Controlling Preanalytical Process in High-Coverage Quantitative Metabolomics: Spot-Sample Collection for Mouse Urine and Fecal Metabolome Profiling

Compared
to conventional MS and NMR techniques, high-performance chemical isotope
labeling (CIL) LC-MS provides accurate relative quantification of
many more metabolites in biological samples. However, to apply this
technique for urine and fecal metabolomics studies of animal models,
the entire workflow, including the preanalytical process, needs to
be strictly controlled to avoid or minimize quantitative errors. In
this study, we report our investigation of the effects of mouse urine
and fecal sample collection methods on CIL LC-MS metabolome analysis.
Metabolic-cage collection and spot-sample collection of urine and
feces were compared in a mouse model of CCl4-induced liver
disease. 13C-/12C-dansylation LC-MS was used
for quantitative profiling of the amine-/phenol-submetabolome changes.
A total of 5026, 4963, 4238, and 4600 peak pairs or metabolites were
detected from spot urine, spot feces, cage-collected urine, and cage-collected
feces, respectively. It was found that samples collected using metabolic
cages, widely used in low coverage metabolomics, could be contaminated
with food as well as cross-specimen (urine in feces or feces in urine)
to the extent that metabolomic comparison of different groups of mice
could be seriously compromised in high-coverage metabolomics. In contrast,
spot urine and spot feces could be collected without contamination
and should be used in CIL LC-MS metabolomics.