Alternative energy efficient membrane bioreactor using reciprocating submerged membrane.

A novel membrane bioreactor (MBR) pilot system, using membrane reciprocation instead of air scouring, was operated at constant high flux and daily fluctuating flux to demonstrate its application under peak and diurnal flow conditions. Low and stable transmembrane pressure was achieved at 40 l/m(2)/h (LMH) by use of repetitive membrane reciprocation. The results reveal that the inertial forces acting on the membrane fibers effectively propel foulants from the membrane surface. Reciprocation of the hollow fiber membrane is beneficial for the constant removal of solids that may build up on the membrane surface and inside the membrane bundle. The membrane reciprocation in the reciprocating MBR pilot consumed less energy than coarse air scouring used in conventional MBR systems. Specific energy consumption for the membrane reciprocation was 0.072 kWh/m(3) permeate produced at 40 LMH flux, which is 75% less than for a conventional air scouring system as reported in literature without consideration of energy consumption for biological aeration (0.29 kWh/m(3)). The daily fluctuating flux test confirmed that the membrane reciprocation is effective to handle fluctuating flux up to 50 LMH. The pilot-scale reciprocating MBR system successfully demonstrated that fouling can be controlled via 0.43 Hz membrane reciprocation with 44 mm or higher amplitude.

Characteristic features of unruptured intracranial aneurysms: predictive risk factors for aneurysm rupture.

BACKGROUND AND PURPOSE: The purpose of this study was to identify the risk factors predisposing to aneurysm rupture and to provide a reliable estimation for likelihood of rupture in unruptured intracranial aneurysms. METHODS: The authors performed a nested case-control study of 290 aneurysms (123 unruptured aneurysms and 167 ruptured aneurysms) occurring during a prospective cohort study in 1493 consecutive patients with newly diagnosed intracranial aneurysm and were treated in a single institute between January 1995 and December 2006. Controls were matched for age, treatment group, number of lesion, sex, region and study period in which the incidence of ruptured and unruptured intracranial aneurysm was equivalently balanced. The authors assessed the predictive risk factors associated with aneurysmal rupture based on the clinical and angiographic findings reported in the patients' medical records. RESULTS: Between January 1997 and December 2002, 167 patients with ruptured intracranial aneurysms were assigned to group 1, and 123 patients with unruptured intracranial aneurysms during the same period were assigned to group 2. Aspect ratio (OR 3.76), maximum diameter of neck (N(max)) < or =3 mm (OR 2.56) and family history of cerebrovascular disease (OR 5.63) were strongly correlated with aneurysm rupture (p<0.05). CONCLUSIONS: There are differences between the clinical and intrinsic characteristics of patients with unruptured and ruptured intracranial aneurysm. It will be helpful to make rational decisions regarding the optimal therapeutic strategy for unruptured intracranial aneurysm.

A functional C-G polymorphism in the CYP7B1 promoter region and its different distribution in Orientals and Caucasians.

Cytochrome P450 (CYP) 7B1 is involved in many metabolic processes including androgen metabolism. Genetic variation in the CYP7B1 gene may play a role in predisposition to prostate cancer. Here, we screened the human CYP7B1 gene for possible polymorphisms. Only one single polymorphism was detected, a C-G change in the promoter -104 base pair from the transcription start site. The allele frequency was investigated in Swedish men and compared to a Korean population, as it is known that the frequency of prostate cancer is low among Orientals. We found that the frequency of the G-allele was 4.04% in Swedes (n=150) but only 0.33% among Koreans (n=153). Computer analysis indicated that the two variants bind with different affinities to a CCAAT-box binding protein. Expression studies with reporter constructs showed significantly higher transcriptional activity of the G variant in Hek293 cells (2.7-fold, P<0.05). In conclusion, we report here for the first time the detection of a single polymorphism in the CYP7B1 gene. This polymorphism is associated with phenotypic differences in an expression system and a widely different allele frequency in two ethnic populations, with great differences in the incidence of prostate cancer.

Disposition of debrisoquine and nortriptyline in Korean subjects in relation to CYP2D6 genotypes, and comparison with Caucasians.

