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PubMed1972

Identification of the sources of nitrous oxide produced by oxidative and reductive processes in Nitrosomonas europaea.

1. Cells of Nitrosomonas europaea produced N(2)O during the oxidation of ammonia and hydroxylamine. 2. The end-product of ammonia oxidation, nitrite, was the predominant source of N(2)O in cells. 3. Cells also produced N(2)O, but not N(2) gas, by the reduction of nitrite under anaerobic conditions. 4. Hydroxylamine was oxidized by cell-free extracts to yield nitrite and N(2)O aerobically, but to yield N(2)O and NO anaerobically. 5. Cell extracts reduced nitrite both aerobically and anaerobically to NO and N(2)O with hydroxylamine as an electron donor. 6. The relative amounts of NO and N(2)O produced during hydroxylamine oxidation and/or nitrite reduction are dependent on the type of artificial electron acceptor utilized. 7. Partially purified hydroxylamine oxidase retained nitrite reductase activity but cytochrome oxidase was absent. 8. There is a close association of hydroxylamine oxidase and nitrite reductase activities in purified preparations.

Ammonia↗
PubMed1976

Viral factors required for interferon induction by Newcastle disease virus in mouse macrophages and chicken embryo cells.

The triggering mechanism for interferon synthesis in mouse peritoneal macrophages and chick embryo (CE) cells by Newcastle disease virus (NDV) exposed to hydroxylamine or homologous antiserum was investigated in relation to the intracellular fate of these agents. Inactivation of NDV at 22 degrees C by I M-hydroxylamine proceeded with first-order kinetics, whereas the interferon-inducing capacity of hydroxylamine-treated virus in macrophages was unimpaired. In contrast to infective NDV, hydroxylamine-inactivated virus produced interferon in CE cells, and such a virus still had partial RNA-dependent RNA polymerase activity. Hydroxylamine-inactivated NDV was adsorbed to and uncoated in both normal and chloroquine diphosphate treated cells, but no viral double-stranded RNA was detected. Hydroxylamine treatment of virion-extracted RNA and neutralization of intact virions by antibody abolished the capacity of the virus to induce interferon. Infective as well as neutralized NDV interacted with macrophages to the same degree, but association between NDV and CE cells was prevented by antibody-coating. In macrophages, the RNA of neutralized NDV became more sensitive to RNase than RNA of infective NDV, but this process was inhibited in chloroquine diphosphate-treated cells. These results suggest that interferon induction by NDV involves components of the virion which are present up to the regular uncoating process.

Animals↗
PubMed1974

Photoinactivation of ammonia oxidation in Nitrosomonas.

Photoinactivation of ammonia oxidation in cells of Nitrosomonas was shown to follow first-order kinetics with a rate constant proportional to incident light intensity. The action spectrum for photoinactivation consisted of a broad peak in the ultraviolet range, where both hydroxylamine and ammonia oxidation were affected, and a shoulder at approximately 410 nm where only ammonia oxidation was affected. In photoinactivated cells, hydroxylamine but not ammonia was oxidized to nitrite and hydroxylamine but not ammonia caused reduction of cytochromes in vivo. The amount per cell of the following constituents was not measurably altered by photoinactivation: cytochromes b, c, a, and P460; ubiquinone; phospholipid; free amino acids; hydroxylamine-dependent nitrite synthetase; nitrite reductase; p-phenylenediamine oxidase; and cytochrome c oxidase. Malonaldehyde or lipid peroxides were not detected in photoinactivated cells. Photoinactivation was prevented (i) under anaerobic conditions, (ii) in the presence of methanol, allylthiourea, thiosemicarbazide, hydroxylamine, ethylxanthate, or CO at concentrations wich caused 100% inhibition of ammonia oxidation, and (iii) at concentrations of ammonia or hydroxylamine which gave a rapid rate of nitrite production. Recovery of ammonia oxidation activity in 90% inactivated cells took place in 6 h, required an energy and/or nitrogen source, and was inhibited by 400 mug of chloramphenicol per ml.

