Plants have to cope with various abiotic stresses including UV-B radiation (280-315 nm). UV-B radiation is perceived by a photoreceptor, triggers morphological responses and primes plant defense mechanisms such as antioxidant levels, photo-repair or accumulation of UV-B screening pigments. As poplar is an important model system for trees, we elucidated the influence of UV-B on overall metabolite patterns in poplar leaves grown under high UV-B radiation. Combining non-targeted metabolomics with gas exchange analysis and confocal microscopy we aimed understanding how UV-B radiation triggers metabolome-wide changes, affects isoprene emission, photosynthetic performance, epidermal light attenuation and finally how isoprene-free poplars adjust their metabolome under UV-B radiation. Exposure to UV-B radiation caused a comprehensive rearrangement of the leaf metabolome. Several hundreds of metabolites were up and down-regulated over various pathways. Our analysis, revealed the up-regulation of flavonoids, anthocyanins and polyphenols and the down-regulation of phenolic precursors in the first 36 hours of UV-B treatment. We also observed a down-regulation of steroids after 12 hours. The accumulation of phenolic compounds leads to a reduced light transmission in UV-B-exposed plants. However, the accumulation of phenolic compounds was reduced in non-isoprene emitting plants suggesting a metabolic- or signaling-based interaction between isoprenoid and phenolic pathways.