Modification of a PAMPA model to predict passive gastrointestinal absorption and plasma protein binding.

European journal of pharmaceutical sciences : official journal of the European Federation for Pharmaceutical Sciences (2015-06-30)
Alban Bujard, Hervé Voirol, Pierre-Alain Carrupt, Julie Schappler
RESUMEN

The Parallel Artificial Membrane Permeability Assay (PAMPA) is a well-known high throughput screening (HTS) technique for predicting in vivo passive absorption. In this technique, two compartments are separated by an artificial membrane that mimics passive permeability through biological membranes such as the dermal layer, the gastrointestinal tract (GIT), and the blood brain barrier (BBB). In the present study, a hexadecane artificial membrane (HDM)-PAMPA was used to predict the binding of compounds towards the human plasma using a mixture of human serum albumin (HSA) and alpha-1-acid glycoprotein (AGP). The ratio of HSA and AGP was equivalent to that found in the human plasma for both proteins (∼20:1). A pH gradient (5.0-7.4) was performed to increase the screening capacity and overcome the issue of passive permeability for acidic and amphoteric compounds. With this assay, the prediction of passive GIT absorption was maintained and the compounds were discriminated according to their permeability (on a no-to-high scale). The plasma protein binding (PPB) was estimated via the correlation of the differences between the amount of compound crossing the artificial membrane in assays conducted with and without protein using only a two end-point measurement. The use of a mixture of HSA and AGP to modulate drug permeation was compared to the use of the same concentrations of HSA and AGP used separately. The addition of HSA alone in the acceptor compartment was sufficient for estimating PPB, while it was demonstrated that AGP alone could enable the estimation of AGP binding.

MATERIALES
Referencia del producto
Marca
Descripción del producto

Sigma-Aldrich
Dimethyl sulfoxide, Hybri-Max, sterile-filtered, BioReagent, suitable for hybridoma, ≥99.7%
Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Methanol, HPLC Plus, ≥99.9%, poly-coated bottles
Sigma-Aldrich
Dimethyl sulfoxide, for molecular biology
Sigma-Aldrich
Dimethyl sulfoxide, anhydrous, ≥99.9%
Sigma-Aldrich
Dimethyl sulfoxide, sterile-filtered, BioPerformance Certified, meets EP, USP testing specifications, suitable for hybridoma
Sigma-Aldrich
Hexane, anhydrous, 95%
Sigma-Aldrich
Potassium phosphate monobasic, powder, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99.0%
Sigma-Aldrich
Dimethyl sulfoxide, ≥99.5% (GC), suitable for plant cell culture
Sigma-Aldrich
Sodium hydroxide solution, BioUltra, for molecular biology, 10 M in H2O
Sigma-Aldrich
Sodium hydroxide solution, 1.0 N, BioReagent, suitable for cell culture
Sigma-Aldrich
Hexadecane, ReagentPlus®, 99%
Sigma-Aldrich
Sodium phosphate dibasic, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Potassium phosphate monobasic, ReagentPlus®
Sigma-Aldrich
Sodium phosphate dibasic, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium phosphate dibasic, for molecular biology, ≥98.5% (titration)
Sigma-Aldrich
Sodium phosphate dibasic, BioReagent, suitable for cell culture, suitable for insect cell culture, ≥99.0%
Sigma-Aldrich
Dimethyl sulfoxide, meets EP testing specifications, meets USP testing specifications
Sigma-Aldrich
Acetic acid, natural, ≥99.5%, FG
Sigma-Aldrich
Potassium phosphate monobasic, for molecular biology, ≥98.0%
Sigma-Aldrich
Sodium phosphate dibasic solution, BioUltra, 0.5 M in H2O
Sigma-Aldrich
Dimethyl sulfoxide, BioUltra, for molecular biology, ≥99.5% (GC)
Sigma-Aldrich
Hexadecane, anhydrous, ≥99%
Sigma-Aldrich
Sodium phosphate dibasic, 99.95% trace metals basis
Sigma-Aldrich
Dimethyl sulfoxide, PCR Reagent
Sigma-Aldrich
Potassium phosphate monobasic, BioUltra, for molecular biology, anhydrous, ≥99.5% (T)
Sigma-Aldrich
Acetic acid, ≥99.5%, FCC, FG
Sigma-Aldrich
5α-Androstan-17β-ol-3-one, ≥97.5%
Sigma-Aldrich
Acetic acid, for luminescence, BioUltra, ≥99.5% (GC)
Sigma-Aldrich
Potassium phosphate monobasic, 99.99% trace metals basis