AIMS: To study the influence of the CYP2D6*10 allele on the disposition of debrisoquine and nortriptyline. METHODS: The pharmacokinetics of debrisoquine and nortriptyline and their main metabolites were determined in ten Koreans with the CYP2D6*1/*1 (n = 5) and CYP2D6*1/*10 (n = 5) genotypes after single oral doses of 20 mg debrisoquine and 25 mg nortriptyline, respectively. The data were compared with previously published findings from 21 Caucasians with 0, one, two, three, four or 13 functional CYP2D6 genes. RESULTS: The AUC0-8 of 4-hydroxydebrisoquine was significantly lower in Koreans with CYP2D6*1/*10 genotype compared with CYP2D6*1/*1[95% confidence interval (CI) for the ratio between means 1.17, 1.85]. No other genotype-related differences were found in the plasma kinetics of nortriptyline and debrisoquine, or their hydroxy metabolites. The AUCnortriptyline/AUC10-hydroxynortriptyline ratio did not differ between the *1/*1 and *1/*10 genotype groups (95% CI for the ratio of means 0.60, 1.26). Similarly, there was no difference between these genotypes with respect to the AUCdebrisoquine/AUC4-hydroxydebrisoquine ratio (95% CI for the ratio of mean values 0.38, 1.46). Both Korean genotype groups had similar AUCs and parent compound/metabolite AUC ratios of debrisoquine and nortriptyline to Caucasians with two functional CYP2D6 genes. CONCLUSIONS: Heterozygosity for CYP2D6*10 decreases the CYP2D6-dependent elimination of nortriptyline and debrisoquine to only a limited degree. Further studies in subjects homozygous for CYP2D6*10 are required to elucidate fully the pharmacokinetic consequences of this CYP2D6 genotype in Orientals.

Plasma concentrations of haloperidol are related to CYP2D6 genotype at low, but not high doses of haloperidol in Korean schizophrenic patients.

AIMS: This study was carried out to evaluate the influence of CYP2D6 genotype on the steady state plasma concentrations of haloperidol and reduced haloperidol in Korean schizophrenic patients. METHODS: One hundred and twenty Korean schizophrenic patients treated with various, clinically determined, doses of haloperidol (range 3-60, median 20 mg day-1) during monotherapy were recruited. CYP2D6 genotypes were determined by analysis of the CYP2D6*10 allele using allele-specific PCR and the CYP2D6*5 allele by long-PCR. Steady state plasma concentrations of haloperidol and reduced haloperidol were analysed by h.p.l.c. RESULTS: Twenty-three (19.2%), 60 (50.0%), 1 (0.8%), 33 (27.5%) and 3 patients (2.5%) possessed the CYP2D6 genotypes *1/*1, *1/*10, *1/*5, *10/*10 and *10/*5, respectively. The allele frequencies of CYP2D6*1, *10 and *5 were 44.6%, 53.8% and 1.7%, respectively. Significant relationships between dose and plasma concentrations of haloperidol (linear; r2 = 0.60, P < 0.0001) and reduced haloperidol (quadratic equation; r(2) = 0.67) were observed. Overall, the concentrations normalized for dose (C/D) of haloperidol were significantly different between the CYP2D6*1/*1, *1/*10 and *10/*10 genotype groups (one-way ANOVA; P = 0.028). No significant differences between the genotype groups were found with respect to the C/D of reduced haloperidol (P = 0.755). However, in patients with daily doses less than 20 mg, significant differences in the C/D of haloperidol (P = 0.003), but not of reduced haloperidol, were found between the three major genotype groups. In patients with doses higher than 20 mg, no differences were found between the genotype groups for either haloperidol or reduced haloperidol. 68 patients (57%) used benztropine, an antimuscarinic agent. All four patients with a *5 allele (one together with *1 and three with *10) were found to use benztropine. The patients homozygous for the *1 allele seemed to need less benztropine than the patients with one or two mutated alleles (Fisher's exact test; P = 0.036). CONCLUSIONS: The dose-corrected steady state plasma concentrations of haloperidol, but not of reduced haloperidol, were significantly different between the CYP2D6*1/*1, *1/*10 and *10/*10 genotype groups when doses lower than 20 mg haloperidol were given. No differences were found at higher doses. These results suggest the involvement of CYP2D6 in the metabolism of haloperidol at low doses of haloperidol (< 20 mg daily), while another enzyme, probably CYP3A4, contributes at higher doses.

