharapov, Kashparov, Ilyina, Bychkova.     

Ammonium and nitrate uptake by Eucalyptus nitens: effects of pH and temperature

Ammonium and nitrate uptake by roots of Eucalyptus nitens was characterised with respect to pH and temperature. Uptake of ammonium and nitrate was measured as depletion from solutions by roots of intact 11 week old solution-cultured seedlings. Uptake rates of ammonium were consistently higher than those of nitrate in all experiments. Uptake rates for ammonium were 200% higher at pH 4 than at pH 6, but for nitrate were unchanged. Uptake rates of ammonium and nitrate were both reduced to a similar extent (70%) with a decrease in temperature from 20 °C to 10 °C. For ammonium uptake, there was rapid (<24 hr) adaptation to a reduction in root temperature. The apparent preference shown here for ammonium over nitrate could be indicative of E. nitens growing in cold, acidic forest soils where ammonium is commonly more available than nitrate. These results suggest that N uptake rates of E. nitens may be maximised under a wide variety of conditions if N is supplied predominantly in the ammonium form. This revised version was published online in June 2006 with corrections to the Cover Date.     

Nitrogen source effects on rhizosphere pH and nutrient accumulation by Pacific Northwest conifers

Growth responses ofCasuarina cunninghamiana to inoculation withFrankia are described in unsterilized field soils at three sites. At Mt Crawford, South Australia, seedlings of three provenances ofC. cunninghamiana were inoculated with a singleFrankia source just prior to planting out. Forty-four months after planting, inoculation had more than doubled wood production by twoC. cunninghamiana provenances, whilst a third provenance grew poorly and did not respond to inoculation. In Zimbabwe, seedlings of one provenance ofC. cunninghamiana were inoculated in the nursery with one of four differentFrankia strains. In an N deficient soil at Kadoma, three of theseFrankia increased tree height 14 months after planting by between 50% and 70% in comparison to the uninoculated seedlings. The fourthFrankia strain resulted in increased tree height to three times that of the uninoculated controls and up to double that of the other threeFrankia strains. At Gympie, Queensland, Australia, seedlings ofC. cunninghamiana raised open-rooted in a nursery bed were inoculated withFrankia seventeen weeks before planting out. During the 22 months following planting in the field, tree growth was limited by soil P status and there was no response in tree height or stem diameter to inoculation withFrankia or to N fertilizer unless P was applied. In the presence of added P there was a significant response both toFrankia inoculation and to N fertilizer. This positive interaction between P application and N treatment was reflected in wood volumes-inoculated trees and those trees supplied N fertilizer produced 34% and 95% more wood volume than did the uninoculated trees. These results demonstrate the potential to increase the productivity of Casuarina plantings by inoculation withFrankia and by alleviation of P deficiency.     

Comparative effects of sodium nitrate and calcium nitrate on the potato

Summary Comparative trials with sodium nitrate and calcium nitrate on the potato indicate a certain superiority of the former, probably due to a greater utilization of potassium and hence improved carbohydrate metabolism.     

Pluronic L81 enhances triacylglycerol accumulation in the cytosol and inhibits chylomicron secretion

Pluronic L81 (PL81) inhibits fat absorption, and other Pluronic copolymers help overcome drug resistance in cancer cells. To understand how PL81 acts, we synthesized a radiolabeled analog, [14C]PL81, and showed that it was structurally similar to PL81 based on (1)H NMR as well as mass spectrometric analysis. [14C]PL81 inhibited the secretion of chylomicrons (CMs), lipoproteins essential for fat absorption, by differentiated Caco-2 cells similar to PL81. Moreover, PL81 competed with the cellular uptake of [14C]PL81. Thus, [14C]PL81 and PL81 behave similarly in these physiologic assays. Uptake of [14C]PL81 by Caco-2 cells was concentration-, time-, and temperature-dependent and occurred mainly from the apical side. Intracellularly, it was assimilated in the cytosol. Cells excreted PL81 toward the apical side via a pathway partially sensitive to verapamil. Small amounts were secreted toward the basolateral side unassociated with CM, and this secretion was unaffected by the inhibition of CM assembly. Nonetheless, PL81 significantly inhibited the secretion of triacylglycerols (TGs) and phospholipids as part of CM. PL81-treated cells showed decreased activity of microsomal triglyceride transfer protein and accumulated more TGs, but not phospholipids, in their cytosol. We propose that Pluronic copolymers act by interfering with the export of molecules from the cytosol. They inhibit fat absorption by decreasing TG transport to the endoplasmic reticulum and increase drug efficacy against cancer cells by competing for their excretion.     

