|The oral administration route is a non-invasive, cost-effective route that is associated with high patient compliance, especially among the elderly and children. Excipients are frequently added to the active ingredient to contribute to a pharmaceutical formulation that is stable, easy to formulate and has increased patient compliance. Ideally, excipients should be chemically inert. Surfactants, in particular, should not alter the permeability or transport of the active pharmaceutical ingredient. However, several studies relating to surfactant characterisation have proven that permeability of tissue could be affected by surfactants. The aim of this study was to determine if selected surfactants had altering effects on intestinal drug permeation. For this purpose, membrane permeability of a model compound was evaluated across excised pig intestinal tissue mounted in a Sweetana-Grass diffusion apparatus. The bidirectional transport of Rhodamine 123 (Rho123), a substrate of the P-glycoprotein (P-gp) efflux transporter with fluorescent properties, was measured in the presence of five surfactants and a bile salt, namely Brij® 58, Tween® 20, Span® 20, Cremophor® CO40 and sodium deoxycholate. The surfactants were added at three different concentrations, 0.1%, 0.5% and 1.0% (w/v), to investigate if the addition of these surfactants had any influence on the membrane permeability of Rho123 and if these effects were concentration dependent. The samples were withdrawn over a period of 120 min, from either the apical or the basolateral chambers, and fluorescence was measured with a microplate reader. The results indicated that polyoxyl 20 cetyl ether (Brij® 58) mediated an increase in Rho123 transport in the apical to basolateral direction at concentrations of 0.5% and 1.0% (w/v), and a decrease in the basolateral to apical direction at concentrations of 0.5% and 1.0% (w/v) when compared to the Rho123 control group, indicating the inhibition of P-gp related efflux in a concentration dependent manner. Polyoxyl 40 hydrogenated castor oil (Cremophor® CO40) and Polysorbate 20 (Tween® 20) mediated no mentionable change in Rho123 transport in the apical to basolateral direction, while a decrease in the transport of the marker was seen in the basolateral to apical direction. This could indicate that P-gp related efflux was inhibited. Sorbitan monolaurate (Span® 20) mediated no effect on Rho123 transport in the apical to basolateral direction but mediated a significant decrease in the P-gp mediated efflux of Rho123. Sodium deoxycholate resulted in an increase in Rho123 transport in the apical to basolateral direction in a concentration dependent manner, and a decrease in transport in the basolateral to apical direction, in a concentration dependent manner when compared to the Rho123 control. P-gp was possibly inhibited and consequently efflux was decreased. The trans-epithelial electrical resistance (TEER) values decreased in the presence of all the surfactants in both directions of transport, indicating that the surfactants possibly opened tight-junctions. The results of the study confirmed that excipients, such as surfactants, can and do have altering effects on drug permeability by means of efflux inhibition and possible opening of tight junctions, which may lead to altered bioavailability of the co-administered drug.