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PubMed2005

Protein nitration in biological aging: proteomic and tandem mass spectrometric characterization of nitrated sites.

Proteomic techniques for the identification of 3-nitrotyrosine-containing proteins in various biological systems are described with emphasis on the direct mass spectrometric detection and sequencing of 3-nitrotyrosine-containing peptides. Strengths and weaknesses of various separation and mass spectrometric techniques are discussed. Some examples for the MS/MS analysis of nitrated peptides obtained from aging rat heart and skeletal muscle are provided, such as nitration of Tyr105 of the mitochondrial electron-transfer flavoprotein and Tyr14 of creatine kinase.

Aging↗
PubMed2006

Evolution of the soluble nitrate reductase: defining the monomeric periplasmic nitrate reductase subgroup.

Bacterial nitrate reductases can be classified into at least three groups according to their localization and function, namely membrane-bound (NAR) or periplasmic (NAP) respiratory and cytoplasmic assimilatory (NAS) enzymes. Monomeric NASs are the simplest of the soluble nitrate reductases, although heterodimeric NASs exist, and a common structural arrangement of NAP is that of a NapAB heterodimer. Using bioinformatic analysis of published genomes, we have identified more representatives of a monomeric class of NAP, which is the evolutionary link between the monomeric NASs and the heterodimeric NAPs. This has further established the monomeric structural clade of NAP. The operons of the monomeric NAP do not contain NapB and suggest that other redox partners are employed by these enzymes, including NapM or NapG predicted proteins. A structural alignment and comparison of the monomeric and heterodimeric NAPs suggests that a difference in surface polarity is related to the interaction of the respective catalytic subunit and redox partner.

Bacterial Proteins↗
PubMed2000

Oxygen uptake, acidification of medium and nitrate uptake induced by blue light in nitrate-starved Chlorella cells.

Blue light-induced oxygen uptake of the colorless mutant of Chlorella kessleri (No. 9.80) was 30-40% higher in the presence of exogenous glycine than in its absence. None of the other amino acids tested had this effect. Moreover, mutant cells in which glutamine synthetase was inhibited by methionine sulphoximine, accumulated approximately 65% more ammonium ions under blue irradiation in the presence of exogenous glycine than in its absence. The protein kinase C inhibitors, staurosporine or K252a, reduced the enhancement of oxygen uptake by approximately 40%. The present results indicate that blue light-dependent deamination of endogenous glycine might be a prerequisite for enhanced oxygen uptake in Chlorella. This blue light-induced oxygen uptake was not influenced by the inhibitors of protein phosphatase, calyculin A or okadaic acid. On the contrary, calyculin A and okadaic acid had a marked effect on the acidification of the suspension medium and nitrate uptake induced by blue light in Chlorella cells. The different responses to the inhibitors of protein kinase and phosphatase suggest the presence of different pathways among the blue light signal transduction operating on oxygen uptake, acidification of the medium and nitrate uptake in Chlorella.

Biological Transport↗
PubMed1992

Formation of active heterologous nitrate reductases between nitrate reductases A and Z of Escherichia coli.

Two nitrate reductases, NRA and NRZ, are present in Escherichia coli. These isoenzymes have the same alpha beta gamma, subunits composition and have similar size and genetic organization. Corresponding subunits of the complexes share at least 75% identity. By subcloning the different genes and expressing them from separate transcriptional units, we have demonstrated (i) that the translation of the subunits and their assembly are not coupled processes, since subunits produced concomitantly but independently can meet efficiently and associate to form active enzymes, and (ii) that the alpha subunit of a given complex can be replaced by its counterpart from the other isoenzyme to yield an active membrane-bound heterologous enzyme. One such heterologous enzyme, alpha A beta Z gamma Z, has been purified; it is less stable than the native enzymes, more susceptible to thermal denaturation, and shows increased sensitivity to proteolysis. It is also less stably bound to the membrane and, consequently, its activity with physiological electron donors is drastically reduced. The possibility that heterologous nitrate reductases could be formed in vivo is discussed with reference to the existence of porin heterotrimers of the outer membrane proteins OmpC, OmpF and PhoE.

Blotting, Western↗
PubMedDate not supplied

Gametogenic count and histopathological effect of thorium nitrate and uranyl nitrate on mice testes.

