• Biomolecules & Therapeutics
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• v.20 no.6
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• pp.562-568
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• 2012

Cytochrome P450 BM3 (CYP102A1) from Bacillus megaterium is a self-sufficient monooxygenase that consists of a heme domain and FAD/FMN-containing reductase domain (BMR). In this report, the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) by BMR was evaluated as a method for monitoring BMR activity. The electron transfer proceeds from NADPH to BMR and then to BMR substrates, MTT and CTC. MTT and CTC are monotetrazolium salts that form formazans upon reduction. The reduction of MTT and CTC followed classical Michaelis-Menten kinetics ($k_{cat}=4120\;min^{-1}$, $K_m=77{\mu}M$ for MTT and $k_{cat}=6580\;min^{-1}$, $K_m=51{\mu}M$ for CTC). Our continuous assay using MTT and CTC allows the simple, rapid measurement of BMR activity. The BMR was able to metabolize mitomycin C and doxorubicin, which are anticancer drug substrates for CPR, producing the same metabolites as those produced by CPR. Moreover, the BMR was able to interact with CYP1A2 and transfer electrons to promote the oxidation reactions of substrates by CYP1A2 and CYP2E1 in humans. The results of this study suggest the possibility of the utilization of BMR as a surrogate for mammalian CPR.

• Korean Journal of Clinical Pharmacy
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• v.20 no.3
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• pp.200-204
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• 2010

The purpose of this study was to investigate the effect of ticlopidine on the pharmacokinetics of nimodipine in rats. Pharmacokinetic parameters of nimodipine were determined in rats after oral administration of nimodipine (16 mg/kg) with or without ticlopidine (3 or 10 mg/kg). Ticlopidine inhibited cytochrome P450 (CYP)3A4 activity. Ticlopidine significantly (p<0.05, 10 mg/kg) increased the area under the plasma concentration-time curve (AUC) of nimodipine and ticlopidine significantly (p<0.05, 10 mg/kg) prolonged the terminal half-life ($t_{1/2}$) of nimodipine. Ticlopidine significantly (p<0.05, 10 mg/kg) decreased the total body clearance ($CL_t$). The absolute bioavailability (AB%) and relative bioavailability (RB%) of nimodipine by presence of ticlopidine were increased by 14% and by 42%, respectively, compared to the control. Based on these results, the increased bioavailability of nimodipine might be due to inhibition of the metabolizing enzyme cytochrome P450 (CYP)3A4 in the liver or intestinal mucosa and/or reducing total body clearance by ticlopidine.

• Korean Journal of Pharmacognosy
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• v.50 no.3
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• pp.159-164
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• 2019

Zanthoxylum Peel is widely used as a common spice for a variety of foods. In the orient, it has also been used as traditional agents for treating diseases such as indigestion. Recently, Zanthoxylum Peel has been reported to have anti-cancer activity, anti-microbial activity, and anti-inflammatory activity. Chemical components are known sanshool compounds and xanthoxylin. In this study, we were carried out to investigate the constituents of inhibiting a drug metabolizing enzyme CYP3A4 from Zanthoxylum Peel. CYP3A4 is known as an enzyme involved in drug metabolism as monooxygenase containing the heme. As a result of experiment, we found that bergapten ($IC_{50}=18.21{\mu}M$) and quercetin ($IC_{50}=17.27{\mu}M$) isolated from EtOAc extract of Zanthoxylum Peel showed remarkable CYP3A4-inhibiting activities. Structures of the isolated active compounds were established by chemical and spectroscopic means.

