Simultaneous determination of reduced glutathione, glutathione disulphide and glutathione sulphonamide in cells and physiological fluids by isotope dilution liquid chromatography–tandem mass spectrometry

Harwood, D. Tim; Kettle, Anthony J.; Brennan, Siobhain; Winterbourn, Christine C.

doi:10.1016/j.jchromb.2009.04.018

Cited by 69 publications

(51 citation statements)

References 45 publications

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“…Analysis of GSH and its oxidation products in BAL GSH, GSSG and GSA was analysed by stable isotope dilution liquid chromatography tandem mass spectrometry assay (LC-MS/MS) as previously described [24]. Aliquots of standards (0?2 mL) or BAL (a volume containing 50 mg total protein made up to 0?2 mL) were mixed with isotopically labelled GSH, GSSG and GSA as internal standards.…”

Section: Biochemical Analysismentioning

confidence: 99%

Oxidation contributes to low glutathione in the airways of children with cystic fibrosis

et al. 2014

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Glutathione is an important antioxidant in the lungs but its concentration is low in the airways of patients with cystic fibrosis. Whether this deficit occurs from an early age or how oxidative stress contributes to lowering glutathione is unknown.We measured glutathione, its oxidation products, myeloperoxidase, and biomarkers of hypochlorous acid in bronchoalveolar lavage from children with cystic fibrosis and disease controls using mass spectrometry and immunological techniques.The concentration of glutathione was lower in bronchoalveolar lavage from children with cystic fibrosis, whereas glutathione sulfonamide, a specific oxidation product of hypochlorous acid, was higher. Oxidised glutathione and glutathione sulfonamide correlated with myeloperoxidase and a biomarker of hypochlorous acid. The percentage of glutathione attached to proteins was higher in children with cystic fibrosis than controls. Pulmonary infections in cystic fibrosis resulted in lower levels of glutathione but higher levels of oxidised glutathione and glutathione sulfonamide in bronchoalveolar lavage.The concentration of glutathione is low in the airways of patients with cystic fibrosis from an early age. Increased oxidation of glutathione by hypochlorous acid and its attachment to proteins contribute to this deficiency. Therapies targeted against myeloperoxidase may boost antioxidant defence and slow the onset and progression of lung disease in cystic fibrosis.

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Section: Biochemical Analysismentioning

confidence: 99%

Oxidation contributes to low glutathione in the airways of children with cystic fibrosis

et al. 2014

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“…GSA was analyzed by stable isotope dilution liquid chromatographytandem mass spectrometry assay (LC-MS/MS) using a Thermo Finnigan LCQ Deca XP Plus Ion Trap mass spectrometer (San Jose, CA) coupled to a Thermo Hypercarb column as described (14). Aliquots of standards (0.2 mL) or ET aspirates (a volume containing 50 g total protein made up to 0.2 mL) mixed with isotopically labeled GSA (10 L of 2 M stock solution) as internal standard.…”

Section: Methodsmentioning

confidence: 99%

“…GSA was prepared by treating GSH with reagent HOCl in phosphate buffer (100 mM, pH 7.4), with the internal standard generated using [glycine 1,2-13 C 2 , 15 N]-GSH. Labeled and unlabeled GSA were purified and used to construct a calibration curve based on mass (14). GSA was stable in buffer or aspirates for at least a year at Ϫ20°C.…”

Section: Methodsmentioning

confidence: 99%

“…Oxidation of intracellular and epithelial lining fluid GSH would be expected if HOCl is generated in the lungs and GSA should be produced, in addition to glutathione disulfide (GSSG). We have developed a sensitive method for detecting GSA in lung lavage fluid (14), but quantitative measurements in clinical samples have not been reported. Also, it has not been established whether GSA is a useful biomarker of MPO activity and neutrophil activation.…”

