Redox Chemiluminescence Research Articles

Regulation of microsomal and cytosolic glutathione S-transferase activities by S-nitrosylation

There is increasing evidence that S-nitrosylation is a mechanism for the regulation of protein function via the modification of critical sulfhydryl groups. The activity of rat liver microsomal glutathione S-transferase (GST) is increased after treatment with N-ethylmaleimide (NEM), a sulfhydryl alkylating reagent, and is also increased under conditions of oxidative stress. In the present study, preincubation of purified rat liver microsomal GST with S-nitrosoglutathione (GSNO) or the nitric oxide (NO) donor, 1,1-diethyl-2-hydroxy-2-nitrosohydrazine (DEA/NO), resulted in a 2-fold increase in enzyme activity. This increase in activity was reversed by dithiothreitol. The initial treatment of microsomal GST with either GSNO or DEA/NO was associated with an 85% loss of free sulfhydryl groups. After removal of the nitrosylating agents over a 6-hr period, approximately 50% of the enzyme was still nitrosylated, as determined by redox chemiluminescence. Furthermore, preincubation of either purified enzyme or hepatic microsomes with GSNO or DEA/NO prevented further enzyme activation by NEM, suggesting that NEM and the NO donors interact with a common population of sulfhydryl groups in the enzyme. In contrast, both NEM and NO donors partially inhibited the activity of cytosolic GST isoforms. The inhibitory activity of NEM and NO donors was much more evident when the GST pi isoform was used instead of a mixture of GST isoforms. These data suggest that there may be differential regulation of microsomal and cytosolic GST activities under conditions of nitrosative stress.

Prolonged decreases in plasma nitrate levels at early postoperative phase after hepato-pancreato-biliary surgery

Nitric oxide (·NO) is known to influence circulatory, neural, immunologic, and metabolic alterations. To evaluate the clinical significance of ·NO production under surgical stress, serial measurements of plasma nitrite plus nitrate levels were performed in 45 surgical patients. Group A included 19 patients who underwent major surgery with uneventful postoperative courses. Group B included 18 patients who underwent laparoscopic cholecystectomy. Group C included 8 patients whose surgery was complicated by intra-abdominal abscesses. Eight healthy volunteers served as controls. Plasma nitrate levels were determined with a redox chemiluminescence ·NO analyzer and coincided with measurements made by high-performance liquid chromatography (r = 0.868, p < 0.0001, 58 samples). During laparotomy, arterial nitrate levels correlated well with peripheral, portal, and hepatic venous nitrate levels (r = 0.966, 0.938, and 0.949, respectively; p < 0.0001). A significant decrease in nitrate from preoperative levels in groups A (postoperative day (POD) 1 and 3; p < 0.0005) and B (POD 1, p < 0.0001) was observed; nitrate levels in group C did not decrease for 14 days after surgery. Plasma nitrate levels in groups A and B were significantly different (POD 1 through 6, p < 0.05) and at POD 3 were significantly lower in group A ( p < 0.005). Plasma nitrate levels measured before and after fasting or food intake were not significantly different. These results suggest that surgical stress leads to a decrease in the end product of ·NO in the whole body, and that the greater the surgical stress the longer the duration of decreased ·NO production.

Interleukin 1 beta markedly stimulates nitric oxide formation in the absence of other cytokines or lipopolysaccharide in primary cultured rat hepatocytes but not in Kupffer cells

To investigate whether a single inflammatory cytokine could stimulate nitric oxide formation in the absence of other cytokines or lipopolysaccharide (LPS), NO was measured by the redox chemiluminescence method in primary cultured rat hepatocytes and in rat Kupffer cells. Interleukin (IL) 1 beta, but neither IL-6, tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), nor LPS stimulated NO formation in a dose-dependent manner and induced half-maximal effects at 30 pmol/L. Maximal stimulation was achieved at 12 to 16 hours after the addition of 1 nmol/L of IL-1 beta, and was 50- to 60-fold above basal levels in rat hepatocytes. The combined effect of these cytokines with LPS or IFN-gamma on NO formation was also examined. Neither LPS nor IFN-gamma affected the IL-1 beta-induced NO formation. TNF-alpha, however, stimulated IL-1 beta-induced NO formation, while IL-6 inhibited it, although independently these cytokines had no effect on NO formation. None of the cytokines tested stimulated NO formation in cultured rat Kupffer cells. In hepatocytes, the NO formation induced by IL-l beta was blocked by both the NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA) and by IL-1 receptor antagonist (IL-1ra). Furthermore, IL-1 beta markedly increased NOS activity, and this increase in activity was accompanied by the expression of inducible NOS (iNOS) messenger RNA (mRNA). This study clearly demonstrated that IL-1 beta markedly stimulates NO formation in hepatocytes, in the absence of other cytokines or LPS. (Hepatology 1996 Apr;23(4):797-802)

