Overcoming Sample Matrix Effect in Quantitative Blood Metabolomics Using Chemical Isotope Labeling Liquid Chromatography Mass Spectrometry

Blood is widely used for discovery metabolomics to search for disease biomarkers. However, blood sample matrix can have a profound effect on metabolome analysis, which can impose an undesirable restriction on the type of blood collection tubes that can be used for blood metabolomics. We investigated the effect of blood sample matrix on metabolome analysis using a high-coverage and quantitative metabolome profiling technique based on differential chemical isotope labeling (CIL) LC-MS. We used ¹²C-/¹³C-dansylation LC-MS to perform relative quantification of the amine/phenol submetabolomes of four types of samples (i.e., serum, EDTA plasma, heparin plasma, and citrate plasma) collected from healthy individuals and compare their metabolomic results. From the analysis of 80 plasma and serum samples in experimental triplicate, we detected a total of 3651 metabolites with an average of 1818 metabolites per run (n = 240). The number of metabolites detected and the precision and accuracy of relative quantification were found to be independent of the sample type. Within each sample type, the metabolome data set could reveal biological variation (e.g., sex separation). Although the relative concentrations of some individual metabolites might be different in the four types of samples, for sex separation, all 66 significant metabolites with larger fold-changes (FC ≥ 2 and p < 0.05) found in at least one sample type could be found in the other types of samples with similar or somewhat reduced, but still significant, fold-changes. Our results indicate that CIL LC-MS could overcome the sample matrix effect, thereby greatly broadening the scope of blood metabolomics; any blood samples properly collected in routine clinical settings, including those in biobanks originally used for other purposes, can potentially be used for discovery metabolomics.