HPLC-Parallel accelerator and molecular mass spectrometry analysis of 14C-labeled amino acids.

Accelerator mass spectrometry (AMS) is the method of choice for quantitation of low amounts of 14C-labeled biomolecules. Despite exquisite sensitivity, an important limitation of AMS is its inability to provide structural information about the analyte. This limitation is not critical when the labeled compounds are well-characterized prior to AMS analysis. However, analyte identity is important in other experiments where, for example, a compound is metabolized and the structures of its metabolites are not known. We previously described a moving wire interface that enables direct AMS measurement of liquid sample in the form of discrete drops or HPLC eluent without the need for individual fraction collection, termed liquid sample-AMS (LS-AMS). We now report the coupling of LS-AMS with a molecular mass spectrometer, providing parallel accelerator and molecular mass spectrometry (PAMMS) detection of analytes separated by liquid chromatography. The repeatability of the method was examined by performing repeated injections of 14C-labeled tryptophan, and relative standard deviations of the 14C peak areas were ≤10.57% after applying a normalization factor based on a standard. Five 14C-labeled amino acids were separated and detected to provide simultaneous quantitative AMS and structural MS data, and AMS results were compared with solid sample-AMS (SS-AMS) data using Bland-Altman plots. To demonstrate the utility of the workflow, yeast cells were grown in a medium with 14C-labeled tryptophan. The cell extracts were analyzed by PAMMS, and 14C was detected in tryptophan and its metabolite kynurenine.