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The Effects of Volatile Salivary Acids and Bases on Exhaled Breath Condensate pH

Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California; and Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin

Correspondence and requests for reprints should be addressed to Richard M. Effros, M.D., LABiomed–Harbor–UCLA Medical Center, 1124 West Carson Street, RB2, Torrance, CA 90502-2064. E-mail: reffros{at}labiomed.org

	ABSTRACT

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Rationale: Recent studies have reported acidification of exhaled breath condensate (EBC) in inflammatory lung diseases. This phenomenon, designated "acidopnea," has been attributed to airway inflammation.

Objectives: To determine whether salivary acids and bases can influence EBC pH in chronic obstructive pulmonary disease (COPD).

Methods: Measurements were made of pH, electrolytes, and volatile bases and acids in saliva and EBC equilibrated with air in 10 healthy subjects and 10 patients.

Results: The average EBC pH in COPD was reduced (normal, 7.24 ± 0.24 SEM; range, 6.11–8.34; COPD, 6.67 ± 0.18; range, 5.74–7.64; p = 0.079). EBCs were well buffered by NH₄⁺/NH₃ and CO₂/HCO₃^– in all but four patients, who had NH₄⁺ concentrations under 60 µmol/L, and acetate concentrations that approached or exceeded those of NH₄⁺. Saliva contained high concentrations of acetate ( 6,000 µmol/L) and NH₄⁺ ( 12,000 µmol/L). EBC acetate increased and EBC NH₄⁺ decreased when salivary pH was low, consistent with a salivary source for these volatile constituents. Nonvolatile acids did not play a significant role in determining pH of condensates because of extreme dilution of respiratory droplets by water vapor ( 1:12,000). Transfer of both acetic acid and NH₃ from the saliva to the EBC was in the gas phase rather than droplets.

Conclusions: EBC acidification in COPD can be affected by the balance of volatile salivary acids and bases, suggesting that EBC pH may not be a reliable marker of airway acidification. Salivary acidification may play an important role in acidopnea.

Key Words: acetate • ammonium • bicarbonate • buffer • exhaled breath condensate

In 2000, Hunt and colleagues reported that exhaled breath condensates (EBCs) are acidified during asthmatic exacerbations (1). They referred to this phenomenon as "acidopnea" and suggested that it reflected excess acid produced in the airways by inflammation. These observations were confirmed in studies of chronic obstructive pulmonary disease (COPD) and other inflammatory lung disorders (2–10). Subsequently, Hunt and colleagues found that EBC NH₄⁺ concentrations were reduced in many patients with asthma (11). They postulated that production of NH₃ in the airways was reduced because of impaired glutaminase activity, and suggested that reductions in airway NH₃ production reduced local buffering, thereby promoting airway acidification. They also argued that concentrations of NH₄⁺ in EBCs did not influence the pH of the EBC samples, which represented an accurate marker of airway acidification (12, 13).

This study has analyzed the acid and base concentrations in saliva and EBC from 10 healthy subjects and 10 patients with COPD to determine whether EBC pH is influenced by volatile or nonvolatile constituents in the saliva.

	METHODS

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Eleven healthy subjects and 10 subjects with COPD were initially selected but condensate from one healthy subject was deleted because high amylase concentrations indicated salivary contamination (Table 1). Spirometry was performed in all subjects (Sensormedics, Yorba Linda, CA). The patients had COPD (FEV₁ < 75% predicted, FEV₁/FVC < 70%) (14) and an average smoking history of 59 ± 43 (SD) pack-yr. None smoked within an hour of the study. All were taking prescribed maintenance bronchodilators, which were not used during the hour before collections.

All samples were stored at –80°C. Before analysis, all samples were allowed to thaw in room air for about 30 min. To minimize potential losses of volatile acids and bases, no attempt was made to remove ambient CO₂ from the EBC or saliva with inert gas (see the online supplement). The buffering capacity of the condensates was determined by sequentially measuring the pH after adding 5-µl aliquots of 0.4 mmol/L NaOH and then 0.4 mmol/L HCl to 0.5 ml samples at room temperature:

(1)

where molality designates the change in molality of NaOH or HCl needed to alter condensate pH one unit in the range between 5 and 7.

