Abstract The stable isotope tracer technique using 13C labeling of substrates followed by NMR spectroscopy of biofluids has been widely used in metabolic investigations, whereas the use of 2H labeling and 2H NMR spectroscopy has been extremely limited. The applicability of the high-field 2H NMR spectroscopy (14.1 T, 92 MHz 2H frequency) in a simple pharmacokinetic problem has now been investigated using selectively deuterated benzoic acid (BA) as a model. [7- 13C,2,6- 2H 2]BA was synthesized for use as a tracer to compare the efficiency and sensitivity of 2H and 13C labeling. The urinary excretion of [7- 13C,2,6- 2H 2]hippuric acid (HA) formed from orally administered [7- 13C,2,6- 2H 2]BA (250 mg) was followed by 92-MHz 2H and 150-MHz 13C NMR spectroscopy (only 10 min accumulation time) following concentration of urine by a factor of 10, using a standard for quantitation. The heights of resonances for 13C7 and 2H2,6 were used to calculate the [7- 13C,2,6- 2H 2]HA concentration. The lower limit of detection using this 2H NMR approach was approximately 60 nmol/ml and was found to be comparable with that of the 13C NMR approach where the quaternary carbon (C7) was labeled. The administered [7- 13C,2,6- 2H 2]BA was found to be quantitatively biotransformed to HA and excreted in urine within 4 h by both NMR approaches. The 2H NMR approach using a high-field NMR spectrometer is potentially useful and practical for pharmacokinetic research on small molecules whose 2H resonances are relatively sharp since the procedures are very simple and convenient.