The study was carried out on milk from cold-blooded Sokólski mares and warm-blooded Polish Halfbred mares. Milk samples were analysed for content of solids, protein, fat, lactose, and ash; density; energy value; percentage of α-lactalbumin (α-La), β-lactoglobulin (β-Lg), serum albumin (SA), immunoglobulin (Ig), lactoferrin (Lf) and lysozyme (Lz) in the total protein; and fatty acid profile. Milk from cold-blooded mares was found to be similar in protein and fat content to that of warm-blooded mares, but had higher content of solids, including lactose and ash. It also had significantly (P≤0.01) higher content of α-La, SA and Lz, and lower content of β-Lg, Ig and Lf in comparison to milk from warm-blooded mares. Milk from Sokólski horses had a significantly (P≤0.01) higher proportion of SFA, including C10:0, C12:0, C14:0, C16:0 (P≤0.01) and C18:0 (P≤0.05), while the milk of warm-blooded mares had a significantly (P≤0.01) higher proportion of MUFA, including C16:1n-9 and C18:n-9, and PUFA, including C18:2n-6 and C18:3n3. The share of n-6 and n-3 PUFA was significantly (P≤0.01) higher in milk from warm-blooded horses, but their ratio (n-6/n-3) was lower (more favourable) in milk from cold-blooded mares. In addition, milk from warm-blooded horses had a lower (P≤0.01) share of HSFA and a higher proportion of DFA compared to cold-blooded horses, as well as lower values for AI and TI. Despite these minor differences in comparison with warmblooded mares (Polish Halfbred), milk from cold-blooded Sokólski mares was shown to be a food product of high nutritional value, which is one argument in favour of the use of this horse breed for dairy purposes.