Biotransformation of diazepam in a clinically relevant flat membrane bioreactor model using primary porcine hepatocytes

Abstract

In vitro biotransformation of drug using commercial culture medium with serum may not be the ideal culture medium for clinical application in extracorporeal bioartificial liver support (BAL) systems. In these systems, patient's blood or plasma is plumbed to primary hepatocytes within a seeded bioreactor, creating interaction between plasma and seeded hepatocytes. To address this situation, we investigated the biotransformation potential of diazepam in primary porcine hepatocytes with a flat membrane bioreactor (FMB); we used human plasma exposure and serum-free media in organotypical double gel culture model for long-term culture. We investigated diazepam clearance and all major metabolites of diazepam, such as oxazepam, temazepam, and desmethyldiazepam, in conventional single gel and organotypical sandwich models and compared them to the FMB model. Diazepam elimination was higher in double gel cultures with exposure to both SF 3 medium conditions and plasma, when compared to the single gel model in a Petri dish. It was observed that in the FMB, diazepam elimination was stable at about 3 pg/h/cell in plasma and SF 3 exposure. Oxazepam synthesis in the bioreactor was approximately one quarter less than in the Petri dish, but there were no differences between N-desmethyldiazepam and temazepam synthesis in double gel culture. In the flat membrane bioreactor, there was no decrease in the biotransformation of diazepam in plasma exposure compared with the control group. Our results suggest that this plasma exposure bioreactor may offer a useful approach in clinical use of extracorporeal BAL, as well as for drug metabolite investigation into toxicological research.