Risperidone metabolism in relation to CYP2D6*10 allele in Korean schizophrenic patients.

OBJECTIVE: Risperidone is known to be biotransformed to its active metabolite, 9-hydroxyrisperidone, by the polymorphic CYP2D6 in Caucasians. This study aimed to investigate the relationship between the CYP2D6*10 allele and the plasma levels of risperidone and 9-hydroxyrisperidone in Korean schizophrenic patients. METHODS: Eighty-two Korean schizophrenic patients in monotherapy with oral doses of risperidone from 1 mg/day to 8 mg/day (mean +/- SD 4.3 +/- 1.9, median 4) participated in this study. Plasma concentrations of risperidone and 9-hydroxyrisperidone were analyzed using high-performance liquid chromatography. The CYP2D6*10 allele, which contains C188T mutation in exon 1, was identified using allele-specific polymerase chain reaction amplification. RESULTS: Seventeen of 82 patients were homozygous for CYP2D6*1, 22 for *10, while the remaining 43 patients were heterozygous for these alleles. The plasma levels of risperidone and 9-hydroxyrisperidone ranged from 1.0 nM to 168 nM and 6.2 nM to 235 nM, respectively. The median concentrations/dose (C/Ds) (range) of risperidone in CYP2D6*1/*1, *1/*10, and *10/*10 groups were 1.7 (0.2-7.9), 2.6 (0.3-27.1), and 6.7 nM/mg (2.4-21.0), respectively. There was a statistically significant difference among the three genotypes (Kruskal-Wallis test, P<0.001). For 9-hydroxyriperidone, the corresponding median C/Ds were 13.1 (3.3-25.4), 11.9 (4.2-30.8), and 13.6 nM/mg (6.5-52.8), respectively, with no significant difference between the genotypes (P=0.54). The medians of the ratios between risperidone and 9-hydroxyrisperidone concentrations were 0.13 (0.01-0.93), 0.28 (0.01-2.77), and 0.46 nM/mg (0.05-1.28) in *1/*1, *1/*10, and *10/*10 genotypes, respectively, and they were significantly different (P=0.004). The active moieties (sum of the C/Ds of risperidone and 9-hydroxyrisperidone) were not significantly different between the genotypes (P=0.063). CONCLUSION: In Korean schizophrenic patients, the metabolism of risperidone is dependent on CYP2D6, and the CYP2D6*10 allele is important for the regulation of the activity of this enzyme. There were no significant differences in the plasma concentration of parent drug plus its active metabolite between the genotypes. This suggests that the clinical significance of this polymorphism is limited. Our study confirms previous studies on risperidone metabolism in Caucasians.

Oxidation of ranitidine by isozymes of flavin-containing monooxygenase and cytochrome P450.

Rat and human liver microsomes oxidized ranitidine to its N-oxide (66-76%) and S-oxide (13-18%) and desmethylranitidine (12-16%). N- and S-oxidations of ranitidine were inhibited by metimazole [flavin-containing monooxygenase (FMO) inhibitor] to 96-97% and 71-85%, respectively, and desmethylation of ranitidine was inhibited by SKF525A [cytochrome P450 (CYP) inhibitor] by 71-95%. Recombinant FMO isozymes like FMO1, FMO2, FMO3 and FMO5 produced 39, 79, 2180 and 4 ranitinine N-oxide and 45, 0, 580 and 280 ranitinine S-oxide pmol x min(-1) x nmol(-1) FMO, respectively. Desmethyranitinine was not produced by recombinant FMOs. Production of desmethylranitidine by rat and human liver microsomes was inhibited by tranylcypromine, a-naphthoflavon and quinidine, which are known to inhibit CYP2C19, 1A2 and 2D6, repectively. FMO3, the major form in adult liver, produced both ranitidine N- and S-oxides at a 4 to 1 ratio. FMO1, expressed primarily in human kidney, was 55- and 13-fold less efficient than the hepatic FMO3 in producing ranitidine N- and S-oxides, respectively. FMO2 and FMO5, although expressed slightly in human liver, kidney and lung, were not efficient producers of ranitidine N- and S-oxides. Thus, urinary contents of ranitidine N-oxide can be used as the in vivo probe to determine the hepatic FMO3 activity.

