Exhaled Breath Condensate Collection Research Articles

Decomposition and solubility of H2O2: implications in exhaled breath condensate

Hydrogen peroxide (H2O2) is one of the metabolic end products present in exhaled breath. High levels of H2O2 found in breath condensate are an indicator of airway inflammation and could be used for monitoring the condition of patients with chronic obstructive pulmonary disease. However, sampling conditions such as breath temperature, condensing temperature, flow rate and collection time can affect the intrinsic properties of H2O2—its solubility, volatility, and decomposition rate. Sudden decreases to H2O2 concentration may be due to the sampling conditions instead of the patient's health status. The decomposition rate and Henry's law constant for saturated H2O2 vapor (RH > 95%) within 22–42 °C, which correlates to room temperature and range of human breath temperatures, are needed for better understanding and standardization of breath collection. In this study, we determined the effects of initial H2O2 concentration, temperature, and sampling time on the decomposition rate by comparing electrochemical measurements of H2O2 in simulated breath samples. The experimental results showed the decomposition rate of H2O2 increased as the breath temperature and sampling time increased and the solubility of H2O2 increased with increasing flow rate and condensing temperature during sampling. Prediction models for H2O2 sensing in exhaled breath sample were developed that could be used in the standardization of exhaled breath condensate collection. These experimental findings need to be further verified with human/animal breath samples.

Asymmetric Dimethylarginine

Asymmetric Dimethylarginine

Asymmetric Dimethylarginine in Exhaled Breath Condensate and Serum of Children With Asthma

Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor and uncoupler of nitric oxide synthase. By promoting the formation of peroxynitrite, ADMA is believed to contribute to several aspects of asthma pathogenesis (ie, airway inflammation, oxidative stress, bronchial hyperresponsiveness, and collagen deposition). The aim of the present study was to compare this mediator in healthy children and children with asthma using the completely noninvasive exhaled breath condensate (EBC) technique. We recruited 77 children with asthma (5-16 years of age) and 65 healthy children (5-15 years of age) who underwent EBC collection and spirometry. Serum ADMA levels and fractional exhaled nitric oxide levels were measured on the same day in a subgroup of children with asthma. EBC was collected using the Turbo-Deccs (Medivac). ADMA levels were measured using the ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) technique. ADMA could be detected in the EBC of 71 subjects with asthma and 64 healthy subjects. ADMA levels in the EBC of children with asthma were significantly higher than in the healthy control subjects (median, 0.12 [interquartile range, 0.05-0.3] vs 0.07 [0.05-0.12]; P = .017), whereas no difference emerged between the children with asthma who were or were not receiving inhaled steroid treatment. No correlation was found between serum and EBC ADMA levels (P > .5). We measured ADMA in EBC by UPLC-MS/MS, a reference analytical technique. Higher ADMA levels were found in children with asthma, supporting a role for this mediator in asthma pathogenesis. This oxidative stress-related mediator also seems to be scarcely affected by steroid therapy. We speculate that ADMA might be a target for new therapeutic strategies designed to control oxidative stress in asthma.

Concentration of exhaled breath condensate biomarkers after fractionated collection based on exhaled CO2 signal

A standard procedure for exhaled breath condensate (EBC) collection is still lacking. The aim of this study was to compare the concentration of several biomarkers in whole (W-EBC) and fractionated EBC (A-EBC), the latter collected starting from CO2 ≥ 50% increase during exhalation. Forty-five healthy non-smokers or asymptomatic light smokers were enrolled. Total protein concentrations in W-EBC and A-EBC were overlapping (median: 0.7 mg l−1 in both cases), whereas mitochondrial DNA was higher in A-EBC (0.021 versus 0.011 ng ml−1), indicating a concentration rather than a dilution of lining fluid droplets in the last portion of exhaled air. H2O2 (0.13 versus 0.08 µM), 8-isoprostane (4.9 versus 4.4 pg ml−1), malondialdehyde (MDA) (4.2 versus 3.2 nM) and 4-hydroxy-2-nonhenal (HNE) (0.78 versus 0.66 nM) were all higher in W-EBC, suggesting a contribution from the upper airways to oxidative stress biomarkers in apparently healthy subjects. NH4+ was also higher in W-EBC (median: 590 versus 370 µM), with an estimated increase over alveolar and bronchial air by a factor 1.5. pH was marginally, but significantly higher in W-EBC (8.05 versus 8.01). In conclusion, the fractionation of exhaled air may be promising in clinical and occupational medicine.