Daily intraperitoneal administration of aqueous solution of 0.05 m mole/100 g b. wt. of uranyl nitrate and thorium nitrate has been observed for 7 days. Marked reduction in the seminiferous tubule diameter and gametogenic count was observed. Signs of testicular necrosis, exfoliation of germ cells, and karyolysis and karyorrhexis were seen in most of the tutubles. Mononucleate and polynucleate giant cells were also observed indicating prolonged pathological condition.

Animals↗
PubMed1996

N-nitrosodimethylamine, nitrate and nitrate-reducing microorganisms in human milk.

Of 54 milk samples from 54 healthy nursing women analysed for volatile N-nitrosamines, 42 appeared negative. Trace amounts (below the detection limit 0.5 microgram l-1) of N-nitrosodimethylamine were detected in the milk of 10 mothers and two samples contained this compound at 1.1 and 1.2 micrograms 1-1 respectively. Almost all samples investigated contained nitrate (mean 2.9 +/- 2.3 mg1-1 and nitrate reducing microorganisms (mean 4.2 +/- 1.0 log ml-1). The recent finding of N-nitrosodimethylamine in human milk gives evidence of the continuous endogenous formation of N-nitrosamines.

Dimethylnitrosamine↗
PubMedDate not supplied

A study on organic nitrates, Part VI. Synthesis and pharmacological activity of 1-isopropylamino-3-(1-naphthyloxy)-2-propyl nitrate.

Basing on the studies on the relationship between the structure and activity, we have synthesized a nitrate analogue of propranolol as potential donor of nitric oxide. The obtained 1-isopropylamino-3-(1-naphthyloxy)-2-propyl nitrate decreases blood pressure more than propranolol and slows the heart rate less than propranolol. It also affects peripheral vascular resistance less than propranolol.

Animals↗
PubMedDate not supplied

Using the late spring nitrate test to reduce nitrate loss within a watershed.

Excessive nitrate leaching from the U.S. Corn Belt has created serious water quality problems and contributed to the expansion of the hypoxic zone in the Gulf of Mexico. We evaluated the effect of implementing the late spring nitrate test (LSNT) for corn (Zea mays L.) grown within a 400-ha, tile-drained subbasin in central Iowa. Surface water discharge and NO3 concentrations from the treated subbasin and two adjacent subbasins receiving primarily fall-applied, anhydrous ammonia were compared. In two of four years, the LSNT method significantly reduced N fertilizer applications compared with the farmers' standard practices. Average corn yield from LSNT fields and nonlimiting N fertilizer check strips was not significantly different. Autoregressive (AR) models using weekly time series in surface water NO3 concentration differences between the LSNT and control subbasins indicated no consistent significant differences during the pre-LSNT (1992-1996) period. However, by the second year (1998) of the treatment period (1997-2000), NO3 concentrations in surface water from the treated subbasin were significantly lower than the concentrations coming from both control basins. Annual average flow-weighted NO3 concentrations for the last two years (1999-2000) were 11.3 mg N L(-1) for the LSNT and subbasin and 16.0 mg N L(-1) for the control subbasins. Based on these values and the AR models, widespread adoption of the LSNT program for managing N fertilizer where fall N application is typically practiced could result in a > or = 30% decrease for NO3 concentrations in surface water.

Agriculture↗
PubMed2004

[Impact of localized compaction and ridge fertilization on field nitrate transport and nitrate use efficiency].

The primary season of nitrate leaching in the Loess Plateau region is the monsoon, which is caused by heavy rainfall during the growth season of corn (Zea mays L.). Nitrate leaching to groundwater is an increasing concern in agriculture, and is one of the major nitrogen losing ways in dryland farming system. Localized compaction and ridge fertilization is a method for nitrogen fertilizer application, by which, less fertilizer leaching would occur. The NO3(-)-N transport in soil profile, corn yields and nitrogen use efficiency under localized compaction and ridge fertilization were investigated through two years field study. The factors that affect NO3(-)-N transport under localized compaction and ridge fertilization were studied, combined with simulated experiment. The results showed that NO3(-)-N was leached to below 90 cm in plat fertilization in the year of about 370 mm rainfall, a mean precipitation during the season, while the NO3(-)-N leakage of the fertilizer zone was reduced by localized compaction and ridge fertilization, as a result that the NO3(-)-N concentration below 60 cm was less than 10 mg.kg-1, and NO3(-)-N accumulated in 20-40 cm with a concentration 80-90 mg.kg-1. There was no significant difference in yield between application methods with 240.0 kg N.hm-2. However, the absorbed amount of nitrogen was improved significantly by localized compaction and ridge fertilization, and the nitrogen use efficiency was increased by 9%. The bulk density of the barriers had an evident effect on NO3(-)-N transport under localized compaction and ridge fertilization, but the effect of ridge slope was insignificant.

