Identification, functional characterization, and regulation of a new cytochrome P450 subfamily, the CYP2Ns.
AuthorsOleksiak MF,Wu S,Parker C,Karchner SI,Stegeman JJ,Zeldin DC
JournalThe Journal of biological chemistry
PubMed ID10644680
The screening of liver and heart cDNA libraries from the teleost Fundulus heteroclitus with degenerate oligonucleotide probes to conserved alpha-helical regions in mammalian P450s resulted in the identification of two cDNAs that together represent a novel P450 subfamily, the CYP2Ns. Northern analysis demonstrated that CYP2N1 transcripts are most abundant in ... More
Microtiter plate assays for inhibition of human, drug-metabolizing cytochromes P450.
AuthorsCrespi CL, Miller VP, Penman BW
JournalAnal Biochem
PubMed ID9177742
Enzymatic analysis of cDNA-expressed human CYP1A1, CYP1A2, and CYP1B1 with 7-ethoxyresorufin as substrate.
AuthorsChang TK, Waxman DJ,
JournalMethods Mol Biol
PubMed ID16719376
'Cytochrome P450 (P450) enzymes belonging to the CYP1 family are highly inducible by polycyclic aromatic hydrocarbons and other environmental chemicals and play a major role in the metabolism of many foreign chemicals and endogenous substances. We describe a spectrofluorometric method for determining 7-ethoxyresorufin O-dealkylation catalyzed by CYP1A1, CYP1A2, and CYPB1. ... More
Stereoselective and regioselective hydroxylation of warfarin and selective O-dealkylation of phenoxazone ethers in human placenta.
'The oxidative metabolism of warfarin and a series of phenoxazone ethers was studied in two groups of human placentas which exhibited high or low levels of aryl hydrocarbon hydroxylase (AHH). Warfarin metabolism was stereoselective (mean R/S = 2.48) for the R-enantiomer and regioselective for the 6- and 8- positions in ... More
Methoxyresorufin and benzyloxyresorufin: substrates preferentially metabolized by cytochromes P4501A2 and 2B, respectively, in the rat and mouse.
AuthorsNerurkar PV, Park SS, Thomas PE, Nims RW, Lubet RA
JournalBiochem Pharmacol
PubMed ID8373445
'The cytochrome P450 isozyme specificity for the O-dealkylation of methoxyresorufin (MTR) and benzyloxyresorufin (BZR) in the rat and mouse was investigated. The induction of various alkoxyresorufin O-dealkylation activities was measured in male F344/NCr rats exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin or 3,4,5,3',4',5'-hexachlorobiphenyl. MTR and ethoxyresorufin (ETR) O-dealkylation activities were induced 30- and 80-fold, ... More
Arachidonic acid inhibits capacitative calcium entry in rat thymocytes and human neutrophils.
AuthorsAlonso-Torre SR, García-Sancho J
JournalBiochim Biophys Acta
PubMed ID9315617
'Emptying the intracellular Ca2+ stores by treatment with the endomembrane Ca2+-ATPase inhibitor thapsigargin activates capacitative Ca2+ entry (CCE). This can be evidenced in fura-2-loaded cells by an increase of [Ca2+]i or by an acceleration of Mn2+ entry. Micromolar concentrations of arachidonic acid inhibited CCE induced by treatment with thapsigargin in ... More
Redox cycling of resorufin catalyzed by rat liver microsomal NADPH-cytochrome P450 reductase.
AuthorsDutton DR, Reed GA, Parkinson A
JournalArch Biochem Biophys
PubMed ID2464338
'The O-dealkylation of 7-alkoxyresorufins to the highly fluorescent compound, resorufin (7-hydroxyphenoxazone), provides a rapid, sensitive, and convenient assay of certain forms of liver microsomal cytochrome P450. The results of this study indicate that NADPH-cytochrome P450 reductase catalyzes the reduction of resorufin (and the 7-alkoxyresorufins) to a colorless, nonfluorescent compound(s). The ... More
Distribution of cytochrome P450 activities towards alkoxyresorufin derivatives in rat brain regions, subcellular fractions and isolated cerebral microvessels.
AuthorsPerrin R, Minn A, Ghersi-Egea JF, Grassiot MC, Siest G
JournalBiochem Pharmacol
PubMed ID2242042
'The regional and subcellular distributions of rat brain cytochrome P450 and cytochrome P450-dependent activities were examined. Cytochrome P450 was found to be mainly localized in mitochondria in all the six cerebral regions studied. The activities of the isoforms mostly implicated in drug metabolism, cytochromes P450 b and c, were measured ... More
7-Alkoxyquinolines: new fluorescent substrates for cytochrome P450 monooxygenases.
'A series of 7-alkoxyquinolines was synthesized and tested as substrates with hepatic microsomes prepared from male Wistar rats. Microsomal O-dealkylation rates and kinetic constants were determined for the 7-alkoxyquinolines with microsomes from control, 3-methylcholanthrene (MC)-pretreated, and phenobarbitone (PB)-pretreated rats. Structure-activity relationship studies indicated that the 7-benzyloxyquinoline was the most rapidly ... More
Evaluation of alkoxyresorufins as fluorescent substrates for cytochrome P450 BM3 and site-directed mutants.