Amino Acids↗
PubMed1963

Phosphorylation by extracts of Nitrosomonas europaea.

Burge, W. D. (University of California, Berkeley), E. Malavolta, and C. C. Delwiche. Phosphorylation by extracts of Nitrosomonas europaea. J. Bacteriol. 85:106-110. 1963.-Cellfree preparations of Nitrosomonas europaea are capable of oxidizing hydroxylamine, but not ammonium ion, to nitrite. The quantity of nitrite formed by our preparations was, at most, equivalent to only 70% of the hydroxylamine added. Although the preparations had a strong phosphatase activity, resulting in a net loss of organic phosphate during the experimental period, P(32)-labeled inorganic phosphate was found to be incorporated into the organic fraction, including adenosine triphosphate (ATP) and adenosine diphosphate (ADP). The provision of hydroxylamine as substrate resulted in the formation of nitrite and an increased incorporation of P(32) into the organic fraction. It is concluded that the chemosynthetic autotroph Nitrosomonas, in common with certain other autotrophic organisms and heterotrophs, is capable of converting energy released in the oxidation of its inorganic substrate into high-energy phosphate units (ATP and ADP) for the mediation of other energy-requiring reactions. The simultaneous formation of ATP and ADP is interpreted as evidence for an adenylate kinase activity. The preparations used exhibited a considerable endogenous incorporation of P(32) into organic phosphate in the absence of added hydroxylamine. Cyanide inhibited both phosphorylation and the oxidation of hydroxylamine. Both the supernatant and particulate fractions of a Nitrosomonas extract subjected to centrifugal fields of 100,000 x g were active in phosphorylation and nitrite formation, but these activities appeared to be uncoupled in the particulate fraction and only partially coupled in the supernatant solution. This most likely reflects a significant endogenous respiration, and not a real lack of coupling between the two reactions.

Cyanides↗
PubMed1995

Dapsone toxicity: some current perspectives.

1. Dapsone is a potent anti-inflammatory and anti-parasitic compound, which is metabolised by cytochrome P-450 to hydroxylamines, which in turn cause methaemoglobinaemia and haemolysis. However, during the process of methaemoglobin formation, erythrocytes are capable of detoxifying the hydroxylamine to the parent drug, which may either reach the tissues to exert a therapeutic effect or return to the liver and be re-oxidised in a form of systemic cycling. This glutathione-dependent effect, combined with the un-ionised state of the drug at physiological pH, may contribute to its efficacy. 2. Paradoxically, other aspects of the glutathione-dependent cycling of the hydroxylamine metabolite may contribute to the major adverse reaction of the drug, agranulocytosis. Erythrocytes exposed to the metabolite and repeatedly washed may still release the hydroxylamine in sufficient concentration to kill mononuclear leucocytes in vitro. Thus, erythrocytes may be a conduit for the hydroxylamine to reach the bone marrow to covalently bind to granulocyte precursors, which may trigger an immune response in certain individuals and may lead to the potentially fatal eradication of granulocytes from the circulation. 3. Attempts to increase patient tolerance to dapsone have been most successful using a metabolic inhibitor to reduce hepatic oxidation of the drug to the hydroxylamine. Methaemoglobin formation in the presence of cimetidine was maintained at 30% below control levels for almost 3 mo, and patients' reported side effects such as headache and lethargy were significantly reduced. 4. As clinical application of new and safer dapsone analogues is years away, the use of cimetidine provides an immediate route to increasing patient compliance during dapsone therapy, especially in those maintained on dapsone dosages in excess of 200 mg/day.

Agranulocytosis↗
PubMed1995

Reductive chemistry of the novel hypoxia-selective cytotoxin 5-[N,N-bis(2-chloroethyl)amino]-2,4-dinitrobenzamide.

