No description available.

In the treatment of arrhythmias, amiodarone is a primary therapeutic agent. Cytochromes P450 (CYP) 1A1 and 1A2 facilitate biotransformation of amiodarone to the biologically active desethylamiodarone. Side effects have been reported during therapy and some are correlated with increased desethylamiodarone levels. Exposure to polycyclic aromatic hydrocarbons (PAH) like -naphthoflavone induces CYP1A1 and CYP1A2 and therefore can increase desethylamiodarone levels. Desethylamiodarone, however, was reported to inactivate human CYP1A1 and therefore can conceivably inhibit its CYP1A1-mediated formation. Our primary objective was to investigate the effect of -naphthoflavone on amiodarone disposition. Since rats were used, CYP isoenzymes involved in desethylamiodarone formation in human were compared to their rat counterparts. The effect of ketoconazole on desethylamiodarone formation, the inactivating potential of desethylamiodarone on CYP1A1 and the mechanism of -naphthoflavone-amiodarone interaction were assessed. Human CYP1A1 and rat CYP2D1 had the highest intrinsic clearance (Clint) for desethylamiodarone. Human and rat CYP1A2 had the lowest Clint. Ketoconazole (18.8 M) inhibited all isoforms except for rat CYP1A2; it potently inhibited human CYP1A1 and CYP3A4 and rat CYP2D2 and CYP1A1. After a single amiodarone dose was administered to control and -naphthoflavone pretreated rats, the plasma area under concentration-time curve (AUC) of desethylamiodarone increased. With multiple doses, amiodarone AUC(0-24h) decreased in -naphthoflavone plasma (30%), lung (35%), liver (48%), kidney (52%), heart (34%), and intestine (43%). Desethylamiodarone AUC(0-24h) increased in -naphthoflavone plasma (36%), lung (56%), liver (101%), kidney (65%), and heart (73%). Desethylamiodarone caused no inactivation of CYP1A1 when preincubations were diluted and nicotinamide adenine dinucleotide phosphate (NADPH) was added in the probe incubation samples. Evidence for reversible mixed-competitive inhibition was apparent. Addition and/or replenishment of NADPH were important factors in maintaining control activity. -naphthoflavone increased desethylamiodarone formation only in lung and kidney microsomes. Desethylamiodarone formation in liver, intestine and heart microsomes was not altered. Body-weight-normalized liver mass was significantly increased (27%) by -naphthoflavone. In conclusion, human CYP1A1 was more efficient in forming desethylamiodarone than rat isoenzyme. Exposure to PAH increased desethylamiodarone levels in vivo. Increased desethylamiodarone levels were partly caused by CYP1A1 induction, and by increased liver mass. Desethylamiodarone did not inactivate CYP1A1 activity. / Pharmaceutical sciences

No description available.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1982. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 138-139).

