Gas chromatography and IDF Standard method (1999) were used to analyze the fatty acid composition of milk fat of mares originating from the following breeds: 10 of Wielkopolska breed, 10 Konik Polski Horses, and 9 Polish Cold-blooded Horses. Eighty-seven mare's milk samples were collected in the years 2000-2002. Unsaturated fatty acids were shown to prevail in the milk fat of mares of Wielkopolska breed (61.32%) and of Konik Polski mares (52.58%) whereas saturated acids prevailed in the milk fat of Cold Blooded mares (54.95%). The study revealed that the fatty acid composition of the investigated groups of mares was breed-specific. Of course, the impact of other uncontrolled factors such as nutrition is not excluded, either.