5-[N,N-Bis(2-chloroethyl)amino]-2,4-dinitrobenzamide (1; SN 23862) is a novel bioreductive drug whose selective toxicity for hypoxic cells appears due to oxygen-inhibited enzymatic reduction of one of the nitro groups to the corresponding amine or hydroxylamine. Radiolytic reduction of 1 using up to four reducing equivalents in 1 N sodium formate was shown to proceed via electron addition to the 4-nitro group, thereby identifying this substituent as the most electron-affinic site in the molecule. The initially-formed 4-hydroxylamine and its N-hydroxytetrahydroquinoxaline half-mustard cyclization product (formed by intramolecular reaction with one arm of the adjacent mustard group) are reduced to the corresponding 4-amines upon further addition of electrons, although reduction of the 2-nitro group leading to 2,4-diamino products begins after addition of only six electron equivalents. Radiolytic reduction of the structurally similar 5-(aziridin-1-yl)-2,4-dinitrobenzamide (2; CB 1954) with six electron equivalents also occurs at the 4-nitro group to give the 4-hydroxylamine and 4-amine. The product mixture from reduction of 2 is less complex, largely because the corresponding 4-hydroxylamine and 4-amine are stable. The major reduction products of 1 were chemically synthesized by unequivocal routes to provide authentic samples for identification of the products of radiolytic reduction and to allow determination of their cytotoxicities. The 2- and 4-amino derivatives of 1 are significantly more cytotoxic than the parent drug, although the toxicity of the 4-amine is moderated by its facile conversion to the corresponding less toxic tetrahydroquinoxaline half-mustard. Although the 2- and 4-hydroxylamino derivatives were prepared by chemical reduction of 1, their toxicity could not be evaluated because of their instability. The 4-hydroxylamine reacts intramolecularly with the 5-mustard group somewhat more rapidly than does the 4-amine, while the 2-hydroxylamine is converted into a 2,2'-azoxy dimer following aerial oxidation to the 2-nitroso derivative. The fully reduced 2,4-diamino derivative of 1 is 10-fold more cytotoxic again than the 2-amine and, surprisingly, does not undergo spontaneous intramolecular alkylation. This elucidation of the reduction chemistry of 1 will facilitate further investigations of the toxic products generated from this compound both by hypoxic tumor cells and by ADEPT enzymes.

Aniline Mustard↗
PubMed2004

2-Amino metabolites are key mediators of CB 1954 and SN 23862 bystander effects in nitroreductase GDEPT.

An important feature of gene-directed enzyme-prodrug therapy is that prodrug activation can provide diffusible cytotoxic metabolites capable of generating a local bystander effect in tumours. Activation of the aziridinyl dinitrobenzamide CB 1954 by E. coli nitroreductase (NTR) provides a bystander effect assumed to be due to the potently cytotoxic 4-hydroxylamine metabolite. We show that there are four cytotoxic extracellular metabolites of CB 1954 in cultures of NTR-expressing tumour cells (the 2- and 4-hydroxylamines and their corresponding amines). The 4-hydroxylamine is the most cytotoxic in DNA crosslink repair defective cells, but the 2-amino derivative (CB 10-236) is of similar potency to the 4-hydroxylamine in human tumour cell lines. Importantly, CB 10-236 has much superior diffusion properties to the 4-hydroxylamine in multicellular layers grown from the SiHa human cervical carcinoma cell line. These results suggest that the 2-amine, not the 4-hydroxylamine, is the major bystander metabolite when CB 1954 is activated by NTR in tumours. The corresponding dinitrobenzamide nitrogen mustard SN 23862 is reduced by NTR to form a single extracellular metabolite (also the 2-amine), which has superior cytotoxic potency and diffusion properties to the CB 1954 metabolites. These results are consistent with the reported high bystander efficiency of SN 23862 as an NTR prodrug in multicellular layers and tumour xenografts.

Aniline Mustard↗
PubMedDate not supplied

Chemical extraction methods to assess bioavailable arsenic in soil and solid media.

