In situ measurements of the free energy difference between the open and closed states of ion channels are challenging due to hysteresis effects and inactivation. Exploiting recent developments in statistical physics, we present a general formalism to extract the free energy difference ΔF between the closed and open states of mechanosensitive ion channels from nonequilibrium work distributions associated with the opening and closing of the channels (gating) in response to ramp stimulation protocols recorded in native patches. We show that the work distributions obtained from the gating of MscS channels in E. coli membrane satisfy the strong symmetry relation predicted by the Crooks fluctuation theorem. Our approach enables the determination of ΔF using patch-clamp experiments, which are often inherently restricted to the nonequilibrium regime.