Hydrogen Peroxide Detection in Wet Air with a Prussian Blue Based Solid Salt Bridged Three Electrode System

We report on a novel electroanalytical system for hydrogen peroxide (H2O2) detection in humidity or droplets of aerosol, formed by air bubbling through a washing chamber; the resulting flow mimics the exhaled human breath. The system is based on a planar three-electrode structure (with a Prussian Blue based H2O2 transducer modified working electrode) bridged by a solid salt-saturated filament material (filter paper, cotton textile). Respective to the hydrogen peroxide content in the washing valve, the response of the aerosol-sensing system is linear in the concentration range of 0.1-10 μM, which overlaps the generally accepted H2O2 content in exhaled breath condensate (EBC), with the sensitivity of 8 A M(-1) cm(-2). The response to the upper limit of the calibration range is stable for more than 50 injection cycles recorded within 3 days. Both the stability and the suitable calibration range allow one to consider the reported aerosol-sensing system as a prototype for a simple (avoiding intermediate EBC collection) noninvasive diagnostic tool for pulmonary patients.

The application of exhaled breath analysis in racing Thoroughbreds and the influence of high intensity exercise and ambient temperature on the concentration of carbon monoxide and pH in exhaled breath

Analyses of exhaled breath (EB) and exhaled breath condensate (EBC) are non-invasive modalities for assessing the lower airways but these methods have not been applied to Thoroughbred racehorses in training. The aims of this study were to determine whether EB and EBC could be obtained from Thoroughbred racehorses in the field and to investigate the effects of exercise per se and during different ambient temperatures and humidity on exhaled concentrations of nitric oxide (eNO), carbon monoxide (eCO) and EBC pH. EB and EBC samples were obtained from 28 Thoroughbred racehorses pre- and post-exercise during warm (n=23) and/or cold (n=19) ambient temperatures.eNO was detected in 19/84 EB samples. eCO was measured in 39/42 EB samples pre-exercise (median 1.3ppm) and concentrations decreased significantly post-exercise (median 0.8ppm, P<0.005) and were associated with ambient temperature. EBC pH was 4.51±0.23 pre-exercise and increased significantly post-exercise (4.79±0.59, P=0.003). The study documented the collection of EB and EBC from Thoroughbred racehorses in a field setting. Alterations in concentrations of volatile gases and EBC pH occurred in response to exercise, and were likely to have been influenced by environmental factors.

Is Health-Related Quality of Life Associated with Upper and Lower Airway Inflammation in Asthmatics?

Background. Allergic diseases impair health-related quality of life (HR-QoL). However, the relationship between airway inflammation and HR-QoL in patients with asthma and rhinitis has not been fully investigated. We explored whether the inflammation of upper and lower airways is associated with HR-QoL. Methods. Twenty-two mild allergic asthmatics with concomitant rhinitis (10 males, 38 ± 17 years) were recruited. The Rhinasthma was used to identify HR-QoL, and the Asthma Control Test (ACT) was used to assess asthma control. Subjects underwent lung function and exhaled nitric oxide (eNO) test, collection of exhaled breath condensate (EBC), and nasal wash. Results. The Rhinasthma Global Summary score (GS) was 25 ± 11. No relationships were found between GS and markers of nasal allergic inflammation (% eosinophils: r = 0.34, P = 0.24; ECP: r = 0.06, P = 0.87) or bronchial inflammation (pH of the EBC: r = 0.12, P = 0.44; bronchial NO: r = 0.27, P = 0.22; alveolar NO: r = 0.38, P = 0.10). The mean ACT score was 18. When subjects were divided into controlled (ACT ≥ 20) and uncontrolled (ACT < 20), the alveolar NO significantly correlated with GS in uncontrolled asthmatics (r = 0.60, P = 0.04). Conclusions. Upper and lower airways inflammation appears unrelated to HR-QoL associated with respiratory symptoms. These preliminary findings suggest that, in uncontrolled asthma, peripheral airway inflammation could be responsible for impaired HR-QoL.

