To provide further insights into ruminant lipid digestion and metabolism, and into cis-9,trans-11 18:2 synthesis, 12 growing Engadine lambs grazing either mountain pasture (2,250 m above sea level; n = 6) or lowland pasture (400 m above sea level; n = 6) were studied. Both pastures consisted exclusively of C(3) plants. Before the experiment, all animals grazed a common pasture for 6 wk. Grasses and perirenal adipose tissues of the sheep were analyzed for fatty acids by gas chromatography. Stable C-isotope ratios (δ(13)C values in ‰ vs. the Vienna Pee Dee Belemnite standard) were determined in the composite samples by elemental analysis-isotope ratio mass spectrometry. The δ(13)C of the individual fatty acids were measured by gas chromatography-combustion-isotope ratio mass spectrometry. The δ(13)C value of the entire mountain pasture grass was -27.5‰ (SD 0.31), whereas that of the lowland pasture grass was -30.0‰ (SD 0.07). This difference was reflected in the perirenal adipose tissues of the corresponding sheep (P < 0.05), even though the δ(13)C values were less in the animals than in the grass. The δ(13)C values for cis-9 16:1 and cis-9 18:1 in perirenal fat differed between mountain and lowland lambs (P < 0.05). The 16:0 in the adipose tissue was enriched in (13)C by 5‰ compared with the dietary 16:0, likely as a result of partly endogenous synthesis. The δ(13)C values of cis-9,trans-11 18:2 (cis-9,trans-11 CLA) in the adipose tissue were smaller than those of its dietary precursors, cis-9,cis-12 18:2 and cis-9,cis-12,cis-15 18:3; conversely, the δ(13)C values of trans-11 18:1 were not, suggesting that large proportions of perirenal cis-9,trans-11 18:2 were of endogenous origin and discrimination against (13)C occurred during Δ(9)-desaturation. The same discrimination was indicated by the isotopic shift between 16:0 and cis-9 16:1 in the mountain grazing group. Furthermore, the δ(13)C values of cis-9,trans-11 18:2 were smaller relative to the precursor fatty acids in the mountain lambs compared with the lowland group. This result suggests a reduced extent of biohydrogenation in lambs grazing on mountain grass in comparison with those grazing on lowland grass. This was supported by the smaller cis-9,trans-11 18:2 concentrations in total fatty acids found in the adipose tissues of the lowland lambs (P < 0.001). The results of this study demonstrate that natural differences between δ(13)C values of swards from different pastures and the adipose tissue fatty acids could be used as tracers in studies of lipid metabolism in ruminants.