Against a background of concentrated production chains and profound changes in the agri-food system, Brazilian agricultural cooperatives are challenged to remain competitive to withstand large multinational companies. This paper proposed to investigate nuances in the behavior of these cooperative organizations due to the changes in their operating environment. It does so by analyzing alterations in efficiency and total factor productivity of a sample of Brazilian agricultural cooperatives, classified according to size, from 2006 to 2010. For this, non-parametric models of Data Envelopment Analysis (DEA) and the Malmquist Index were used. Through the DEA approach, it was seen that larger cooperatives faced decreasing efficiency, while smaller ones experienced the opposite. Moreover, as demonstrated by the Malmquist Indices, the cooperatives, on average, presented negative technological variations, and smaller cooperatives underwent positive changes in technical efficiency. The results suggest directions which public policies could take to strengthen Brazilian agricultural cooperatives in the face of new challenges.