Characterization of a portable method for the collection of exhaled breath condensate and subsequent analysis of metal content

Using exhaled breath condensate (EBC) as a biological media for analysis of biomarkers of exposure may facilitate the understanding of inhalation exposures. In this study, we present method validation for the collection of EBC and analysis of metals in EBC. The collection method was designed for use in a small scale longitudinal study with the goal of improving reproducibility while maintaining economic feasibility. We incorporated the use of an Rtube with additional components as an assembly, and trained subjects to breathe into the apparatus. EBC was collected from 8 healthy adult subjects with no known elevated exposures to Mn, Cr, Ni, and Cd repeatedly (10 times) within 7 days and analyzed for these metals via ICP-MS. Method detection limits were obtained by mimicking the process of EBC collection with ultrapure water, and resulted in 46-62% of samples falling in a range less than the method detection limit. EBC metal concentrations were found to be statistically significantly associated (p < 0.05) with room temperature and relative humidity during collection, as well as with the gender of the subject. The geometric mean EBC metal concentrations in our unexposed subjects were 0.57 μg Mn per L, 0.25 μg Cr per L, 0.87 μg Ni per L, and 0.14 μg Cd per L. The overall standard deviation was greater than the mean estimate, and the major source in EBC metals concentrations was due to fluctuations in subjects' measurements over time rather than to the differences between separate subjects. These results suggest that measurement and control of EBC collection and analytical parameters are critical to the interpretation of EBC metals measurements. In particular, rigorous estimation of method detection limits of metals in EBC provides a more thorough evaluation of accuracy.

Vest Chest Physiotherapy Airway Clearance is Associated with Nitric Oxide Metabolism.

Background. Vest chest physiotherapy (VCPT) enhances airway clearance in cystic fibrosis (CF) by an unknown mechanism. Because cilia are sensitive to nitric oxide (NO), we hypothesized that VCPT enhances clearance by changing NO metabolism. Methods. Both normal subjects and stable CF subjects had pre- and post-VCPT airway clearance assessed using nasal saccharin transit time (NSTT) followed by a collection of exhaled breath condensate (EBC) analyzed for NO metabolites (NOx). Results. VCPT shorted NSTT by 35% in normal and stable CF subjects with no difference observed between the groups. EBC NOx concentrations decreased 68% in control subjects after VCPT (before = 115 ± 32 μM versus after = 37 ± 17 μM; P < 0.002). CF subjects had a trend toward lower EBC NOx. Conclusion. We found an association between VCPT-stimulated clearance and exhaled NOx levels in human subjects. We speculate that VCPT stimulates clearance via increased NO metabolism.

Exhaled breath condensate: a comprehensive update

Since the late 1990s, a surge in interest in the analysis of exhaled breath condensate (EBC) resulted in the American Thoracic Society and European Respiratory Society (ATS/ERS) organising a Task Force in 2001 to develop guidelines on EBC collection and measurement of biomarkers. This Task Force published their guidelines in 2005 based on literature and expert opinions at that time, and multiple shortcomings and knowledge deficits were also identified. The clinical application of EBC collection and its biomarkers are currently still limited by several of these knowledge gaps, hence further guidelines for standardisation are required to ensure external validity. Using related articles produced since the publication of the ATS/ERS Task Force report, this paper attempts to provide a comprehensive update to the original guideline and review the methodological shortcomings identified. This review can hopefully serve as a yardstick for future studies involving this emerging clinical tool.