The nucleus may be the critical site for the activation of chemical carcinogens, and subsequently the initiation of neoplasia. However, isolated nuclei may be contaminated with endoplasmic reticulum, the major site of the drug metabolizing enzymes. One of the objectives of the present study was to determine whether the enzymes in isolated rat hepatic nuclei were of nuclear origin and, if so, to compare these enzymes with those in the microsomal fraction. The selective manipulation of nuclear enzymes would be a useful tool in determining their role in cellular toxicity. Recentrifugation experiments, with aryl hydrocarbon hydroxylase (AHH) activity as a marker, showed that isolated nuclei were not contaminated with endoplasmic reticulum in the form of microsomes formed upon homogenization. However, small "tags" of endoplasmic reticulum, continuous with the nuclear membrane, and indiscernable in electron micrographs, could remain following centrifugation and account for all of the measurable enzyme activity in the isolated nuclei. It was reasoned that if endoplasmic reticulum accounted for all of the activity, then the ratio of nuclear to microsomal activity for all enzymes determined should be the same. The ratios of epoxide hydrolase and AHH were found to differ in the two fractions. The simplest interpretation of these data was that drug metabolizing enzymes existed in the nuclei. However, the distribution of drug metabolizing enzymes throughout the endoplasmic reticulum is known to be heterogeneous and these "tags" could differ from the total endoplasmic reticulum (microsomes) in their enzyme make-up. Whether these enzymes are in the nuclear membrane, nucleoplasm, or as "tags" of endoplasmic reticulum, they represent activities in close proximity to potential target sites in the nucleus. The inhibition, induction, and activation characteristics of nuclear and microsomal enzymes were studied with the goal of selective manipulation of nuclear enzymes. The enzymes in the nuclear and microsomal fractions were found to differ' only in quantitative inducibility, and were identical in all other respects. Therefore, the selective manipulation of nuclear enzymes was not achieved. The induction of hepatic drug metabolizing enzymes is a measure of altered genetic expression in the liver. Inducers of drug metabolizing enzymes have also been shown to promote neoplasia in the liver. Therefore, studying the induction of such enzymes may lead to a further understanding of the mechanism of tumour promotion. Phenobarbital, 3-methylcholanthrene and pregnenolone-16α-carbonitrile produce three distinct induction responses. In the present study, spironolactone and trans-stiIbene oxide were shown to produce distinct induction responses, also. Spironolactone was shown to be a different inducer based on the protein band patterns observed following SDS-polyacrylamide gel electrophoresis of liver microsomes. trans-Stilbene oxide was found to produce a significantly different maximal level of AHH activity. The observation of five distinct induction responses suggests at least five recognition sites (receptors) mediating the pleiotropic actions of exogenous compounds in the liver. / Pharmaceutical Sciences, Faculty of / Graduate

Thesis (M. Phil.)--University of Hong Kong, 1986.

published_or_final_version / Biochemistry / Master / Master of Philosophy

Induction of the cytochrome P-450 mixed-function oxidase and specifically the cytochrome P-450 IVA1 isoenzyme by seven phenoxy acid herbicides in rat liver and kidney, have been studied. results using liver microsomes demonstrated that the 12-hydroxylation of lauric acid was significantly induced by all compounds (3-8-fold), 4-chlorophenoxyacetic acid (CPA) (300 mg/kg) being the weakest and 2,4,5-trichlorophenoxypropionic acid (2,4,5-TP) (200 mg/kg) the most potent inducers respectively. This increase in lauric acid 12-hydroxylase-activity was accompanied by an increase in the hepatic content of cytochrome P-450 IVA1 as assessed by both a qualitative Western blot procedure and a quantitative ELISA method. Furthermore, there was a parallel increase in cytochrome P-450 IVA1 mRNA and a similar increase in peroxisomal B-oxidation subsequent to exposure to these compounds. In addition, benzphetamine-N-demethylase, a marker of cytochrome P-450 IIBl and IIB2 activities, was not affected by any of the herbicides, whereas cytochrome P-450 IA1 and IA2, as assayed by ethoxyresorufin-O-deethylase activity, was significantly increased (up to 2.2-fold) by some of the compounds. Kidney microsomal parameters were not affected by any of these compounds. My in vivo studies using antipyrine, pentobarbital and zoxazolamine indicated that the metabolism of these substrates was marginally affected by only some of the compounds. In order to highlight the possible involvement of a metabolite of the chlorophenoxy acids in the induction of cytochrome P-450, I investigated four related chlorophenols. There was no significant change in cytochrome P-450 isoenzyme levels in rat liver and kidney microsomes nor was there any increase in peroxisomal beta-oxidation. Taken collectively, the results presented in this thesis indicate that the chlorinated phenoxy acid herbicides studied preferentially induce the cytochrome P-450 IVA1 isoenzyme and peroxisomal beta-oxidation in a pattern similar to the typical inducers of this isoenzyme such as clofibrate. A scheme is presented whereby induction of catalytically competent cytochrome P-450 IVA1 is required for the phenomenon of peroxisome proliferation by these chlorophenoxy acid derivatives.

Thesis (M. Phil.)--University of Hong Kong, 1993.

published_or_final_version / Biochemistry / Master / Master of Philosophy