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PubMed1979

Nitrite reduction in Veillonella alcalescens.

Nitrite reduction was examined in Veillonella alcalescens C-1, and obligate anaerobe with an ATP-yielding nitrate-reducing system. Hydrogen donors for nitrite reduction included hydrosulfite, hydrogen gas, and pyruvate, but not pyridine nucleotides, in the presnece or absence of flavins. Pyruvate-linked nitrite reduction was not inhibited by 4,4,4-trifluoro-1-(2-thienyl) 1,3-butanedione, dicoumarol, or 2-heptyl-4-hydroxy-quinoline-N-oxide. The noninvolvement of membrane-bound factors was supported by the fact that 100% of pyruvate-linked activity remained in the soluble fraction after fractionation of crude extracts by ultracentrifugation. Using DEAE-cellulose column chromatography, however, the participation of ferredoxin in nitrite reduction was demonstrated. The product of nitrite reduction appeared to be ammonia, as determined from H2-to-NO2- ratios. Nitrite reductase was induced by nitrate or nitrite and was repressed by increased levels of reduced nitrogenous compounds.

Anaerobiosis↗

PubMed1976

A reduced pyridine nucleotides-diaphorase activity associated to the assimilatory nitrite reductase complex from Neurospora crassa.

The Neurospora crassa assimilatory NAD(P)H-nitrite reductase complex has associated a NAD(P)H-diaphorase activity. 1. This NAD(P)H-diaphorase activity can use either mammalian cytochrome c, 2,6--dichlorophenol-indophenol, ferricyanide, or menadione as electron acceptor from the reduced pyridine nucleotides, and requires flavin adenine dinucleotide for maximal activity. 2. It is inhibited by p-hydroxymercuribenzoate, 1 muM, and it is unaffected by cyanide, sulfite, or arsenite at concentrations which completely inhibit the NAD(P)H-nitrite reductase activity. 3. Flavin adenine dinucleotide specifically protects the NAD(P)H-diaphorase activities, but not the NAD(P)H-nitrite reductase activities, against thermal inactivation. 4. In vitro preincubation of the Neurospora crassa nitrite reductase complex with reduced pyridine nucleotides plus flavin adenine dinucleotide inactivates the NAD(P)H-nitrite reductase activities, but does not affect the NAD(P)H-diaphorase activities, indicating that this nitrite reductase inactivation occurs in the part of the enzyme that contain the nitrite reducing center.

Cytochrome Reductases↗

PubMed1975

Studies on the in vitro inactivation of the Neurospora crassa assimilatory nitrite reductase in the presence of reduced pyridine nucleotides plus flavin.

In vitro inactivation of Neurospora crassa nitrite reductase (NAD(P)H: nitrite oxidoreductase, EC 1.6.6.4) can be obtained by preincubation of the enzyme with reduced pyridine nucleotide plus FAD. The presence of nitrite or hydroxylamine, electron acceptors for the N. crassa nitrite reductase, or cyanide, sulfite or arsenite, competitive inhibitors with respect to nitrite of this enzyme, protects the enzyme against this inactivation. Anaerobic experiments reveal that oxygen is required in order to obtain complete inactivation of nitrite reductase by preincubation with reduced pyridine nucleotide plus FAD. Also, inactivation is prevented if catalase is included in the preincubation mixture. The presence of hydrogen peroxide in the preincubation mixture increases the sensitivity of nitrite reductase to the in vitro FAD-dependent NAD(P)H inactivation. Neither electron acceptors, competitive inhibitors nor catalase, agents which protect the enzyme against the FAD-dependent NAD(P)H inactivation, can reverse this process once it has occurred.

Anaerobiosis↗

PubMedDate not supplied

The influence of oxygen on nitrite reduction in a reconstituted system.