Exhaled Breath Condensate Detects Baseline Reductions in Chloride and Increases in Response to Albuterol in Cystic Fibrosis Patients

Impaired ion regulation and dehydration is the primary pathophysiology in cystic fibrosis (CF) lung disease. A potential application of exhaled breath condensate (EBC) collection is to assess airway surface liquid ionic composition at baseline and in response to pharmacological therapy in CF. Our aims were to determine if EBC could detect differences in ion regulation between CF and healthy and measure the effect of the albuterol on EBC ions in these populations. Baseline EBC Cl−, DLCO and SpO2 were lower in CF (n = 16) compared to healthy participants (n = 16). EBC Cl− increased in CF subjects, while there was no change in DLCO or membrane conductance, but a decrease in pulmonary-capillary blood volume in both groups following albuterol. This resulted in an improvement in diffusion at the alveolar-capillary unit, and removal of the baseline difference in SpO2 by 90-minutes in CF subjects. These results demonstrate that EBC detects differences in ion regulation between healthy and CF individuals, and that albuterol mediates increases in Cl− in CF, suggesting that the benefits of albuterol extend beyond simple bronchodilation.

Differences in IL-8 in serum and exhaled breath condensate from patients with exacerbated COPD or asthma attacks.

The collection of exhaled breath condensate (EBC) is a noninvasive method that can be used to monitor the inflammatory status of patients with chronic airway diseases. We aimed to study differences in cytokine expression between patients with exacerbations of chronic obstructive pulmonary disease (COPD) and patients with asthma attacks. Using a custom-made device and methods based on American Thoracic Society (ATS)/European Respiratory Society (ERS) recommendations, EBC samples were collected from nine COPD patients, 12 asthma patients and 10 healthy individuals. Cytokine concentrations in serum and EBC were measured via commercial ELISA kits. Of four cytokines measured in EBC [interleukin-8 (IL-8), IL-17, IL-4 and tumor necrosis factor-α (TNF-α)], only IL-8 was significantly higher in COPD than in asthma patients (5.27 ± 0.18 vs. 4.36 ± 0.34 pg/mL, p = 0.001). Moreover, COPD patients had higher serum IL-8 than asthma patients (10.57 ± 0.55 vs. 5.15 ± 0.24 pg/mL, p < 0.001). No significant correlation between serum and EBC cytokine concentrations was observed in each subgroup of patients. Compared with patients with asthma attacks, patients with exacerbated COPD had increased IL-8 expression in both serum and EBC. These results suggest that IL-8 may be more important in airway and systemic inflammation in COPD exacerbations than in asthma attacks.

Effect of Nebulized Beclomethasone on Airway Inflammation and Clinical Status of Children with Allergic Asthma and Rhinitis: A Randomized, Double-Blind, Placebo-Controlled Study

We aimed to evaluate the therapeutic effect of nebulized beclomethasone dipropionate (nBDP) on both allergic asthma and rhinitis. In a randomized, double-blind, placebo-controlled study, 40 children (mean age 10.7 ± 2.1 years) with allergic asthma and rhinitis received either nBDP (daily dose of 800 µg, administered twice daily) or placebo for 4 weeks (with a face mask), after a 2-week run-in period of clinical assessment. Nasal and oral fractional exhaled nitric oxide (FeNO) measurements together with pulmonary function tests, nasal and oral exhaled breath condensate (EBC) collection for pH and interleukin-5 (IL-5) measurements as well as nasal and bronchial symptom scores were obtained at baseline and after 4-week treatment. A significant improvement in oral FeNO, oral and nasal EBC IL-5 and nasal EBC pH was observed in the nBDP group when comparing the values with baseline, together with an improvement in symptom score of the visual analogue scale, nasal obstruction, sneezing, rhinorrhea, breathing difficulty, cough, wheezing and sleep disturbance (nBDP end treatment vs. baseline, Wilcoxon signed-rank test). nBDP was more effective than placebo (ANCOVA test) in improving [difference Δ = response after treatment at the last visit (active or placebo) – value at baseline] nasal pH, oral IL-5, oral FeNO, forced expiratory volume in 1 s, forced expiratory volume in 1 s/forced vital capacity, peek expiratory flow, visual analogue scale, breathing difficulty, cough, wheezing and sleep disturbance scores. No differences were observed between the nBDP and the placebo group for symptom score of rhinitis. nBDP is a useful treatment for airway inflammation and clinical status in children with concomitant allergic asthma and rhinitis.

