Direct quantification of fossil fuel CO₂ (CO₂ff) in atmospheric samples can be used to examine several carbon cycle and air quality questions. We collected in situ CO₂, CO, and CH₄ measurements and flask samples in the boundary layer and free troposphere over Sacramento, California, USA, during two aircraft flights over and downwind of this urban area during spring of 2009. The flask samples were analyzed for Δ¹⁴CO₂ and CO₂ to determine the recently added CO₂ff mole fraction. A suite of greenhouse and other trace gases, including hydrocarbons and halocarbons, were measured in the same samples. Strong correlations were observed between CO₂ff and numerous trace gases associated with urban emissions. From these correlations we estimate emission ratios between CO₂ff and these species, and compare these with bottom-up inventory-derived estimates. Recent county level inventory estimates for carbon monoxide (CO) and benzene from the California Air Resources Board CEPAM database are in good agreement with our measured emission ratios, whereas older emissions inventories appear to overestimate emissions of these gases by a factor of two. For most other trace species, there are substantial differences (200–500%) between our measured emission ratios and those derived from available emission inventories. For the first flight, we combine in situ CO measurements with the measured CO:CO₂ff emission ratio of 14 ± 2 ppbCO/ppmCO₂ to derive an estimate of CO₂ff mole fraction throughout this flight, and also estimate the biospheric CO₂ mixing ratio (CO₂bio) from the difference of total and fossil CO₂. The resulting CO₂bio varies dramatically from up to 8 ± 2 ppm in the urban plume to −6 ± 1 ppm in the surrounding boundary layer air. Finally, we use the in situ estimates of CO₂ff mole fraction to infer total fossil fuel CO₂ emissions from the Sacramento region, using a mass balance approach. The resulting emissions are uncertain to within a factor of two due to uncertainties in wind speed and boundary layer height. Nevertheless, this first attempt to estimate urban-scale CO₂ff from atmospheric radiocarbon measurements shows that CO₂ff can be used to verify and improve emission inventories for many poorly known anthropogenic species, separate biospheric CO₂, and indicates the potential to constrain CO₂ff emissions if transport uncertainties are reduced.