Objective-Estimate the transmucosal CO2 and O2 rate-constants for adult middle ears (MEs).Methods-Ten adults with healthy MEs had a unilateral myringotomy. A custom-fitted acrylic mold with a valved line to a Mass Spectrometer (MS) and central tube coupled to a 3-way valve and connected to a pressure transducer (the probe) was sealed with adhesive glue within the ipsilateral ear-canal. A second 3-way valve was attached to the probe valve, a flow-regulated tank gas source and paired syringes. Volumes of the ME and probe were measured. On sequential days, the probe+ME was washed for 15-minutes with 6% O2, Balance N2 and 25% O2, 6% CO2, Balance N2 to create transmucosal CO2 and O2 gradients, respectively. After washing, the probe +ME was isolated from the gas source, and baseline and 10-minute gas samples were obtained for MS analysis of gas partial-pressures. The rates of change in ME CO2 and O2 pressures were divided by their established transmucosal gradients to yield CO2 and O2 rate-constants.Results-The average (±std) transmucosal CO2 and O2 rate-constants were 0.062±0.034 (N=10, range: 0.032-0.119) and 0.011±0.009 (N=8, range: 0.002-0.032) mmHg/min/mmHg, respectively. The average half-life for the CO2 and O2 gradient was 11.1 and 61.6 minutes. The average CO2:O2 rate-constant ratio was 8.1±4.0 (N=8, range 3.6-14.6).Conclusions-For adult human MEs, transmucosal CO2 exchange is rapid and much faster than transmucosal O2 exchange. The estimated CO2/O2 rate-constant ratio for the human ME is not consistent with that predicted for diffusion-limited gas exchange across a water-based barrier.