Soil ingestion by children is an important pathway in assessing public health risks associated with exposure to arsenic-contaminated soils. Soil chemical methods are available to extract various pools of soil arsenic, but their ability to measure bioavailable arsenic from soil ingestion is unknown. Arsenic extracted by five commonly used soil extractants was compared with bioavailable arsenic measured in vivo by immature swine (Sus scrofa) dosing trials. Fifteen contaminated soils that contained 233 to 17 500 mg kg(-1) arsenic were studied. Soil extractants were selected to dissolve surficially adsorbed and/or readily soluble arsenic (water, 1 M sodium acetate, 0.1 M Na2HPO4/0.1 M NaH2PO4) and arsenic in Fe and Mn oxide minerals (hydroxylamine hydrochloride, ammonium oxalate). The mean percent of total arsenic extracted was: ammonium oxalate (53.6%) > or = hydroxylamine hydrochloride (51.7%) > phosphate (10.5%), acetate (7.16%) > water (0.15%). The strongest relationship between arsenic determined by soil chemical extraction and in vivo bioavailable arsenic was found for hydroxylamine hydrochloride extractant (r = 0.88, significant at the 0.01 probability level). Comparison of the amount of arsenic extracted by soil methods with bioavailable arsenic showed the following trend: ammonium oxalate, hydroxylamine hydrochloride > in vivo > phosphate, acetate > water. The amount of arsenic dissolved in the stomach (potentially bioavailable) is between surficially adsorbed (extracted by phosphate or acetate) and surficially adsorbed + nonsurficial forms in Fe and Mn oxides (extracted by hydroxylamine hydrochloride or ammonium oxalate). Soil extraction methods that dissolve some of the amorphous Fe, such as hydroxylamine hydrochloride, can be designed to provide closer estimates of bioavailable arsenic.

Animals↗
PubMed1988

Metabolism of chlorpromazine and promazine in vitro: isolation and characterization of N-oxidation products.

1. The syntheses of the secondary hydroxylamines of nor1chlorpromazine and nor1promazine via their corresponding primary hydroxylamines and oximes are described. 2. The N-oxidation products are unstable to analysis by g.l.c. without prior derivatization; the decomposition products and the structures of the trimethylsilyl (TMS) and trifluoroacetyl (TFA) derivatives were characterized by g.l.c.-mass spectrometry. 3. Chlorpromazine, promazine and their demethylated products were shown to undergo metabolic N- and alpha-C-oxidation, to yield hydroxylamines and carboxylic acids, on incubation with fortified 9000 g liver homogenates of male New Zealand white rabbits. 4. A condensation product, an artifact formed by reaction of the metabolically derived primary hydroxylamines with acetaldehyde, an impurity in the extraction solvent, diethyl ether, was identified. 5. N-hydroxynor1- and N-hydroxynor2chlorpromazine undergo metabolic reduction to the parent amines, and the secondary hydroxylamine undergoes N-demethylation to yield the corresponding primary hydroxylamine.

Animals↗
PubMed1993

Nitric oxide induces calcium-dependent [3H]dopamine release from striatal slices.

Hydroxylamine (0.01-30 mM), a nitric oxide (NO) generator, produced a concentration-dependent release of [3H]dopamine ([3H]DA) from rat striatal slices. Hemoglobin (10 microM), a NO scavenger, reduced basal [3H]DA release and blocked hydroxylamine (100 microM)-stimulated [3H]DA efflux. Tetrodotoxin (0.5 microM) had no significant effect. Sodium cyanide was used as a model compound to test the possibility that NO acted through blockade of mitochondrial electron transport. Calcium-free experimental buffer (1 mM EGTA) reduced basal release and the hydroxylamine response, while sodium cyanide-induced release did not change under these experimental conditions. Cadmium (200 microM), a non-selective inhibitor of voltage-dependent calcium channels, reduced the hydroxylamine response by 69%. Methylene blue (10 microM), an inhibitor of guanylate cyclase, produced a 3-fold increase in the basal release but had no significant effect on the hydroxylamine response. These data suggest that NO induces calcium-dependent [3H]DA release from the striatum via a mechanism which is independent of blockade of electron transport or activation of guanylate cyclase.

Animals↗
PubMed1991

Rapid analysis of glycolipid anchors in amphiphilic dimers of acetylcholinesterases.

