New lines of Brassica carinata, which were developed recently as oil crops for bio-fuel production in the dry western Canadian prairie, could also be regarded as a potential new feed source for animals. To our knowledge, there is little research on metabolic characteristics of protein in the new yellow and brown lines of carinata seeds. The objective of this study was to determine the differences in metabolic characteristics of protein from the new lines of carinata seeds in yellow and brown coats for dairy cows compared with commercial canola seeds. The new carinata lines were bred by Agriculture and Agri-Food Canada (AAFC). The experiment of rumen and intestinal digestion was carried out in U of S dairy research facility. Four cannulated lactating dairy cows were used for rumen degradation trial. The nutrient supply to dairy cattle and feed milk value (FMV) were determined based on the DVE/OEB system with data from rumen and intestinal digestion experiments. Statistical analyses were performed using PROC MIXED procedure of SAS 9.3 with significance declared at P < 0.05. The results showed that yellow carinata seed line had higher (P < 0.05) rumen undegraded feed crude protein (RUP) and effective degradability of crude protein (EDCP) compared with canola seeds. From in vitro intestinal digestion results, carinata seeds had higher total digestible protein (TDP) than canola seeds (P < 0.05). Based on the DVE/OEB system, carinata seeds had higher truly absorbed rumen-synthesized microbial protein, total truly digested protein (DVE) in small intestine and feed milk value (FMV) than canola seeds (P < 0.05). Yellow carinata seed line had higher protein supply than the brown seed line. In conclusion, the new lines of carinata seeds could be regarded as a potential feed protein supplement for dairy cows, and yellow carinata seed line had greater protein digestibility than brown carinata seeds.