| Assay Detail |
PAMPA was used to determine the passive permeability of these compounds with modifications and calculations as described in Naylor et al. and here. Briefly, pure compound stocks (150 µL, 1 µM, 5% DMSO in phosphate-buffer saline (PBS) pH 7.4) were added to a 96-well donor plate with 0.45 µm hydrophobic Immobilon-P membranes (Millipore MAIPNTR10) pre-soaked with a 1% solution of lecithin in n-dodecane. The donor plate was loaded in to a 96-well Teflon acceptor plate (Millipore MSSACCEPTOR) with 300 µL of PBS with 5% DMSO. After a ~15 h, the acceptor and donor plates were separated, and 50 µL was collect from each well. Samples were diluted with 50 µL methanol and analyzed by LC/MS as described above. Samples were run in triplicate with 1 µM propranolol as a control. Exact time in seconds was used in the calculations. Error was calculated as standard deviation.
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