Inside the Personalized Medicine Toolbox: GCxGC-Mass Spectrometry for High-Throughput Profiling of the Human Plasma Metabolome

Personalized medicine advocates have been frustrated by the issue of analyte component resolution in biomolecular profiling. Because complex human biological samples such as plasma, serum, or urine contain ~10³–10⁷ unique molecular entities, the analyte capacity of any high-throughput platform is typically exceeded. In our attempt to overcome this problem, we employed the use of comprehensive two-dimensional gas chromatography (GC×GC) combined with time-of-flight mass spectrometry (TOF-MS) for high-throughput profiling of the human plasma metabolome. The profiling experiments demonstrated the capability of a GC×GC-TOF-MS approach with regard to component resolution as well as overall utility for comparative metabolomic analyses employing plasma samples from a "control" cohort (81 samples) versus a cardiovascular-compromised cohort (15 samples) of individuals. The use of the GC×GC separation has resulted in a platform that provides more than an order of magnitude greater resolving power while at the same time not affecting the overall experimental run time. Additionally, the high-throughput approach provides information-rich datasets that can be used to distinguish the control and disease sample cohorts. Such results suggest that high-throughput, high-coverage profiling capabilities, such as those afforded by GC×GC-TOF-MS techniques, can impact the development of personalized medicine in which future disease prevention, diagnosis, and treatment are tailored to an individual's unique molecular makeup. Challenges to full implementation of high-throughput metabolite profiling by GC×GC-TOF-MS as well as future directions are discussed.

Figure 1

Having noted the difficulties in characterizing the metabolome, it is important to consider the current analytical methods for individual sample profiling. The most widely used techniques include ¹H nuclear magnetic resonance (NMR) spectroscopy, direct infusion mass spectrometry (MS), Fourier transform-infrared (FT-IR) spectroscopy, and liquid chromatography (LC) and gas chromatography (GC) coupled with MS (10,11). Each of these techniques has its advantages as well as its limitations and for this reason, metabolomic profiling often requires the use of multiple approaches (10,12–17). The relatively recent technique of comprehensive two-dimensional (2D) GC (GC×GC) (18–23) is gaining acceptance as a powerful tool for the rapid profiling of complex biological samples (24,25). In addition to the increased peak capacity, the advantage of the approach is that the extra separation dimension reduces the congestion of eluted peaks, thereby allowing the resolution of lower signal species (24).

The work reported here presents an evaluation of a GC×GC-TOF-MS analytical approach for human plasma metabolite profiling. A total of 96 plasma samples have been analyzed using a rapid throughput approach requiring approximately 20 min (the separation time is ~20 min and the cycle time is ~30 min; see the following) per sample. The analytical platform is evaluated based upon robustness, resolving power, and metabolome coverage, and to some degree, the quality of the data. We present a comparative metabolomics study in which the plasma metabolite profiles of 81 control cohort samples are compared to those from a cardiovascular-compromised patient sample cohort (15 total). The limitations of the approach as they pertain to profiling efficacy and personalized medicine as well as future directions for the technique are discussed in the following.