From 1 to 8 ml of the EBC samples were lyophilized (freeze-dried) at –55°C to dryness to remove volatile solutes such as NH₃, CO₂, and acetic acid. These samples were reconstituted in 2.1 ml of deionized water and measurements were repeated of conductivity and pH. Corrections were made for the differences in volumes used in lyophilization. EBC pH, conductivity, and amylase and ionic concentrations (by ion chromatography) were determined before and/or after they were lyophilized, as described in the online supplement.

Dilution (D) of respiratory droplets (epithelial lining fluid) by water was then calculated from conductivity measurements:

(2)

The asterisk indicates lyophilized samples (15, 16).

The coefficient of variance of repeated (5 to 10) measurements of electrolyte concentrations in the same sample at 5 µmol/L was less than 2%. The lower limits of detection were 0.5 µmol/L for each of the ions, 2.5 µmol/L NaCl for conductivity, and 0.2 mU/ml for amylase.

Statistical analyses were conducted with SigmaStat version 2 software (Jandel, San Rafael, CA). Statistical differences between mean values of normal and COPD parameters were compared by unpaired t test. Linear regression and correlation coefficients were used to compare EBC parameters. Electrolyte concentrations were compared by ranked analysis of variance repeated measurements and a Student-Newman-Keuls test of differences in mean values.

These studies were approved by the institutional human research review committees, and consent was obtained from each subject before each study. More details on the methods used in this study are available on the online supplement.

	RESULTS

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pH, Buffer, NH₄⁺, and Volatile Acids in Unlyophilized Condensates (Equilibrated in Room Air)
EBC pH averaged 7.24 ± 0.24 SEM in normal subjects, and 6.67 ± 0.18 in patients who had COPD (p = 0.079; Figure 1). Differences in EBC NH₄⁺ were less obvious, but it will be noted that all four of the EBC samples with NH₄⁺ concentrations that were below 60 µmol/L were collected from patients with COPD.

View larger version (14K): [in this window] [in a new window]   Figure 1. (A) Mean exhaled breath condensate (EBC) pH in chronic obstructive pulmonary disease (COPD) was less than that of healthy subjects (p = 0.075). (B) Differences in mean EBC NH4+ concentrations were less obvious, but NH4+ concentrations below 60 µmol/L were only observed in four patients with COPD. Rectangles indicate mean and SEM.

View larger version (23K): [in this window] [in a new window]   Figure 2. Significant buffering was seen in some EBC samples from both healthy subjects (A) and patients with COPD (B). Buffering was maximal at about 6.3 (indicated by horizontal line), which is the pKa of CO2/HCO3– at room temperature. Note the resemblance of these titration curves to those of solutions of NH4HCO3 (C). No evidence was found for buffers with other pKa values.

View larger version (15K): [in this window] [in a new window]   Figure 3. The buffer capacity of the EBC solutions between pH 5 and 7 was well correlated with the NH4+ concentrations. (The buffer capacity was calculated with Equation 1). This reflects the fact that the concentration of NH4+ in the solution effectively determines the concentration of ambient CO2 that is trapped in the condensate. mM designates mmol/L.

View larger version (25K): [in this window] [in a new window]   Figure 4. (A) Volatile ion concentrations in the nonlyophylized samples. Average values of NH4+ exceeded those of acetate. Average values of acetate were higher in the COPD samples than the normal samples, but significance was at p = 0.07. (B) Salivary concentrations of both NH4+ and acetate were more than 50 times greater than those in the EBC. This suggests that the oral cavity is the source for much of these constituents in the EBC.

View larger version (8K): [in this window] [in a new window]   Figure 5. Acetate concentrations were usually well below those of NH4+. However, the ratio of acetate to NH4+ approached or exceeded 1.0 in four patients with COPD.

