BACKGROUND: The nutritional profile of human milk varies significantly between women, and the impact of maternal diet on these variations is not well understood. OBJECTIVE: We analyzed breast-milk vitamin B-12 concentration and vitamin B-12 supplement use pattern among women who adhered to different dietary patterns: vegan, vegetarian, and nonvegetarian. DESIGN: A total of 74 milk samples, 29 from vegan, 19 from vegetarian, and 26 from nonvegetarian breastfeeding mothers, were analyzed. RESULTS: The prevalences of low vitamin B-12 (<310 pmol/L) were 19.2% for vegans, 18.2% for vegetarians, and 15.4% for nonvegetarians, which was not significant by diet group (P = 1.00). The median (quartile 1, quartile 3) vitamin B-12 values were 558 pmol/L (331, 759 pmol/L) for vegans, 509 pmol/L (368, 765 pmol/L) for vegetarians, and 444 pmol/L (355, 777 pmol/L) for nonvegetarians (P = 0.890). The use of individual vitamin B-12 supplements was higher in vegans (46.2%) than in vegetarians (27.3%) and nonvegetarians (3.9%) (P = 0.001). In linear regression analysis, the use of individual vitamin B-12 supplements was a significant positive predictor of milk vitamin B-12 concentration (β ± SE: 172.9 ± 75.2; standardized β = 0.263; P = 0.024; R2 = 0.069), the use of a multivitamin had a significant negative relation with milk vitamin B-12 concentrations (β ± SE -222.0 ± 98.7; standardized β = -0.258; P = 0.028, R2 = 0.067;), whereas the use of a B-complex vitamin and prenatal vitamin were not predictive of vitamin B-12 milk concentration (P > 0.05). CONCLUSIONS: Almost 20% of our study participants were classified as having low breast-milk vitamin B-12 concentrations (<310 pmol/L), independent of maternal diet pattern. Approximately 85% of participants categorized as having low vitamin B-12 were taking vitamin B-12 supplements at doses in excess of the Recommended Dietary Allowance, which suggests that more research is needed to determine breast-milk adequacy values.