BackgroundLow-calorie sweetener (LCS) consumption is prevalent among lactating women, yet infants’ exposure to LCS in human milk is not well-characterized. ObjectiveConduct a pharmacokinetic study of sucralose and ace-K in mothers’ milk and plasma over 72 hours, and in infants’ plasma. MethodsFollowing baseline blood and milk collection, mothers (n=40) consumed 20-ounces of diet cranberry juice, containing sucralose and ace-K. Blood samples were collected from the mother 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, 24, 48, and 72 hours after beverage ingestion, and milk was expressed at 1, 2, 3, 4, 6, 8, 12, and 24 hours post-ingestion. One blood sample was collected from each infant, the timing of which was determined using pharmacokinetics model-based simulation. Concentration-time profiles of LCS from mother’s plasma and milk were analyzed using non-compartmental methods. ResultsAce-K rapidly entered human milk with the largest observed concentration of 373.0 (CV 69%) ng/ml first detected 4 hours following diet beverage ingestion. Sucralose appeared in human milk 1-2 hours after diet beverage ingestion with the largest observed concentration of 7.2 (CV 63%) ng/ml first detected 7 hours post-ingestion. The mean 24-hour milk to plasma ratio (MPR) of ace-K was 1.75 (SD 1.37) with a mean relative infant dose (RID) of 1.59% (SD 1.72%). ace-K was detected in all infants’ plasma with an average concentration of 9.2 (SD% 14.8) ng/ml approximately 6 hours after maternal beverage ingestion. The mean 24-hour MPR of sucralose was 0.15 (SD 0.06) with a mean RID of 0.04% (SD 0.02%). Sucralose was detected in only fifteen infants’ plasma, and the average concentration was 5.0 (SD% 7.1) ng/ml, approximately 5 hours after diet beverage ingestion. ConclusionsAce-K rapidly transfers from human milk into infants’ circulation whereas sucralose was detected at much lower concentrations and in some but not all infants. Future research should investigate effects of early life sucralose and ace-K exposure via human milk on infants’ health. Trial RegistrationNCT05379270, https://classic.clinicaltrials.gov/ct2/show/NCT05379270