A correlation was found between salivary pH and the concentration of NH₄⁺ in the condensate (r² = 0.383, p = 0.004; Figure 6A), suggesting that salivary acidification reduces the release of NH₃ from the mouth and the amount of NH₄⁺ recovered in the EBC. In contrast, EBC concentrations of acetate were increased when saliva was acidic (r² = 0.509, p < 0.001; Figure 6B).

View larger version (17K): [in this window] [in a new window]   Figure 6. (A) Acidification of the saliva increased acetate concentrations in the EBC. (B) Acidification of the saliva reduced NH4+ concentrations in the EBC.

View larger version (29K): [in this window] [in a new window]   Figure 7. (A) EBC concentrations of NH4+ and nonvolatile cations and anions. NH4+ concentrations are shown as prior to lyophilization (PRE) and after lyophilization (POST). Most of the NH4+ was removed by this procedure. All other measurements were made in the lyophylized EBC. Average values of NH4+ in the nonlyophilized samples exceeded those of any other ion by a factor of more than 20. Ion concentrations did not differ between normal and COPD samples. For the entire population, the relative molar magnitudes of the nonvolatile ionic concentrations were Na+ = Cl– = Ca2+ > K+ > lactate > SO42– > NO3– > NO2– = NH4+ > phosphate > Mg2+ (p < 0.05). (B) Total cation concentrations exceeded those of the anions and the total ionic concentration estimated from conductivity (Conduc; p < 0.05). The total concentration of nonvolatile cations (Na+, K+, Ca2+, and Mg2+) slightly exceeded total measured by conductivity and the total anion concentrations.

View larger version (17K): [in this window] [in a new window]   Figure 8. Lyophilization caused similar reductions in EBC NH4+ and conductivity, suggesting that NH4+ was the principal cation lost during lyophilization.

View larger version (17K): [in this window] [in a new window]   Figure 9. Although the values of conductivity and total cations were variable in the lyophilized samples, they were well correlated, suggesting that either could be used to calculate the dilution of respiratory droplets by water vapor.

No differences were detected between the normal and COPD EBC concentrations of individual ions or measures of total ions, cations, or anions that might influence the pH of the respiratory fluid (Figure 7). When compared with comparable plasma ratios, EBC concentrations of K⁺, Ca²⁺, lactate, and SO₄^2– were disproportionately high relative to EBC Na⁺ (Table 3), but no differences were found between normal and COPD samples.

View larger version (25K): [in this window] [in a new window]   Figure 10. Effect of ambient CO2 on the pH of solutions of NH4OH. Room air was bubbled at 20 ml/min through 30 ml of solutions of NH4OH in an open 100-ml beaker until pH stabilized. The fall in pH reflects diffusion of room air into the solution with increases in HCO3– to more than 97% of those of NH4+ in the solutions. These curves are consistent with an ambient fractional volume of CO2 (FCO2) of about 0.001. If CO2 could be removed from these solutions by flushing with inert gases, the pH would return to that measured before exposure to air. Total concentration of NH3 + +4 remains relatively unchanged (upper panel).

	DISCUSSION

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These observations concerning EBC pH are consistent with many others, which indicate that pH of the EBC is frequently acidic in patients with COPD and other inflammatory lung diseases (1–11). Although the suggestion of Hunt and coworkers (1) that EBC acidification reflects acidification of the airways seemed plausible, there are a number of problems associated with this hypothesis. It has been shown in this and previous studies that NH₄⁺ is by far the most abundant cationic buffer in the EBC of healthy subjects and in many patients with COPD (see Figure 7 and References 15 and 16). Most of this NH₄⁺ is derived from saliva, which normally contains abundant NH₄⁺; when the mouth is "bypassed" with an endotracheal tube or by tracheostomy, EBC NH₄⁺ concentrations fall by about 80% (12, 15–17). EBC NH₄⁺ can also be reduced by 90% by simply washing the mouth with acidic solutions, which tend to trap NH₃ as NH₄⁺ in the saliva (18). Thus, much of the NH₄⁺ found in the EBC reflects contamination by salivary NH₃, some of which is derived from bacterial degradation of urea.