Interleukin 1 β markedly stimulates nitric oxide formation in the absence of other cytokines or lipopolysaccharide in primary cultured rat hepatocytes but not in Kupffer cells

To investigate whether a single inflammatory cytokine could stimulate nitric oxide formation in the absence of other cytokines or lipopolysaccharide (LPS), NO was measured by the redox chemiluminescence method in primary cultured rat hepatocytes and in rat Kupffer cells. Interleukin (IL) 1 beta, but neither IL-6, tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), nor LPS stimulated NO formation in a dose-dependent manner and induced half-maximal effects at 30 pmol/L. Maximal stimulation was achieved at 12 to 16 hours after the addition of 1 nmol/L of IL-1 beta, and was 50- to 60-fold above basal levels in rat hepatocytes. The combined effect of these cytokines with LPS or IFN-gamma on NO formation was also examined. Neither LPS nor IFN-gamma affected the IL-1 beta-induced NO formation. TNF-alpha, however, stimulated IL-1 beta-induced NO formation, while IL-6 inhibited it, although independently these cytokines had no effect on NO formation. None of the cytokines tested stimulated NO formation in cultured rat Kupffer cells. In hepatocytes, the NO formation induced by IL-l beta was blocked by both the NO synthase (NOS) inhibitor NG-monomethyl-l-arginine (l-NMMA) and by IL-1 receptor antagonist (IL-1ra). Furthermore, IL-1 beta markedly increased NOS activity, and this increase in activity was accompanied by the expression of inducible NOS (iNOS) messenger RNA (mRNA). This study clearly demonstrated that IL-1 beta markedly stimulates NO formation in hepatocytes, in the absence of other cytokines or LPS.

Nitric oxide raises cytosolic concentrations of Ca 2+ in cultured nodose ganglion neurons from rabbits

The effects of donors of nitric oxide (NO), namely, S-nitroso- N-acetylpenicillamine (SNAP) and E-methyl-2- E-hydroxyimino-5-nitro-6-methoxy-3-hexeneamide (NOR1), on cytosolic concentrations of Ca 2+ ([Ca 2+] i) were studied in cultured nodose ganglion neurons from newborn and young rabbits by loading with fura-2AM and microfluorometry. Application of SNAP (5 μM-1 mM) increased [Ca 2+] i in 46% of neurons tested. The threshold dose of the response was 10 μM and the response increased in a dose-dependent fashion. The increase in [Ca 2+] i at 50 μM averaged 74 ± 8% above the control value. [Ca 2+] i rose immediately after injection of SNAP and the plateau level was maintained in the presence of SNAP. NOR1, another donor of NO, increased [Ca 2+] i with an average increase of 82 ± 7% at 50 μM. Quantitation of NO gas in the solution of NOR1 by a redox chemiluminescence method revealed the constant release of 0.06 ppm NO from 5 ml of a 20 μM solution of NOR1. These data suggest that NO released from donors of NO induced an increase in [Ca 2+] i in nodose ganglion neurons and, therefore, that NO might play a role as a transmitter or a modulator in autonomic primary sensory systems.

Direct evidence for nitric oxide formation from glyceryl trinitrate during incubation with intact bovine pulmonary artery.