The effects of E2 supplementation from the early proliferative phase to the late secretory phase of the endometrium in hMG-stimulated IVF-ET.

PURPOSE: Our purpose was to determine if pregnancy rates (PRs) for hMG (human menopausal gonadotropin)-stimulated IVF-ET (in vitro fertilization--embryo transfer) can be increased by estradiol (E2) supplementation from the early proliferative phase to the late secretory phase of the endometrium. METHOD: Eighty-one infertile women with pure tubal factor were randomized into two groups. One group received no E2 supplementation (control group) and the other received oral E2 supplementation (2 mg two times daily) from the early proliferative phase starting on the third day of the menstrual cycle to the late secretory phase of the endometrium, with hMG stimulation for ovulation induction starting on the sixth day of the menstrual cycle. RESULTS: In 85 cycles, at least one embryo was transferred. Compared with the control group (n = 27 cycles), the E2 supplementation group (n = 58 cycles) had a significantly higher PR (control, 25.9%, versus E2 supplementation, 48.3%) and IR per ET (control, 10%, versus E2 supplementation, 26%), but FRs per retrieved oocytes were not statistically different between the two groups (control, 74%, versus E2 supplementation group, 73%). Four spontaneous abortions occurred in the E2 supplementation group, and one case in the control group. Ectopic pregnancy occurred in one case in the control group. CONCLUSIONS: Clinical PRs and IRs in the E2 supplementation group were significantly higher than in the control group, while FRs in the control group did not differ statistically from the E2 supplementation group. This suggests that E2 supplementation from the early proliferative phase to the late secretory phase of the endometrium in hMG-stimulated IVF-ET increases the receptivity of the endometrium for transferred embryos and clinical PRs.

Phenotypes of flavin-containing monooxygenase activity determined by ranitidine N-oxidation are positively correlated with genotypes of linked FM03 gene mutations in a Korean population.

A non-invasive urine analysis method to determine the in-vivo flavin-containing mono-oxygenase (FMO) activity catalysing N-oxidation of ranitidine (RA) was developed and used to phenotype a Korean population. FMO activity was assessed by the molar concentration ratio of RA and RANO in the bulked 8 h urine. This method was used to determine the FMO phenotypes of 210 Korean volunteers (173 men and 37 women, 110 nonsmokers and 100 smokers). Urinary RA/RANO ratio, representing the metabolic ratio and the reciprocal index of FMO activity, ranged from 5.67-27.20 (4.8-fold difference) and was not different between men and women (P = 0.76) or between smokers and nonsmokers (P = 0.50). The frequencies of RA/RANO ratios were distributed in a trimodal fashion. Among the 210 Korean subjects, 93 (44.3%) were fast metabolizers, 104 (49.5%) were intermediate metabolizers and 13 (6.2%) were slow metabolizers. Subsequently, the relationship between the ranitidine N-oxidation phenotypes and FMO3 genotypes, determined by the presence of two previously identified mutant alleles (Glu158Lys: FMO3/Lys158 and Glu308Gly: FMO3/Gly308 alleles) commonly found in our Korean population was examined. The results showed that subjects who were homozygous and heterozygous for either one or both of the FMO3/Lys158 and FMO3/Gly308 mutant alleles had significantly lower in-vivo FMO activities than those with homozygous wild-type alleles (FMO3/Glu158 and FMO3/Glu308) (P < 0.001, Mann-Whitney U-test). Furthermore, the FMO activities of subjects with either FMO3/Lys158 or FMO3/Gly308 mutant alleles were almost identical to those having both FMO3 mutant alleles (FMO3/Lys158 and FMO3/Gly308). These two mutant alleles located, respectively, at exons 4 and 7 in the FMO3 gene appeared to be strongly linked by cis-configuration in Koreans. Therefore, we concluded that presence of FMO3/Lys158 and FMO3/Gly308 mutant alleles in FMO3 gene is responsible for the low ranitidine N-oxidation (FMO3 activity) in our Korean population.