Although most of the NH₄⁺ in the EBC is generated in saliva, very little of it is delivered to the EBC in salivary droplets. This conclusion is supported by several observations:

1. EBC concentrations of amylase determined with a very sensitive assay were disproportionately lower than those in the saliva.
   
2. Relative concentrations of nonvolatile cations in the EBC differed significantly from those in the saliva. In the EBC, Na⁺ concentrations were greater than those of K⁺, which were similar to those of Ca²⁺. In the saliva, Ca²⁺ concentrations are much lower than those of sodium, whereas sodium concentrations are much lower than those of potassium (Figure 7 and Reference 19).
   
3. Concentrations of NH₄⁺ in the EBC are much greater than those of the total nonvolatile cations (Table 4). In contrast, the concentrations of NH₄⁺ in saliva are lower than those of total nonvolatile cations. Had the NH₄⁺ in the EBC been delivered in salivary droplets, then the total nonvolatile cation concentrations in the EBC would have been greater than the NH₄⁺ concentrations.
   
4. Although NH₃ can be virtually eliminated from EBC samples and artificial samples containing NH₄HCO₃ by overnight lyophilization, it cannot be removed in this fashion from solutions of NH₄Cl. It must therefore be concluded that most of the NH₄⁺ in the EBC arrived as NH₃ gas released from the saliva. Additional evidence that NH₄⁺ in EBC was added in the form of NH₃ gas rather than droplets of saliva containing NH₄⁺ is provided in a previous study (15).
   

(3)

where g indicates the gas phase. The observation that the pH of the EBC ( 7–8) is normally much lower than the dissociation constant of NH₄⁺ (9.3 at room temperature) indicates that acid must have also been added to the EBC during collection. Because the concentration of acid involved would have to be comparable to that of NH₄⁺, it must presumably be delivered to the EBC as a gas, because the concentrations of nonvolatile solutes are much lower (see above). The most likely source for this volatile acid is CO₂, which forms carbonic acid, and which is present at high concentrations in the exhaled air and low concentrations in the environment. As indicated in Figure 10, solutions containing NH₄⁺ and OH^– equilibrate with ambient CO₂, significantly reducing the pH of these solutions. It was impossible to directly measure HCO₃^– in our solutions by ion chromatography because the anionic eluent used for this purpose contains both HCO₃^– and CO₃^2–. However, we were able to titrate the EBC with NaOH and HCl to show that maximal buffering was present at a pH of about 6.3, which equals the pK_a of CO₂/HCO₃^– at room temperature. This observation suggests that NH₃ entering the EBC is neutralized by CO₂:

(4)

At the pH of the EBC samples, concentrations of HCO₃^– approach those of NH₄⁺. This explains why buffering capacities of normal EBCs are closely correlated with the NH₄⁺ concentrations (Figure 3). These observations are consistent with the hypothesis that NH₄⁺/NH₃ and CO₂/HCO₃^– are the principal buffer systems that determine the pH of the EBC in healthy subjects and many patients with COPD.

