A mass spectrometric method to simultaneously measure a biomarker and dilution marker in exhaled breath condensate

Esther, Charles R.; Jasin, H. Matias; Collins, Leonard B.; Swenberg, James A.; Boysen, Gunnar

doi:10.1002/rcm.3408

Cited by 45 publications

(59 citation statements)

References 10 publications

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“…Sixty-one samples from RES19044 and 102 from ECLIPSE were analyzed using a previously published method 6,15 ( Fig 1A ) modifi ed to account for variable volumes. As previously described, 15 ]) were lyophilized, resuspended, and subjected to mass spectrometric analysis of urea, adenosine, and AMP.…”

Section: Mass Spectrometric Analysismentioning

confidence: 99%

“…As previously described, 15 ]) were lyophilized, resuspended, and subjected to mass spectrometric analysis of urea, adenosine, and AMP. A second aliquot of up to 500 m L was processed as described here for replicate analysis if suffi cient volume was available.…”

Section: Mass Spectrometric Analysismentioning

confidence: 99%

“…3 orders of magnitude above the mean); seven with urea concentrations below reliable quantifi cation threshold of 1 m M as determined in a previous study. 15 Samples with subthreshold urea measurements were excluded since we could not determine the extent of dilution of airway secretions within the condensate. Two samples with AMP concentrations below quantifi able limits were assigned a value of one-half of the lowest measured concentration.…”

Section: Mass Spectrometric Analysismentioning

confidence: 99%

“…Age and gender distribution was generally similar among cohorts, whereas FEV 1 was decreased signifi cantly in COPD ( Table 1 ). Use of inhaled corticosteroids was common in the COPD To assess airway purine concentrations noninvasively in COPD, we applied a previously reported mass spectrometric method 6,15 to measure adenosine, AMP, and phenylalanine, plus urea as a dilution marker, in EBC collected from participants in two multicenter observational studies. Variable dilution of airway secretions within EBC was controlled by assessing biomarkers as ratios to urea or using serum to EBC urea-based dilution factors.…”

Section: Adenosine and Other Biomarkers Measured In Ebcmentioning

confidence: 99%

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Elevated Airway Purines in COPD

Esther

Lazaar

Bordonali

et al. 2011

Chest

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Purinergic signaling pathways regulate airways defense mechanisms, including mucociliary clearance and infl ammatory responses. These pathways are activated by extracellular purines, which include adenosine triphosphate (ATP) and its metabolites adenosine diphosphate, adenosine monophosphate (AMP), and adenosine. Adenosine, in particular, has been linked to infl ammatory airways diseases, 1 including twofold to fourfold elevations in blood 2,3 and BAL fl uid 4 of subjects with asthma. Similarly, adenosine concentrations in exhaled breath condensate (EBC) are elevated in subjects with stable asthma, 5,6 increased with asthma exacerbations, 7 and decreased with successful treatment. Findings from animal models, particularly the adenosine deaminase knockout mouse, 8,9 further support a link between airway infl ammation and lung adenosine.These data suggest that purinergic signaling pathways may be active in infl ammatory lung diseases such as COPD and may be targets for therapeutic intervention. 10 Indeed, elevated airway adenosine concentrations in COPD have been inferred from studies of human airway adenosine receptors, 11 and several reviews have touted potential benefi ts of pharmacologic blockade of adenosine receptors. 10,12 Although these benefi ts are a reasonable extrapolation from the animal 8,13 and human studies, 4,5 a recent investigation suggested that airway adenosine concentrations (as measured in sputum) may actually be lower in COPD. 14 However, adenosine concentrations in respiratory samples such as sputum or BAL can be altered unpredictably by extracellular metabolism during processing or by mechanical and osmotic forces that trigger purine release during collection.

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Section: Mass Spectrometric Analysismentioning

confidence: 99%

Section: Mass Spectrometric Analysismentioning

confidence: 99%

Section: Mass Spectrometric Analysismentioning

confidence: 99%

Section: Adenosine and Other Biomarkers Measured In Ebcmentioning

confidence: 99%

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Elevated Airway Purines in COPD

Esther

Lazaar

Bordonali

et al. 2011

Chest

Self Cite

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“…Urea derivatization is a time and labor-intensive procedure and requires the use of hazardous materials. Methods used in other studies also involve a 20-to 250-fold concentration of the EBC samples (13,30), requiring relatively large volumes of EBC samples, which typically are difficult to obtain from young subjects. Indeed, Patel et al (31) had some difficulty getting reliable urea measurements from the small volumes of EBC they collected from infants and preschoolers.…”

Section: Introductionmentioning

confidence: 99%

Determining Urea Levels in Exhaled Breath Condensate with Minimal Preparation Steps and Classic LC-MS

Pitiranggon¹,

Perzanowski²,

Kinney³

et al. 2013

Journal of Chromatographic Science

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Exhaled breath condensate (EBC) provides a relatively easy, noninvasive 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 mL). 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 mg/mL.

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A universal surrogate matrix assay for urea measurement in clinical pharmacokinetic studies of respiratory diseases

Tang,

Van Parys,

Walker

et al. 2023

Biomedical Chromatography

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In pharmacokinetic studies for respiratory diseases, urea is a commonly used dilution marker for volume normalization of various biological matrices, owing to the fact that urea diffuses freely throughout the body and is minimally affected by disease states. In this study, we developed a convenient liquid chromatography–tandem mass spectrometry (LC–MS/MS) surrogate matrix assay for accurate urea quantitation in plasma, serum and epithelial lining fluid. Different mass spectrometer platforms and ionization modes were compared in parallel. The LC method and mass spectrometer parameters were comprehensively optimized to reduce interferences, to smooth the baseline and to maximize the signal‐to‐noise ratio. Saline was selected as the surrogate matrix, and its suitability was confirmed by good parallelism and accurate quality control sample measurements. Reliable and robust assay performance was demonstrated by precision and accuracy, dilution integrity, sensitivity, recovery and stability, all of which met bioanalysis requirements to support clinical studies. The assay performance was also verified and better understood by comparing it with a colorimetric assay and to a surrogate analyte assay. The newly developed surrogate matrix assay has the potential to be further expanded for urea quantitation in numerous physiological matrices.

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A mass spectrometric method to simultaneously measure a biomarker and dilution marker in exhaled breath condensate

Cited by 45 publications

References 10 publications

Elevated Airway Purines in COPD

Elevated Airway Purines in COPD

Determining Urea Levels in Exhaled Breath Condensate with Minimal Preparation Steps and Classic LC-MS

A universal surrogate matrix assay for urea measurement in clinical pharmacokinetic studies of respiratory diseases

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