The objective of the study was to assess the composition of fatty acids (FA) in the mare’s milk fat and to compare them with the composition of FA in the cow’s milk fat, with a focus on the FA appearing essential in human nutrition. The research material consisted of 10 mare’s milk samples and 10 cow’s milk samples taken from the milk of the animals during the winter and summer feeding periods, in the Mazovia region. In each sample, the fatty matter content was determined using a Rose-Gottlieb method, as was the composition of FA using a gas chromatography (GC). The analysis of FA composition in the mare’s milk showed a high content of saturated FA: caprylic C8:0 (from 2.81 to 5.17 %), capric C10: 0 (from 6.30 to 11.34 %), lauric C12:0 (from 6.94 to 9.79 %). The unsaturated FA of the mare’s milk were represented mainly by the linoleic acid C18:2 9c12c (from 12.29 to 13.78 %) and the α-linolenic acid C18:3 9c12c15c (from 3.74 to 5.10 %). The cow’s milk was characterized by a higher content of the following saturated FA: butyric acid C4:0 (from 2.37 to 2.63 %), caproic acid C6:0 (from 1.78 to 2.06 %), and stearic acid C18:0 (from 10.77 to 10.89 %), as well as by a significantly lower content of polyunsaturated FA: linolenic (from 1.17 to 1.22 %) and α-linolenic acid (from 0.38 to 0.48 %). Particularly, specific FA synthesized in the bio-hydrogenation process were found in the cow’s milk, i.e. the vaccenic acid C18:1 11t (from 1.08 to 1.65 %) and CLA C18:2 9c11t (from 0.43 to 0.67 %). The content of FA in the milk of those two animal species was characterized by the high season-dependent variation.