Nitrogen-13- ([13N]) ammonia is a widely used tracer for PET myocardial blood flow studies. Quantification of blood flow using tracer kinetic principles requires accurate determination of [13N]ammonia activity in blood. Since [13N] ammonia is rapidly metabolized, the arterial input function may be contaminated by labeled metabolites. We, therefore, characterized the 13N-labeled metabolites in blood after intravenous (i.v.) injection of 20 mCi [13N]ammonia in nine healthy volunteers. Utilizing a series of ion exchange resins, 13N-labeled compounds were separated into four groups: ammonia, neutral amino acids, acidic amino acids, and urea. Analysis of the metabolic fate of [13N]ammonia indicates that over 90% of the blood activity within the first two minutes after injection is present as [13N]ammonia. However, there is considerable contamination of the blood activity at 3-5 min by [13N]glutamine (amide) and urea, which collectively represent 18%-50% of the blood activity. Thus, correction of the arterial input function for 13N-metabolites is required to accurately quantify the arterial input function of [13N]ammonia in myocardial blood flow studies.