Changes in fatty acid profile of woman, cow and sow colostrum and milk fat during lactation were investigated. Colostrum samples were taken 2 times: a few hours postpartum and then 2–3 days postpartum. Also, milk samples were collected 2 times. The first collection from each 3 investigated species took place in 2nd week of lactation and the second collection – in 4th, 6th and 3rd month of lactation from women, cows and sows, respectively. The level of linoleic acid in the colostrum and milk of women gradually decreased during lactation, whereas linolenic acid gradually increased until 2nd week of lactation, after which its level slighty decreased. In cow milk fat, the linoleic acid was stable during the two first weeks of lactation, then its content decreased. The level of linolenic acid increased (from 0.74 to 1.14%) within the investigated 6 months of lactation. The percentage of arachidonic acid also slightly increased (from 0.41% to about 0.50%). High linoleic content in sow colostrum (2.6–3.1%) decreased considerably in milk fat (1.8%). The lowest (0.3%) linolenic acid content in colostrum increased gradually with time of lactation up to 0.8% in 3rd month postpartum.