One problem associated with measurements of EBC pH is uncertainty concerning the PCO₂ of these samples. The pH of normal EBC samples measured immediately after collection tends to be relatively acidic (pH 6.0) (7, 10). This presumably reflects the presence of an end-tidal CO₂ of about 40 mm Hg in the exhaled air, which is equivalent to 1,200 µmol/L in the condensate. Because end-tidal CO₂ can be influenced by changes in arterial PCO₂, ventilation/perfusion ratios, and dead space of the lungs, Hunt and coworkers adopted the procedure of removing CO₂ by flushing the samples with inert gas (argon) (1). In earlier studies, they exposed 1.0 ml to a flow of argon at 350 ml/min (1). This increases the pH of the EBC to approximately 7.65. More recently, they have suggested that 0.2 ml can be used for this purpose, increasing the EBC pH to 7.9 (12). They assumed that virtually all of the CO₂ was removed from the samples when the pH stabilized. However, in the absence of some other acids, removal of all of the CO₂ from the samples should increase the pH of solutions containing NH₄⁺ to about 9.0 when NH₄⁺ concentrations are as high as those reported in normal samples. This is far higher than that observed when the EBC samples are flushed with argon. The effect on EBC pH of purging the samples with argon to remove the effects of CO₂ on the pH would be comparable to reversing the effect of atmospheric CO₂ in Figure 10. Although acetic acid is found in EBC samples collected from some patients with asthma and patients with COPD (see below), we have been unable to detect any other acids other than CO₂ in concentrations comparable to those of NH₄⁺ in healthy subjects. This suggests that flushing the EBC with inert gases for 10 min does not remove all of the CO₂. We have tried both of the procedures recommended to remove CO₂ from defined solutions of NH₄HCO₃ using dry nitrogen as the inert gas, but the EBC pH did not increase above 8 (see the online supplement), indicating that much of the CO₂ remained. Furthermore, there was loss of some water and NH₃ from these solutions. This indicates that purging EBC with inert gases is nonselective and other important volatile constituents of the EBC can be lost. We reasoned that the PCO₂ of the samples could be decreased to ambient levels and loss of other volatile acids and bases minimized by exposing our samples to room air for 30 min before measuring pH. It also would not have been practical to expose samples of saliva to argon without foaming.

Hunt and colleagues (11) also found that EBC NH₄⁺ concentrations are decreased in subjects with asthma. Because most of the NH₄⁺ is derived from the mouth and saliva and salivary NH₄⁺ concentrations are unchanged in COPD (see Figure 4), this suggests that decreases in EBC NH₄⁺ reflect acidification of the saliva, which decreases the concentrations of NH₃ relative to those of NH₄⁺ in the saliva.

The observation that EBC pH can fall to values of 5 and below cannot be attributed to either the CO₂/HCO₃^– (pK_a = 6.3) or NH₄⁺/NH₃ (pK_a = 9.3) buffer systems; nor is it possible to attribute this acidification to any nonvolatile buffers that could be delivered in respiratory or salivary droplets. The extreme dilution of nonvolatile constituents in the EBC by water vapor ( 1:12,000) (15, 18–21) would make the contribution of even strong acids negligible. For example, the dilution of respiratory or gastric droplets containing a 0.1 M solution of HCl would result in a pH of greater than 6.0.

Preliminary evidence has been reported that acetic acid concentrations (pK_a = 4.75) are elevated in the EBC of some patients with asthma (22). We also detected increased acetic acid concentrations in some of our patients with COPD. However like NH₄⁺, acetic acid is present in the saliva in much higher concentrations than those found in the EBC or plasma, and acetic acid represents the most abundant volatile anion in the saliva (23). This suggests that at least some of this acid may be derived from the saliva rather than the lungs. Like salivary NH₃, much of this acetate may reflect oral bacterial metabolism (24). The role of bacterial metabolism on the concentration of acetate in the extracellular fluid is illustrated by the observation that acetate levels in the serum increase from about 70 to about 2,000 µmol/L with ingestion of bran, because of bacterial metabolism in the colon (24). The most persuasive evidence that much of the EBC acetate, like EBC NH₄⁺, represents a salivary contaminant is based on the observation that when the saliva is acid, the concentration of acetate in the EBC increases (Figure 6). In contrast, the concentration of NH₄⁺ decreases when the saliva is acid. This inverse relationship is due to the fact that acidification of the saliva increases the concentration of acetic acid (which is volatile) relative to acetate (which is not volatile). In contrast, acidification reduces the concentrations of NH₃ relative to that of NH₄⁺. We therefore have concluded that the pH of the EBC in these individuals is affected by the relative amounts of acetic acid and ammonia that have been added to the EBC. Although addition of NH₃ to the EBC increases the concentration of HCO₃^– in the EBC, acetic acid reduces the concentration of HCO₃^– in accordance with the reaction:

(5)

If the PCO₂ is kept constant by exposing the EBC to room air, and if losses of NH₃ and acetic acid are minimized, then the pH will vary in accordance with the relative amounts of acetic acid and ammonia that have been absorbed by the EBC. Because most of these constituents are in the ionic form at the pH of the EBC, the ratio of acetate to NH₄⁺ was used to estimate the balance of volatile acids and bases in the EBC (Figure 5).

Definitive evidence that much of the EBC acetate, like EBC NH₄⁺, is derived from saliva rather than the lungs would require collections of EBC from patients before and after intubation. This may not be practical, because high acetate concentrations are observed among patients with lung disease, in whom elective intubation might be unwise. To the extent that extrapulmonary acids and bases influence the EBC pH, acidopnea cannot provide a reliable index of airway acidification or inflammation.

The present study indicates that "acidopnea" may reflect decreases in salivary rather than respiratory pH. Acidification of the saliva could be due to gastroesophageal reflux disease, which is quite common in patients with various forms of obstructive lung disease (25, 26). It would seem prudent to measure salivary pH when measurements are made of EBC pH. Furthermore, it would be of interest to determine whether proton pump inhibitors decrease the incidence of acidopnea. Because acetic acid aerosols can cause both cough and bronchospasm (27, 28), and because acidopnea has been linked to the deleterious effects of air pollution on lung growth in children (29, 30), it can be speculated that saliva may prove to be a source of an agent that is injurious to the lungs.

It would theoretically be advantageous to lyophilize the samples and reconstitute them with deionized water before measuring pH, to minimize the effect of volatile salivary constituents, which can be derived from the mouth. However, the low buffer capacity of the lyophilized samples complicates these measurements. Alternatively, the presence of increased concentrations of specific nonvolatile ions could be used to detect airway acidification. As indicated in Figure 7, no difference was found in the nonvolatile anion concentrations of EBC between our limited COPD and healthy populations. Nevertheless, attention to individual constituents in the EBC may prove more useful than pH, which is a dependent variable that is a function of the concentrations and dissociation constants of multiple acids and bases.

	FOOTNOTES

 
Supported in part by National Institutes of Health grants HL55268, HL074407, and DK25731.

This article has an online supplement, which is accessible from this issue's table of contents at www.atsjournals.org

Originally Published in Press as DOI: 10.1164/rccm.200507-1059OC on November 10, 2005

Conflict of Interest Statement: None of the authors have a financial relationship with a commercial entity that has an interest in the subject of this manuscript.

Received in original form July 10, 2005; accepted in final form November 9, 2005

	REFERENCES

TOP ABSTRACT METHODS RESULTS DISCUSSION REFERENCES

 