• Journal of Animal Reproduction and Biotechnology
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• v.35 no.4
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• pp.329-337
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• 2020

Cytochrome P450 1A2 (CYP1A2) is a member of the cytochrome P450 superfamily enzymes in mammals and plays a major role in metabolizing endogenous hormones in the liver. In recent days, CYP1A2 expression has been found in not only the liver but also other tissues including the pancreas and lung. However, little information is available regarding the expression of CYP1A2 in the ovary, in spite of the facts that the ovarian follicle growth and atresia are tightly associated with controls of endocrine hormonal networks. Therefore, the expression of CYP1A2 in the ovaries of prepubertal and pubertal rats was investigated to assess its expression pattern and puberty-related alteration. It was demonstrated that the expression level of CYP1A2 was significantly (p < 0.01) higher in the pubertal ovaries than prepubertal counterparts. At the ovarian follicle level in both groups, whereas CYP1A2 expression was less detectable in the primordial, primary and secondary follicles, the strongly positive expression of CYP1A2 was localized in the granulosa cell layers in the antral and pre-ovulatory follicles. However, the ratio of CYP1A2-positive ovarian follicle was significantly (p < 0.01) higher in the ovary of pubertal group (73.1 ± 3.1%) than prepubertal one (41.0 ± 10.5%). During the Immunofluorescence, expression of CYP1A2 was mainly localized in Fas-positive follicles, indicating the atretic follicles. In conclusion, these results suggested that CYP1A2 expression was mainly localized at the atretic follicular cells and affected by the onset of puberty. Further study is still necessary but we hypothesize that CYP1A2 expresses in the atretic follicles to metabolize residue of the reproductive hormones. These findings may have important implications for the fields of reproductive biology of animals.

• Parasites, Hosts and Diseases
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• v.51 no.2
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• pp.165-175
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• 2013

The fear that schistosomes will become resistant to praziquantel (PZQ) motivates the search for alternatives to treat schistosomiasis. The antimalarials quinine (QN) and halofantrine (HF) possess moderate antischistosomal properties. The major metabolic pathway of QN and HF is through cytochrome P450 (CYP) 3A4. Accordingly, this study investigates the effects of CYP3A4 inhibitor, ketoconazole (KTZ), on the antischistosomal potential of these quinolines against Schistosoma mansoni infection by evaluating parasitological, histopathological, and biochemical parameters. Mice were classified into 7 groups: uninfected untreated (I), infected untreated (II), infected treated orally with PZQ (1,000 mg/kg) (III), QN (400 mg/kg) (IV), KTZ (10 mg/kg)+QN as group IV (V), HF (400 mg/kg) (VI), and KTZ (as group V)+HF (as group VI) (VII). KTZ plus QN or HF produced more inhibition (P<0.05) in hepatic CYP450 (85.7% and 83.8%) and CYT b5 (75.5% and 73.5%) activities, respectively, than in groups treated with QN or HF alone. This was accompanied with more reduction in female (89.0% and 79.3%), total worms (81.4% and 70.3%), and eggs burden (hepatic; 83.8%, 66.0% and intestinal; 68%, 64.5%), respectively, and encountering the granulomatous reaction to parasite eggs trapped in the liver. QN and HF significantly (P<0.05) elevated malondialdehyde levels when used alone or with KTZ. Meanwhile, KTZ plus QN or HF restored serum levels of ALT, albumin, and reduced hepatic glutathione (KTZ+HF) to their control values. KTZ enhanced the therapeutic antischistosomal potential of QN and HF over each drug alone. Moreover, the effect of KTZ+QN was more evident than KTZ+HF.

• Biomolecules & Therapeutics
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• v.21 no.6
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• pp.487-492
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• 2013