Biological Transport↗
PubMedDate not supplied

Study on organic nitrates, part VIII. Pharmacological activity and nitric oxide generation capacity of nitrate derivatives of piperazine.

Organic nitrates, derivatives of piperazine, incubated with L-cysteine hydrochloride in phosphate buffer at appropriately high pH and temperature underwent decomposition, releasing nitric oxide (NO). NO generated in this reaction was quantitated by polarographic method using a selective ISO NO sensor. Spontaneously hypertensive rats were used in the tests of hypotensive activity of both investigated organic nitrates and their hydroxyl analogues. The experiments with both groups of compounds were performed under identical conditions. The results of the study demonstrated that the ability of individual compounds to release NO influenced in a different manner their pharmacological activity. Only the compound which released the largest quantity of NO during in vitro tests exhibited in vivo the most prolonged hypotensive effect and significant decrease in all arterial blood pressure parameters.

Animals↗
PubMed1990

Organic nitrates. O-(2,3-dihydroxypropyl)-1,4:3,6-dianhydro-hexitol-nitrates.

Several hybrid molecules have been synthesized which contain both the 1,4:3,6-dianhydro-hexitol (isohexide) and glycerol partial structures, with nitrate ester groups at different positions. Alkylation of the isohexide ring system with suitable 3-carbon units generates compounds with the 2,3-epoxypropyl side chain as key intermediates. Ring opening of these oxiranes under various conditions yield primary and secondary nitrates as well as dinitrates. Pharmacological screening shows that the new structures are much less active than the parent compounds.

Cardiotonic Agents↗
PubMed1988

Mechanisms for the pharmacologic interaction of organic nitrates with thiols. Existence of an extracellular pathway for the reversal of nitrate vascular tolerance by N-acetylcysteine.

Recent reports have shown that the coadministration of N-acetylcysteine (NAC) potentiated the hemodynamic actions of i.v. nitroglycerin (NTG) and reversed NTG tolerance in humans. This study has investigated the feasibility of various pharmacokinetic and biochemical mechanisms for the thiol-organic nitrate interaction, using the rat as an animal model. In order to establish that the potentiating interaction between NAC and NTG can be reproduced in the rat, NTG dose-blood pressure response curves were determined before and during concurrent thiol infusion. The hypotensive effect of NTG was enhanced significantly by NAC and glutathione, but not by N-acetylserine, showing clearly that the potentiating effect of NAC was due specifically to its thiol functional group. The systemic clearance of NTG was not affected significantly by NAC coinfusion. In addition, the intracellular metabolism of NTG in thoracic aorta segments from rats infused previously with NAC or N-acetylserine was similar, both with respect to total production of metabolites and their distribution. Thus, the enhancement of NTG action could not be attributed apparently to an effect of NAC on NTG systemic pharmacokinetics or vascular metabolism of NTG. Because glutathione, which does not enter cells readily, also potentiated the effects of NTG, the possibility of an extracellular pathway for the thiol-organic nitrate interaction was examined. In vitro degradation of NTG in plasma and blood was accelerated in the presence of NAC (or glutathione). NAC also promoted the formation of S-nitroso-N-acetylcysteine from NTG in rat and human plasma and human blood.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcysteine↗
PubMed1990

Impact of bimetallic combinations of Cu, Ni and Fe on growth rate, uptake of nitrate and ammonium, 14CO2 fixation, nitrate reductase and urease activity of Chlorella vulgaris.

The toxicity of Cu, Ni and Fe individually, as well as in combination (Cu + Ni, Cu + Fe, Ni + Fe), on growth-rate depression, uptake of NO3- and NH4+, photosynthesis, nitrate reductase and urease activity of Chlorella vulgaris has been studied. All the test metals when used individually showed pronounced toxicity on all the parameters studied. However, their interactive effect was mostly antagonistic except for Cu + Ni (synergism). Pre-addition of Fe offered more protection to the cells against copper and nickel toxicity. The data of statistical analysis reconfirmed that 14CO2 uptake is the most sensitive parameter (significant at P less than 0.005, both for time and treatment) than others in metal toxicity assessment. However, these results suggest further that exposure time and sequence of metal addition are very important in biomonitoring of heavy metal toxicity.