Phenotyping of flavin-containing monooxygenase using caffeine metabolism and genotyping of FMO3 gene in a Korean population.

Flavin-containing monooxygenase (FMO) activity was determined in 82 Korean volunteers by taking molar concentration ratio of theobromine and caffeine present in the 1 h urine (between 4 and 5 h) samples collected after administration of a cup of coffee containing 110 mg of caffeine. Among 82 volunteers, there were 19 women and 63 men (30 smokers and 52 non-smokers). Volunteers were divided into two groups comprising low (0.53-2.99) and high (3.18-11.95) FMO activities separated by an antimode of 3.18. Peripheral bloods were sampled from these volunteers and their genomic DNAs were amplified by polymerase chain reaction with oligonucleotides designed from intronic sequences of human FMO3 gene. Comparing nucleotide sequences of the amplified FMO3 gene originating from randomly selected individuals with low and high FMO activities, nine point mutations were identified in the open reading frame sequences. Among these nine mutations, three FMO3 mutant types (FMO3/Stop148, Lys158 and Gly308) were selected and correlated with FMO activities observed in our Korean population. A rare FMO3/Stop148 mutant allele originating from FMO3/Gly148 occurred by substitution of G442T in exon 4 and yielded a premature TGA stop codon. The stop codon was detected in one individual having the second lowest FMO activity and he had the mutation in heterozygous state. In a pedigree study, he was found to have inherited the mutation from his mother who also had a heterozygous stop codon and equally low FMO activity. In our volunteers, two other common mutations were detected in exons 4 and 7. The one in exon 4 resulted from a G472A change eliminating a HinfI restriction site and produced an amino acid substitution from Glu158 to Lys. The other mutation in exon 7 resulted from an A923G change generating a DraII restriction site and produced a non-conservative replacement of Glu308 to Gly. Based on the secondary structure maps of FMO3 enzyme proteins for these two mutant types, FMO3/Gly308 mutation transformed the helix structure into a sheet shape and indicated that dysfunctional FMO3 may be produced. FMO3/Lys158 mutation did not alter the secondary structure. Approximately 80% of volunteers with homozygous and/or heterozygous mutations on either one or two of these mutations had low FMO activities. Thus, individuals with these FMO3 gene mutations may have defective metabolic activity for many clinically used drugs and dietary plant alkaloids which are oxidized primarily by hepatic FMO3.

Decreased formation of ethoxyacetic acid from ethylene glycol monoethyl ether and reduced atrophy of testes in male rats upon combined administration with toluene and xylene.

Male painters are commonly exposed to ethylene glycol monoethyl ether (EGE), a well known reproductive toxic agent causing testicular atrophy, in the form of solvent mixture containing toluene (TOL) and xylene (XYL). This study was carried out to determine the effect of exposing male rats to solvent mixture containing TOL and XYL on the EGE (200 mg/kg) on testicular atrophy and production of toxic metabolite, ethoxyacetic acid (EAA) from EGE. Compared to the extent of testes atrophy observed upon EGE administration alone, the combined administration of EGE (200 mg/kg) with TOL (250 mg/kg) and XYL (500 mg/kg) for 4 weeks has reduced the extent of testes atrophy by 25%. The combined administration delayed the time for appearance of the highest plasma concentration (t(max)) of EAA from 3 to 6 h and also decreased the highest concentration (Cmax) as well as the total amount of plasma EAA (AUC(0-18 h)) by 45 and 29%, respectively. This explained the diminished testicular atrophy in male rats observed when EGE was administered in a solvent mixture containing TOL and XYL. This study suggested that testicular toxicity observed in male painters caused by EGE may be decreased when they are exposed to EGE in the form of solvent mixture containing TOL and XYL.

