Studying microRNA (miRNAs) in certain agri-food products is attractive because (1) they have potential as biomarkers that may allow traceability and authentication of such products; and (2) they may reveal insights into the products' functional potential. The present study evaluated differences in miRNAs levels in fat and cellular fractions of tank milk collected from commercial farms which employ extensive or intensive dairy production systems. We first sequenced miRNAs in three milk samples from each production system, and then validated miRNAs whose levels in the cellular and fat fraction differed significantly between the two production systems. To accomplish this, we used quantitative PCR with both fractions of tank milk samples from another 20 commercial farms. Differences in miRNAs were identified in fat fractions: overall levels of miRNAs, and, specifically, the levels of bta-mir-215, were higher in intensive systems than in extensive systems. Bovine mRNA targets for bta-miR-215 and their pathway analysis were performed. While the causes of these miRNAs differences remain to be elucidated, our results suggest that the type of production system could affect miRNAs levels and potential functionality of agri-food products of animal origin.