'In this study, the first fluorescent assay for bacterial cytochrome P450 BM3 (BM3) and mutants is described. BM3 mutants are potentially very versatile biocatalysts for the production of fine chemicals. A fluorescent assay would be very useful for the identification of nonnatural ligands in high-throughput inhibition assays. Because of the ... More
Human adult hepatocytes in primary monolayer culture. Maintenance of mixed function oxidase and conjugation pathways of drug metabolism.
'The stabilities of several drug oxidation and conjugation pathways in human adult hepatocytes have been investigated during 72 hr culture. Cytochrome P-450-dependent mixed function oxidase was measured by the O-dealkylations of ethoxyresorufin (EROD), pentoxyresorufin (PROD) and benzyloxyresorufin (BROD), which are probes for different isozymes of cytochrome P-450 in the rat. ... More
Polycyclic aromatic hydrocarbons cause hepatic porphyria in iron-loaded C57BL/10 mice: comparison of uroporphyrinogen decarboxylase inhibition with induction of alkoxyphenoxazone dealkylations.
AuthorsFrancis JE, Smith AG
JournalBiochem Biophys Res Commun
PubMed ID3606611
'Multiple doses of beta-naphthoflavone to iron-loaded C57BL/10ScSn mice for 6 weeks caused inhibition of hepatic uroporphyrinogen decarboxylase and a porphyria indistinguishable from that previously only reported for polyhalogenated aromatic chemicals. beta-Naphthoflavone and other polycyclic aromatic hydrocarbon inducers of cytochrome P1-450-mediated ethoxyphenoxazone deethylation (ethoxyresorufin deethylase), benzo[a]pyrene, benz[a]anthracene, dibenz[ah]anthracene, 3-methylcholanthrene and alpha-naphthoflavone, ... More
Differential effects of phenobarbitone and 3-methylcholanthrene induction on the hepatic microsomal metabolism and cytochrome P-450-binding of phenoxazone and a homologous series of its n-alkyl ethers (alkoxyresorufins).
AuthorsBurke MD, Mayer RT
JournalChem Biol Interact
PubMed ID6883573
'The metabolism and cytochrome P-450-binding of phenoxazone and a homologous series of its n-alkyl ethers (1-8C) was studied in hepatic microsomes of control, phenobarbitone-pretreated (PB) and 3-methylcholanthrene-pretreated (3MC) C57/BL10 mice. Phenoxazone and its ethers were hydroxylated and O-dealkylated respectively to a common metabolite, resorufin. The three categories of microsomes differed ... More
Ethoxy-, pentoxy- and benzyloxyphenoxazones and homologues: a series of substrates to distinguish between different induced cytochromes P-450.
'The individual members of a homologous series of phenoxazone ethers related to ethoxyresorufin were O-dealkylated, and the parent compound phenoxazone was ring-hydroxylated, each at different rates with hepatic microsomes of untreated rats. A structure-activity relationship (SAR) was plotted, relating the rate of O-dealkylation to the length and type of the ... More
Further characterization of the expression in liver and catalytic activity of CYP2B6.
AuthorsEkins S, Vandenbranden M, Ring BJ, Gillespie JS, Yang TJ, Gelboin HV, Wrighton SA
JournalJ Pharmacol Exp Ther
PubMed ID9732386
'Previous studies in this laboratory have determined the lack of specificity of several antibody and substrate probes of CYP2B6. The goals of the current study were to examine the expression of CYP2B6 in a bank of human liver microsome (HLM) samples using a new specific monoclonal antibody (MAb 49-10-20) and ... More
Use of 7-alkoxyphenoxazones, 7-alkoxycoumarins and 7-alkoxyquinolines as fluorescent substrates for rainbow trout hepatic microsomes after treatment with various inducers.
'Various fluorescent substrates have been used as specific indicators of induction or activity of different cytochrome P450 isozymes in both fish and mammalian species. In an attempt to identify additional definitive fluorescent substrates for use in fish, we examined a series of 7-alkoxyphenoxazones, 7-alkoxycoumarins and 7-alkoxyquinolines as substrates in O-dealkylation ... More
Induction of liver monooxygenases by annatto and bixin in female rats.
AuthorsDe-Oliveira AC, Silva IB, Manhaes-Rocha DA, Paumgartten FJ
JournalBraz J Med Biol Res
PubMed ID12532234
'Annatto or urucum is an orange-yellow dye obtained from Bixa orellana seeds. It has been used as a natural dye in a variety of food products, drugs and cosmetics, and also in Brazilian cuisine as a condiment (''colorau''). Bixin, a carotenoid devoid of provitamin A activity, is the main pigment ... More
Multiple forms of cytochrome P-450 related to forms induced marginally by phenobarbital. Differences in structure and in the metabolism of alkoxyresorufins.
