ABSTRACT
PURPOSE: The aims of the study are to develop and evaluate an in vitro rat intestine segmental perfusion model for the prediction of the oral fraction absorbed of compounds and to assess the ability of the model to study intestinal metabolism. METHODS: The system consisted of a perfusion cell with a rat intestinal segment and three perfusion circulations (donor, receiver, and rinsing circulation). Lucifer yellow (LY) was applied as internal standard together with test compounds in the donor circulation. To validate the model, the permeability of eight noncongeneric passively absorbed drugs was determined. Intestinal N-demethylation of verapamil into norverapamil was followed in the donor and receiver circulations by high-performance liquid chromatography analysis. RESULTS: The in vitro model allowed ranking of the tested compounds according to their in vivo absorption potential. The Spearman's correlation coefficient between the oral fraction absorbed in humans and the ratio of permeation coefficient of test compound to the permeation coefficient of LY within the same experiment was 0.98 (P < 0.01). Moreover, intestinal N-demethylation of verapamil, its permeation, and the permeation of its metabolite norverapamil could be assessed in parallel. CONCLUSIONS: Up to six permeation kinetics can be obtained per rat, and the method has shown to be a valuable tool to estimate human oral absorption.
Subject(s)
Intestinal Absorption , Jejunum/metabolism , Perfusion/methods , Pharmaceutical Preparations/metabolism , Animals , Antipyrine/metabolism , Dealkylation , In Vitro Techniques , Male , Models, Animal , Naproxen/metabolism , Perfusion/instrumentation , Permeability , Rats , Rats, Sprague-Dawley , Testosterone/metabolism , Verapamil/analogs & derivatives , Verapamil/metabolismABSTRACT
The aim of our study was to develop a magnetic resonance (MR)-compatible in vitro model containing freshly isolated rat hepatocytes to study the transport of hepatobiliary contrast agents (CA) by MR imaging (MRI). We set up a perfusion system including a perfusion circuit, a heating device, an oxygenator, and a hollow fiber bioreactor (HFB). The role of the porosity and surface of the hollow fiber (HF) as well as the perfusate flow rate applied on the diffusion of CAs and O2 was determined. Hepatocytes were isolated and injected in the extracapillary space of the HFB (4 x 10(7) cells/mL). The hepatocyte HFB was perfused with an extracellular CA, gadopentetate dimeglumine (Gd-DTPA), and gadobenate dimeglumine (Gd-BOPTA), which also enters into hepatocytes. The HFB was imaged in the MR room using a dynamic T1-weighed sequence. No adsorption of CAs was detected in the perfusion system without hepatocytes. The use of a membrane with a high porosity (0.5 microm) and surface (420 cm2), and a high flow rate perfusion (100 mL/min) resulted in a rapid filling of the HFB with CAs. The cellular viability of hepatocytes in the HFB was greater than 85% and the O2 consumption was maintained over the experimental period. The kinetics of MR signal intensity (SI) clearly showed the different behavior of Gd-BOPTA that enters into hepatocytes and Gd-DTPA that remains extracellular. Thus, these results show that our newly developed in vitro model is an interesting tool to investigate the transport kinetics of hepatobiliary CAs by measuring the MR SI over time.
Subject(s)
Bioreactors , Cell Culture Techniques/instrumentation , Contrast Media/pharmacokinetics , Hepatocytes/cytology , Hepatocytes/metabolism , Magnetic Resonance Imaging/methods , Meglumine/analogs & derivatives , Meglumine/pharmacokinetics , Organometallic Compounds/pharmacokinetics , Animals , Cell Culture Techniques/methods , Cell Survival , Cells, Cultured , Equipment Design , Equipment Failure Analysis , Male , Membranes, Artificial , Rats , Rats, Sprague-DawleyABSTRACT
Hepatocytes carry out many vital biological functions, such as synthetic and catabolic reactions, detoxification and excretion. Due to their ability to restore a tissue-like environment, hollow-fibre bioreactors (HFBs) show great potential among the different systems used to culture hepatocytes. Several designs of HFBs have been proposed in which hepatocytes or hepatocyte-derived cell lines can be cultured in suspensions or on a solid support. Currently the major use of hepatocyte HFBs is as bioartificial livers to sustain patients suffering from acute liver failure, but they can also be used to synthesize cell products and as cellular models for drug metabolism and transport studies. Here, we present an overview of the set-up of hepatocyte HFBs and aim to provide potential users with the basic knowledge necessary to develop their own system. First, general information on HFBs is given, including basic principles, transport phenomena, designs and cell culture conditions. The importance of the tests necessary to assess the performance of the HFBs, i.e. the viability and functionality of hepatocytes, is underlined. Special attention is paid to drug metabolism studies and to adequate analytical methods. Finally, the potential uses of hepatocyte HFBs are described.
Subject(s)
Bioreactors , Biotechnology/methods , Cell Culture Techniques/methods , Hepatocytes/cytology , Animals , Cell Culture Techniques/instrumentation , Cells, Cultured , Humans , Reproducibility of ResultsABSTRACT
Eye drops are multiple dosage forms protected against microbial contamination by means of preservatives. However, the ocular tolerance of these chemicals can vary and this may result in adverse toxic or allergic reactions. This overview presents the pharmacopoeial requirements for the preservation of eye drops, the factors affecting ocular tolerance as well as the adverse external ocular effects induced by preservatives. The alternatives to the use of preservatives are also discussed, including the recent progress in eye drops packaging.