Data regarding the role of oxygen in nitrite reduction are presented. In an NADPH-generating system including homogeneously purified ferredoxin-NADP reductase, ferredoxin (or flavodoxin) and nitrite reductase from the alga Bumilleriopsis filiformis, oxygen and nitrite can be reduced simultaneously. In air, rates of 1.2 mumol nitrite reduced-min-1-mg-1 nitrite reductase are obtained, which are physiologically feasible. Ferredoxin is inhibited non-competitively by oxygen during nitrite reduction. Oxygen uptake due to the oxidase reaction of ferredoxin-NADP reductase mediated by flavodoxin from Chlorella fusca and ferredoxin from Bumilleriopsis involves superoxide and is inhibited by the nitrite reducing system.

Anaerobiosis↗

PubMed1976

Damage and repair of rat liver DNA by simultaneous oral administration of amines and nitrite.

The effect of simultaneous oral administration of dimethylamine or aminopyrine and sodium nitrite on the damage and repair of rat liver DNA was studied by the use of centrifugation in alkaline sucrose gradient. Fragmentation of the intact DNA was observed significantly shortly after combined treatment with aminopyrine and nitrite when no liver necrosis occurred yet and the damaged DNA at the lower dosage level of aminopyrine with nitrite was found to be repaired in contrast to the increased activity of serum aminotransferase. The lowest effective dose on DNA damage at 2 hr after administration was about 40 mg/kg of aminopyrine and 80 mg/kg of sodium nitrite. This effect was equal to that caused by single oral administration of 2 to 10 mg/kg of dimethylnitrosamine, while dimethylamine at 160 or 320 mg/kg together with 80 mg/kg of sodium nitrite produced no pronounced DNA damage. The significant damage of rat liver DNA by aminopyrine and nitrite correlates well with the easier nitrosation of aminopyrine and the occurrence of high incidence of malignant liver tumors in rats fed aminopyrine together with nitrite already reported by Lijinsky.

Alanine Transaminase↗

PubMed1977

Esophageal spasm: clinical and manometric response to nitroglycerine and long acting nitrites.

The effect of nitroglycerine and long acting nitrites was studied in a group of 8 normal control subjects and 12 patients with esophageal spasm. The objective response of the esophagus to these drugs was recorded by obtaining esophageal manometric studies and was correlated with response in clinical symptoms. In 7 patients who had significant gastroesophageal reflux associated with spasm, the response to nitroglycerine was unpredictable. But in the group of 5 patients with diffuse esophageal spasm without gastroesophageal reflux, the response was uniformly good. All of the patients who responded to nitroglycerine also responded to long acting nitrites. These 5 patients, who were placed on long term management with long acting nitrites, remained symptom-free from 6 months to 4 years. None of them had recurrence of symptoms while they were on long acting nitrite therapy. The study suggests that if esophageal spasm is associated with reflux, the use of nitrites is less effective in controlling spasm than it is in those who do not show this association, and that diffuse esophageal spasm can be effectively managed with long acting nitrites on a long term basis in the absence of reflux. If there is esophageal spasm associated with reflux esophagitis, nitrites may be beneficial as an adjunct to antireflux therapy.

Adult↗

PubMed1979

Cyanide intoxication in sheep: enhancement of efficacy of sodium nitrite, sodium thiosulfate, and cobaltous chloride.

For treatment of cyanide intoxication of ruminants, the present recommended doses of sodium nitrite (5 mg/kg of body weight) and sodium thiosulfate (25 to 50 mg/kg) are smaller than those recommended for other animals; the decrease is partially attributed to the greater susceptibility of ruminants to the toxic effects of sodium nitrite. Based on the high tissue concentration and activity rate of rhodanese in ruminants, sulfur donors such as sodium thiosulfate could be utilized more efficaciously. Doses of sodium nitrite and sodium thiosulfate (up to 22 and 660 mg/kg, respectively) were evaluated in the present studies. Adjustment of the antidotal combination provided almost three times the protection afforded by the previously recommended doses. Moreover, under the conditions tested, the newly adjusted dose levels of sodium thiosulfate alone were more effective than the previously used antidotal combination of sodium nitrite and sodium thiosulfate and this protective effect was enhanced by cobaltous chloride (10.6 mg/kg) or sodium nitrite. The present recommended therapeutic approach to cyanide intoxication in sheep should be based primarily on administration of a much higher dose of sodium thiosulfate in combination with sodium nitrite or cobaltous chloride (or both).