1. We describe two simple procedures for the rapid identification of certain structural features of glycolipid anchors in acetylcholinesterases (AChEs). 2. Treatment with alkaline hydroxylamine (that cleaves ester-linked acyl chains but not ether-linked alkyl chains) converts molecules possessing a diacylglycerol, but not those with an alkylacylglycerol, into hydrophilic derivatives. AChEs in human and bovine erythrocytes possess an alkylacylglycerol (Roberts et al., J. Biol. Chem. 263:18766-18775, 1988; Biochem. Biophys. Res. Commun. 150:271-277, 1988) and are not converted to hydrophilic dimers by alkaline hydroxylamine. Amphiphilic dimers of AChE from Drosophila, from mouse erythrocytes, and from the human erythroleukaemia cell line K562 also resist the treatment with hydroxylamine and likely possess a terminal alkylacylglycerol. This indicates that the cellular pool of free glycolipids used as precursors of protein anchors is distinct from the pool of membrane phosphatidylinositols (which contain diacylglycerols). 3. Pretreatment with alkaline hydroxylamine is required to render the amphiphilic AChE from human erythrocytes susceptible to digestion by Bacillus thuringiensis phosphatidylinositol-specific phospholipase C (PI-PLC) (Toutant et al., Eur. J. Biochem. 180:503-508, 1989). We show here that this is also the case for the AChE from mouse erythrocytes, which therefore likely possesses an additional acyl chain in the anchor that prevents the action of PI-PLC. 4. In two sublines of K562 cells (48 and 243), we observed that AChE either was directly susceptible to PI-PLC (243) or required a prior deacylation by alkaline hydroxylamine (48). This suggests that glycolipid anchors in AChE of K562-48 cells, but not those in AChE of K562-243 cells, contain the additional acylation demonstrated in AChE from human erythrocytes. These observations illustrate the cell specificity (and the lack of species-specificity) of the structure of glycolipid anchors.

Acetylcholinesterase↗
PubMed1980

NMR relaxivity changes in chloroplast suspensions. Effects of NH2OH and of treatments altering the redox state of the photosynthetic electron transport chain.

Treatments (illumination, chemical oxidation or reduction) which are potentially capable of producing paramagnetic centers in chloroplast thylakoid membranes do not produce enhancements of the proton magnetic relaxivities of these preparations. However, exposure of thylakoid membranes to varying concentrations of hydroxylamine induces a time-dependent increase in relaxivity for which the steady-state magnitude is dependent on hydroxylamine concentration. The appearance of relaxivity is correlated kinetically with inactivation of oxygen-evolving centers; in addition both processes show a threshold effect with respect to hydroxylamine concentration. Kinetic analyses of these hydroxylamine-induced effects suggest that at low (less than or equal to 100 microM) and at intermediate (200--500 microM) concentrations, hydroxylamine extraction is partially counteracted by a reverse process that reactivates oxygen-evolving centers in the dark.

Chloroplasts↗
PubMed1994

The effect of nitric oxide donors on insulin secretion, cyclic GMP and cyclic AMP in rat islets of Langerhans and the insulin-secreting cell lines HIT-T15 and RINm5F.

The aim of this study was to investigate whether short-term treatment with nitric oxide donors could mimic cytokine inhibition of insulin secretion. We tested the nitric oxide generating compounds 3-morpholinosydnonimine (SIN-1), S-nitroso-N-penicillamine (SNAP), S-nitrosoglutathione and hydroxylamine for their ability to inhibit insulin secretion, raise cyclic GMP and lower cyclic AMP levels in isolated rat islets of Langerhans and the insulin-secreting cell lines HIT-T15 and RINm5F. In islets, all nitric oxide donors inhibited glucose-induced insulin secretion and raised cyclic GMP levels. SIN-1 and S-nitrosoglutathione also reduced cyclic AMP, while SNAP and hydroxylamine had no effect. Insulin secretion in HIT-T15 cells was inhibited by SIN-1, SNAP and hydroxylamine and in RINm5F cells by hydroxylamine. Inhibition of HIT-T15 and RINm5F cell insulin secretion was not accompanied by an increase in cyclic GMP levels. The degree of inhibition of insulin secretion was unrelated to the extent of release of nitric oxide by the compounds as measured by nitrite and nitrate production. More effective inhibition by S-nitrosoglutathione and hydroxylamine versus SIN-1 and SNAP may be related to intracellular versus extracellular site of nitric oxide generation.

