Standardization procedures for real-time breath analysis by secondary electrospray ionization high-resolution mass spectrometry

摘要

Despite the attractiveness of breath analysis as a non-invasive means to retrieve relevant metabolic information, its introduction into routine clinical practice remains a challenge. Among all the different analytical techniques available to interrogate exhaled breath, secondary electrospray ionization high-resolution mass spectrometry (SESI-HRMS) offers a number of advantages (e.g., real-time, yet wide, metabolome coverage) that makes it ideal for untargeted and targeted studies. However, so far, SESI-HRMS has relied mostly on lab-built prototypes, making it difficult to standardize breath sampling and subsequent analysis, hence preventing further developments such as multi-center clinical studies. To address this issue, we present here a number of new developments. In particular, we have characterized a new SESI interface featuring real-time readout of critical exhalation parameters such as CO2, exhalation flow rate, and exhaled volume. Four healthy subjects provided breath specimens over a period of 1month to characterize the stability of the SESI-HRMS system. A first assessment of the repeatability of the system using a gas standard revealed a coefficient of variation (CV) of 2.9. Three classes of aldehydes, namely 4-hydroxy-2-alkenals, 2-alkenals and 4-hydroxy-2,6-alkedienals?hypothesized to be markers of oxidative stress?were chosen as representative metabolites of interest to evaluate the repeatability and reproducibility of this breath analysis analytical platform. Median and interquartile ranges (IQRs) of CVs for CO2, exhalation flow rate, and exhaled volume were 3.2 (1.5), 3.1 (1.9), and 5.0 (4.6), respectively. Despite the high repeatability observed for these parameters, we observed a systematic decay in the signal during repeated measurements for the shorter fatty aldehydes, which eventually reached a steady state after three/four repeated exhalations. In contrast, longer fatty aldehydes showed a steady behavior, independent of the number of repe