Secondhand smoke exposure induces acutely airway acidification and oxidative stress

Previous studies have shown that secondhand smoke induces lung function impairment and increases proinflammatory cytokines. The aim of the present study was to evaluate the acute effects of secondhand smoke on airway acidification and airway oxidative stress in never-smokers. In a randomized controlled cross-over trial, 18 young healthy never-smokers were assessed at baseline and 0, 30, 60, 120, 180 and 240min after one-hour secondhand smoke exposure at bar/restaurant levels. Exhaled NO and CO measurements, exhaled breath condensate collection (for pH, H(2)O(2) and NO(2)(-)/NO(3)(-) measurements) and spirometry were performed at all time-points. Secondhand smoke exposure induced increases in serum cotinine and exhaled CO that persisted until 240min. Exhaled breath condensate pH decreased immediately after exposure (p<0.001) and returned to baseline by 180min, whereas H(2)O(2) increased at 120min and remained increased at 240min (p=0.001). No changes in exhaled NO and NO(2)/NO(3) were observed, while decreases in FEV(1) (p<0.001) and FEV(1)/FVC (p<0.001) were observed after exposure and returned to baseline by 180min. A 1-h exposure to secondhand smoke induced airway acidification and increased airway oxidative stress, accompanied by significant impairment of lung function. Despite the reversal in EBC pH and lung function, airway oxidative stress remained increased 4h after the exposure. Clinical trial registration number (EudraCT): 2009-013545-28.

Asthma severity in childhood and metabolomic profiling of breath condensate

Asthma is a heterogeneous disease and its different phenotypes need to be better characterized from a biochemical-inflammatory standpoint. This study aimed to apply the metabolomic approach to exhaled breath condensate (breathomics) to discriminate different asthma phenotypes, with a particular focus on severe asthma in children. In this cross-sectional study, we recruited 42 asthmatic children (age, 8-17years): 31 with nonsevere asthma (treated with inhaled steroids or not) and 11 with severe asthma. Fifteen healthy children were enrolled as controls. Children performed exhaled nitric oxide measurement, spirometry, exhaled breath condensate (EBC) collection. Condensate samples were analyzed using a metabolomic approach based on mass spectrometry. A robust Bidirectional-Orthogonal Projections to Latent Structures-Discriminant Analysis (O2PLS-DA) model was found for discriminating both between severe asthma cases and healthy controls (R(2) =0.93; Q(2) =0.75) and between severe asthma and nonsevere asthma (R(2) =0.84; Q(2) =0.47). The metabolomic data analysis leads to a robust model also when the 3 groups of children were considered altogether (K=0.80), indicating that each group is characterized by a specific metabolomic profile. Compounds related to retinoic acid, adenosine and vitamin D (Human Metabolome Database) were relevant for the discrimination between groups. The metabolomic profiling of EBC could clearly distinguish different biochemical-metabolic profiles in asthmatic children and enabled the severe asthma phenotype to be fully discriminated. The breathomics approach may therefore be suitable for discriminating between different asthma metabolic phenotypes.