Lack of association between debrisoquine 4-hydroxylase (CYP2D6) gene polymorphisms and schizophrenia.

Debrisoquine 4-hydroxylase (CYP2D6) is a cytochrome P450 enzyme involved in the metabolism of most neuroleptics, which are the drugs of choice for the treatment of psychotic symptoms. CYP2D6 in the brain was suggested to be functionally similar to the dopamine transporter, thus possibly influencing a neurotransmitter system involved in schizophrenia. Swedish schizophrenic patients (n = 124) and control individuals (n = 85) were investigated for two CYP2D6 polymorphisms, responsible for approximately 90% of mutations leading to poor debrisoquine metabolism. No significant CYP2D6 allele or genotype difference was found between schizophrenic patients and control individuals. Taken together with previous results, no major effect appears to be caused by the CYP2D6 gene on schizophrenia.

Involvement of CYP3A1, 2B1, and 2E1 in C-8 hydroxylation and CYP 1A2 and flavin-containing monooxygenase in N-demethylation of caffeine; identified by using inducer treated rat liver microsomes that are characterized with testosterone metabolic patterns.

Caffeine (CA) is oxidized by rat liver microsomal enzymes to theobromine (TB), paraxanthine (PX), and theophylline (TP) by N-demethylation and to trimethylurate (TMU) by C-8 hydroxylation, In order to identify the specific enzymes responsible for productions of these primary CA metabolites, liver microsomes enriched with various isoforms of cytochrome P450 (CYP) and flavin-containing monooxygenase (FMO) are prepared by pretreatment of rats with several inducers. The specific increases in various CYP or FMO activities are identified with the diagnostic testosterone metabolic patterns or the thiobenzamide S-oxidation assay. They are then employed to metabolize the CA. Liver microsomes isolated from rats pretreated with phenobarbital (PB-microsomes) did not have increased FMO activity but had increased activities for hydroxylating the testosterone at 6 beta-(CYP3A1), 16 beta-(CYP2B1), and 2 beta-(CYP3A1) positions. This PB-microsomes had increased activity for TMU production from CA (result of C-8 hydroxylation). Liver microsomes isolated from rats pretreated with acetone (AC-microsomes) had a normal level of FMO activity but had enhanced rates of 6 beta-(CYP3A1) and 2 beta-(CYP3A1) hydroxylations of testosterone. The AC-microsomes again had increased activity for production of TMU. Similarly, the liver microsomes isolated from rats pretreated with dexamethasone (DEX-microsomes) had a normal level of FMO activity but had enhanced rates of forming 6 beta-and 2 beta-hydroxytestosterone (Cyp3A1) as well as androstenedione (CYP3A1). The DEX-microsomes again had increased activity for production of TMU only. Liver microsomes isolated from rats pretreated with 3-methylcholanthrene (MC-microsomes), however, had increased FMO activity and also enhanced rates of forming the 7 alpha-(CYP1A1/2, and 2A1), 6 beta-(CYP3A1), and 2 beta-(CYP3A1) hydroxytestosterone. The MC-microsomes had increased activity for producing all of the four primary metabolites of CA, i.e. the N-demethylation metabolites like TB, PX. and TP, as well as the C-8 hydroxylation metabolite TMU. By the process of association of the obtained results, liver microsomes with increased contents of CYP2B1, 3A1, and 2E1 could catalyze the C-8 hydroxylation at an increased rate producing increased amount of TMU. Increased productions of CA N-demethylation metabolites (TB, PX, and TP) are, however, catalyzed by the increased activities of CYP1A2 and FMO which are associated uniquely with the MC-microsomes.

Induction of flavin-containing monooxygenase (FMO1) by a polycyclic aromatic hydrocarbon, 3-methylcholanthrene, in rat liver.