The influence of pH and external H+ concentration on caesium toxicity and accumulation inEscherichia coli andBacillus subtilis

Summary Toxicity screening ofEscherichia coli NCIB 9484 andBacillus subtilis 007, NCIB 168 and NCIB 1650 has shown Cs⁺ to be the most toxic Group 1 metal cation. However, toxicity and accumulation of Cs⁺ by the bacteria was affected by two main external factors; pH and the presence of other monovalent cations, particularly K⁺. Over the pH range 6–9 bothE. coli andB. subtilis showed increasing sensitivity towards caesium as the pH was raised. The presence of K⁺ and Na⁺ in the laboratory media used lowered caesium toxicity and lowered acumulation of the metal. In order to assess accurately Cs⁺ toxicity towards the bacterial strains it was therefore necessary to define the K⁺:Cs⁺ ratio in the external medium. The minimum inhibitory K⁺:Cs⁺ concentration ratio for theBacillus strains tested was in the range 12–13 whileE. coli had a minimum inhibitory K⁺:Cs⁺ concentration ratio of 16.     

高、低氮浓度对2株真眼点藻的生长和油脂积累的影响

【目的】研究氮浓度对真眼点藻纲(Eustigmatophyceae)的2株高产油微藻大真眼点藻(Eustigmatos magnus,EM)和波氏真眼点藻(Eustigmatos polyphem,EP)的细胞形态、生长、总脂含量、脂质组成和脂肪酸组成与含量的时序变化规律。【方法】利用高氮(18.0 mmol/L NO3?-N)和低氮(3.6 mmol/L NO3?-N)浓度培养微藻。【结果】形态观察结果表明,大真眼点藻(E. magnus)和波氏真眼点藻(E. polyphem)营养细胞具有1个周生的裂叶状叶绿体,细胞质中有液泡,内含能够振动的颗粒物,以及一个较为明显的红色色素体;生殖方式通过形成2个D形或4个四角形的似亲孢子;随着培养周期的延伸和营养盐的消耗,细胞中油体逐步形成,其数量不断增加,体积不断增大。实验结果表明,初始氮浓度对2种微藻的总脂积累及生长均有显著影响(P<0.05),低氮浓度下2种微藻的生物质浓度分别为9.0 g/L和8.5 g/L,均低于高氮浓度下的生物质浓度。而低氮浓度下2种微藻的总脂、中性脂和总脂肪酸的含量以及总脂、中性脂与总脂肪酸的单位体积产率均明显高于高氮浓度组,其最高值分别为：59.10%、51.90%、46.95%和0.28、0.24、0.22 g/(L·d) (EM);64.20%、56.80%、50.01%和0.32、0.28、0.25 g/(L·d) (EP)。脂肪酸分析结果表明,两种微藻的脂肪酸主要成分均为棕榈酸(C16:0)、棕榈油酸(C16:1)、油酸(C18:1)和二十碳五烯酸(C20:5,EPA),四者的总含量(占总脂肪酸)分别达到85.83%和85.48%,其中棕榈油酸的含量最高。【结论】低氮浓度胁迫有利于大真眼点藻和波氏真眼点藻细胞内油脂的积累,两种微藻均为适合于生产生物柴油的油脂生产藻株。     

Effect of exogenous and endogenous nitrate concentration on nitrate utilization by dwarf bean

The effect of the exogenous and endogenous NO₃⁻ concentration on net uptake, influx, and efflux of NO₃⁻ and on nitrate reductase activity (NRA) in roots was studied in Phaseolus vulgaris L. cv. Witte Krombek. After exposure to NO₃⁻, an apparent induction period of about 6 hours occurred regardless of the exogenous NO₃⁻ level. A double reciprocal plot of the net uptake rate of induced plants versus exogenous NO₃⁻ concentration yielded four distinct phases, each with simple Michaelis-Menten kinetics, and separated by sharp breaks at about 45, 80, and 480 micromoles per cubic decimeter.

Influx was estimated as the accumulation of ¹⁵N after 1 hour exposure to ¹⁵NO₃⁻. The isotherms for influx and net uptake were similar and corresponded to those for alkali cations and Cl⁻. Efflux of NO₃⁻ was a constant proportion of net uptake during initial NO₃⁻ supply and increased with exogenous NO₃⁻ concentration. No efflux occurred to a NO₃⁻-free medium.

The net uptake rate was negatively correlated with the NO₃⁻ content of roots. Nitrate efflux, but not influx, was influenced by endogenous NO₃⁻. Variations between experiments, e.g. in NO₃⁻ status, affected the values of K_m and V_max in the various concentration phases. The concentrations at which phase transitions occurred, however, were constant both for influx and net uptake. The findings corroborate the contention that separate sites are responsible for uptake and transitions between phases.

Beyond 100 micromoles per cubic decimeter, root NRA was not affected by exogenous NO₃⁻ indicating that NO₃⁻ uptake was not coupled to root NRA, at least not at high concentrations.