An isotope dilution assay for the determination of N,N-dimethyltryptamine in human plasma has been developed. Dideutero-N,N-dimethyltryptamine is added to the plasma as a carrier, and also serves as an internal standard in a gas chromatographic-mass spectrometric analysis of the final isolate, Mass spectrometric isotope intensity ratios for two ions are used to calculate the amount of endogenous N,N-dimethyltryptamine in the isolate at a sensitivity limit of 0.5 ± 0.2 ng/ml plasma.