1. Hunt JF, Fang K, Malik R, Snyder A, Malhotra N, Platts-Mills TAE, Gaston B. Endogenous airway acidification: implications for asthma pathophysiology. Am J Respir Crit Care Med 2000;161:694–699.[Abstract/Free Full Text]
2. Borrill Z, Starkey C, Vestbo J, Singh D. Reproducibility of exhaled breath condensate pH in chronic obstructive pulmonary disease Eur Respir J 2005;25:269–274.[Abstract/Free Full Text]
3. Carpagnano GE, Barnes PJ, Francis J, Wilson N, Bush A, Kharitonov SA. Breath condensate pH in children with cystic fibrosis and asthma: a new noninvasive marker of airway inflammation? Chest 2004;125:2005–2010.[Abstract/Free Full Text]
4. Chung KF. Reduced pH and chloride levels in exhaled breath condensate of patients with chronic cough. Thorax 2004;59:608–612.[Abstract/Free Full Text]
5. Gessner C, Hammerschmidt S, Kuhn H, Seyfarth HJ, Sack U, Engelmann L, Schauer J, Wirtz H. Exhaled breath condensate acidification in acute lung injury. Respir Med 2003;97:1188–1194.[CrossRef][Medline]
6. Kostikas K, Papatheodorou G, Ganas K, Psathakis K, Panagou P, Loukides S. pH in expired breath condensate of patients with inflammatory airway diseases. Am J Respir Crit Care Med 2002;165:1364–1370.[Abstract/Free Full Text]
7. Moloney ED, Mumby SE, Gajdocsi R, Cranshaw JH, Kharitonov SA, Quinlan GJ, Griffiths MJ. Exhaled breath condensate detects markers of pulmonary inflammation after cardiothoracic surgery. Am J Respir Crit Care Med 2004;169:64–69.[Abstract/Free Full Text]
8. Niimi A, Nguyen LT, Usmani O, Mann B, Chung KF. Reduced pH and chloride levels in exhaled breath condensate of patients with chronic cough. Thorax 2004;59:608–612.[Abstract/Free Full Text]
9. Ojoo JC, Mulrennan SA, Kastelik JA, Morice AH, Redington AE. Exhaled breath condensate pH and exhaled nitric oxide in allergic asthma and in cystic fibrosis. Thorax 2005;60:22–26.[Abstract/Free Full Text]
10. Tate S, MacGregor G, Davis M, Innes JA, Greening AP. Airways in cystic fibrosis are acidified: detection by exhaled breath condensate. Thorax 2002;57:926–929.[Abstract/Free Full Text]
11. Hunt JF, Erwin E, Palmer L, Vaughan J, Malhotra N, Platts-Mills T, Gaston B. Expression and activity of pH-regulatory glutaminase in the human airway epithelium. Am J Respir Crit Care Med 2002;165:101–107.[Abstract/Free Full Text]
12. Wells K, Vaughan J, Pajewski TN, Hom S, Ngamtrakulpanit L, Smith A, Nguyen A, Turner R, Hunt J. Exhaled breath condensate pH assays are not influenced by oral ammonia. Thorax 2005;60:27–31.[Abstract/Free Full Text]
13. Vaughan J, Ngamtrakulpanit L, Pajewski TN, Turner R, Nguyen TA, Smith A, Urban P, Hom S, Gaston B, Hunt J. Exhaled breath condensate pH is a robust and reproducible assay of airway acidity. Eur Respir J 2003;322:889–894.
14. Crapo RO, Morris AH, Gardner RM. Reference spirometric values using techniques and equipment that meet ATS recommendations. Am Rev Respir Dis 1981;123:659–664.[Medline]
15. Effros RM, Wahlen K, Bosbous M, Castillo D, Foss B, Dunning M, Gare M, Lin W, Sun F. Dilution of respiratory solutes in exhaled condensates. Am J Respir Crit Care Med 2002;165:663–669.[Abstract/Free Full Text]
16. Effros RM, Biller J, Foss B, Hoagland K, Dunning MB, Castillo D, Bosbous M, Sun F, Shaker R. A simple method for estimating respiratory solute dilution in exhaled breath condensates. Am J Respir Crit Care Med 2003;168:1500–1505.[Abstract/Free Full Text]
17. Vass G, Huszár E, Barát E, Valyon M, Kiss D, Pénzes I, Augusztinovicz M, Horváth I. Comparison of nasal and oral inhalation during exhaled breath condensate collection. Am J Respir Crit Care Med 2003;167:850–855.[Abstract/Free Full Text]
18. Norwood DM, Wainman T, Lioy PJ, Waldman JM. Breath ammonia depletion and its relevance to acidic aerosol exposure studies. Arch Environ Health 1992;47:309–313.[Medline]
19. Effros RM, Peterson B, Casaburi R, Su J, Dunning M III, Torday J, Biller J, Shaker R. Epithelial lining fluid concentrations in chronic obstructive lung disease patients and normal subjects. J Appl Physiol 2005;99:1286–1292.[Abstract/Free Full Text]
20. Effros RM, Dunning M, Biller J, Shaker R. The promise and perils of exhaled breath condensates. Am J Physiol 2004;287:L1073–L1080.
21. Dwyer TM. Sampling airway surface liquid: nonvolatiles in exhaled breath condensate. Lung 2004;182:241–250.[CrossRef][Medline]
22. Vaughan W, Gaston B, MacDonald T, Erwin E, Malhotra N, Zaman K, Platts-Mills TAE, Hunt J. Acetic acid contributes to breath condensate acidity in asthma [abstract]. Eur Respir J 2001;18:463S.
23. Chen ZF, Darvell BW, Leung VW. Human salivary anionic analysis using ion chromatography. Arch Oral Biol 2004;49:863–869.[Medline]
24. Bridges SB, Anderson JW, Deakins DA, Dillon DW, Wood CL. Oral bran increases serum acetate of hypercholesterolemic men. Am J Clin Nutr 1992;56:455–459.[Abstract/Free Full Text]
25. Harding SM. Recent clinical investigations examining the association of asthma and gastroesophageal reflux. Am J Med 2003;115:137–143.[CrossRef]
26. Effros RM, Bosbous M, Foss B, Shaker R, Biller J. Exhaled breath condensates: a potential novel technique for detecting aspiration. Am J Med 2003;115:137–143.
27. Ricciardolo FLM, Gaston B, Hunt J. Acid stress in the pathology of asthma. J Allergy Clin Immunol 2004;113:610–619.[CrossRef][Medline]
28. Mitsuhashi M, Mochizuki H, Tokuyama K, Morikawa A, Kuroume T. Hyperresponsiveness of cough receptors in patients with bronchial asthma. Pediatrics 1985;75:855–858.[Abstract/Free Full Text]
29. Gauderman WJ, Gilliland GF, Vora H, Avol E, Stram D, McConnell R, Thomas D, Lurmann F, Margolis HG, Rappaport EB, et al. Association between air pollution and lung function growth in southern California children: results from a second cohort. Am J Respir Crit Care Med 2002;166:76–84.[Abstract/Free Full Text]
30. Gauderman WJ, Avol E, Gilliland F, Vora H, Thomas D, Berhane K, McConnell R, Kuenzli N, Lurmann F, Rappaport E, et al. The effect of air pollution on lung development from 10 to 18 years of age. N Engl J Med 2004;351:1057–1067.[Abstract/Free Full Text]