Ammonia↗
PubMed1989

Pharmacokinetics of isosorbide dinitrate, isosorbide-2-nitrate and isosorbide-5-nitrate in renal insufficiency after repeated oral dosage.

The pharmacokinetics of isosorbide dinitrate (ISDN) and its two active metabolites isosorbide-2-nitrate (IS-2-N) and isosorbide-5-nitrate (IS-5-N) were studied in 20 patients with normal and impaired renal function after repeated oral doses of standard 20 mg tablets ISDN t.i.d. Blood samples were taken in the steady-state on days 2 and 14, and the plasma concentrations were measured by electron capture capillary gas chromatography. We found a wide variation of pharmacokinetic parameters (AUCss0-8 and t1/2) of ISDN, IS-2-N, and IS-5-N in our patients. No correlation was detected between AUCss0-8 or t1/2 and the degree of renal insufficiency. No drug accumulation was observed after 14 days of administration.

Administration, Oral↗
PubMed2005

Nitrate radical quantum yield from peroxyacetyl nitrate photolysis.

Peroxyacetyl nitrate (PAN, CH3C(O)OONO2) is a ubiquitous pollutant that is primarily destroyed by either thermal or photochemical mechanisms. We have investigated the photochemical destruction of PAN using a combination of laser pulsed photolysis and cavity ring-down spectroscopic detection of the NO3 photoproduct. We find that the nitrate radical quantum yield from the 289 nm photolysis of PAN is Phi(NO3)PAN = 0.31 +/- 0.08 (+/-2 sigma). The quantum yield is determined relative to that of dinitrogen pentoxide, which is assumed to be unity, under identical experimental conditions. The instrument design and experimental procedure are discussed as well as auxiliary experiments performed to further characterize the performance of the optical cavity and photolysis system.

Journal Article↗
PubMed1992

Effects of nitrite, chlorate, and chlorite on nitrate uptake and nitrate reductase activity.

Effects of NO(2) (-), ClO(3) (-), and ClO(2) (-) on the induction of nitrate transport and nitrate reductase activity (NRA) as well as their effects on NO(3) (-) influx into roots of intact barley (Hordeum vulgare cv Klondike) seedlings were investigated. A 24-h pretreatment with 0.1 mol m(-3) NO(2) (-) fully induced NO(3) (-) transport but failed to induce NRA. Similar pretreatments with ClO(3) (-) and ClO(2) (-) induced neither NO(3) (-) transport nor NRA. Net ClO(3) (-) uptake was induced by NO(3) (-) but not by ClO(3) (-) itself, indicating that NO(3) (-) and ClO(3) (-) transport occur via the NO(3) (-) carrier. At the uptake step, NO(2) (-) and ClO(2) (-) strongly inhibited NO(3) (-) influx; the former exhibited classical competitive kinetics, whereas the latter exhibited complex mixed-type kinetics. ClO(3) (-) proved to be a weak inhibitor of NO(3) (-) influx (K(i) = 16 mol m(-3)) in a noncompetitive manner. The implications of these findings are discussed in the context of the suitability of these NO(3) (-) analogs as screening agents for the isolation of mutants defective in NO(3) (-) transport.

Journal Article↗
PubMed1982

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

The effect of the exogenous and endogenous NO(3) (-) concentration on net uptake, influx, and efflux of NO(3) (-) and on nitrate reductase activity (NRA) in roots was studied in Phaseolus vulgaris L. cv. Witte Krombek. After exposure to NO(3) (-), an apparent induction period of about 6 hours occurred regardless of the exogenous NO(3) (-) level. A double reciprocal plot of the net uptake rate of induced plants versus exogenous NO(3) (-) 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 (15)N after 1 hour exposure to (15)NO(3) (-). The isotherms for influx and net uptake were similar and corresponded to those for alkali cations and Cl(-). Efflux of NO(3) (-) was a constant proportion of net uptake during initial NO(3) (-) supply and increased with exogenous NO(3) (-) concentration. No efflux occurred to a NO(3) (-)-free medium.The net uptake rate was negatively correlated with the NO(3) (-) content of roots. Nitrate efflux, but not influx, was influenced by endogenous NO(3) (-). Variations between experiments, e.g. in NO(3) (-) 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 NO(3) (-) indicating that NO(3) (-) uptake was not coupled to root NRA, at least not at high concentrations.

Journal Article↗