Reduction Of Xenobiotics Research Articles

Simultaneous Absolute Quantification of 11 Cytochrome P450 Isoforms in Human Liver Microsomes by Liquid Chromatography Tandem Mass Spectrometry with In Silico Target Peptide Selection

Cytochrome P450 (CYP) proteins are involved in the biological oxidation and reduction of xenobiotics, affecting the pharmacological efficiency of drugs. This study aimed to establish a method to simultaneously quantify 11 CYP isoforms by multiplexed-multiple reaction monitoring analysis with liquid chromatography tandem mass spectrometry and in silico peptide selection to clarify CYP isoform expression profiles in human liver tissue. CYP1A2, 2A6, and 2D6 target peptides were identified by shot-gun proteomic analysis, and those of other isoforms were selected by in silico peptide selection criteria. The established quantification method detected target peptides at 10 fmol, and the dynamic range of calibration curves was at least 500-fold. The quantification value of CYP1A2 in Supersomes was not significantly different between the established method and quantitative immunoblot analysis. The absolute protein expression levels of 11 CYP isoforms were determined from one pooled and 10 individual human liver microsomes. In the individual microsomes, CYP2C9 showed the highest protein expression level, and CYP1A2, 2A6, 2C19, and 3A4 protein expression exhibited more than a 20-fold difference among individuals. This highly sensitive and selective quantification method is a useful tool for the analysis of highly homologous CYP isoforms and the contribution made by each CYP isoform to drug metabolism.

P450 SUBFAMILY CYP2J AND THEIR ROLE IN THE BIOACTIVATION OF ARACHIDONIC ACID IN EXTRAHEPATIC TISSUES*

Historically, there has been intense interest in P450 metabolic oxidation, peroxidation, and reduction of xenobiotics. More recently, there has been a growing appreciation for the role of P450s in the oxidation of lipophilic endobiotics, such as bile acids, fat-soluble vitamins, and eicosanoids. This review details the emerging CYP2J subfamily of P450s and their role as catalysts of arachidonic acid metabolism.

Oxygen and Xenobiotic Reductase Activities of Cytochrome P450

The oxygen reductase and xenobiotic reductase activities of cytochrome P450 (P450) are reviewed. During the oxygen reductase activity of P450, molecular oxygen is reduced to superoxide anion radicals (O2-.) most likely by autooxidation of a P450 ferric-dioxyanion complex. The formation of reactive oxygen species (O2-., hydrogen peroxide, and, notably, hydroxyl free radicals) presents a potential toxication pathway, particularly when effective means of detoxication are lacking. Under anaerobic conditions, P450 may also be involved in the reduction of xenobiotics. During the xenobiotic reductase activity of P450, xenobiotics are reduced by the ferrous xenobiotic complex. After xenobiotic reduction by P450, xenobiotic free radicals are formed that are often capable of reacting directly with tissue macromolecules. Unfortunately, the compounds that are reductively activated by P450 have little structural similarity. The precise molecular mechanism underlying the xenobiotic reductase activity of P450 is, therefore, not yet fully understood. Moreover, description of the molecular mechanisms of xenobiotic and oxygen reduction reactions by P450 is limited by the lack of knowledge of the three-dimensional (3D) structure of the mammalian P450 proteins.

Xenobiotic reduction and clinical improvements in capacitor workers: A feasible method

Abstract Eleven capacitor workers, occupationally exposed to PCBs and other industrial chemicals, were treated with the detoxification method developed by Hubbard while thirteen co‐workers served as controls. The mean pretreatment PCB levels were 28.0 mg/Kg in adipose and 188.0 μg/L in serum. At post‐treatment, PCBs were reduced in serum by 42% (p<0.05) and in adipose by 30% for the six patients without concurrent disease. Patients with concurrent disease had a 10% reduction in adipose levels, while serum levels remained unchanged. Both adipose and serum PCB levels increased in controls. These differences were maintained at a 4‐month follow‐up examination, though the mean adipose PCB values in all groups were higher than at post‐treatment. Changes in cholesterol, triglycerides, serum AST, ALT, and gGT did not correlate with changes of PCB values. Clinically, at post‐treatment all patients reported a marked improvement in symptoms, with most of these improvements retained at follow‐up. No such improvements...

Survey of drug-metabolizing enzymes.

The ‘drug-metabolizing enzymes’ comprise those enzymes which facilitate the biotransformation of drugs and other xenobiotics, generally rendering the lipophilic substrates more polar and hence more readily excretable. They are most abundant in mammalian liver, but also occur, at much lower activities, in the gastrointestinal tract, kidney, lung, and many other tissues. They have been classified into phase 1 enzymes (mixed-function oxidases, oxidoreductases, reductases, esterases), which modify the lipophilic xenobiotic substrates by oxidative, reductive or hydrolytic catabolism with the creation of functional groups, and phase 2 enzymes (conjugases), which transfer endogenous moieties (glucuronate, sulphate, acetate, glycine, glutathione, etc.) to the xenobiotics or their metabolites, thereby making them more polar and more readily eliminated from the cell (Parke, 1982~) . The mixed-function oxidases are the most extensively studied of these different enzymes, and comprise a NADPH-dependent flavoprotein (FAD, FMN), cytochrome P-450 reductase, and the terminal oxygen-activating/transferring enzyme, cytochrome P-450. This is an ubiquitous system, found in microorganisms, yeasts, higher animals and plants, and in many different tissues in mammals. In yeasts and higher organisms it is membrane-bound (endoplasmic reticulum and mitochondria) and generally functions to insert oxygen into xenobiotic and certain endogenous substrates (cholesterol, steroids, fatty acids). This enzyme system may also catalyse the reduction of xenobiotics (nitro and azo compounds), effect reductive dehalogenation (carbon tetrachloride, halothane, etc.) with formation of free radicals, and form ligand and covalent complexes with certain reaction products (radicals, carbenes, etc.). This complex-formation may result in the activation of oxygen by cytochrome P-450 to form superoxide anion and peroxide, without insertion of oxygen into organic substrates (Parke, 1982~) . The mixed-function oxidases also effect the oxygenation of numerous endogenous substrates and are involved in the o-hydroxylation of fatty acids, the biosynthesis and catabolism of cholesterol, steroid hormones, and the conversion of arachidonate into prostaglandins (Kupfer, 1980). The mixed-function oxidases, once thought to be a single, multi-functional enzyme system have, during the last decade, been solubilized, and shown by reconstitution to comprise a diverse three component system, namely cytochrome P-450, the flavoprotein reductase, and a phospholipid (phosphatidylcholine) electron coupler. The cytochrome P-450 fraction has been fractionated and it is now considered that tissues may contain several different cytochrome P-450 species with different, but overlapping, substrate specificities (Lu & West, 1980).