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confidence: 99%

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Biomarkers of Neutrophil-Mediated Glutathione and Protein Oxidation in Tracheal Aspirates From Preterm Infants: Association With Bacterial Infection

et al. 2011

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Bronchopulmonary dysplasia is associated with neutrophil infiltration into the lungs and oxidative injury. However, the pathological importance of neutrophil oxidants is still not clear. Nosocomial pneumonia is also implicated, but the evidence is limited, in part because of the difficulty of distinguishing genuine infection from bacterial colonization. Good biomarkers of neutrophil oxidant activity and lung infection are needed. We tested whether glutathione sulfonamide, a product of glutathione oxidation by myeloperoxidase-derived hypochlorous acid (HOCl) and a potential new neutrophil oxidant biomarker, is detectable in endotracheal aspirates from ventilated preterm infants. As infectious organisms stimulate neutrophils to generate HOCl, we determined whether levels of HOCl-specific biomarkers were increased in samples that were bacterial culture positive. Glutathione sulfonamide was detected in 66 of 87 endotracheal aspirate samples. Levels correlated with myeloperoxidase activity and another HOCl-specific marker, chlorotyrosine. Median levels of glutathione sulfonamide (4-fold) and other biomarkers (2-fold) were significantly higher in culture positive aspirates. Staphylococcus epidermidis, a frequent colonizer, was associated with glutathione sulfonamide levels no different from those in negative samples. Glutathione sulfonamide showed good sensitivity and specificity for detecting bacterial growth and has promise for detecting lung infection. (Pediatr Res 69: 28-33, 2011)

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“…Reduced (GSH) and oxidized (GSSG) glutathione were determined by ultraperformance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) following the procedure of Harwood et al (39) with some modifications based on Nishiyama and Kuninori's method of determination of thiols (40). Briefly, lung tissue was homogenized in the presence of N-ethylmaleimide 10 mmol/l dissolved in phosphate-buffered saline at a 4:1 (vol/wt).…”

Section: Glutathione Determinationmentioning

confidence: 99%

Sex-dependent changes in the pulmonary vasoconstriction potential of newborn rats following short-term oxygen exposure

et al. 2012

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Background: chronic exposure to supplemental oxygen (O 2 ) induces lung damage and mortality in a sex-dependent manner. The effect of short-term hyperoxia on the newborn pulmonary vasculature is unknown but is, however, of clinical significance in the neonatal resuscitation context. We hypothesize that short-term hyperoxia has a sex-dependent effect on the pulmonary vasculature. Methods: Following 1-h 100% O 2 exposure, the pulmonary arteries and lung tissues of newborn rats were evaluated. results: superoxide dismutase 3 (sOD3) expression in female pups' lungs was increased as compared with that in the lungs of male pups. as compared with air-treated pups, the response of male pups to thromboxane was increased by O 2 , whereas the opposite effect was documented in the vessels of female pups. The enhanced force of hyperoxia-exposed arteries of the male pups was suppressed with superoxide or peroxynitrite scavengers, and increased lung sOD activity and hydrogen peroxide content were seen in female, but not in male, rats. hyperoxia induced an increase in lung tissue oxidative products and Rho-kinase (ROcK) activity in male, but not in female, pups. conclusion: a lower lung sOD content and failure to upregulate sOD activity facilitates peroxynitrite generation and ROcK activation in hyperoxia-exposed males, predisposing them to pulmonary vasoconstriction. These observations, if relevant to humans, may explain the increased mortality and higher incidence of pulmonary hypertension in male neonates.

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Simultaneous determination of reduced glutathione, glutathione disulphide and glutathione sulphonamide in cells and physiological fluids by isotope dilution liquid chromatography–tandem mass spectrometry

Cited by 69 publications

References 45 publications

Oxidation contributes to low glutathione in the airways of children with cystic fibrosis

Oxidation contributes to low glutathione in the airways of children with cystic fibrosis

Biomarkers of Neutrophil-Mediated Glutathione and Protein Oxidation in Tracheal Aspirates From Preterm Infants: Association With Bacterial Infection

Sex-dependent changes in the pulmonary vasoconstriction potential of newborn rats following short-term oxygen exposure

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