The effects of gestational age at delivery (GA), postnatal age (PNA) and post-menstrual age (PMA=PNA+GA, an indicator of autonomous developmental processes not affected by the moment of birth) on macronutrient composition of very preterm milk were studied. Total N, fat, lactose and carbohydrate concentrations, energy density and 24 h volume were determined in 282 24 h milk samples collected at weekly intervals (days 7–55 of lactation) from seventy-nine women delivering their babies between 25 and 29 weeks of gestation. GA related differences were found for carbohydrate concentration only: carbohydrate concentration was lower with increasing GA. PNA was related to a decrease in total N and an increase in lactose concentration. PMA was not related to milk composition. Our data indicate that PNA strongly influences the development of the composition of very preterm human milk, while GA affects carbohydrate content with a negligible effect on the nutritional value of the milk. We conclude that in accordance with current opinion in paediatrics, human milk is the best source of nutrients even for very preterm (< 30 weeks GA) infants.