Abstract In alkaline freshwater systems, the apparent absence of carbon limitation to gross primary production (GPP) at low CO2 concentrations suggests that bicarbonates can support GPP. However, the contribution of bicarbonates to GPP has never been quantified in lakes along the seasons. To detect the origin of the inorganic carbon maintaining GPP, we analyze the daily stoichiometric ratios of CO2–O2 and alkalinity–O2 in a deep hardwater lake. Results show that aquatic primary production withdraws bicarbonate from the alkalinity pool for two-thirds of the year. Alkalinity rather than CO2 is the dominant inorganic carbon source for GPP throughout the stratified period in both the littoral and pelagic environments. This study sheds light on the neglected role of alkalinity in the freshwater carbon cycle throughout an annual cycle.