Cytochrome P450 4A11 (CYP4A11) is a fatty acid hydroxylase enzyme expressed in human liver. It catalyzes not only the hydroxylation of saturated and unsaturated fatty acids, but the conversion of arachidonic acid to 20-hydroxyeicosatetraenoic acid (20-HETE), a regulator of blood pressure. In this study, we performed a directed evolution analysis of CYP4A11 using the luminogenic assay system. A random mutant library of CYP4A11, in which mutations were made throughout the entire coding region, was screened with luciferase activity to detect the demethylation of luciferin-4A (2-[6-methoxyquinolin-2-yl]-4,5-dihydrothiazole-4-carboxylic acid) of CYP4A11 mutants in Escherichia coli. Consecutive rounds of random mutagenesis and screening yielded three improved CYP4A11 mutants, CP2600 (A24T/T263A), CP2601 (T263A), and CP2616 (A24T/T263A/V430E) with ~3-fold increase in whole cells and >10-fold increase in purified proteins on the luminescence assay. However, the steady state kinetic analysis for lauric acid hydroxylation showed the significant reductions in enzymatic activities in all three mutants. A mutant, CP2600, showed a 51% decrease in catalytic efficiency ($k_{cat}/K_m$) for lauric acid hydroxylation mainly due to an increase in $K_m$. CP2601 and CP2616 showed much greater reductions (>75%) in the catalytic efficiency due to both a decrease in $k_{cat}$ and an increase in Km. These decreased catalytic activities of CP2601 and CP2616 can be partially attributed to the changes in substrate affinities. These results suggest that the enzymatic activities of CYP4A11 mutants selected from directed evolution using a luminogenic P450 substrate may not demonstrate a direct correlation with the hydroxylation activities of lauric acid.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.16
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• pp.6575-6580
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• 2014

This study aimed to evaluate the antimutagenic and anticarcinogenic activity of turmeric essential oil as well as to establish biochemical mechanisms of action. Antimutagenicity testing was accomplished using strains and known mutagens with and without microsomal activation. Anticarcinogenic activity was assessed by topical application of 7, 12 - dimethylbenz[a]anthracene (DMBA) as initiator and 1% croton oil as promoter for the induction of skin papillomas in mice. Inhibition of p450 enzymes by TEO was studied using various resorufins and aminopyrene as substrate. Turmeric essential oil (TEO) showed significant antimutagenic activity (p<0.001) against direct acting mutagens such as sodium azide ($NaN_3$), 4-nitro-O-phenylenediamine (NPD) and N-methyl-N-nitro N'nitrosoguanine (MNNG). TEO was found to have significant antimutagenic effect (>90%) against mutagen needing metabolic activation such as 2-acetamidoflourene (2-AAF). The study also revealed that TEO significantly inhibited (p<0.001) the mutagenicity induced by tobacco extract to Salmonella TA 102 strain. DMBA and croton oil induced papilloma development in mice was found to be delayed and prevented significantly by TEO application. Moreover TEO significantly (P<0.001) inhibited isoforms of cytochrome p450 (CYP1A1, CYP1A2, CYP2B1/2, CYP2A, CYP2B and CYP3A) enzymes in vitro, which are involved in the activation of carcinogens. Results indicated that TEO is antimutagenic and anticarcinogenic and inhibition of enzymes (p450) involved in the activation of carcinogen is one of its mechanisms of action.

• Archives of Pharmacal Research
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• v.28 no.10
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• pp.1196-1202
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• 2005

Omeprazole (OMP) is a proton pump inhibitor used as an oral treatment for acid-related gastrointestinal disorders. In the liver, it is primarily metabolized by cytochrome P-450 (CYP450) isoenzymes such as CYP2C19 and CYP3A4. 5-Hyroxyomeprazole (5-OHOMP) and omeprazole sulfone (OMP-SFN) are the two major metabolites of OMP in human. Cimetidine (CMT) inhibits the breakdown of drugs metabolized by CYP450 and reduces, the clearance of coad-ministered drug resulted from both the CMT binding to CYP450 and the decreased hepatic blood flow due to CMT. Phenobarbital (PB) induces drug metabolism in laboratory animals and human. PB induction mainly involves mammalian CYP forms in gene families 2B and 3A. PB has been widely used as a prototype inducer for biochemical investigations of drug metabolism and the enzymes catalyzing this metabolism, as well as for genetic, pharmacological, and toxicological investigations. In order to investigate the influence of CMT and PB on the metabolite kinetics of OMP, we intravenously administered OMP (30 mg/kg) to rats intraperitoneally pretreated with normal saline (5 mL/kg), CMT (100 mg/kg) or PB (75 mg/kg) once a day for four days, and compared the pharmacokinetic parameters of OMP. The systemic clearance ($CL_{t}$) of OMP was significantly (p<0.05) decreased in CMT-pretreated rats and significantly (p<0.05) increased in PB-pretreated rats. These results indicate that CMT inhibits the OMP metabolism due to both decreased hepatic blood flow and inhibited enzyme activity of CYP2C19 and 3A4 and that PB increases the OMP metabolism due to stimulation of the liver blood flow and/or bile flow, due not to induction of the enzyme activity of CYP3A4.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.22 no.4
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• pp.373-382
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• 2000