It has been proposed that the mechanism of the vasodilator action of glyceryl trinitrate (GTN) involves biotransformation to nitric oxide. A sensitive chemiluminescence method for nitric oxide determination was used to test this hypothesis. In four experiments, bovine pulmonary artery (BPA) was incubated with GTN (0.1 mM) in Krebs' solution (2 mL) containing 30 mM KCl, and in anaerobic conditions using 95% Ar - 5% CO2, in a sealed micro-Fernbach flask (6.2-mL volume). After incubation for 2, 5, 10, or 20 min at 37 degrees C, 400-microL aliquots of headspace gas were removed and injected into a redox chemiluminescence detector. Nitric oxide formation was first measurable at 5 min (76 +/- 53 pmol/g wet wt. BPA), and increased with incubation time (174 +/- 46 pmol/g wet wt. BPA after 10 min and 310 +/- 67 pmol/g wet wt. BPA after 20 min). This is the first direct chemical measurement of nitric oxide formation during interaction of GTN with vascular smooth muscle. These data support the concept that GTN is a nitrovasodilator prodrug acting via the formation of nitric oxide.

Quantitation of nitric oxide formation from nitrovasodilator drugs by chemiluminescence analysis of headspace gas

A rapid and reliable method has been developed for the quantitation of nitric oxide by chemiluminescence analysis of headspace gas. Aqueous nitric oxide standards are used to calibrate the method. There is a linear relationship between the amount of nitric oxide and the redox chemiluminescence detector response over the range of 52–2445 pmol of nitric oxide in 2.0 mL of deionized water contained in a sealed 6.2-mL flask. The intra-day and inter-day coefficient of variation values of the method do not exceed 4% and 9%, respectively. The lower limit of quantitative sensitivity and the lower limit of qualitative detection are 52 pmol and 26 pmol of nitric oxide, respectively. This method has been used to measure nitric oxide formation during the incubation of glyceryl trinitrate or sodium nitroprusside in the presence of cysteine. This method has two major advantages over the currently available procedures for the quantitation of nitric oxide, viz., no artifactual formation of nitric oxide during sample preparation and decreased instrumental contamination.

Superconductor metal oxide catalyst in a chemiluminescence chromatography detector

The superconducting oxygen-deficient perovskite, YBa 2Cu 3O 7, was used to catalyze redox reactions between organic species and NO 2 in a chromatography detector based upon chemiluminescence. The reactions were studied at catalyst temperatures ranging from 200°C to 350°C and at make-up gas flow-rates of 3.5–36.5 ml/min. By changing the flow-rate or the catalyst temperature, one can alter the sensitivity of analyte response in redox chemiluminescence and the selectivity ratios. Alcohols and alkenes responded the most strongly, with ethanol being detected at the level of 0.5 ng. Ammonia, acetaldehyde, acetone, methylethylketone, acetonitrile and nitromethane were also sensitively detected. Chromatographic analyses of oxygenates, e.g., methyl tert.-butyl ether or alcohols, in paraffinic matrices such as gasoline were demonstrated.

Coupling liquid chromatography with redox chemiluminescence detection

The post-column oxidation of many chemical species with dilute nitric acid, nitrous acid, or nitrite ion produces nitric oxide, which is subsequently detected by its chemiluminescence reaction with ozone. Organic compounds, such as sugars, react with dilute nitric acid at elevated temperature and pressure to produce a surrogate nitric oxide pulse for measurement on a chromatographically useful time scale. In addition, certain inorganic reducing agents, e.g., iron(II), react under similar conditions and are sensitively detected. Ultraviolet irradiation can initiate the redox reactions of easily oxidized inorganic species with aq. nitric acid at 25°C, and it also enhances the reactivity of analytes at elevated temperatures. Aqueous nitrate ion is not effective as a redox reagent, while nitrous acid and nitrite ion are less potent oxidizing agents than nitric acid. A post-column reaction system is described and the characteristics of redox reactions of potential analytes with aq. nitric acid and other reagents are discussed. Illustrative chromatograms which show the high-performance liquid chromatographic separation and chemiluminescence measurement of sugars at ppm levels are presented.

Supercritical fluid chromatography with redox chemiluminescence detection

The new selective and sensitive redox chemiluminescence detector (RCD), originally developed for gas chromatography, has been successfully interfaced with a capillary column supercritical fluid chromatograph. Interfacing the SFC-RCD was accomplished simply through utilization of the redox reaction chamber presently used for both capillary and packed column GC. The feasibility of this new separation-detection scheme was demonstrated with SFC chromatograms of antioxidant isomers and some compounds of biological relevance. Mobile phase compatibility and signal quenching are also discussed.