Animals↗

PubMed1978

Purification and properties of nitrite reductase from Escherichia coli K12.

NADH-nitrite oxidoreductase (EC 1.6.4) was purified to better than 95% homogeneity from batch cultures of Escherichia coli strain OR75Ch15, which is partially constitutive for nitrite reductase synthesis. Yields of purified enzyme were low, mainly because of a large loss of activity during chromatography on DEAE-cellulose. The quantitative separation of cytochrome c-552 from nitrite reductase activity resulted in an increase in the specific activity of the enzyme: this cytochrome is not therefore an integral part of nitrite reductase. The subunit molecular weights of nitrite reductase and of a haemoprotein contaminant, as determined by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, were 88000 and 80000 respectively. The sedimentation coefficient was calculated to be in the range 8.5-9.5S, consistent with a mol.wt. of 190000. It is suggested therefore that the native enzyme is a dimer with two identical or similar-sized subunits. Purest samples contained 0.4 mol of flavin/mol of enzyme, but no detectable haem. Catalytic activity was totally inhibited by 20 micron-p-chloromercuribenzoate and 1 mM-cyanide, slightly inhibited by 1 micron-sulphite and 10mM-arsenite, but insensitive to 1 mM-2,2'-bipyridine, 4mM-1,10-phenanthroline and 10mM-NaN3. Three molecules of NADH were oxidized for each NO2-ion reduced: the product of the reaction is therefore assumed to be NH4+. The specific activity of hydroxylamine reductase increased at each step in the purification of nitrite reductase, and the elution profiles for these two activities during chromatography on DEAE-Sephadex were coincident. It is likely that a single enzyme is responsible for both activities.

Centrifugation, Density Gradient↗

PubMed1978

Nitrite studies in oesophageal cancer.

As nitrate consumption may have considerable importance for the in vivo formation of nitrites and potentially carcinogenic N-Nitroso compounds, we have studied salivary nitrite levels in patients with oesophageal cancer and adult volunteers before and after administering 100 ml beet juice containing 160 mg nitrate. Initial salivary nitrite levels were slightly lower in the cancer patients, perhaps because of previous malnutrition. In both groups there was a marked increase in salivary nitrite levels 90 minutes after ingesting beet juice and the attained levels in the two groups were similar. The results imply that formation of salivary nitrite is highly dependent on exogenous dietary nitrate and that there is no difference in the capacity to form nitrites between oesophageal cancer patients and healthy adult subjects.

Adult↗

PubMed1978

Effect of nitrite on microsomal cytochrome P-450.

1. Addition of nitrite to anaerobic rat liver microsomes leads to the appearance of a difference spectrum similar to the spectrum of the ferrous cytochrome P-450-NO complex. A Soret band is found at 444 nm in phenobarbital-stimulated microsomes but at 442 nm in 3-methylcholanthrene-stimulated microsomes. An alpha-band is located at 583 nm in both types of microsome. 2. The initial nitrite-induced difference spectrum is converted into a spectrum lacking a Soret band but with a prominent absorbance minimum at 417 nm. This is more rapid in microsomes from phenobarbital-treated animals where it is completed in 8 min than in microsomes from 3-methylcholanthrene-treated animals. A similar spectrum can be obtained by addition of nitrite to urea-treated microsomes in which cytochrome P-450 has been converted to cytochrome P-420. 3. Azo cleavage of neoprontosil in anaerobic microsomes is markedly inhibited by 1 mM nitrite. In contrast, oxidative drug metabolism is affected only by very high nitrite concentrations around 100 mM. It is concluded that in anaerobic microsomes, NO formed from nitrite complexes with ferrous cytochrome P-450 and thereby inhibits reductive drug metabolism.

Animals↗

PubMed1979

Death associated with nitrite ingestion: report of a case.

