PERFORMANCE OF HUMAN MASS BALANCE METABOLITE IDENTIFICATION STUDIES USING STABLE ISOTOPE (C-13, N-15) LABELING AND CONTINUOUS-FLOW ISOTOPE-RATIO MASS-SPECTROMETRY AS AN ALTERNATIVE TO RADIOACTIVE LABELING METHODS

Stable isotope labeling in therapeutic and subtherapeutic quantities of drug ((N213C)N-15-phenobarbital) can be quantitated in biological matrices (urine) and high performance liquid chromatography (HPLC) peaks from urine using continuous-flow isotope-ratio mass spectrometry (CF-IRMS). Standard curves for (N213C)-N-15-phenobarbital were reproducible and linear (R2 > 0.985) over the ranges of 3-100 mug/mL for whole urine (N-15(2) or C-13 labeling) and 0.1-8.0 mug/mL for HPLC peaks derived from urine (N-15(2) labeling). The lower limit of quantitation values for urine drug concentration was 0.46-2.62 mug/mL in whole urine and 0.10-0.70 mug/mL in HPLC peaks. Validation samples quantitated with these standard curves yielded close to expected values. These data suggest stable isotope labeling and CF-IRMS may be used as an alternative to C-14 labeling and radioactivity counting methods in mass balance/metabolite identification and other biomedical studies.