Introduction: Diesel fuel (D) is used in a variety of applications for several industries, including transportation, agriculture, railroads, construction, and mining. In addition to being non-renewable, combustion of diesel fuel (D) leads to negative occupational health outcomes in mining. Currently the Mine Safety and Health Administration (MSHA) regulates diesel exhaust exposure with an 8-hour, time-weighted average permissible exposure limit (PEL) of 160 µg/m3 respirable (<1.0 µm in size) diesel particulate matter (rDPM). Alternative fuels such as biodiesel (B) and a natural gas/diesel blend (GDiesel® [G]) are considered promising alternatives. While the former fuel has been extensively investigated, the latter has not. Objectives: The aims of this dissertation were: (1) to evaluate and compare D and G exhaust exposures from operation of a Wagner and a (2) JCI load-haul-dump (LHD) at the University of Arizona San Xavier Underground Mining Laboratory (SX); and (3) to synthesize existing peer-reviewed literature comparing D emission exposures to those of B and/or G. Methods: For Aims 1 and 2, operator-location and area exposure samples were collected for 200 minutes in an underground mining laboratory while an LHD with oxidation catalyst was operated with D and then G fuel. Analytes of interest included total diesel particulate matter (tDPM) and rDPM, total and respirable elemental and organic carbon (tEC, rEC, tOC, rOC, respectively), as well as the carbonyl compound (CC) formaldehyde (CH2O), nitric oxide (NO), and nitrogen dioxide (NO2). Exposure assessment was conducted within the guidelines of the National Institute of Occupational Safety and Health’s (NIOSH) Manual of Analytical Methods. Specifically, methods #5040 (tDPM, tEC, tOC, rDPM, rEC, rOC), #6014 (NO, NO2), and #2016 (CH2O). Reported laboratory results were time-weighted over an 8-hour period. Between-fuel comparisons were performed using Wilcoxon rank sum testing. Results: For Aim 1, twenty-three D and 12 G samples were collected. Use of G in the Wagner LHD showed statistically and practically significant reductions in rDPM, tDPM, elemental and organic carbons, NO, NO2, and CH2O. For Aim 2, twenty D and 16 G samples were collected. Use of G in the JCI LHD was associated with a significant decrease in NO2 (p=0.012), and significant increase in rEC (p=0.024). After removing outliers, tEC also showed significant increase (p=0.023). Most of the 20 scholarly works reviewed utilized a laboratory setting (75%), while just 15% were conducted in the field, and 10% simulated field conditions. Twenty percent (4) of studies specifically focused on the mining industry. In addition, most evaluated soy-based B (56%) but did not utilize pollution controls (70%) on equipment. Generally, literature showed that use of B decreased DPM and increased oxides of nitrogen (NOx) emission exposures. While more studies (5) showed increases in CCs, two showed decreases. Discussion: Our studies show that: 1) the use of G has potential for statistically and practically significant reductions in several D exhaust contaminants regulated by MSHA; and 2) variability in exposure and emission concentrations across engine, pollution control and operation configurations exists for B and likely exists for G. Differences observed across fuels and studies are also likely due to fuel composition and characteristics, and combustion temperatures. Further occupational health research is needed to evaluate G emissions under controlled conditions with various equipment configurations, as well as in-field settings to determine whether G exhaust exposures are reduced and actually less toxic than those of D. The impact of this work is substantial and timely. Recent increases in respiratory disease prevalence among miners, including young miners, concerns occupational health and industrial hygiene professionals. In addition, MSHA has requested information regarding diesel exhaust controls and is considering future revisions to the rDPM standard. Efforts to reduce D exhaust emissions will also impact occupational and environmental health worldwide.

