Metabolic profiling of carnitine, acylcarnitines, amino acids, and other metabolites in plasma samples by monolithic and HILIC HPLC-ESI-TOF-MS

ESTELA SOLEDAD CERUTTI; TIMOTHY GARRETT; PEGGY BORUM; JODIE V. JOHNSON; RICHARD A. YOST; DAVID H. POWELL

Denver, Colorado, Estados Unidos

Conferencia; 56th ASMS Conference on Mass Spectrometry and Allied Topics; 2008

American Society for Mass Spectrometry

Acidic conditions were used to promote the protonation of some of the compounds under study. Thus, the positive ESI and APCI normal mass spectra were dominated by the presence of the [M+H]+ ions with minimal fragmentation. On the contrary, ammonium acetate was used to enhance deprotonation of those metabolites more easily ionized in negative mode ESI and APCI. In addition, in order to facilitate a better spectral correlation, LC methodologies not including any buffer addition were evaluated. In addition, two different column approaches were compared, a small particle packed column and a silica-based monolithic column. Similar chromatographic resolution was obtained with both columns. The chromatographic profile of eluting compounds was analogous for both column approaches. Rapid, comprehensive, and high-resolution metabolite separation was achieved. Our studies involved a large number of samples that required powerful data analysis capabilities. In this sense, the raw instrument data were processed in several steps by the instrument software. From the obtained chromatograms, features, molecular entities obtained using mass and retention times, were extracted, aligned, normalized, statistically processed, and finally identified. This process resulted in the extraction of > 1,000 feature groups, from which approximately >300 compounds had a S/N ratio greater than nine. Calibration curves have been obtained for a suite of acylcarnitines. Amino acids, lipids, and phospholipids will be included in the data presented. The analytical separation/detection strategy and the analysis steps were also used to evaluate technical variability (sample preparation, instrumental response, etc.). The methodology will be discussed and the results of these analyses will be presented.