Animals↗
PubMed1982

ADP-ribosyl-protein conjugate subclasses in various tissues. Specific influence of thyroid hormone on liver conjugates.

The amounts of endogenous mono (ADP ribose)-protein conjugates and their hydroxyl-amine-sensitive and hydroxylamine-resistant subfractions in various tissues of the mouse were determined and compared with poly (ADP-ribose) conjugates. Total mono-(ADP-ribose) conjugates did not correlate with poly (ADP-ribose) residues or the cellularity of a tissue, but were related to the protein content and the amounts of NAD+ +NADH. The hydroxylamine-resistant subfraction, which in liver is mainly associated with the mitochondria [Adamietz, Wielckens, Bredehorst, Lengyel & Hilz (1981) Biochem. Biophys. Res, Commun. 101, 96-103], did not correlate with the tissue content of cytochrome c oxidase. In hypothyroid mice hydroxylamine-resistant mono (ADP-ribose) conjugates of the liver were increased by a factor of two while the hydroxylamine-sensitive conjugates did not change significantly under these conditions. Upon administration of thyroxine the hydroxylamine-resistant subfraction returned to normal.

Adenosine Diphosphate Ribose↗
PubMed1988

Structural beta-cell changes and transient hyperglycemia in mice treated with compounds inducing inhibited citric acid cycle enzyme activity.

An initial transient hyperglycemia was seen in mice injected with asparagine, fluoroacetate, hydroxylamine, or malonate plus methionine, whereas an initial triphasic blood glucose response and a transient "secondary" hyperglycemia were exhibited in those injected with hydroxylamine plus arsenite, and a delayed hypoglycemia was observed in those treated with fluoroacetate or arsenite. The glucose-induced insulin secretion was significantly decreased in isolated pancreatic islets incubated with hydroxylamine plus arsenite. Light and electron microscopy, pyroantimonate technique, and X-ray microanalysis disclosed mitochondrial damage, degeneration, and necrosis among the beta-cells in the islets of mice injected with hydroxylamine plus arsenite. Glycogen depletion and microvesicular fatty change were seen in the liver of mice treated with fluoroacetate, arsenite, or hydroxylamine plus arsenite. These observations support the view that inhibition of the activity of citric acid cycle enzymes and associated reactions in the beta-cells play a role in the induction of diabetic features.

Animals↗
PubMed1987

Cytotoxicity of oxidative metabolites of procainamide.

Indirect evidence suggests that metabolism of procainamide (PA) may be necessary for generation of its autoimmunity-inducing capacity. Reactive metabolites of PA produced by hepatic mixed function oxidases have been identified by their capacity to bind proteins covalently. The present study extends these findings by comparing the toxicity of various chemically synthesized metabolites of PA to a variety of cell lines and primary cultures. PA and its N-acetyl- and N-acetyl-N-oxide-derivatives were nontoxic to S49.1 cells at concentrations up to 5 mM, whereas nitro-Pa (4-nitro-N-(diethylaminoethyl)benzamide hydrochloride) and azoxy-PA (bis-N,N'-(diethylaminoethyl)-4,4'-azoxydibenzamide) displayed partial cytotoxicity at 1 and 0.1 mM, respectively. (The therapeutic range of PA is 0.02-0.05 mM.). In contrast, hydroxylamine-PA (4-hydroxylamino-N-(diethylaminoethyl)benzamide hydrochloride) was highly cytotoxic, producing a TD50 between 0.002 and 0.045 mM among the eight cell lines tested. Hydroxylamine-PA sensitivity correlated with cell reducing capacity, suggesting that redox cycling contributes to cytotoxicity. The hydroxylamine metabolite of PA undergoes spontaneous air oxidation to the nitroso-derivative (nitroso-PA). When this oxidation was minimized by rapid manipulations or inhibited by ascorbic acid, cytotoxicity was reduced or eliminated, suggesting that the nitroso-derivative may be the toxic metabolite. Quiescent peripheral blood lymphocytes, especially T-cells, were relatively insensitive to hydroxylamine-PA when evaluated by dye exclusion 1 day after drug exposure. However, massive DNA strand breaks were detectable in these cells immediately after drug exposure, and resting lymphocytes treated with 0.01 mM hydroxylamine-PA died slowly during the subsequent week.(ABSTRACT TRUNCATED AT 250 WORDS)

