Determination of UWL Permeabilities using Stirring Speed Dependence

3 Determination of UWL Permeabilities using Stirring Speed Dependence 

Caco-2 assay permeabilities corrected for the UWL usually include Pu determined as a function of the stirring speed (since the cells are not stable over a wide pH range), as in Eq. (7.57) [511-514,552,578] 

Measurements of Pe in fixed-pH solutions but at various different stirring speeds need to be made. The double-reciprocal analysis, HPe versus 1/v , for Caco-2 permeability measurements in the Transwell (Corning Costar) system produced a linear plot for x- 0.8 [514]. The intercept yields the membrane permeability for the particular pH value in the study the slope determines the k constant. From the analysis of testosterone transport, for the stirring speed of 25 rpm (planar rotating shaker), the thickness of each UWL (assuming symmetric geometry) was calculated to be 465 pm at 150 rpm, haq= 110 pm [514], Karlsson and Artursson [512] found x = 1.0 to best represent their stirring-based analysis of the UWL permeability. 

TABLE 7.15 Intrinsic Permeabilities and Unstirred Water Layer Permeabilities Determined from Iso-pH Dependence of Effective Permeabilities 2% DOPC in Dodecane 

Similar analysis can be applied to side-by-side diffusion cell systems, where stirring is effected by bubbling an 02/C02 gas mixture. For a bubbling rate of 40 mL gas/min, each UWL was estimated to be 282 pm [515]. 

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