One of the most prevalent pollutants in urban cities is diesel exhaust particulate (DEP). Air pollution has been linked with increased risk of recurrent myocardial infarction (MI) and MI related death (Brook, 2008). This may be due, in part, to effects on atherosclerotic plaque stability and blood clotting tendency. Whether exposure to DEP changes the response of the heart to ischaemia, resulting in increased damage after MI is less well documented. The work described in this thesis was designed to investigate the hypothesis that pulmonary instillation of DEP would increase vulnerability of the heart to subsequent myocardial reperfusion injury secondary to activation of a systemic inflammatory response, endothelial dysfunction and triggering of transient receptor potential vanilloid 1 (TRPV1) mediated autonomic reflexes in the lung. Examination of bronchoalveolar lavage (BAL) fluid revealed pulmonary inflammation 6 h after exposure to DEP, characterised by neutrophil infiltration, raised levels of the inflammatory mediator interleukin-6 (IL-6) and an increase in alveolar permeability demonstrated by increased levels of protein in the lavage fluid. Pulmonary inflammation was largely resolved 24 h after exposure. While there was no indication of systemic inflammation at 6 h after DEP instillation, the levels of two inflammatory mediators, IL-6 and tumour necrosis factor alpha (TNFα) were increased in the plasma by 24 h after exposure. DEP had no affect on blood flow responses to the endothelium dependent dilator acetylcholine (ACh) in rat hind-limb vasculature in vivo at 6 or 24 h. In summary, while exposure of rats to DEP can induce both pulmonary and systemic inflammation, it does not modify endothelium-dependent vasodilatation. Ischaemia-reperfusion (I/R) was induced in vivo in anaesthetised rats and ex vivo in buffer perfused hearts from rats that had received DEP in vivo 6 h earlier. In both in vivo and ex vivo I/R models, infarct size (unstained by triphenyltetrazolium choride) was significantly increased in hearts from DEP-instilled rats relative to hearts from saline-instilled or non-instilled rats. Baseline oxidant stress, determined by electron paramagnetic spin resonance (EPR) in heart perfusate, was also significantly higher in perfusate of hearts from DEP-instilled rats. In summary, a single exposure of the lung to DEP leads to priming of the myocardium for I/R injury. As the results cited above illustrated, priming of hearts appeared unlikely to be due to either coronary vascular endothelial dysfunction or systemic inflammation. At 6 h post exposure, DEP was associated with increased blood pressure and myocardial hypersensitivity to ischaemia-induced arrhythmias, both suggestive of sympathetic activation. The beta 1 (β1) selective blocker metoprolol was used to investigate the role of the sympathetic nervous system (SNS) in transmitting the influence of DEP in the lung to the myocardium via β1 adrenoceptor activation. Administration of metoprolol (10 mg/kg, intraperotineal) at the time of DEP instillation into the lung was found to protect the heart from potentiation of ex vivo reperfusion injury. Metoprolol was also effective in reducing oxygen free radical generation from these hearts. The TRPV1 antagonist AMG 9810 was also used to study the role of TRPV1 receptors in mediating the priming influence of pulmonary DEP to the myocardium since activation of sensory receptors have been reported to modify sympathetic output via feedback to the central nervous system (Widdicombe et al., 2001). Coadministration of AMG 9810 (30 mg/kg) in vivo with DEP into the lung was found to prevent enhancement of ex vivo reperfusion injury associated with DEP instillation alone. Collectively these results have demonstrated that a single exposure of the lung to DEP leads to priming of the myocardium for I/R injury. Furthermore, this priming occurs via activation of a pulmonary sensory reflex that is likely to involve secondary activation of systemic β1 adrenoceptors.

Increases in air pollution, especially the particulate component, are associated with increased cardiovascular mortality, possibly through increases in thrombogenic mechanisms. The research presented in this thesis addresses the hypothesis that diesel exhaust particulates (DEP) increase thrombogenicity by impairing the release of tissue plasminogen activator (t-PA) from vascular endothelial cells, thus inhibiting the endogenous fibrinolytic mechanisms that promote thrombus breakdown. The initial aims of this work were to develop an in vivo model of thrombosis, to determine whether exposure to DEP did alter clotting. Initial attempts to develop the Folts’ model (which stimulates thrombus formation via arterial stenosis and mechanical injury), first in male C57/Bl6 mice and later in male Wistar rats, were unsuccessful. An alternative approach, using ferric chloride (FeCl3) to induce chemical injury to the rat carotid artery was found to produce reliable and reproducible thrombotic occlusion: this model was used for all subsequent experiments. The effects of DEP on thrombus formation were assessed in vivo by applying the FeCl3 model. DEP were administered via intratracheal instillation or tail vein injection 2, 6 or 24 hours prior to induction of thrombosis. The effects of DEP were compared with vehicle and suitable controls: carbon black (a clean carbon nanoparticle); quartz (a large non-carbon particle that causes well-characterised pulmonary inflammation). The time to thrombotic occlusion was significantly reduced 6h after intra-pulmonary instillation of DEP (0.5ml of a 1mg/ml suspension). In contrast, instillation of carbon black or quartz had no significant effect on thrombosis, despite causing greater pulmonary (increased neutrophils and levels of interleukin-6, tumour necrosis factor-α and C-reactive protein in bronchoalveolar lavage fluid) and systemic (C-reactive protein in plasma) inflammation than DEP. Direct administration of DEP (0.5mg/kg) to the blood stream resulted in an acute (2 hours after injection) increase in time to thrombotic occlusion in the absence of pulmonary inflammation. A similar (but less pronounced) effect was observed following administration of carbon black (0.5mg/kg). These data suggest that the DEP-mediated increase in thrombosis is independent of pulmonary and systemic inflammation. The mechanisms involved were addressed by measuring platelet-monocyte interactions (flow cytometry) and markers of the endogenous fibrinolytic system (ELISA). Exposure (either instillation of injection) to DEP significantly increased platelet-monocyte aggregation. Carbon black and quartz produced no such effect (but did increase platelet-platelet aggregation). t-PA antigen and activity were reduced, whilst PAI-1 and fibrinogen were increased, following either instillation or injection of DEP. The final aim was to develop a suitable dispersant for use in cell culture to determine whether DEP alter the expression (real-time polymerase chain reaction; rtPCR) and generation (enzyme-linked immunosorbent assay; ELISA) of t-PA and plasminogen activator inhibitor (PAI-1). Cell culture medium containing bovine serum albumin (0.5mg/ml; BSA) provided the best combination for DEP dispersal and maintenance of small particle size (<200nM), without detrimental effects on human umbilical endothelial cells (HUVECs). Exposure (6 and 24 hours) of HUVECs to DEP resulted in reduced basal and thrombin stimulated t-PA and PAI-1 expression. This was mirrored by reduced detection of t-PA and PAI-1 in culture medium. In conclusion, these investigations confirm that exposure to DEP is capable of increasing the rate of thrombus formation and that this is, in part, mediated by an alteration in the endogenous fibrinolytic system. These changes did not appear to be secondary to pulmonary or systemic inflammation. Whilst cell culture experiments suggested DEP could directly alter endogenous fibrinolytic activity in endothelial cells, there was no evidence from these experiments of DEP translocation into the systemic circulation. Thus, this work suggests that DEP is capable of increasing thrombus formation in vivo via several mechanisms. Similar changes may account for the increased thrombus formation in humans exposed to diesel exhaust in air pollution.

No description available.

Fine particulate matter (PM2.5) constitutes a significant component of ambient air pollution that has been implicated in the pathogenesis of metabolic disorders, including insulin resistance and type 2 diabetes. Among PM2.5 constituents, diesel exhaust particles (DEP) are prevalent particulates that infiltrate the bloodstream to drive systemic pathologies. The purpose of this study was to characterize the metabolic response of adipose tissue to DEP. We aimed to provide a comprehensive understanding by exploring mitochondrial bioenergetics, characterizing the inflammatory marker profile, including adipokines, and conducting a detailed histological analysis of adipocytes to provide valuable insights to the evolving understanding of the intricate interplay between pollution and adipose tissue function. Following daily inhalation exposure to DEP in mice, we observed a selective increase in adipose tissue mass and altered mitochondrial respiration in the adipose tissue. Furthermore, we observed increased pro-inflammatory cytokines, changes in adipokine secretion, and alterations in adipose histology reflective of adipocyte hypertrophy. In conclusion, exposure to DEP disrupts adipose tissue function by altering adipocyte mitochondrial function and contributing to inflammation. These novel findings provide valuable insights that may facilitate the development of therapeutic interventions addressing metabolic disorders in the future.

