SHORT COMMUNICATION Plasma nitrite concentration decreases after hyperoxia-induced oxidative stress in healthy humans Darko Modun 1 , Mladen Krnic 2 , Jonatan Vukovic 1 , Visnja Kokic 1 , Lea Kukoc-Modun 3 , Dimitrios Tsikas 4 and Zeljko Dujic 5 1 Department of Pharmacology, University of Split School of Medicine, Split, Croatia, 2 Department of Endocrinology, University Hospital Split, Split, Croatia, 3 Department of Analytical Chemistry, Faculty of Chemistry and Technology, University of Split, Split, Croatia, 4 Institute of Clinical Pharmacology, Hannover Medical School, Hannover, Germany, and 5 Department of Physiology, University of Split School of Medicine, Split, Croatia Summary Correspondence Darko Modun, Department of Pharmacology, Uni- versity of Split School of Medicine, Soltanska 2, 21000 Split, Croatia E-mail: drmodun@gmail.com Accepted for publication Received 31 December 2011; accepted 15 March 2012 Key words arterial stiffness; endothelium; nitric oxide; oxygen; vasoconstriction The aim of this study was to measure plasma nitrite, the biochemical marker of endothelial nitric oxide ( • NO) synthesis, before and after hyperoxia, in order to test the hypothesis that hyperoxia-induced vasoconstriction is a consequence of reduced bioavailability of • NO caused by elevated oxidative stress. Ten healthy men breathed 100% normobaric O 2 for 30 min between 15th and 45th min of the 1-h study protocol. Plasma nitrite and malondialdehyde (MDA), arterial stiff- ness (indicated by augmentation index, AIx) and arterial oxygen (P tc O 2 ) pressure were measured at 1st, 15th, 45th and 60th minute of the study. Breathing of nor- mobaric 100% oxygen during 30 min caused an increase in P tc O 2 (from 75 ± 2 to 412 ± 25 mm Hg), AIx (from À63 ± 4 to À51 ± 3%) and MDA (from 152 ± 13 to 218 ± 15 nM) values and a decrease in plasma nitrite (from 918 ± 58 to 773 ± 55 nM). During the 15-min recovery phase, plasma nitrite, AIx and MDA values remained altered. This study suggests that the underlying mechanism of hyperoxia-induced vasoconstriction may involve reduced • NO bioavailability caused by elevated and sustained oxidative stress. Introduction Molecular oxygen (O 2 ) is widely used in the treatment of a diverse range of acute medical conditions. Commonly, O 2 is often administered empirically, without prior knowledge of arterial oxyhaemoglobin saturation (Waring et al., 2003). As a result, patients may be exposed to significant periods of hyperoxia. However, hyperoxia causes vasoconstriction in both coronary and peripheral blood vessels (Crawford et al., 1997; McNulty et al., 2005; Rossi & Boussuges, 2005; Bak et al., 2007), and acutely increases arterial stiffness, as mani- fested by elevation of augmentation index (AIx), in healthy subjects (Waring et al., 2003). A potential mechanism by which hyperoxia may affect vas- cular reactivity is the generation of reactive oxygen species (ROS), notably the superoxide anion O ÀÁ 2 . In vitro, O ÀÁ 2 produced during hyperoxia reacts rapidly with • NO to form peroxynitrite (ONOO À ) (Rubanyi & Vanhoutte, 1986). This irreversible reaction decreases • NO bioavailability. ROS such as O ÀÁ 2 may also oxidize tetrahydrobiopterin (BH 4 ), an essential and crucial cofactor of endothelial • NO synthase (eNOS), thus impairing eNOS functionality (Mayer & Andrew, 1998). Clinical studies have shown that antioxidants, such as ascor- bic acid (Mak et al., 2002; McNulty et al., 2007; Gao et al., 2012) and uric acid (Vukovic et al., 2009), are protective against hyperoxia-induced deterioration of vascular reactivity in healthy and ill subjects. These findings are supportive of the hypothesis that hyperoxia-induced ROS formation is the culprit responsible for the reduction of • NO bioavailability. However, no solid evidence has been provided by the above-mentioned studies. Thus, in none of these studies was • NO bioavailability measured in the circulation before and after hyperoxia. Direct measurement of • NO within human blood vessels is very difficult because of the very short half-life of • NO (<0·1 s). Instead, circulating nitrite (NO À 2 ), the • NO autoxida- tion product, is susceptible for analysis and has been shown to reflect endothelial NO synthesis (Grau et al., 2007). The aim of the present study was to test the hypothesis that hyperoxia-induced vasoconstriction is a consequence of reduced • NO bioavailability caused by hyperoxia-induced Clin Physiol Funct Imaging (2012) 32, pp404–408 doi: 10.1111/j.1475-097X.2012.01133.x 404 © 2012 The Authors Clinical Physiology and Functional Imaging © 2012 Scandinavian Society of Clinical Physiology and Nuclear Medicine 32, 5, 404–408