Atmospheric CO2 concentration is hypothesized to influence vegetation distribution via tree–grass competition, with higher CO2 concentrations favouring trees. The stable carbon isotope (δ13C) signature of vegetation is influenced by the relative importance of C4 plants (including most tropical grasses) and C3 plants (including nearly all trees), and the degree of stomatal closure – a response to aridity – in C3 plants. Compound-specific δ13C analyses of leaf-wax biomarkers in sediment cores of an offshore South Atlantic transect are used here as a record of vegetation changes in subequatorial Africa. These data suggest a large increase in C3 relative to C4 plant dominance after the Last Glacial Maximum. Using a process-based biogeography model that explicitly simulates 13C discrimination, it is shown that precipitation and temperature changes cannot explain the observed shift in δ13C values. The physiological effect of increasing CO2 concentration is decisive, altering the C3/C4 balance and bringing the simulated and observed δ13C values into line. It is concluded that CO2 concentration itself was a key agent of vegetation change in tropical southern Africa during the last glacial–interglacial transition. Two additional inferences follow. First, long-term variations in terrestrial δ13Cvalues are not simply a proxy for regional rainfall, as has sometimes been assumed. Although precipitation and temperature changes have had major effects on vegetation in many regions of the world during the period between the Last Glacial Maximum and recent times, CO2 effects must also be taken into account, especially when reconstructing changes in climate between glacial and interglacial states. Second, rising CO2 concentration today is likely to be influencing tree–grass competition in a similar way, and thus contributing to the "woody thickening" observed in savannas worldwide. This second inference points to the importance of experiments to determine how vegetation composition in savannas is likely to be influenced by the continuing rise of CO2 concentration.