The arbuscular mycorrhizal (AM) fungal symbiosis is widely hypothesized to have promoted the evolution of land plants from rootless gametophytes to rooted sporophytes during the mid-Palaeozoic (480-360 Myr, ago), at a time coincident with a 90% fall in the atmospheric CO(2) concentration (CO(2)(a)). Here we show using standardized dual isotopic tracers ((14)C and (33)P) that AM symbiosis efficiency (defined as plant P gain per unit of C invested into fungi) of liverwort gametophytes declines, but increases in the sporophytes of vascular plants (ferns and angiosperms), at 440 p.p.m. compared with 1,500 p.p.m. CO(2)(a). These contrasting responses are associated with larger AM hyphal networks, and structural advances in vascular plant water-conducting systems, promoting P transport that enhances AM efficiency at 440 p.p.m. CO(2)(a). Our results suggest that non-vascular land plants not only faced intense competition for light, as vascular land floras grew taller in the Palaeozoic, but also markedly reduced efficiency and total capture of P as CO(2)(a) fell.