Low plasma protein nitrotyrosine levels distinguish primary Raynaud's phenomenon from scleroderma.

OBJECTIVE: To investigate the hypothesis that increased formation of reactive nitrogen species may contribute to the vascular pathology that develops in patients with connective tissue disease such as scleroderma. PATIENTS AND METHODS: The level of protein-bound nitrotyrosine in plasma was measured by stable isotope dilution gas chromatography/negative ion chemical ionisation mass spectrometry in 11 patients with primary Raynaud's phenomenon, 37 with scleroderma, 13 with chronic renal impairment, and in 23 healthy controls. RESULTS: Plasma protein-bound nitrotyrosine was markedly decreased in patients with primary Raynaud's phenomenon (mean (SEM) 0.60 (0.06) ng/mg dry protein) compared with patients with scleroderma (1.78 (0.21) ng/mg protein), chronic renal impairment (1.42 (0.17) ng/mg protein) or healthy controls (1.63+/-0.15 ng/mg protein, ANOVA p<0.001). CONCLUSION: These data suggest that there is decreased nitration of plasma proteins, or increased degradation of nitrated proteins from the circulation of patients with primary but not secondary Raynaud's phenomenon.

Diagnostic monitoring of a changing environment: an alternative UK perspective.

Adaptive management of the marine environment requires an understanding of the complex interactions within it. Establishing levels of natural variability within and between marine ecosystems is a necessary prerequisite to this process and requires a monitoring programme which takes account of the issues of time, space and scale. In this paper, we argue that an ecosystem approach to managing the marine environment should take direct account of climate change indicators at a regional level if it is to cope with the unprecedented change expected as a result of human impacts on the earth climate system. We discuss the purpose of environmental monitoring and the importance of maintaining long-term time series. Recommendations are made on the use of these data in conjunction with modern extrapolation and integration tools (e.g. ecosystem models, remote sensing) to provide a diagnostic approach to the management of marine ecosystems, based on adaptive indicators and dynamic baselines.

The ingestion of inorganic nitrate increases gastric S-nitrosothiol levels and inhibits platelet function in humans.

Platelets play an important role in the development of vascular disease, while vegetarian diets, which are rich in inorganic nitrate, protect against it. This study was performed to assess the effect of potassium nitrate (KNO(3)) ingestion on platelet function in humans. Oral KNO(3) (2 mmol) was given to healthy volunteers and its effect on platelet function assessed by measuring the aggregant effect of collagen. Blood samples were taken for measurement of plasma S-nitrosothiols (RSNO) and platelet cyclic GMP and nitrotyrosine levels. Gastric juice samples were taken for measurement of RSNO. In a separate study, the effect of oral KNO(3) on portal RSNO levels in patients with intrahepatic porto-systemic shunts was assessed. KNO(3) caused a significant increase in gastric RSNO levels, from 0.46 +/- 0.06 to 3.62 +/- 2.82 microM (t(max) 45 min; P < 0.001), and significantly inhibited platelet function (t(max) 60 min; P < 0.001). There was no effect on systemic or portal RSNO, platelet cGMP or platelet nitrotyrosine levels. Oral KNO(3) inhibits platelet aggregation. The time course suggests that gastric RSNO production may be involved in this effect. The protection against vascular events associated with a high intake of vegetables may be due to their high nitrate content.

Analysis of free and protein-bound nitrotyrosine in human plasma by a gas chromatography/mass spectrometry method that avoids nitration artifacts.

Measurement of nitrotyrosine in biological fluids and tissues is increasingly being used to monitor the production of reactive nitrogen species in vivo. The detection of nitrotyrosine in vivo has been reported with the use of a variety of methods including immunoassay, HPLC and GLC/MS. The validity of HPLC and immunoassays have been questioned with regard to their selectivity and sensitivity limits. In principle, the measurement of nitrotyrosine by GLC/MS permits a highly specific, highly sensitive and fully quantitative assay. The nitration of tyrosine under acidic conditions in the presence of nitrite is well documented. Derivatization for the full quantification of nitrotyrosine by using GLC/MS can lead to the artifactual nitration of tyrosine if performed under acidic conditions in the presence of nitrite. We describe a novel alkaline method for the hydrolysis and derivatization of nitrotyrosine and tyrosine, and demonstrate its applicability to the measurement of plasma concentrations of both free and protein-bound nitrotyrosine and tyrosine. A detection limit of 1 pg for nitrotyrosine and 100 pg for tyrosine has been achieved. Our method allows, for the first time, the analysis of free and protein-bound nitrotyrosine and tyrosine in biological samples. The plasma concentrations (means+/-S.E.M.) of free tyrosine and nitrotyrosine in eight normal subjects were 12+/-0.6 microg/ml and 14+/-0.7 ng/ml respectively. Plasma proteins contained tyrosine and nitrotyrosine at 60.7+/-1.7 microg/mg and 2.7+/-0.4 ng/mg respectively.