'The properties of five structurally related forms of cytochrome P-450 (PB1a, PB1b, PB2a, PB2b and PB2d) isolated from rats treated with phenobarbital have been compared with two forms isolated previously now termed ''PB1c'' and ''PB2c''. These enzymes were characterized by their marginal inducibility by phenobarbital and are clearly distinguishable from ... More
Inhibition of hepatic and cutaneous biotransformation of resorufin ethers following intraperitoneal administration of 1-aminobenzotriazole.
AuthorsOldfield NF, Mortillo M, Garland WA, Mico BA
JournalPharm Res
PubMed ID1409385
O-dealkylation of resorufin ethers as an indicator of hepatic cytochrome P-450 isoenzyme induction in the cotton rat (Sigmodon hispidus): a method for monitoring environmental contamination.
AuthorsElangbam CS, Qualls CW, Lochmiller RL
JournalBull Environ Contam Toxicol
PubMed ID1932860
Metabolism of alkoxyphenoxazones by channel catfish liver microsomes: effects of phenobarbital, Aroclor 1254 and 3-methylcholanthrene.
AuthorsAnkley GT, Reinert RE, Mayer RT, Burke MD, Agosin M
JournalBiochem Pharmacol
PubMed ID3109440
Alkoxyresorufin O-dealkylases: association with the murine Ah locus.
AuthorsLum PY, Burke MD, Mayer RT, Ioannides C
JournalCancer Lett
PubMed ID3768851
The hepatic microsomal dealkylation of a series of alkoxyresorufins and the oxidation of phenoxazone to resorufin were investigated in C57BL/6 and DBA/2 mice of both sexes. In both strains of mice and in both sexes the dealkylation rate decreased with increasing length of the alkyl chain. With all alkoxyresorufins the ... More
Probing the active site of cytochrome P450 2B1: metabolism of 7-alkoxycoumarins by the wild type and five site-directed mutants.
AuthorsKobayashi Y, Fang X, Szklarz GD, Halpert JR
JournalBiochemistry
PubMed ID9578551
A series of 7-alkoxycoumarins (chain length of 1-7 carbon atoms) was utilized as active site probes of purified Escherichia coli-expressed cytochrome P450 2B1 wild type and five site-directed mutants (I114V, F206L, V363A, V363L, and G478S). The production of 7-hydroxycoumarin, the O-dealkylation product, by the wild-type enzyme exhibited a rank order ... More
Inhibition of cytochromes P450 by nitric oxide and a nitric oxide-releasing agent.
AuthorsWink DA, Osawa Y, Darbyshire JF, Jones CR, Eshenaur SC, Nims RW
JournalArch Biochem Biophys
PubMed ID8424643
The effect of nitric oxide (NO) on cytochrome P450-mediated benzyloxyresorufin and ethoxyresorufin O-dealkylase activity of rat hepatic postmitochondrial (S-9) or microsomal subfractions, or purified rat liver CYP2B1, was examined. Two distinct inhibitory phases were observed regardless of whether the NO was added prior to initiation of the reactions with NADPH ... More
A comparative study of constitutive and induced alkoxyresorufin O-dealkylation and individual cytochrome P450 forms in cynomolgus monkey (Macaca fascicularis), human, mouse, rat and hamster liver microsomes.
AuthorsWeaver RJ, Thompson S, Smith G, Dickins M, Elcombe CR, Mayer RT, Burke MD
JournalBiochem Pharmacol
PubMed ID8135852
The expression of constitutive and inducible cytochrome P450 forms was measured in cynomolgus monkey liver and compared with man, rat, mouse and hamster. Four alkoxyresorufin O-dealkylation (AROD) activities widely used as indicators of P450 induction were measured: methoxyresorufin O-demethylation (MROD), ethoxyresorufin O-deethylation (EROD), pentoxyresorufin O-dealkylation (PROD) and benzyloxyresorufin O-dealkylation (BROD). ... More
Effects of polychlorinated biphenyls, kanechlor-500, on serum thyroid hormone levels in rats and mice.
AuthorsKato Y, Haraguchi K, Yamazaki T, Ito Y, Miyajima S, Nemoto K, Koga N, Kimura R, Degawa M
JournalToxicol Sci
PubMed ID12660360
Effects of a commercial polychlorinated biphenyls mixture, Kanechlor-500 (KC500), on the levels of serum thyroid hormones such as total thyroxine (T4) and triiodothyronine (T3) were examined comparatively in male Wistar rats and ddy mice. Serum T4 levels were significantly decreased in both rats and mice 4 days after a single ... More
Assessment of regional cytochrome P450 activities in rat liver slices using resorufin substrates and fluorescence confocal laser cytometry.
Characterizing constitutive activities and inducibility of various cytochrome P450 isozymes is important for elucidating species and individual differences in susceptibility to many toxicants. Although expression of certain P450s has been studied in homogenized tissues, the ability to assess functional enzyme activity without tissue disruption would further our understanding of interactive ... More