This review analyzes the potential role and long-term effects of field perennial polycultures (mixtures) in agricultural systems, with the aim of reducing the trade-offs between provisioning and regulating ecosystem services. First, crop rotations are identified as a suitable tool for the assessment of the long-term effects of perennial polycultures on ecosystem services, which are not visible at the single-crop level. Second, the ability of perennial polycultures to support ecosystem services when used in crop rotations is quantified through eight agricultural ecosystem services. Legume-grass mixtures and wildflower mixtures are used as examples of perennial polycultures, and compared with silage maize as a typical crop for biomass production. Perennial polycultures enhance soil fertility, soil protection, climate regulation, pollination, pest and weed control, and landscape aesthetics compared with maize. They also score lower for biomass production compared with maize, which confirms the trade-off between provisioning and regulating ecosystem services. However, the additional positive factors provided by perennial polycultures, such as reduced costs for mineral fertilizer, pesticides, and soil tillage, and a significant preceding crop effect that increases the yields of subsequent crops, should be taken into account. However, a full assessment of agricultural ecosystem services requires a more holistic analysis that is beyond the capabilities of current frameworks.