Diffusion of carbon dioxide through lipid bilayer membranes

Diffusion of ¹⁴C-labeled CO₂ was measured through lipid bilayer membranes composed of egg lecithin and cholesterol (1: 1 mol ratio) dissolved in n-decane. The results indicate that CO₂, but not HCO₃⁻, crosses the membrane and that different steps in the transport process are rate limiting under different conditions. In one series of experiments we studied one-way fluxes between identical solutions at constant pCO₂ but differing [HCO₃⁻] and pH. In the absence of carbonic anhydrase (CA) the diffusion of CO₂ through the aqueous unstirred layers is rate limiting because the uncatalyzed hydration-dehydration of CO₂ is too slow to permit the high [HCO₃⁻] to facilitate tracer diffusion through the unstirred layers. Addition of CA (ca. 1 mg/ml) to both bathing solutions causes a 10-100-fold stimulation of the CO₂ flux, which is proportional to [HCO₃⁻] over the pH range 78. In the presence of CA the hydration-dehydration reaction is so fast that CO₂ transport across the entire system is rate limited by diffusion of HC03 through the unstirred layers. However, in the presence of CA when the ratio [HCO₃⁻ + CO₃⁼]:[CO₂] > 1,000 (pH 9-10), the CO₂ flux reaches a maximum value. Under these conditions the diffusion of CO₂ through the membrane becomes rate limiting, which allows us to estimate a permeability coefficient of the membrane to CO₂ of 0.35 cm s^-1. In a second series of experiments we studied the effects of CA and buffer concentration on the net flux of CO₂. CA stimulates the net CO₂ flux in well buffered, but not in unbuffered, solutions. The buffer provides a proton source on the upstream side of the membrane and proton sink on the downstream side, thus allowing HCO₃⁻ to facilitate the net transport of CO₂ through the unstirred layers.