Many studies have found that future predicted CO2 levels (< 800 ppm) can increase plant mass but dilute N content in leaves, impacting antiherbivore compounds. Nitrogen-fixing plants may balance leaf C:N ratio under elevated CO2, counteracting this dilution effect. The aim of this study was to look at how nitrogen-fixing plants grow and respond to herbivore damage at different CO2 levels. Alnus incana ssp. rugosa was grown at 400, 800, or 1600 ppm CO2 in soil collected from the field, inoculated with Frankia and exposed to herbivores (Orgyia leucostigma). Elevated CO2 increased nodulated plant biomass and stimulated the nitrogen fixation rate in the early growth stage. However, nitrogen-fixing plants were not able to balance the C:N ratio under elevated CO2 after grown 19 weeks. When plant were grown at 400 and 1600 ppm CO2, herbivores preferred to feed on leaves of nodulated plants. At 800 ppm CO2, nodulated plants accumulated more total phenolic compounds in response to herbivore damage than plants in the non-Frankia and non-herbivore treatments. Our results suggest that plant leaf defence, not leaf nutritional content, is the dominant driver of herbivory and nitrogen fixing plants have limited ability to balance C:N ratios at elevated CO2 in natural soil.