[1-13C]- and [1-14C]glycine were infused into chronically catheterized fetal lambs via a brachial vein. At tracer glycine steady state, samples were collected from the fetal abdominal aorta, umbilical vein, and fetal hepatic vein and from the maternal femoral artery and uterine vein. The samples were analyzed for plasma glycine and serine, for glycine and serine 13C atom% excess (APE), and for whole blood 14CO2 and O2 concentrations. Fetal plasma glycine disposal rate (DR) was 12.4 +/- 0.8 mumol.min-1.kg fetus-1.CO2 production from decarboxylation of fetal plasma glycine was 1.63 +/- 0.16 mumol.min-1.kg fetus-1 and represented 12.3 +/- 0.7% of DR. Approximately 50% of infused tracer glycine was taken up by the fetal liver with the release of labeled serine and CO2 in the fetal circulation. There was no detectable efflux of tracer glycine from the placenta into the maternal circulation. The tracer production of serine and CO2 accounted for 23 and 17%, respectively, of the hepatic tracer glycine uptake. The labeled CO2 released by the liver was a large fraction (approximately 70%) of the labeled CO2 produced by the fetus. The serine-to-glycine APE ratio in fetal plasma was approximately 5%. These results indicate that the fetal liver is the major site of fetal plasma glycine decarboxylation and of serine synthesis from plasma glycine.