BACKGROUND: Common ragweed has been spreading as a neophyte in Europe. Elevated CO(2) levels, a hallmark of global climate change, have been shown to increase ragweed pollen production, but their effects on pollen allergenicity remain to be elucidated. METHODS: Ragweed was grown in climate-controlled chambers under normal (380 ppm, control) or elevated (700 ppm, based on RCP4.5 scenario) CO(2) levels. Aqueous pollen extracts (RWE) from control- or CO(2) -pollen were administered in vivo in a mouse model for allergic disease (daily for 3-11 days, n = 5) and employed in human in vitro systems of nasal epithelial cells (HNECs), monocyte-derived dendritic cells (DCs), and HNEC-DC co-cultures. Additionally, adjuvant factors and metabolites in control- and CO(2) -RWE were investigated using ELISA and untargeted metabolomics. RESULTS: In vivo, CO(2) -RWE induced stronger allergic lung inflammation compared to control-RWE, as indicated by lung inflammatory cell infiltrate and mediators, mucus hypersecretion, and serum total IgE. In vitro, HNECs stimulated with RWE increased indistinctively the production of pro-inflammatory cytokines (IL-8, IL-1β, and IL-6). In contrast, supernatants from CO(2) -RWE-stimulated HNECs, compared to control-RWE-stimulated HNECS, significantly increased TNF and decreased IL-10 production in DCs. Comparable results were obtained by stimulating DCs directly with RWEs. The metabolome analysis revealed differential expression of secondary plant metabolites in control- vs CO(2) -RWE. Mixes of these metabolites elicited similar responses in DCs as compared to respective RWEs. CONCLUSION: Our results indicate that elevated ambient CO(2) levels elicit a stronger RWE-induced allergic response in vivo and in vitro and that RWE increased allergenicity depends on the interplay of multiple metabolites.