The present work intends to obtain and characterize a milk by-product: the human whey (HW), in order to avoid the excessive disposal of human milk (HM). The characterization was performed through analyzes of the chemical composition and fatty acids (FAs) and the triacylglycerol (TAG) lipid profile. The results indicated that the chemical composition altered significantly after the HW obtained, except for the percentage of ash and carbohydrates. About the FA composition, it was observed that strictly essential FA, essential FA and other FAs found in HM remained present in HW. For the nutritional lipids quality, the atherogenicity and the thrombogenicity indices presented desirable values, and the TAGs lipid profile revealed a variation in the analyzed samples. Therefore, it can be concluded that the characterized HW has potential for application in isolated form or to be used as an ingredient in other foods.