The tissue concentrations of nitrite and nitrate found at autopsy of a case of intentional ingestion of nitrite salts have been reported. The percentage of methemoglobin and the serum nitrite concentrations are consistent with those reported for acute overdoses. We conclude that noth nitrite and nitrate salts may be identified in tissues from persons ingesting only nitrite salts and that a significant conversion to nitrate may result from oxidation of nitrite during the conversion of heme iron to Fe3+.

Adult↗

PubMed1979

Effect of processing variables on the outgrowth of Clostridium sporogenes PA 3679 spores in comminuted meat cured with sorbic acid and sodium nitrite.

The effects of the initial pH and a "short pump" on the outgrowth of Clostridium sporogenes PA 3679 spores in comminuted cured pork were studied. Fresh ground pork was cured with salt, sugar, phosphate, ascorbate, and varying amounts of sodium nitrite and sorbic acid. The product was comminuted and inoculated with 1,000 spores of C. sporogenes per g. The meat was stuffed into 1-ounce (ca. 28.4-g) aluminum tubes, cooked to 58.5 degrees C, cooled, and incubated at 27 degrees C to observe for swells. Product cured with 0.2% sorbic acid in combination with 40 ppm sodium nitrite (40 microgram/g) had better clostridium inhibition than did product cured with 120 ppm nitrite within a pH range of 5.0 to 6.7. The sorbic acid-40 ppm nitrite combination also gave better clostridial protection than did the 120 ppm nitrite alone when reduced amounts of curing ingredients were present.

Animals↗

PubMed1975

Enzymatic and non-enzymatic reduction of nitrite by extracts of Neurospora crassa.

Two activites causing nitrite disappearance are found in extracts of Neurospora; one, inducible by nitrate or nitrite and present only in nitrite-utilizing strains, catalyze the stoichiometric reduction of nitrite to ammonia; the other, present in all strains under all conditions, causes the disappearance of nitrite to something other than ammonia. The latter activity has a molecular weight of about 600 and may contain an oligopeptide, a metal, and an SH group(s). It has no known physiological function.

Ammonia↗

PubMed1975

Tumor induction in rats by feeding aminopyrine or oxytetracycline with nitrite.

Sprague-Dawley rats were given combinations of aminopyrine or oxytetracycline and sodium nitrite in drinking water. Of 30 animals receiving 0.1% (1,000 ppm) of aminopyrine and sodium nitrite for 30 weeks, 29 died with hemangioendothelial sarcomas of the liver. The same tumor caused death in 26 of 30 animals that received 0.025% (250 ppm) of both aminopyrine and sodium nitrite for 50 weeks. No animals in a control group of the same size that received 0.1% aminopyrine for 30 weeks developed this tumor, although one-half of them were still alive 2 years after the experiment was begun. After feeding a comparable dose (0.1%) of oxytetracycline and sodium nitrite for 60 weeks, liver tumors were present in 4 of 30 rats (3 hepatocellular tumors and 1 cholangioma). Since aminopyrine has been widely used for medicinal purposes in the human population, it is possible that many people have been exposed to a potent carcinogen (dimethylnitrosamine) by its formation in vivo. It is not certain whether the result of feeding oxytetracycline and sodium nitrite indicates significant carcinogenicity of this combination.

Adenoma, Bile Duct↗

PubMed1978

Comparison of heart rate and blood pressure response to amyl nitrite, isoproterenol, and standing before and during acute beta-adrenergic blockade with intravenous propranolol.

Heart rate responses to three different procedures (amyl nitrite inhalation, standing up, and isoproterenol infusion), were studied before and during acute adrenergic beta blockade with intravenous propranolol in three normal and six hypertensive subjects. Propranolol decreased, but did not completely abolish, the heart rate increase produced by amyl nitrite and standing, probably because of vagal participation (withdrawal) in heart rate increase produced by baroreceptor hypotension (amyl nitrite) and on assuming the upright posture. Heart rate responses to amyl nitrite varied greatly from patient to patient (from 27 to 97%), but the drug proved to be the most potent stimulus for heart rate increase as a result of its marked hypotensive effect. However, this vasodilator-induced acute hypotension was well tolerated, and without deleterious hemodynamic consequences, despite the presence of beta blockade. Different degrees of correlation in heart rate increase were observed with the three procedures, reflecting probably the varying sympathetic-parasympathetic participation in reflex heart rate control. It is concluded that from the clinical stand point, neither amyl nitrite administration nor standing up can be used as a test to assess acurately the degree of beta blockade, because both procedures activate vagal withdrawal which increases heart rate regardless of the degree of beta blockade.