This thesis describes an investigation into the spatial species conversion profiles of a Cu-zeolite SCR under engine conditions at low exhaust gas temperatures; this was then compared with a CFD model that models the catalyst via a porous medium measuring 5 x 5 x 91 cells assuming a uniform cross-sectional flow distribution. Species conversion rates were sampled at fixed points in the axial direction. The analysis of the spatial conversion profiles is a more rigorous method in assessing the ability of a mathematical model to predict the experimental data. It can also assist in the optimisation of the catalyst size, minimising packaging requirements and manufacturing costs. The experiments were undertaken on a light-duty diesel engine at a speed of 1500rpm, and at a load of 6bar BMEP; this provided exhaust gas temeraqtures between 200 and 220°C. NO2:NOx ratios were controlled by changing the size and position of the diesel oxidation catalyst, the inlet NH3: NOx ratio was also also varied, ammonia gas was used instead of urea for the purposes of simlicity. The advantage of testing on an actual engine over lab-babed studies is that the conditions such as exhaust gas composition are more realistic. A 1D CFD model was constructed using the ‘porous medium approach’ with kinetics obtained from open literature. Results from the simulations were then compared with the experimental data for the same engine conditions. It was observed that the majority of the NOx conversion took place in the first half of the brick for all NH3: NOx ratios investigated, and that the formation of N2O via NO2 and ammonia had the same influence as the ‘fast’ SCR reaction just after the inlet, which the CFD model failed to predict for the base case analyses. The influence of the inlet ammonia on the model was also noticed to be greater than in the experiments. Simple transient analyses were also undertaken on the short SCR bricks for NO2: NOx ratios of 0.6 and 0.07, and it was observed that the response time to steady-state was noticeably higher in the experiments than in the model. Modifications made to the model, including decreasing the influence of the ‘fast’ SCR reaction, and the addition of an empirical term onto the ammonia adsorption provided a noticeably better agreement for different NH3: NOx injection ratios. The desorption kinetics in the model were also altered by increasing the strength of the bonding of the ammonia onto the adsorption sites. This improved the transient agreement between the model and the experiments, but reduced the steady-state concentrations at the exit of the brick for all NH3:NOx ratios investigated.

Exposure to air pollution, particularly fine and ultrafine particulate matter derived from combustion sources, has been consistently associated with increased cardiovascular morbidity and mortality. Recent controlled exposure studies demonstrate that short-term exposure to diesel exhaust, which can contribute up to 40% of urban particulate air pollution, results in impaired vascular endothelial and fibrinolytic function in healthy volunteers, and increased exercise-induced myocardial ischaemia in patients with coronary heart disease. These observations may, in part, explain the observed increase in cardiovascular events following exposure to air pollution. Despite these observations there remain uncertainties regarding the key constituents of the air pollution mixture that mediate these adverse effects, and the underlying physiological and biological pathways involved. In these studies, using two controlled exposure facilities, I explored the vascular effects of the most prevalent gaseous component of the air pollution mixture – nitrogen dioxide – and the mechanisms responsible for impaired vasomotor function following exposure to diesel exhaust. Furthermore, I investigated the effect of acute exposure to “real-world” urban air pollution in both healthy volunteers and patients with coronary heart disease, and the effect of reducing that exposure using a simple facemask. In total, 10 healthy volunteers were exposed to nitrogen dioxide, and 29 healthy volunteers exposed to dilute diesel exhaust in a series of doubleblind randomised crossover studies. Exposure to nitrogen dioxide had no effect on either vasomotor function or endogenous fibrinolysis, providing indirect evidence that the adverse vascular effects are predominantly driven by particulate components. Following exposure to diesel exhaust there was no up regulation of endothelin-1 production, although there was increased vasoconstriction to intra-arterial infusion of endothelin-1. Following endothelin A receptor antagonism, there was attenuated vasodilatation following exposure to diesel exhaust as compared to air, an effect abrogated by endothelin B receptor antagonism. My findings suggest that the endothelin system does not play a central role in the adverse vascular effects of air pollution, but given the tonic interaction between the endothelin and nitric oxide systems, these observations could be explained by reduced nitric oxide bioavailability. Following diesel exhaust inhalation, plasma nitrite concentrations (as a marker for nitric oxide generation) are markedly increased without changes in haemodynamics or basal blood flow consistent with increased nitric oxide consumption. In the presence of a nitric oxide clamp, and without endogenous nitric oxide release, the vascular responses to vasodilators are similar. This perturbation of nitric oxide consumption and release appears to underlie the observed vascular endothelial effects. Fifteen healthy volunteers and 98 patients with coronary artery disease were recruited in Beijing, China. Subjects walked along a predefined city centre route for 2 hours in the presence and absence of a highly efficient facemask to reduce personal particulate air pollution exposure in an open label randomised crossover study. When wearing a facemask, there was an attenuation of exercise-induced increases in blood pressure, an improvement in heart rate variability, reduced myocardial ischaemia and subjects reported fewer symptoms. My findings have identified the biological mechanisms underlying the adverse vascular effects of exposure to diesel exhaust, and have helped to clarify the components responsible for these effects. Moreover, I have identified important benefits of reducing personal exposure to particulate matter using a simple facemask that have the potential to reduce cardiovascular events in patients living in urban or industrialised areas. Ongoing research in this area will provide further insight into the underlying vascular mechanisms, and the potential benefits of reducing particulate air pollution exposure, and may result in important targeted interventions to reduce the impact of air pollution on cardiovascular health.

