title: In vitro investigation on the impact of the surface-active excipients Cremophor EL, Tween 80 and Solutol HS 15 on the metabolism of midazolam.
abstract: The impact of the surface-active formulation ingredients Cremophor EL, Tween 80 and Solutol HS 15 on the intrinsic clearance (Clint) of midazolam (MDZ) was investigated in rat hepatocytes and microsomes. In rat hepatocytes with 0.003%, 0.03% and 0.3% (w/v) Solutol HS 15 already present in the incubation medium, the Clint was significantly reduced in a dose-dependent manner by about 25%, 30% and 50%, respectively. In the presence of Cremophor EL and Tween 80 a significant reduction in Clint by about 30% and 25%, respectively, was observed at 0.03% surfactant concentration. At 0.3% of Cremophor EL and Tween 80, Clint was reduced by about 50% and 20%, respectively. A reduction in Clint was also observed in experiments with rat liver microsomes. At surfactant concentrations up to 0.03%, cytotoxicity assays (lactate dehydrogenase release, adenosine triphosphate content) as well as light microscope investigations did not reveal any cytotoxic impact of the surfactants on the hepatocyte monolayer. A potential interaction of the surfactants with biological membranes was determined using phosphatidylcholine-cholesterol liposomes loaded with self-quenching concentrations of carboxyfluorescein. No marked release of carboxyfluorescein from the liposomes (that would be an indication for a surfactant-dependent disruption of membrane integrity) was observed up to concentrations of 0.03% of the different surfactants. It is concluded that cytochrome P450 3A mediated metabolism of MDZ seems to be prevented by all surfactants at concentrations above 0.03%. In our experiments the surfactants did not show toxic effects at concentrations that resulted in a decreased Clint of MDZ. Thus, a direct inhibition of the metabolizing enzymes, a molecular interaction with the microsomes as well as an alteration of membrane properties that did not yet result in a release of LDH have to be taken into consideration as reasons for the observed changes in the metabolism of MDZ.
authors.affiliation: Discovery DMPK, F. Hoffmann-La Roche Ltd, Grenzacher Strasse 124, Basel, Switzerland.
authors.forename: Roberto C
authors.initials: RC
authors.lastname: Bravo González
authors.forename: Jörg
authors.initials: J
authors.lastname: Huwyler
authors.forename: Franziska
authors.initials: F
authors.lastname: Boess
authors.forename: Isabelle
authors.initials: I
authors.lastname: Walter
authors.forename: Beate
authors.initials: B
authors.lastname: Bittner
mesh_terms: D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003470:Culture Media / Q000737:chemistry; D051544:Cytochrome P-450 CYP3A; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004347:Drug Interactions; D005079:Excipients / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D005990:Glycerol / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism; D008874:Midazolam / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011136:Polysorbates / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013229:Stearic Acids / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D013501:Surface-Active Agents / Q000009:adverse effects* / Q000592:standards; D013678:Technology, Pharmaceutical / Q000379:methods / Q000639:trends
chemical_list: D003470:Culture Media; D005079:Excipients; D008565:Membrane Proteins; D011136:Polysorbates; D013229:Stearic Acids; D013501:Surface-Active Agents; D011092:Polyethylene Glycols; C067028:Solutol HS 15; C000515:cremophor EL; D001189:Aryl Hydrocarbon Hydroxylases; C465260:Cyp3a2 protein, rat; D051544:Cytochrome P-450 CYP3A; D005990:Glycerol; D008874:Midazolam