The factors of variation for vitamin B9 and B12 content in cow's milk have not been studied intensively. Nevertheless, these vitamins are of great nutritional interest and the consumption of a large glass of milk could provides up to 15% of the recommended daily allowances for vitamin B9 and 90% for vitamin B12. The objective of this study was therefore to describe the variability of the levels of vitamins B9 and B12 in 100 bulk milk samples produced under four French major production systems (plain or mountain and forage system based on grass or maize silage). For each production system, milk was sampled and analyzed at five periods over the year 2008. The characteristics of these milks differed according to the production system. The highest levels of vitamin B9 were generally associated with diets based on hay in the winter period and with diets based on pasture during the grazing season. Conversely, the highest levels of vitamin B12 were principally measured in milks produced with diets based on maize silage.