Protein binding of thiopental was studied in 21 samples of neonatal serum (from placental blood) and compared with protein binding in ten healthy adult volunteers. These infants ranged between 32 and 43 weeks of gestational age (mean, 37.7 weeks) and the adult age range was from 27 to 54 years (mean, 35.4 years). Because the unbound fraction of the drug is responsible for its pharmacologic effect, a marked difference in the protein binding between neonates and adults may be relevant to the clinician. Blood obtained from freshly delivered placentas or from adult volunteers was allowed to clot and the serum separated from the sample. A portion of the serum was sent for protein and bilirubin analysis and the remainder retained for study. This latter serum was combined with four concentrations of thiopental. These specimens were then ultrafiltered and the amount of thiopental in the ultrafiltrate (unbound) compared with the prefiltered amount (total), as measured by reversephase high-performance liquid chromatography. The binding studies were repeated atpH 7.2, 7.4, and 7.6 in both the adult and neonatal serum. Total protein and albumin are significantly less in neonatal serum, whereas bilirubin (total and direct) is significantly higher in neonatal serum than in adult serum (P< 0.01). Neonatal serum was associated with significantly more unbound thiopental than adult serum at all levels ofpH studied (P< 0.005). Increasing thepH resulted in less free drug in both groups, but this reached statistical significance only in the adult group (P< 0.025). Drug concentration had no effect on binding in the range examined. This increased unbound thiopental (almost twice as much as in adults) may result in increased sensitivity to thiopental in the neonatal age group.