Ex vivo permeability study of poorly soluble drugs across gastrointestinal membranes: acceptor compartment media composition

Introduction
In ex vivo drug permeability studies using permeation systems, such as the
Ussing chamber, excised tissue (usually small intestine from rodents, pigs, or humans) is mounted between the donor (mucosal) and acceptor (serosal) compartments, which are filled with appropriate buffer media. The acceptor media compartment represents the blood and should provide the highest transmembrane gradients, in order to establish the sink conditions, thus, mimicking drug distribution in vivo. In laboratory settings, achieving sink conditions for permeated poorly soluble drugs in the relatively low and/or limited media volumes of the permeability system's acceptor compartment is crucial. Adding solubility-enhancing agents to the acceptor media addresses this challenge. The aim of this work is to compile and critically evaluate the compositions of the acceptor compartment media used in ex vivo permeability studies for poorly soluble drugs, in relation to their physicochemical, biopharmaceutical, and pharmacokinetic properties. Media compositions in the permeability system’s acceptor compartments has not been reviewed in detail before.
Materials and Methods
A literature search was conducted using the Scopus database, focusing on contributions from leading pharmacy schools and research groups in the field of drug permeability studies. The keywords used for the search were “ex vivo,” “permeability,” and “Ussing.”

Results and Discussion
The most commonly reported acceptor media were Krebs Ringer bicarbonate buffer (KBR), Hank's balanced salt solution (HBSS), and phosphate-buffered saline (PBS). However, the components and their compositions varied across studies. In most cases, no additives were used to establish sink conditions. Additives not only provide sink conditions but also reduce non-specific binding of drug molecules to the walls of the permeability system and the permeable membrane itself. Only nine studies reported the use of solubility-enhancing additives in acceptor media for ex vivo studies. Bovine serum albumin (BSA, 0.1-4.5%) was used in acceptor media where poorly soluble drugs exhibited high plasma protein binding (e.g., aprepitant 99%, diazepam 98%, ibuprofen 99%) or binding to tissue proteins (e.g., digoxin). The ionic surfactant sodium lauryl sulphate (SLS), with a high hydrophile-lipophile balance (HLB40), was added in a study with the lipophilic drug exemestane (LogP 3.5). The non-ionic surfactant Poloxamer 407 (HLB 22) was used to establish acceptor compartment sink
conditions in studies with lipophilic dapivirine (LogP 5.6). Tween 80 (HLB 15) was applied in studies with lipophilic fenretinide (LogP 6.3) and docetaxel (LogP 4.1), as well as the hydrophilic drug cisplatin (LogP -2.3) (Figure 2). The use of surfactants and BSA impacts the analysis of drug concentrations, requiring additional purification steps. The potential impact of surfactant addition on drug permeation in the serosal to mucosal direction has not been reported. An accompanying problem, foaming due to continuous oxygenation in ex vivo systems, was reported for BSA and may also occur with surfactants. Anti-foaming agents helped resolve this issue, although their impact on permeability study results was not discussed. In addition to BSA, SLS, and Tween 80, in vitro permeability studies have reported the use of the following additives to establish sink conditions: d-α-tocopheryl polyethylene glycol succinate (TPGS), caprylocaproyl polyoxyl-8 glycerides, β-cyclodextrin, and sodium taurocholate. These additives could also be used in ex vivo permeability studies for the same purpose, provided they are used at appropriate and safe concentrations for the cells of the excised intestinal tissue.

Conclusion
The compositions of acceptor compartment media vary among laboratories, with Krebs Ringer Bicarbonate buffer (KBR) and Hank's balanced salt solution (HBSS) being the most common, albeit with slight composition variations. Despite the clear necessity, the addition of solubility-enhancing additives to increase sink conditions was rarely reported in the published literature. However, in ex vivo studies where these additives were applied, they included permeability assessments of aprepitant, cisplatin, dapivirine, diazepam, digoxin, docetaxel, exemestane, fenretinide, and ibuprofen. In contrast, in vitro studies reported the use of these additives more frequently, primarily including
BSA, TPGS, Tween 80, and cyclodextrins. The importance of using solubility-enhancing additives for poorly soluble drugs in acceptor media of permeability systems is underscored by this literature study.