Flavin-containing monooxygenase (FMO), known not to be induced by xenobiotics, has been induced by a polycyclic aromatic hydrocarbon, 3-methylcholanthrene (3MC). We have found a prominent augmentation of hepatic FMO1 both at transcription and translation levels by pretreatment of rats with 3MC. Liver tissues were used to study the inductive effect of 3MC on the FMO1 isoform, the major form present in rat liver. Evidence for significant induction of rat FMO1 was observed in mRNA production (3.5 fold) identified from reverse transcription-polymerase chain reaction (RT-PCR) results. Induction was also seen in the catalytic activity of the enzyme (2.9 fold) as measured by the thiobenzamide S-oxidation assay using induced rat liver microsomes. Our finding is the first report to indicate that hepatic FMO1 can be induced with a polycyclic aromatic hydrocarbon compound. FMO plays crucial roles in the oxidation of N- and S-containing drugs. If FMO is also inducible with other environmental polyaromatic hydrocarbon compounds in general, this finding will have important consequences in understanding the altered half-lives of many clinically useful drugs.

CYP2C19 genotype and phenotype determined by omeprazole in a Korean population.

Omeprazole (20 mg orally) was given to 103 healthy Korean subjects and blood was taken 3 h after administration. The plasma concentration ratio of omeprazole and hydroxyomeprazole, used as an index of CYP2C19 activity, was bimodally distributed. Thirteen subjects (12.6%) were identified as poor metabolizers (PMs) with an omeprazole hydroxylation ratio of 6.95 or higher. Among the 206 CYP2C19 alleles, CYP2C19*2 and CYP2C19*3 were found in 43 alleles (21%) and 24 alleles (12%), respectively. Twelve subjects (12%) carried two defect alleles (*2/*2, *2/*3 or *3/*3), 43 subjects (42%) were heterozygous for a mutated (*2 or *3) and a wild type (*1) allele, and the remaining 48 subjects (47%) were homozygous for the wild type allele. The distributions of the metabolic ratio between these three genotype groups were significantly different (Kruskal-Wallis test: p < 0.0001). The genotypes of 19 additional Korean PMs has been identified in a previous mephenytoin study. From a total of 32 PMs, 31 were genotypically PMs by analysis of the CYP2C19*2 and *3 alleles and only one PM subject was found to be heterozygous for the *1 and *2 alleles. At present it cannot be judged whether this subject has a defective allele with a so-far unidentified mutation or a true wild type allele. We thus confirm a high incidence (12.6%) of PMs of omeprazole in Koreans and of the 32 Korean PMs 97% could be identified by the genotype analysis.

Acceleration of the alcohol oxidation rate in rats with aloin, a quinone derivative of Aloe.

Aloe contains abundant aloin, a C-glycoside derivative of anthraquinone. Based on recent reports indicating that the water extract of Aloe enhances the ethanol oxidation rate and also that quinones, in general, have a functional role in elevating the alcohol oxidation rate in vivo, we have attempted to identify the quinone derivative contained in Aloe that could increase the alcohol oxidation rate. Upon oral administration of aloin (300 mg/kg) given 12 hr prior to the administration of alcohol (3.0 g/kg), the blood alcohol area under the curve (AUC) was found to be decreased significantly (by 40%). This was supported by increases in the rates of blood alcohol elimination and the disappearance of alcohol from the body by 45 and 50%, respectively. Analysis of hepatic triglyceride (TG) levels revealed that both the ethanol and the aloin treatment alone significantly increased the TG levels in a comparable manner; however, the level obtained by the combined treatment of aloin and ethanol was not statistically different from that produced by either treatment alone. The levels of serum L-aspartate:2-oxoglutarate aminotransferase (AST) and L-alanine:2-oxoglutarate aminotransferase (ALT) activities were not increased by acute alcohol intoxication, aloin alone, or by the combined treatment of alcohol and aloin. Pretreatments with aloe-emodin, the anthraquinone aglycone of aloin, resulted in a significantly decreased blood alcohol AUC and an increase in the rate of ethanol disappearance. These results suggested that when the aloin localized primarily in the skin of Aloe is ingested, aloe-emodin (the quinone aglycone) may be released, and the released quinone may produce acceleration of the ethanol metabolism rate in vivo.