Background: Air pollution is associated with negative health effects. Exposure to combustion-derived particulate matter (PM) air pollution has been related to increased incidence of cardiovascular and respiratory morbidity and mortality, specifically in susceptible populations. Ambient particles, with a diameter of less than 2.5 mm, have been suggested to be the strongest contributor to these health effects. Diesel exhaust (DE) is a major source of small combustion-derived PM air pollution world wide.  In healthy volunteers, exposure to DE, has been associated with airway inflammation and impaired vasomotor function and endogenous fibrinolysis. The aims of this thesis were to further elucidate the underlying mechanisms to the reported cardiovascular effects following exposure to DE, with specific focus on endothelin-1 (ET-1). Additionally, the vascular effects of the major gaseous component of DE, nitrogen dioxide (NO2), were assessed together with the impact of an exhaust particle trap to reduce the observed negative vascular effects after DE exposure. Methods: In all studies healthy, non-smoking male volunteers were included and exposed for one hour during intermittent exercise in a randomised double-blind crossover fashion. In studies I-III, subjects were exposed to DE at a particulate matter concentration of approximately 300 μg/m3 and filtered air, on two different occasions. In study V an additional exposure was employed, during which DE was filtered through an exhaust particle trap. In study IV subjects were exposed to nitrogen dioxide (NO2) at 4 ppm or filtered air. In study I, thrombus formation and platelet activation were assessed using the Badimon ex vivo perfusion chamber and flow cytometry. Study II comprised the determination of arterial stiffness including pulse wave analysis and velocity. In studies III-V, vascular assessment was performed using venous occlusion plethysmography. In studies IV and V, the vascular responses to intra-arterially infused endothelial-dependent and endothelial-independent vasodilatators were registered. In study III, vascular responses to intra-arterial infusion of Endothelin-1 (ET-1) and ET-1-receptor antagonists were assessed. Venous occlusion phlethysmography was in all cases performed 4-6 hours following exposures. Blood samples for markers of inflammation, coagulation and platelet activation were collected before and throughout the study periods in studies III and V. Results: Exposure to DE increased ex vivo thrombus formation and arterial stiffness, in terms of augmentation index. DE inhalation impaired vasomotor function and endogenous fibrinolysis. The exhaust particle trap reduced the particle concentration by 98% and abolished the effects on vasomotor function, endogenous fibrinolysis and ex vivo thrombus formation. Plasma concentrations of ET-1 and its precursor big-ET-1 were unchanged following exposure. Dual endothelial receptor antagonism caused similar vasodilatation after both exposures, although vasodilatation to the endothelin-A receptor alone was blunted after DE exposure. ET-1 infusion induced vasoconstriction only following DE exposure. Exposure to nitrogen dioxide did not affect vascular function. Conclusion: Inhalation of diesel exhaust in young healthy men impaired important and complementary aspects of vascular function in humans; regulation of vascular tone and endogenous fibrinolysis as well as increased ex vivo thrombus formation. The use of an exhaust particle trap significantly reduced particle emissions and abolished the DE-induced vascular and prothrombotic effects. The adverse vascular effects following DE exposure do not appear to be directly mediated through the endothelin system. Neither is NO2 suggested to be a major arbiter of the DE-induced cardiovascular responses. Arterial stiffness is a non-invasive and easily accessible method and could thus be employed to address vascular function in larger field studies. Taken together, this thesis has given further knowledge about the mechanisms underlying the DE-induced vascular effects.