Individual genetic susceptibilities to cancers may result from several factors including differences in xenobiotics metabolism to chemical carcinogens, DNA repair, altered oncogenes and suppressor genes, and environmental carcinogen exposures. Among them, genetic polymorphisms of metabolizing enzymes to chemical carcinogens have been recognized as a major important host factors in human cancers. They have two main types of enzymes: the phase I cytochrome P-450 mediating enzymes (CYPs) and phase II conjugating enzymes. The purpose of this study is to determine the frequencies of genotypes of phase I (CYP1A1 and CYP2E1) and phase II (NAT2) metabolizing enzymes in healthy control and head and neck cancer patients of Korean and to identify the relative high risk genotypes of these metabolizing enzymes to head and neck cancer in Korean. The author has analyzed 132 head and neck cancer patients and 113 healthy controls using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The results were as following; 1. The frequencies of genotypes of CYP1A1, CYP2E1 and NAT2 in healthy control were as following; CYP1A1 exon 7 polymorphism; Ile/Ile: Ile/Val: Val/Val = 59.3%: 36.3%: 4.4% CYP2E1 Pst I polymorphism, C1/C1: C1/C2: C2/C2 = 61.1%: 32.1%: 6.2% NAT2 polymorphism; F/F: F/S: S/S = 43.4%: 48.7%: 8.0% 2. In analysis of phase I enzyme, Val/Val genotype in CYP1A1 exon 7 polymorphism and C2/C2 genotype in CYP2E1 Pst I polymorphism were associated with relative high risks to head and neck cancers (Odds' ratio: 2.09 and 1.37, respectively). 3. Among the genotypes of NAT2 enzyme polymorphism, S/S genotype of NAT2 enzyme had 1.03 times of relative risk to head and neck cancers. 4. In combined genotyping of CYP1A1, CYP2E1, and NAT2 enzymes polymorphisms, the patients with Val/Val and C1/C1, C2/C2 and fast acetylator, and Val/Val and fast acetylator had higher relative risks than the patients with each baseline of combined genotypes (Odds' ratio: 2.82, 1.98 and 2.1, respectively). These results suggest the combined genotypes of Val/Val and C1/C1, C2/C2 and fast acetylator, and Val/Val and fast acetylator were more susceptible to head and neck cancers in Korean. And genotyping of metabolizing enzymes could be useful for predicting individual susceptibility to head and neck cancer.

• Mass Spectrometry Letters
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• v.12 no.4
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• pp.172-178
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• 2021

HYIpro-3-1 is an adjuvant for preventing or treating inflammatory growth diseases. In this study, we identified the metabolic pathway of HYIpro-3-1 in human liver microsomes (HLMs) by quadrupole-orbitrap high-resolution mass spectrometry (HR-MS) and characterized the major human cytochrome P450 (CYP). Ten metabolites were identified, including one O-demethylation (M1), two O-demethylation and monohydroxylation (M2 and M3), and seven monohydroxylation metabolites (M4-M10). Based on the HR-MS² spectra, the metabolites are divided into two groups of monohydroxylated metabolites according to the hydroxylation position. We verified that HYIpro-3-1 is metabolized by CYP in HLMs, CYP2B6 is mainly involved in O-demethylation, and various CYPs are involved in the monohydroxylation of HYIpro-3-1.