Ascorbic Acid↗
PubMed2003

Nitrogen isotopomer site preference of N2O produced by Nitrosomonas europaea and Methylococcus capsulatus Bath.

The relative importance of individual microbial pathways in nitrous oxide (N(2)O) production is not well known. The intramolecular distribution of (15)N in N(2)O provides a basis for distinguishing biological pathways. Concentrated cell suspensions of Methylococcus capsulatus Bath and Nitrosomonas europaea were used to investigate the site preference of N(2)O by microbial processes during nitrification. The average site preference of N(2)O formed during hydroxylamine oxidation by M. capsulatus Bath (5.5 +/- 3.5 per thousand) and N. europaea (-2.3 +/- 1.9 per thousand) and nitrite reduction by N. europaea (-8.3 +/- 3.6 per thousand) differed significantly (ANOVA, f((2,35)) = 247.9, p = 0). These results demonstrate that the mechanisms for hydroxylamine oxidation are distinct in M. capsulatus Bath and N. europaea. The average delta(18)O-N(2)O values of N(2)O formed during hydroxylamine oxidation for M. capsulatus Bath (53.1 +/- 2.9 per thousand) and N. europaea (-23.4 +/- 7.2 per thousand) and nitrite reduction by N. europaea (4.6 +/- 1.4 per thousand) were significantly different (ANOVA, f((2,35)) = 279.98, p = 0). Although the nitrogen isotope value of the substrate, hydroxylamine, was similar in both cultures, the observed fractionation (delta(15)N) associated with N(2)O production via hydroxylamine oxidation by M. capsulatus Bath and N. europaea (-2.3 and 26.0 per thousand, respectively) provided evidence that differences in isotopic fractionation were associated with these two organisms. The site preferences in this study are the first measured values for isolated microbial processes. The differences in site preference are significant and indicate that isotopomers provide a basis for apportioning biological processes producing N(2)O.

Binding Sites↗
PubMed1977

The relationship between the activity of chloroplast photosystem II and the midpoint oxidation-reduction potential of cytochrome b-559.

The role of cytochrome b-559 in Photosystem II reactions has been investigated using hydroxylamine treatment of chloroplast membranes. Incubation of chloroplasts with hydroxylamine in darkness resulted in inhibition of water oxidation and a decrease in the amplitude of cytochrome b-559 reducible by hydroquinone. The loss of water oxidizing activity perfectly correlated with the decrease in amplitude of cytochrome b-559 reduction. Potentiometric titration of cytochrome b-559 after hydroxylamine treatment revealed a component with Em7.8 at +240 mV in addition to a lower potential species at +90 mV. This compared to control chloroplasts in which cytochrome b-559 exists in the typical high potential state, Em7.8 = +383 mV, in addition to some of the low potential (Em7.8 = +77 mV) form. Photosystem II activity could be further inhibited by incubation with hydroxylamine in the light. In these chloroplasts only low rates of photooxidation of artificial electron donors were observed compared to 'dark' chloroplasts. In addition, the hydroxylamine light treatment caused a further change in cytochrome b-559 redox properties; a single component, Em7.8 = 90 mV is seen in titration curves. The role of cytochrome b-559 in Photosystem II functioning is discussed on the basis of these observations which suggest a dependence of photooxidizing ability of Photosystem II on the redox properties of this cytochrome.

Chloroplasts↗