Abstract. Linseed is an excellent source of PUFA, especially linolenic acid (18:3n-3), but it is not commonly used in dairy diet in Hungary. Aim of this study was to investigate the effect of the supplemented whole linseed on fatty acid profile of milk fat, with particular reference to health promoting components (α-linolenic acid). A total of thirty multiparous Holstein cows were used in two dairy farms to determine the effect of feeding whole linseed rich in linolenic (C18:3) fatty acids on fatty acid composition of milk fat. The experiment was conducted on two large-scale dairy farms located in HajdĂș-Bihar County (referred to as farm "A" and "B"). Fifteen multiparous Holstein-Friesian (165 average days in milk) on farm A and fifteen multiparous Holstein-Friesian (200 average days in milk) on farm B were randomly allocated to treatment groups to investigate the effect of whole linseed supplementation on the composition of fatty acids of milk fat. The group on farm A and B fed TMR based maize silage without whole linseed during the control-period. In the experimental-period the cows received the same TMR on both farms, but it was supplemented with 2.60% DM of whole linseed on farm A, and 2.16% DM of whole linseed on farm B. The experimental period was four weeks in both farms. The supplementation of whole linseed had no effect (P > 0.05) on the short chain saturated fatty acid (SFA), and most of the medium chain saturated fatty acid. During the trial the oleic acid (18:1c) content in milk fat significantly increased (21.33% at the start, 24.71% after 2 weeks and 22.51% after 4 weeks on farm B, respectively). The proportion of α-linolenic acid (18:3n-3) increased (P > 0.05) during the experimental period compared to the control period (0.23% at the start, 0.58% after 2 weeks and 0.59% after 4 weeks on farm B). The concentration of long-chain polyunsaturated fatty acids (eicosapentaenoic acid and docosapentaenoic acid) significantly increased in milk fat after 4 weeks on both farms. The n-6/n-3 ratio narrowed from 13.52:1 to 3.83:1 and 3.69:1 on farm B, respectively, but this result was not significant. This study concluded that dietary supplementation of whole linseed decreased the proportion of some SFA and increased the proportion of some MUFA and PUFA.