Sustainable production and biodiversity conservation can be mutually supportive in providing multiple ecosystem services to farmers and society. This study aimed to determine the contribution of agroforestry systems, as tested by family farmers in the Brazilian Rainforest region since 1993, to tree biodiversity and evaluated farmers’ criteria for tree species selection. In addition, long-term effects on microclimatic temperature conditions for coffee production and chemical and biological soil characteristics at the field scale were compared to full-sun coffee systems. A floristic inventory of 8 agroforests and 4 reference forest sites identified 231 tree species in total. Seventy-eight percent of the tree species found in agroforests were native. The variation in species composition among agroforests contributed to a greater γ-diversity than α-diversity. Monthly average maximum temperatures were approximately 6 °C higher in full-sun coffee than in agroforests and forests. Total soil organic C, N mineralization and soil microbial activity were higher in forests than in coffee systems, whereas the chemical and biological soil quality in agroforests did not differ significantly from full-sun coffee after 13 years. Given its contribution to the conservation of biodiversity and its capacity to adapt coffee production to future climate change, coffee agroforestry offers a promising strategy for the area.