Karst spring measurements assess biogeochemical processes occurring within groundwater contributing areas to springs (springsheds) but can only provide aggregated information. To better understand spatially distributed processes that comprise these aggregated measures, we investigated aquifer denitrification evidence in groundwater wells (n = 16) distributed throughout a springshed in the Upper Floridan aquifer in northern Florida. Aquifer geochemistry, nitrate isotopes, and dissolved gases were compared against similar measurements at the spring outlet to evaluate spatial heterogeneity of denitrification evidence in relation to land surface–aquifer connectivity. Sample locations spanned spatial variation in recharge processes (i.e., diffuse vs. focused recharge) and proximity to sources of denitrification reactants (e.g., wetlands). Although no distinct spatial pattern in denitrification was uncovered, excess dissolved N₂ gas measurements were only above detection in the unconfined springshed, with some evidence of a wetland proximity effect. Measured oxidation–reduction potential and dissolved oxygen poorly predicted denitrification, indicating that measured denitrification may be occurring upgradient from sampled wells. Despite dramatic spatial chemical heterogeneity across wells, mean values for recharge nitrate concentrations (0.02 to 5.56 mg N L⁻¹) and excess N₂ from aquifer denitrification (below detection to 1.37 mg N L⁻¹) corresponded reasonably with mean spring outlet measurements for initial nitrate (0.78 to 1.36 mg N L⁻¹) and excess N₂ (0.15 to 1.04 mg N L⁻¹). Congruence between groundwater and spring measurements indicates that combining sampling at the spring outlet and across the springshed is useful for understanding spatial aquifer denitrification. However, this approach would be improved with a high‐density sampling network with transects of wells along distinct groundwater flow paths.