Exhaled breath condensate nitric oxide end products and pH in controlled asthma

ObjectivesAsthma imposes a growing burden on the society in terms of morbidity, quality of life, and healthcare costs. It has the highest morbidity amongst inflammatory lung diseases and its prevalence continues to increase over the world. Inquiry into recent day or nighttime symptoms alone underestimates the burden of asthma and may lead to inadequate treatment of asthma. The aim of the present work is to evaluate the role of nitric oxide (NO) and hydrogen ion concentration (pH) levels in exhaled breath condensate (EBC) in cases of controlled bronchial asthma. Patients and methodsThe present study was conducted on 49 controlled asthmatic patients and 12 control subjects. All patients were subjected to thorough history taking, complete clinical examination and plain postero-anterior chest X-ray. All asthmatics and control subjects were subjected to routine laboratory investigations, spirometric study, EBC collection, processing and analysis for its content of both nitric oxide end products: nitrite and nitrate (NOx) and pH. ResultsAll asthmatics represented Group IT which was further divided into Group Ia: 34 patients on regular inhaled corticosteroid (ICS) therapy and Group Ib: 15 patients on no regular therapy. The control subjects represented Group II. The forced vital capacity (FVC), forced expiratory volume in the first second (FEV1), FEV1/FVC, peak expiratory flow rate (PEFR) and forced expiratory flow during the middle portion of a forced expiration (FEF 25–75%) were significantly lower in asthmatics than control subjects. The EBC-NOx mean±SD in μmol/L in Group IT (5.99±1.63), in Group Ia (5.27±1.26) and in Group Ib (7.63±1.15) were significantly higher than in Group II (3.66±0.67) with, respectively (p=0.000), (p=0.000) and (p=0.000); and was significantly higher in Group Ib than Group Ia (p=0.000). The EBC-pH mean±SD in Group IT (7.32±0.27), in Group Ia (7.35±0.25) and in Group Ib (7.27±0.3) were significantly lower than in Group II (7.82±0.09) with, respectively (p=0.000), (p=0.000) and (p=0.000); with no significant difference between Group Ia and Group Ib. The EBC-NOx was significantly directly correlated to eosinophils count (p=0.017) and neutrophils count (p=0.002); and inversely correlated to FEV1 (p=0.016), FEV1/FVC (p=0.001), PEFR (p=0.030) and EBC-pH (p=0.003). The EBC-pH was significantly inversely correlated to eosinophils count (p=0.017) and neutrophils count (p=0.036); and directly correlated to FVC (p=0.004), FEV1 (p=0.004) and PEFR (p=0.000). ConclusionEBC-NOx is significantly higher and EBC-pH is significantly lower in asthmatic patients than in control subjects. Asthmatics receiving ICS have a lower EBC-NOx level than those not. EBC-NOx and EBC-pH were significantly correlated and both of them showed significant correlations with spirometric parameters of airway obstruction.

Molecular and microscopic analysis of bacteria and viruses in exhaled breath collected using a simple impaction and condensing method.

Exhaled breath condensate (EBC) is increasingly being used as a non-invasive method for disease diagnosis and environmental exposure assessment. By using hydrophobic surface, ice, and droplet scavenging, a simple impaction and condensing based collection method is reported here. Human subjects were recruited to exhale toward the device for 1, 2, 3, and 4 min. The exhaled breath quickly formed into tiny droplets on the hydrophobic surface, which were subsequently scavenged into a 10 µL rolling deionized water droplet. The collected EBC was further analyzed using culturing, DNA stain, Scanning Electron Microscope (SEM), polymerase chain reaction (PCR) and colorimetry (VITEK 2) for bacteria and viruses.Experimental data revealed that bacteria and viruses in EBC can be rapidly collected using the method developed here, with an observed efficiency of 100 µL EBC within 1 min. Culturing, DNA stain, SEM, and qPCR methods all detected high bacterial concentrations up to 7000 CFU/m3 in exhaled breath, including both viable and dead cells of various types. Sphingomonas paucimobilis and Kocuria variants were found dominant in EBC samples using VITEK 2 system. SEM images revealed that most bacteria in exhaled breath are detected in the size range of 0.5–1.0 µm, which is able to enable them to remain airborne for a longer time, thus presenting a risk for airborne transmission of potential diseases. Using qPCR, influenza A H3N2 viruses were also detected in one EBC sample. Different from other devices restricted solely to condensation, the developed method can be easily achieved both by impaction and condensation in a laboratory and could impact current practice of EBC collection. Nonetheless, the reported work is a proof-of-concept demonstration, and its performance in non-invasive disease diagnosis such as bacterimia and virus infections needs to be further validated including effects of its influencing matrix.

