Abstract Background Radical cure with an 8-aminoquinoline is required to prevent relapses from Plasmodium vivax . Single-dose treatment with tafenoquine offers significant advantages over primaquine, but the amount of tafenoquine excreted in breastmilk is unknown. As tafenoquine can cause serious haemolysis in G6PD deficient individuals, the amount of drug in breastmilk must be determined in order to advise on safe and effective treatment for lactating women. Methods Six healthy lactating volunteers ≥ 1 year postpartum were recruited for this mother-only pilot pharmacokinetic study. Paired breastmilk and venous samples were taken on day 0, 1, 7 and 14. Breastmilk samples were collected in serial 5 ml aliquots to test foremilk and hindmilk drug concentrations. Results Tafenoquine concentrations in milk were similar to concentrations in blood (milk/plasma ratio 1.09, range: 0.58–1.63). Concentrations were higher in hindmilk and in lower volume samples. Estimated drug passed to infants was calculated using a standard estimate of milk intake (150 ml per kg per day) and a high intake scenario (200 ml per kg per day). The maximum estimated dose that would be passed to the infant on a single day was 0.050 mg/kg in the first 24 h (range 0.032–0.062 mg/kg). Median weight-adjusted proportion of a therapeutic dose that infants would ingest (relative infant dose, RID) in total ranged from 5.07% (assuming of 150 ml milk per kg per day and 10 mg/kg therapeutic dose) to 13.52% (assuming 200 ml milk per kg per day and 5 mg/kg therapeutic dose). The drug was well tolerated by mothers. Discussion This pilot study demonstrated measurable tafenoquine in breastmilk, with very small amounts of drug passing to babies on any given day but overall estimated RIDs around the conventional 10% threshold for safety concerns. Further studies are needed to determine with greater certainty the excretion of tafenoquine in breastmilk and drug concentrations in infant blood to inform recommendations for breastfeeding women.