Chemistry occurring in redox chemiluminescence detectors for chromatography

Several aspects of the gold catalyzed reactions of NO2 with oxidizable compounds were investigated in order to understand the chemistry of selective redox chemiluminescence detectors. The interactions of NO2 and NO with gold have been studied using temperature programmed desorption techniques. NO2 is observed to bond with gold surfaces, and undergo rapid reaction with certain analytes (e.g., CO, H2, H2S, CS2, SO2 and various other organic compounds) to produce NO which is subsequently detected by chemiluminescence. The selectivity of the detector is dependent on the substrate supporting the layer of gold catalyst. With soda-lime glass beads coated with gold, n-alkanes do not react with NO2 to exhibit a response, while with borosilicate glass beads coated with gold, the alkanes react to produce NO. The gold-catalyzed oxidation products of citronellol and citronellal with NO2 include NO and a variety of cyclic compounds resulting from oxidative dehydrogenation.

Gas chromatographic measurement of carbon monoxide in hydrocarbon matrices with a redox chemiluminescence detector

The rapid measurement of trace levels of carbon monoxide in ethylene by gas chromatography with redox chemiluminescence detection is described. Linear response for carbon monoxide over three decades and a detection limit in the sub-parts per million by volume (ppmv) concentration range were observed without methanation or preconcentration of the sample. Samples containing 0.2 ppmv of carbon monoxide in ethylene were rapidly quantitated.

Chemistry occurring in redox chemiluminescence detectors for chromatography

Several aspects of the gold catalyzed reactions of NO 2 with oxidizable compounds were investigated in order to understand the chemistry of selective redox chemiluminescence detectors. The interactions of NO 2 and NO with gold have been studied using temperature programmed desorption techniques. NO 2 is observed to bond with gold surfaces, and undergo rapid reaction with certain analytes ( e.g., CO, H 2, H 2S, CS 2, SO 2 and various other organic compounds) to produce NO which is subsequently detected by chemiluminescence. The selectivity of the detector is dependent on the substrate supporting the layer of gold catalyst. With soda-lime glass beads coated with gold, n-alkanes do not react with NO 2 to exhibit a response, while with borosilicate glass beads coated with gold, the alkanes react to produce NO. The gold-catalyzed oxidation products of citronellol and citronellal with NO 2 include NO and a variety of cyclic compounds resulting from oxidative dehydrogenation.

Selectivity of the redox chemiluminescence detector for complex sample analysis

The redox chemiluminescence detector is a new selective detector for gas chromatography, based on catalysed redox reactions of NO 2 or HNO 3 with reducing agents to form NO, which is subsequently detected by the ozone chemiluminescence technique. By change of some of the operating parameters of the redox chemiluminescence detector, including the active metal used as the catalyst and the temperature of the catalyst bed, the selectivity of the detector can be tailored to a particular analysis. When an elemental gold catalyst is used at a relatively low temperature (200–300°C), only those compounds that can be easily oxidized, such as alcohols, are detected, while at higher temperatures (>350°C), wider ranges of compounds produce a response, including aldehydes and ketones. When gold is used as the catalyst, alkanes are not detected except at quite high catalyst temperatures (>450°C). When palladium is used in the catalyst bed, even at lower temperatures (<300°C) more compounds produce a redox chemiluminescence detector response, including alkanes. Thus the selectivity of the redox chemiluminescence detector can be controlled to permit selective analysis for a wide range of sample types.

Redox chemiluminescence detector: application to gas chromatography

Ce detecteur est base sur une reaction redox couplee avec la mesure de la chimiluminescence. Ce detecteur repond aux composes qui ne sont pas sensiblement detectes par les detecteurs FID

ChemInform Abstract: THE SPIN‐STATISTICAL CONTRIBUTION IN REDOX CHEMILUMINESCENCE QUANTUM EFFICIENCY

Chemischer InformationsdienstVolume 6, Issue 23 Physical Inorganic Chemistry ChemInform Abstract: THE SPIN-STATISTICAL CONTRIBUTION IN REDOX CHEMILUMINESCENCE QUANTUM EFFICIENCY CSABA P. KESZTHELYI, CSABA P. KESZTHELYISearch for more papers by this author CSABA P. KESZTHELYI, CSABA P. KESZTHELYISearch for more papers by this author First published: June 10, 1975 https://doi.org/10.1002/chin.197523020Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume6, Issue23June 10, 1975 RelatedInformation