Metabolic Profiling Using In Vivo High Field Flow NMR.

In vivo NMR (nuclear magnetic resonance) has the potential to monitor and record metabolic flux in close to real time, which is essential for better understanding the toxic mode of action of a contaminant and deciphering complex interconnected stress-induced pathways impacted inside an organism. Here, we describe how to construct and use a simple flow system to keep small aquatic organisms alive inside the NMR spectrometer. In living organisms, magnetic susceptibility distortions lead to severe broadening in conventional NMR. Two main approaches can be employed to overcome this issue: (1) use a pulse sequence to reduce the distortions, or (2) employ multidimensional NMR in combination with isotopic enrichment to provide the spectral dispersion required to separate peaks from overlapping resonances. Both approaches are discussed, and protocols for both approaches are provided here in the context of small aquatic organisms.