title: Determining urea levels in exhaled breath condensate with minimal preparation steps and classic LC-MS.
abstract: Exhaled breath condensate (EBC) provides a relatively easy, non-invasive method for measuring biomarkers of inflammation and oxidative stress in the airways. However, the levels of these biomarkers in EBC are influenced, not only by their levels in lung lining fluid but also by the volume of water vapor that also condenses during EBC collection. For this reason, the use of a biomarker of dilution has been recommended. Urea has been proposed and utilized as a promising dilution biomarker due to its even distribution throughout the body and relatively low volatility. Current EBC urea analytical methods either are not sensitive enough, necessitating large volumes of EBC, or are labor intensive, requiring a derivatization step or other pretreatment. We report here a straightforward and reliable LC-MS approach that we developed that does not require derivatization or large sample volume (∼36 µL). An Acclaim mixed-mode hydrophilic interaction chromatography column was selected because it can produce good peak symmetry and efficiently separate urea from other polar and nonpolar compounds. To achieve a high recovery rate, a slow and incomplete evaporation method was used followed by a solvent-phase exchange. Among EBC samples collected from 28 children, urea levels were found to be highly variable, with a relative standard deviation of 234%, suggesting high variability in dilution of the lung lining fluid component of EBC. The limit of detection was found to be 0.036 µg/mL.
authors.affiliation: Lamont-Doherty Earth Observatory of Columbia University, Palisades, NY, USA Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY, USA.
authors.forename: Masha
authors.initials: M
authors.lastname: Pitiranggon
authors.affiliation: Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY, USA.
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Perzanowski
authors.affiliation: Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY, USA.
authors.forename: Patrick L
authors.initials: PL
authors.lastname: Kinney
authors.affiliation: Institute for Environmental Health and Related Product Safety, Chinese Center for Disease Control and Prevention, Beijing, PR China.
authors.forename: Dongqun
authors.initials: D
authors.lastname: Xu
authors.affiliation: Lamont-Doherty Earth Observatory of Columbia University, Palisades, NY, USA.
authors.forename: Steven N
authors.initials: SN
authors.lastname: Chillrud
authors.affiliation: Lamont-Doherty Earth Observatory of Columbia University, Palisades, NY, USA yanbz@ldeo.columbia.edu.
authors.forename: Beizhan
authors.initials: B
authors.lastname: Yan
mesh_terms: D015415:Biomarkers / Q000032:analysis; D001944:Breath Tests / Q000379:methods*; D002648:Child; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D045853:Exhalation; D006801:Humans; D057230:Limit of Detection; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods*; D014508:Urea / Q000032:analysis*
chemical_list: D015415:Biomarkers; D014508:Urea