The effect of temperature on exhaled breath condensate collection

Exhaled breath condensate (EBC) collection is an innovative method of non-invasively sampling the lung, and can detect a variety of volatile and non-volatile biomarkers, but the disadvantage is the small volume of sample collected. It was hypothesized that a collection system at a lower temperature would increase the volume collected, but may alter the relative concentration of the biomarkers of interest. EBC was collected in a cross-over study using a custom-made collection system, cooled using either wet (4 °C) or dry ice (−20 °C) in randomized order in normal non-smoking volunteers. The volume of the EBC collected per unit time was determined as were conductivity, the concentrations and total amount of protein, hydrogen peroxide, and nitrite/nitrate concentrations. Dry ice was associated with a 79% greater volume of EBC than the wet ice (1387 ± 612 µL; 773 ± 448 µL respectively, p < 0.0001). Conductivity was influenced by the temperature of collection (18.78 ± 6.71 µS cm−1 for wet ice and 15.32 ± 6.28 µS cm−1 for dry ice, p = 0.02) as was hydrogen peroxide (1.34 ± 0.88 µg mL−1 for wet ice and 0.68 ± 0.32 µg mL−1 for dry ice, p = 0.009) while the concentrations and total values for protein and nitrate/nitrite were not significantly different (p > 0.05). This pilot study suggests that lower collection temperatures facilitate the collection of a larger sample volume. This larger volume is not simply more dilute, with increased water content, nor is there a simple correction factor that can be applied to the EBC biomarkers to correct for the different methods.

Non-invasive collection of exhaled breath condensate in rats: Evaluation of pH, H2O2 and NOx in lipopolysaccharide-induced acute lung injury

The analysis of exhaled breath condensate (EBC) offers the potential for identifying lung disease markers in humans and animals, but methodological issues and standardised procedures need to be addressed before the technique can be considered for use in applications to help understand the role of environmental pollution in respiratory diseases. The purpose of this study was to develop and implement a new device using a glass-chamber for collecting EBC non-invasively from rats in order to analyse EBC markers in lipopolysaccharide (LPS)-induced acute lung injury. Eighty-four adult rats were used in five different series of experiments to determine the source of EBC formation, intra-day and inter-day variability, and the influence of environmental parameters on EBC markers. The hypothesis that inflammation induces an oxidative stress was assessed by measuring pH, nitrogen oxides (NOx) and hydrogen peroxide (H(2)O(2)) in EBC. The results confirmed that EBC fluid was generated at the level of the respiratory tract. The repeatability studies of disease markers indicated higher concentrations of NOx and H(2)O(2) at midday compared to the morning, but there were no significant difference between measurements on consecutive days. EBC volume was influenced by both ambient temperature and humidity. Moreover, 3h after LPS challenge, significantly increased concentrations of both NOx and H(2)O(2) were observed in EBC of the LPS group compared with controls (P=0.005 and P=0.027, respectively). These results suggested that EBC collection may be a valuable tool to monitor the presence of markers, such as NOx and H(2)O(2), in an animal model of LPS-induced acute lung injury.

Assessment of airway inflammation in chronic obstructive pulmonary disease: Biomarkers in exhaled breath condensate

Airway inflammation plays a central role in the pathophysiology of chronic obstructive pulmonary disease. Exposure to cigarette smoke induces the recruitment of inflammatory cells in the airways, which in turn produces various cytokines, chemokines, proteases and pro-inflammatory mediators leading ultimately to increased oxidative stress, a protease/anti-protease imbalance and progressive lung tissue injury. Biomarkers may be useful in monitoring airway inflammation and oxidative stress, defining different phenotypes of the disease and evaluating the response of therapies. Exhaled breath condensate collection is a simple and completely non-invasive method of sampling the lower respiratory tract in humans. Exhaled breath condensate may be a rich source of pulmonary biomarkers including hydrogen peroxide, cytokines, metabolites of the arachidonic acid, nitric oxides and the pH. However, the concentration of these biomarkers is often very low, which may cause several problems in their detection. The clinical applicability of exhaled breath condensate biomarkers cannot be assessed until methods of sample collection and analysis have been standardized.