Calcium salts (CS) of fatty acids (FAs) are involved as fat sources in dairy nutrition. In this supplement dissociation of FAs and calcium is affected by the nature of fatty acids (saturated or unsaturated), so this process led to the release oil in the whole gastrointestinal tract. This study was designed to evaluate the oil releasing extent (ORE) from CS of FAs in a simulated culture of the gastrointestinal tract. Rumen fluid was collected from fistulated cows 4 hours after feeding and transferred to the laboratory. A simulation of the digestion environment at three sites (rumen, abomasums, and small intestine) pH (6.4, 1.2, 6.8) was used to incubate the rumen fluid. Treatments are represented by 1) CS of fish oil (CFO), 2) CS of flaxseed oil (CFL), 3) CS of herbal n-6 and n-3 (CHO) and 4) CS of fish and herbal n-3 (CFH). The ORE in the rumen was affected by treatments, with CS of flaxseed having the greatest rate of release (P<0.01). Differences between treatments in the abomasum were nonsignificant (P=0.1), however, the greatest ORE was in this part in comparison to other sections of the tract. The treatments did affect ORE in the small intestine (P<0.01). According to our results, CS of FAs supplied less than 200 mg/g ORE in the rumen which is acceptable and could be used with minimum adverse effects on rumen function. تفاصيل المقالة