A workflow for the metabolomic/metabonomic investigation of exhaled breath using thermal desorption GC–MS

Abstract

Confounding factors in the analysis of human breath by thermal desorption GC-MS are reviewed, with special emphasis on the high water levels encountered in human breath samples. Multilinear regression optimization of breath sampling factors, along with the selection of ubiquitous sample components used as retention-time standards, enabled data registration based on retention indexing and mass spectral alignment. This was done on a component-by-component basis. The methodology developed reconciled participant safety, artefacts from accelerated hydrolysis of the stationary phase and the destructive nature of thermal desorption. Furthermore, using ubiquitous methylated cyclic-siloxanes in the thermal desorption-GC-MS chromatograms enabled secondary retention indexing for each chromatogram. This methodology enables the creation of a 'breath matrix' that is based on a combination of retention indexing and the mass spectral registration of isolated peaks. This approach facilitated more efficient data modeling and a case study from a 22-participant (10 male, 12 female) stress-intervention experiment. Principal component analysis of data registered by retention indexing did not classify successfully stressed from unstressed states. By contrast, adoption of a breath matrix approach enabled 95% separation.