Adult↗

PubMed1978

The mechanism of the control of carbon fixation by the pH in the chloroplast stroma. Studies with nitrite-mediated proton transfer across the envelope.

1. CO2 fixation of intact spinach chloroplasts is inhibited by nitrite in a pH-dependent mode. At pH 7.3 in the medium 1 mM NaNO2 and at pH 7.9 5 mM NaNO2 were required for 50% inhibition. 2. The addition of nitrite leads to an acidificiation in the stroma. It appears that nitrite renders the envelope permeable for protons resulting in a breakdown of the pH gradient between the external space and the stroma. 3. In view of earlier results on the pH sensitivity of C02 fixation it is concluded that this pH shift in the stroma is responsible for the observed inhibition of CO2 fixation by nitrite. 4. Octanoate and to some extent also high concentrations of bicarbonate and acetate have a similar effect as nitrite in inhibiting CO2 fixation through an acidification in the stroma. 5. The levels of the intermediates of the CO2 fixation cycle were measured. A strong rise of the levels of fructose- and sedoheptulose biphosphates and a concomitant decrease of the corresponding monophosphates was observed during inhibition of CO2 fixation. It appears that the enzymatic steps of the CO2 fixation cycle responsible for the overall inhibition of CO2 fixation caused by lowering of the H+ concentration in the stroma are fructose- and sedopheptulose bisphosphatase. These two enzymes have an important function in the light regulation of CO2 fixation.

Ammonium Chloride↗

PubMed1977

Effect of nitrite upon leghemoglobin and interaction with nitrogen fixation.

Nitrite (0.4 mM) added to soybean bacteroid preparations strongly inhibited C2H2 reduction. In the presence of leghemoglobin (0.1mM), a 3-fold enhancement of nitrogen fixation occurred but the inhibitory effect of nitrite was delayed. Spectra of leghemoglobin showed a rapid disappearance of the 574 nm and 541 nm peaks of oxyleghemoglobin the presence of nitrite. Concomitant oxidation of this hemoprotein gave ferric leghemoglobin as the single final product. High nitrite levels could depress nitrogen fixation both by inactivation of nitrogenase and by conversion of leghemoglobin into an inactive form. Nitrite present at low concentrations reacts with this hemoprotein and is then no longer able to penetrate into bacteroids.

Hemeproteins↗

PubMed1978

Effects of low concentrations of bisulfite-sulfite and nitrite on microorganisms.

A wide range of microorganisms was tested to determine their sensitivity to low concentrations of bisulfite-sulfite and nitrite, solubility products of SO2 and NO2, respectively. Photosynthesis by blue-green algae (cyanobacteria) was more strongly inhibited by 0.1 mM bisulfite-sulfite and 1 mM nitrite at pH 6.0 than photosynthesis by eucaryotic algae and respiration of bacteria, fungi, and protozoa. At pH 7.7, blue-green algae were still more sensitive to bisulfite-sulfite and nitrite than eucaryotic algae, but the toxicity of bisulfite-sulfite and nitrite decreased as the pH increased. Photosynthesis by Anabaena flos-aquae at pH 6.0 was inhibited 25% by a bisulfite-sulfite concentration of 10 micrometer and 15% by a nitrite concentration of 50 micrometer. Photosynthesis by the blue-green alga, Lyngbya sp., was not exceptionally sensitive to chlorate and thiosulfate. Acetylene-reducing activity of Beijerinckia indica was completely inhibited by 0.1 mM bisulfite-sulfite at pH 4.0, the suppression being decreased with increasing pH.

Animals↗