Once-daily administration of aminoglycoside antibiotics has become the most acceptable dosing schedule for the majority of patients. There are few published data on the impact of post-natal age on aminoglycoside concentrations in preterm infants receiving once-daily dosage regimens. Netilmicin was administered as a once-daily dose of 4 mg/kg. In 141 episodes of suspected sepsis in 123 babies, trough netilmicin concentrations ranged from undetectable to 4.0 mg/l. Netilmicin concentrations were above a level of 2 mg/l in 10.6% of episodes. Netilmicin concentrations decreased with increasing post-natal age and weight. Levels were higher in males compared to females. Increased creatinine concentrations were associated with higher netilmicin concentrations. This study emphasises the importance of post-natal age as a determinant of aminoglycoside concentrations with a once-daily dosing regimen in a neonatal intensive care population. Trough levels should be carefully monitored and consideration given to extending dosage intervals particularly when netilmicin is administered once daily to preterm infants in the first week of life.