In vivo metabolism studies

B. R., SB-242235, a selective inhibitor of p-38 mitogen-activated protein kinase. II in vitro and in vivo metabolism studies and pharmacokinetic extrapolation to man, Xenobiotica 2002, 32, 235—250. 

Salicylic acid and Its metabolite were separated by two methods. The first was thin layer chromatography on cellulose with BAW solvent as for the In vivo metabolism studies. A quicker separation was achieved with a polyamide column. The entire 400 pL from an individual assay was placed on top of a 0.8 x 2.0 cm column packed with Polyamide-6 (Accurate Chemical and Scientific Corp.). The salicylic acid metabolite was eluted with 6 mL water but salicylic acid was retained. 3a70B scintillation fluid (Research Product International Corp.) was used to determine the radioactive content of the entire 6 mL of eluant. Separation of salicylic acid and its metabolite by polyamide column chromatography was verified by thin layer chromatography. 

Both in vitro and in vivo metabolic studies have shown that some drug metabolism occurs or can be induced in placental tissues (20). Further degradation of parent chloroquine that reaches the placenta would result in the appearance of less chloroquine and more of the nonpolar metabolites in the fetal or neonatal circulation. (17). 

Several other in vitro and in vivo metabolic studies of vitamin Bg have been published (79-86). 

Turteltaub, K.W. Livermore National Laboratory and University of California at San Francisco San Francisco, CA In vivo metabolic studies characterization of DNA and protein adducts in bone marrow Health Effects Institute... 

Both in vitro and in vivo metabohsm has to be studied. Most of the results reviewed are from in vitro studies. The metabolites are generated by the use of recombinant enzymes, e.g., related to the cytochrome P450 (CYP) complex, by the use of subcellular fractions, e.g., microsomes, cytosols, or S9 fractions, or by the use of hepatocytes or liver slices. In the case of in vivo metabolism studies, the metabohtes are generated in living animals or humans, and analysis is performed in urine, plasma, bile, and/or faeces. 

The combination of electron capture ionization with liquid chromatography/mass spectrometry produces a valuable technique for metabolism studies of fluorinated compounds. Selective ionization of halogena-ted components simplifies sample purification since most biological constituents are transparent to detection. This report details the application of this technique to in vitro and in vivo metabolism studies of fluorinated herbicides. 

This report describes several applications of negative ion LC/MS to in vitro and in vivo metabolism studies of these fluorinated herbicides. 

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. 

The metabolic activation of AAF has been shown by several studies ultimately to involve the esterification of A -hydroxy-AAF (Fig. 1). While in vivo metabolism studies coupled with comparative bioassays for carcinogenicity were largely responsible for revealing this mechanism of action, in vitro metabolism of aromatic amines provided some essential details on their metabolic activation (223, 224). 

Sulphotransferase activity is present in hepatopancreas of H, americanus (Table 20) and has been indicated in other crustacean species (Khan et al. 1974). The activity in H. americanus utilizes PAPS to conjugate / -nitrophenol and is mainly cytosolic (Elmamlouk and Gessner 1978). Methylase and acetylase activities are indicated from in vivo metabolism studies (Table 21). 

In vivo metabolism smdies entail analysis and determination of dmgs and metabolites in blood, urine, and feces. These matrices contain a larger amount of endogenous compounds that could coelute and interfere with LC-MS analysis. A good sample preparation technique and capable chromatographic separation (LC, UPLC) and a selective detection technique (MS/MS) are therefore important for successful in vivo metabolism studies. 

An example of the application of LC-MS and UPLC-MS in vitro and in vivo metabolism studies is the study of the metabolism of prazosin (antihypertensive agent being investigated for the treatment of posttraumatic stress disorder). The in vivo metabolism of prazosin in rat was first reported in 1977,[28] metabolites were characterized, though at... 

The analysis of samples in biological matrices is extremely powerful compared with drawbacks such as narrow concentration range for quantitative measurements and long time required for spot detection. Digital autoradiography can be used in hyphenated off-line and online methods. A powerful example is TLC/DAR/FABMS/MS with applications in in vitro and in vivo metabolism studies. 

When stable isotopes are used, for example, as tracers for in vivo metabolic studies, typically the tracer flux is being measured and inferences are made about the flux of the material being traced ( tracee ). When there is no isotope fractionation, the flux of the heavy tracer is equal to the flux of the lighter major isotope and hence the tracee flux. However, when there is isotope fractionation the fluxes of the heavy tracer and tracee are not equal and thus the flux derived from the kinetic analysis of the tracer is not equal to that of the tracee. For example, is fractionated between water and carbon dioxide (a = 1.041 at 25°C) thus the rate of removal of by CO2 is 4.1% greater than the rate of removal. Because comprises 99.8% of the oxygen pool, the rate of CO2 production is essentially equal to the rate of removal by CO2, which will be less by 4.1% than that measured from 0. Without taking this correction into account, the calculated tracee flux will be in error. On the other hand, the isotope effect becomes negligible in the distribution of the isotopes of heavier elements therefore, no corrections are necessary in the tracer experiments. 

Four oxidative metabolites 1-hydroxycarvedilol, 8-hydroxycarvedilol, 4 -hydroxycarvedilol, and O-desmethylcarvedilol were formed by incubation of R +)- and S(-)-carvedilol with rat liver microsomes (Fujimaki 1994). As expected from in vivo metabolism studies, 1-hydroxycarvedilol and 8-hydroxycarvedilol were the major products for both enantiomers used as a substrate. The S/R enantiomeric ratios for intrinsic clearance (Vmax/KM) of 1-hydroxycarvedilol, 8-hydroxycarvedilol, 0-desmethyl-carvedilol and 4 -hydroxycarvedilol were 0.40,1.99, 0.77, and 2.71, respectively, showing that stereospe-cific oxidation occurs in this species. The cause of... 

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