It is well established that air pollution has detrimental effects on both human health as well as the environment. Exposure to ozone and particulate matter pollution, is associated with an increase in cardiopulmonary mortality and morbidity. Asthmatics, elderly and children have been indicated as especially sensitive groups. With a global increase in use of vehicles and industry, ambient air pollution represents a crucial health concern as well as a political, economical and environmental dilemma. Both ozone (O3) and diesel exhaust (DE) trigger oxidative stress and inflammation in the airways, causing symptoms such as wheezing, coughing and reduced lung function. The aim of this thesis was to further examine which pro-inflammatory signaling pathways that are initiated in the airways by ozone, as compared to diesel exhaust. Furthermore, to study the effects of these two ambient air pollutants in a sequential exposure, thus mimicking an urban profile. In order to investigate this in healthy as well as asthmatic subjects, walk-in exposure chambers were utilized and various airway compartments were studied by obtaining induced sputum, endobronchial biopsies, or airway lavage fluids. In asthmatic subjects, exposure to 0.2 ppm of O3 induced an increase in the cytokines IL 5, GM-CSF and ENA-78 in the bronchial epithelium six hours post-exposure. The healthy subjects, however, displayed no elevations of bronchial epithelial cytokine expression in response to the ozone exposure. The heightened levels of neutrophil chemoattractants and Th2 cytokines in the asthmatic airway epithelium may contribute to symptom exacerbations following air pollution exposure. When examining an earlier time point post O3 exposure (1½ hours), healthy subjects exhibited a suppression of IL-8 as well as of the transcription factors NFκB and c-jun in the bronchial epithelium, as opposed to after filtered air exposure. This inhibition of early signal transduction in the bronchial epithelium after O3 differs from the response detected after exposure to DE. Since both O3 and DE are associated with generating airway neutrophilia as well as causing direct oxidative damage, it raises the query of whether daily exposure to these two air pollutants creates a synergistic or additive effect. Induced sputum attained from healthy subjects exposed in sequence to 0.2 ppm of O3 five hours following DE at a PM concentration of 300 µg/m3, demonstrated significantly increased neutrophils, and elevated MPO levels, as compared to the sequential DE and filtered air exposure. O3 and DE interactions were further investigated by analyses of bronchoalveolar lavage and bronchial wash. It was demonstrated that pre-exposure to DE, as compared to filtered air, enhances the O3-induced airway inflammation, in terms of an increase in neutrophil and macrophage numbers in BW and higher EPX expression in BAL. In conclusion, this thesis has aspired to expand the knowledge of O3-induced inflammatory pathways in humans, observing a divergence to the previously described DE initiated responses. Moreover, a potentially adverse airway inflammation augmentation has been revealed after exposure to a relevant ambient combination of these air pollutants. This provides a foundation towards an understanding of the cumulative airway effects when exposed to a combination of ambient air pollutants and may have implications regarding future regulations of exposure limits.

Air pollutants, such as ozone (O3) and diesel exhaust particles, elicit oxidative stress in the lung. Antioxidants within the respiratory tract lining fluid (RTLF) protect the underlying tissue from oxidative injury. Supplementation with vitamins has been shown to modulate the acute ozone-induced effects, but the mechanisms behind this have not been fully clarified. The aim of this thesis was to investigate the airway responses to diesel exhaust and ozone exposure in healthy humans, with the emphasis on inflammatory and antioxidant responses. Furthermore, to study whether oral supplementation with vitamin C could increase ascorbate concentration in the RTLF and whether vitamin supplementation could modulate the negative effects induced by ozone exposure. Diesel exhaust (100 µg/m3 PM10 for 2h), evaluated 18 hours post exposure (PE), induced a neutrophilic airway inflammation and an increase in bronchoalveolar (BAL) urate and reduced glutathione. During O3 exposure (0.2 ppm for 2h), significant losses of nasal RTLF urate and ascorbate concentrations were observed. Six hours PE, a neutrophilic inflammation was evident in the bronchial wash (BW), together with enhanced concentrations of urate and total glutathione. In the bronchoalveolar lavage (BAL), where vitamin C, urate and glutathione concentrations were augmented, no inflammatory response was seen. In alveolar lavage leukocytes, there was a significant loss of glutathione and cysteine, whereas an increase in ascorbate was found in bronchial tissue samples. Following supplementation with increasing doses of vitamin C (60-1,000 mg/day, for 14 days), evaluated 24 hours after the last dose, ascorbate concentrations were unchanged in the nasal RTLF, despite elevated concentrations in plasma and urine. In contrast, following a single dose of 1g of vitamin C, vitamin C concentrations increased significantly in both plasma and nasal lavage two hours post supplementation, before returning to baseline levels at 24 hours. Notably, dehydroascorbate (DHA) accounted for the largest part of RTLF vitamin C and a number of control experiments were performed to ensure the authenticity of this finding. Healthy O3 responders were exposed to O3 (0.2 ppm for 2 h) and air, following seven days of supplementation with vitamin C and E or placebo. No protective effect on lung function or airway inflammation was observed following supplementation. BW and BAL-DHA were enhanced after O3, with further increases following supplementation. In conclusion, oxidative air pollutants induce airway inflammation, as well as a broad spectrum of antioxidant adaptations, which could ultimately limit the airway inflammatory responses. Oral vitamin supplementation was shown to augment RTLF-vitamin C concentrations, but it did not provide protection from the ozone-induced airway responses following a single insult of ozone. The finding of high concentrations of DHA in the RTLF could indicate that DHA represents an important transport form of vitamin C onto the surface of the lung.