(182b) Tandem Mass Spectrometry: A Novel Approach for Metabolic Flux Analysis

Recent progress in methods for metabolic flux analysis using stable-isotope labeling experiments and gas chromatography/mass spectrometry (GC/MS) and nuclear magnetic resonance (NMR) has allowed us to estimate fluxes in medium-sized and large scale network models. Here, we introduce a novel method for quantifying isotopic labeling and estimating metabolic fluxes based on new technique, tandem mass spectrometry (MS/MS), that can dramatically increase the resolution with which fluxes can be determined in vivo. Tandem mass spectrometry is a relatively new tool that provides an order of magnitude more sensitive labeling information than can be obtained from GC-MS. However, the experimental and computational methods for measuring and analyzing data from tandem mass spectrometry have not been fully developed. Here, we present the theoretical framework that allows us to take full advantage of the additional labeling information from MS/MS, in particular for estimating metabolic fluxes. In this novel modeling framework the isotopic labeling of is represented in terms of tandem mass isotopomer distributions that can be measured using daughter ion scanning and multiple reaction monitoring (MRM). The resulting tandem mass isotopomer matrix contains all possible parent-to-daughter ion transitions, which can be simulated using the computational tools that we have developed. We will illustrate that our modeling methods allow us to trace isotopic labeling in complex metabolic networks and ultimately estimate fluxes with much a much higher degree of confidence.