Purpose <p>Modern agriculture faces the dual challenge of sustainably increasing food production while mitigating the environmental impacts of intensive monocultures. Mixed cropping, which is the cultivation of multiple species or varieties, may provide ecological benefits that address productivity and environmental sustainability challenges. However, evaluating multifunctionality in conventional agricultural field experiments is costly and labour-intensive. Furthermore, small sample sizes and high field heterogeneity often limit the ability to detect statistically significant mixed cropping effects.</p> Methods <p>This study aims to introduce and validate a high-throughput field phenotyping (HTP) framework that integrates aerial imagery obtained from unmanned aerial vehicles (UAVs) to efficiently assess the multifunctionality of mixed cropping systems. We conducted a field experiment comparing monocultures of oat, rye, and barley; intraspecific mixed cropping combining three oat varieties; and interspecific mixed cropping combining oat, rye, and barley. Using UAV-derived data across the entire field, including vegetation cover, plant height, and the normalised difference vegetation index, we evaluated five multifunctionalities (biomass production, biomass variability, early canopy closure, lodging resistance, and lodging resilience).</p> Results <p>This framework revealed that on average, mixedcropping outperforms monocultures in biomass production, biomass variability, early canopy closure, and lodging resilience, althoughmonocultures showed higher lodging resistance compared to interspecific mixtures.</p> Conclusion <p>The proposed UAV-based HTP approachenables cost-effective, robust, and scalable evaluation of mixed cropping systems, facilitating their optimisation for multifunctionality andcontributing to the advancement of sustainable agriculture.</p> Graphical abstract <p></p>