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				G. Horvath, N. Schmid, M. A. Fragoso, A. Schmid, G. E. Conner, M. Salathe, and A. Wanner Epithelial Organic Cation Transporters Ensure pH-Dependent Drug Absorption in the Airway Am. J. Respir. Cell Mol. Biol., January 1, 2007; 36(1): 53 - 60. [Abstract] [Full Text] [PDF]

				Z. L. Borrill, J. A. Smith, J. Naylor, A. A. Woodcock, and D. Singh The effect of gas standardisation on exhaled breath condensate pH. Eur. Respir. J., July 1, 2006; 28(1): 251 - 252. [Full Text] [PDF]

				P. J. Barnes, B. Chowdhury, S. A. Kharitonov, H. Magnussen, C. P. Page, D. Postma, and M. Saetta Pulmonary Biomarkers in Chronic Obstructive Pulmonary Disease Am. J. Respir. Crit. Care Med., July 1, 2006; 174(1): 6 - 14. [Abstract] [Full Text] [PDF]

				R. M. Effros Exhaled Breath Condensate pH. Am. J. Respir. Crit. Care Med., May 1, 2006; 173(9): 1047b - 11048. [Full Text] [PDF]

				J. Hunt Exhaled Breath Condensate pH: Reflecting Acidification of the Airway at All Levels Am. J. Respir. Crit. Care Med., February 15, 2006; 173(4): 366 - 367. [Full Text] [PDF]

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