As a result of human nutritional recommendations, experiments have been performed to modify milk fatty acid (FA) composition in order to increase milk content of C18:3 n-3 and to decrease that of both deleterious saturated FA and trans FA. The modifications observed when cow’s diet is supplemented with a source rich in C18:3 n-3, such as linseeds, are largely dependent on rumen biohydrogenation processes. Indeed, milk trans FA and C18:3 n-3 are a consequence of the ruminal biohydrogenation of dietary C18:3 n-3. However, little is known about the effect of increasing amount of extruded linseeds intake by cows on the kinetics of rumen biohydrogenation. This may impact human health because it could affect both the amount of C18:3 n-3 and of trans FA available for milk fat production. Trans FA profile may be important to consider because their side effect on human health may be due to specific trans FA. The aim of this study was thus to assess the effects of increasing intakes of extruded linseeds on kinetics of rumen FA profile, especially trans FA.