An electrochemical method has been developed for the in situ determination of concentration gradients in the electrolyte of sealed Li-ion batteries by measuring the potential difference between microreference electrodes. Formulas relating the concentration gradient and the potential difference between the microreference electrodes were derived from the Nernst-Planck equation. The concentration gradients in Li-ion batteries are theoretically and experimentally proven to be linear at steady state under galvanostatic charging and discharging conditions. Based on this finding, both the diffusion coefficient of the lithium ions in the electrolyte and the diffusion overpotential related to the concentration gradient have been calculated. It was found that the concentration gradient is proportional to the applied current over a wide current range and that the obtained diffusion coefficient of lithium ions is almost constant. For the batteries studied in this work, the diffusion overpotential is already noticeable at 0.30 A and the limiting current is estimated to be 1.1 A, corresponding to a C-rate of 3.7.