© 2003 American Thoracic Society
Further Discussion on Breath Condensate AnalysisSchool of Hygiene and Public Health Johns Hopkins University Baltimore, Maryland In this issue of AJRCCM (pp. 1380–1386), Corradi and coworkers (1) report a new method for determining aldehydes in exhaled breath condensates based on derivatization with 2,4-dinitrophenylhydrazine and analysis by liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometry. This novel analytical method was used to determine the concentrations of aldehydes (malondialdehyde, 1-hexanal, 1-heptanal, 1-nonanal, 4-hydroxynonenal, and 4-hydroxyhexenal) in exhaled breath condensates collected from patients with chronic obstructive pulmonary disease (COPD), and smoking and nonsmoking control subjects (1). Aldehydes were selected because they are produced by the reaction of reactive oxygen species with membrane polyunsaturated fatty acids and because patients with COPD, smokers, or both are known to have elevated levels of reactive oxygen species (oxidative stress). The goal of this report was to identify whether one or more of these products of lipid peroxidation could be used to identify those smokers who were susceptible for the development of COPD. Malondialdehyde was the only aldehyde that exhibited the potential as a biomarker of susceptibility in that it was elevated and the difference between its concentrations in patients with COPD versus smoking control subjects was statistically significant. The remaining saturated aldehydes did not show comparable statistically significant elevations, although they were elevated in both the patients with COPD and smoking control subjects. Hydroxyaldehydes were detected only in a few samples of exhaled breath condensate, although they have been reported to be elevated in the lung tissue of patients with COPD (2). In a related study, some of the investigators from the current study (1) observed that malondialdehyde was the only aldehyde that was elevated in children with exacerbation of asthma (3). The collection and analysis of breath condensate remains controversial. When these two articles (1, 3) are examined they illustrate one of the sources of this controversy. In the present report (1) the concentrations of aldehydes are reported in units of nanomoles of aldehyde per liter of condensate, whereas the previous article (3) reports the levels of aldehydes in units of nanomoles. Clearly it is difficult to compare the data from these two publications. Normalizing data to the volume of breath condensate is not ideal and it has been suggested by Hunt (4) that data should be normalized to an as yet unidentified "gold standard." Reporting the concentrations of compounds found in exhaled breath condensate in units of nanograms, nanomoles, and so on is not unique to these researchers and a review has listed many studies that report their data in this manner (5). Data that are not normalized cannot be compared from patient to patient and from study to study; this makes analysis of exhaled breath condensate less attractive. Another difficulty that is found in these articles (1, 3) is that the concentrations of 1-hexanal, 1-heptanal, and 1-nonanal are increased for smoking control subjects and patients with COPD as compared with nonsmoking control subjects. In the study of children with exacerbations of asthma, the levels of 1-hexanal and 1-heptanal were comparable to those for the control subjects, the level of 1-nonanal in the control children was almost double the level found in the children with asthma (3). On the basis of the data in these two articles it is difficult to support the hypothesis that 1-hexanal, 1-heptanal, and 1-nonanal are products of lipid peroxidation found in exhaled breath condensate. The vapor pressures of these saturated aldehydes at physiological temperatures (1–10 mm Hg) may account for the concentration of aldehydes found in exhaled breath condensate. Moreover, no differences in the levels of aldehydes were observed in exhaled breath condensate collected before and immediately after smoking one cigarette, which is surprising because aldehydes are among the more than 3,000 compounds that have been determined to be present in cigarette smoke (for review see Hoffmann and Hecht [6]). This report also found that the expiratory flow (50–200 ml/second) and time (10 or 20 minutes) of sampling of exhaled breath did not affect the concentration of molecules in exhaled breath condensate. In addition, the authors found comparable concentrations of aldehydes in exhaled breath condensate when breath was sampled with their homemade breath collection device versus with a commercially available device. It should be noted that in this comparison study the subjects were asked to breathe tidally. The mechanism for the production of aerosolization of species found in the respiratory tract remains poorly understood and the results presented in this article support the proposition that these species are produced by the popping open of closed respiratory bronchioles and alveoli (S. Kharitonov, oral communication, June 2001). This hypothesis must be corroborated by peer-reviewed studies. Finally, if biomarkers found in exhaled breath condensate are to be accepted then it is essential that a task force be established, under the auspices of the American Thoracic Society or the European Respiratory Society, that defines the equipment and protocol for the collection of exhaled breath condensate. A standardized procedure for the measurement of breath nitric oxide is helping this novel volatile breath biomarker gain clinical acceptance (7). REFERENCES
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