The nutritional composition of milk from donkeys of the Nordestina breed was characterised during lactation from animals raised in the Brazilian semiarid region, in a semi-confinement and fed with native Caatinga vegetation. The contents of lactose, total solids, melatonin, minerals, fatty acids varied during lactation, while, the concentrations of protein and lipids remained stable. Medium chain and long chain fatty acids made up more than 80% of total fatty acids in this dairy matrix. Lysozyme, β-lactoglobulin and α-lactalbumin were present, and essential amino acid content was reported. The high concentration of lactose, along with protein, amino acid and mineral profile constitutes the Nordestina donkey milk an expressive nutritional value, presenting health benefits, as well as an important ingredient in the formulation of food products.