Penicillin concentrations in cerebrospinal fluid (CSF) were measured at various hours and days of treatment in 163 infants undergoing therapy for congenital syphilis. The CSF levels were compared for three treatment regimens. Aqueous penicillin G (A-PEN), 100,000 U/kg per day, was used in 23 infant, and a dosage of 200,000 U/kg per day was used in 40 patients; procaine penicillin G (P-PEN), 50,000 U/kg per day, was used in 100 children. Mean CSF penicillin levels were 0.416, 0.493, and 0.077 microgram/ml, respectively, in the three treatment groups. The mean CSF penicillin concentration among the 63 infants treated with either of the A-PEN regimens (0.465 microgram/ml) was significantly greater than the mean concentration (0.077 microgram/ml) among those treated with P-PEN (p < 0.001). Among those who received A-PEN, the difference in dosage was not associated with a significant difference in mean CSF penicillin concentration (p = 0.68). All the specimens obtained from patients who received A-PEN, but only 82% of those from patients who received P-PEN, had treponemicidal concentrations (> or = 0.018 microgram/ml). However, 33.3% (9/27) of specimens from infants who received P-PEN, tested between 18 and 24 hours after a dose, had CSF penicillin concentrations < 0.018 microgram/ml. These data suggest that administration of A-PEN may be the preferred therapy if CSF levels > 0.018 microgram/ml are desired, especially for infants with severe disease or congenital neurosyphilis.