Animal studies quantitative drug metabolism

Cryopreserved hepatocytes in suspension were successfully applied in short-term metabolism studies and as metabolizing system in mutagenicity assays (Hengstler 2000), providing qualitative metabolic information and quantitative pharmacokinetic parameters from key animal species and human at the early stage of drug discovery and drug development. 

Experts in drug metabolism carry out time-course studies with radioactive samples. They give a variety of doses and, at intervals, measure plasma radioactivity rather than therapeutic effect. Sensitive instruments aUow for quantitative measurements for example, of the time and concentration at which radioactivity reaches a maximum. To lay a basis for seiecting the animal species used for long-term toxicology studies, these pilot probes of duration may employ several mammalian species. Different species metabolize a single drug dissimilarly, so a suitable animal species handles it as the human one does. 

The FDA suggests that the metabolic profile of a drug be identified during the drug development process. In vitro studies utilise liver microsomes, liver slices or hepatocytes from animals and humans before initiation of clinical trials. Results from in vivo metabolism studies should be available early in the drug development process since they confirm results from in vitro studies or reveal quantitative or qualitative differences in metabolism across species, which would cause a safety concern. Human in vivo metabolism is carried out at a later stage. 

ADME studies provide information on absorption, distribution, metabolism, and excretion for the compound of interest in animals and humans. In drug development, these studies are performed with either C-14- or tritium-labeled material to provide detailed quantitative information on the circulating metabolites, the extent of metabolism and routes of excretion for drug and its metabolites. Readers are encouraged to refer Chapter 18 of this book for more detailed discussion on ADME study design and data presentation. 

Liquid chromatography is routinely used for the quantitative analysis of a drug and its metabolites in radiolabeled in vitro metabolism experiments and in humans and animal ADME studies. Online radio-flow detection (RFD) and offline microplate scintillation counting (MSC) are the most commonly used radiochromatographic techniques in radiolabeled metabolite profiling and quantification (Boernsen et al., 2000 Nassar et ah, 2003 Bruin et al., 2006). LC/RFD is compatible with ESI mass spectrometry and provides high analytical speed and excellent separation resolution (Athersuch et al., 2008). 

Vogel et al., 2004). In biochemistry and drug development, this sensitivity is applied to increasing precision, to lowering radiactive doses presented to human volunteers or animal subjects, and to isolating ever more specific metabolic products to obtain more quantitative detail in absorption, distribution, metabolism, and elimination (ADME) studies. 

An important feature of drug development is the estimation of pharmacokinetic parameters in animal models. Pharmacokinetics is the study of the time dependence and mechanism of absorption of a compound dosed into the body, its distribution throughout the fluids and other body tissues, the sequential metabolic transformations of the compound and its first-generation metabolites, and the elimination of the original compound and its metabolites (whence the common abbreviation ADME studies). The usual experimental raw data consist of concentrations of the test compound (and sometimes of its metabolites) in body tissues and body fluids (blood plasma, urine) as a function of time following a single dose. Extraction of quantitative parameters characterizing this behavior is determined by the theoretical model used to interpret the data. For example, if the dose is administered intravenously and the compound concentrations are measured in the blood, there will be an immediate drop of compound concentration with time as the compound is re-distributed, metabolized and excreted, but if an oral dose is used (as... 

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