Debrisoquine and S-mephenytoin hydroxylation phenotypes and genotypes in a Korean population.

One hundred and fifty-two healthy Korean volunteers were phenotyped with debrisoquine and mephenytoin and genotyped with respect to CYP2D6. The debrisoquine metabolic ratio (MR) varied between 0.09 and 6.3, and all subjects were thus classified as extensive metabolizers of debrisoquine. Polymerase chain reaction (PCR)-based amplification of genomic DNA with primers specific for the C188-->T mutation present in exon 1 of the CYP2D6*10B allele was performed and revealed an allele frequency of 0.51 in this Korean population. Forty-three subjects (28%) were homozygous for CYP2D6*10B, 69 subjects (45%) were heterozygous for this allele, while in 40 subjects (26%) no exon 1 mutation could be found. All subjects except one homozygous for the wild type allele had MRs below 0.75 whereas the MR was higher than 0.99 in all subjects homozygous for the CYP2D6*10B allele. The MRs in the three genotype groups were significantly different (p < 0.0001; Kruskal-Wallis test). Eco RI RFLP analysis of DNA from six subjects with debrisoquine MRs < or = 0.11 revealed that only one (MR 0.09) carried a duplicated CYP2D6*Z-gene (CYP2D6*2X2) as indicated by the Eco RI 12.1 kb haplotype. It is concluded that, as shown earlier for Chinese and Japanese populations, the CYP2D6*10B-allele containing the C188-->T mutation is the major cause of diminished CYP2D6 activity in Koreans. In this Korean population, the MR of debrisoquine was shifted towards higher values (lower CYP2D6 activity) compared with Caucasian populations but the shift appeared to be less pronounced than earlier shown for Chinese. Twenty-four subjects (16%) were poor metabolizers of S-mephenytoin as indicated by the S/R mephenytoin ratio of about 1. Twenty-three of these were genotyped with respect to the defect CYP2C19-alleles CYP2C19*2 and CYP2C19*3. Of the 46 poor metabolizer alleles, 32 (70%) were CYP2C19*2 and the remaining 14 (30%) were CYP2C19*3. Thus, the defect CYP2C19*2 and CYP2C19*3-alleles explained 100% of the 23 Korean poor metabolizers of S-mephenytoin.

Genetic analysis of the CYP2D locus in relation to debrisoquine hydroxylation capacity in Korean, Japanese and Chinese subjects.

The CYP2D6 genotype and the debrisoquine and mephenytoin hydroxylation phenotypes were studied in 63 Oriental subjects including 21 Chinese, 21 Japanese and 21 Koreans. All subjects were extensive metabolizers of debrisoquine. The incidence of the S-mephenytoin poor metabolizer phenotype was 14% in the Chinese, 24% in the Japanese and 24% in the Korean population, respectively, which is similar to previous reports. The CYP2D6 genotype was analysed by Xba I and Eco RI RFLP, and by allele-specific PCR analysis for the presence of several allelic variants of the CYP2D locus. No CYP2D6A or CYP2D6B alleles, two of the most common defect alleles among Caucasians, were found among the Oriental subjects. The frequency of the CYP2D6D allele was similar to that in Caucasian populations and consistent with the low incidence of the poor metabolizer phenotype in all three Oriental populations. The CYP2D6L2-allele with duplication of an active CYP2D6L gene was identified in one Korean and one Chinese allele in association with high CYP2D6 activity. The CYP2D6Ch alleles CYP2D6Ch1 and Ch2, identified by RFLP and PCR for the -1338C-->T and 188C-->T mutations, were the most frequent allelic variants in all three populations studied, and were related to a decreased CYP2D6 activity as previously shown in Chinese. In conclusion, the present pilot study revealed major similarities in the polymorphic CYP2D locus between Korean, Japanese and Chinese populations.

Induction of physical dependence by chronic alcohol treatment and enhancement of hepatic metabolism of 7-ethoxycoumarin and subsequent conjugation of its metabolite in perfused rat livers.

Physical dependence to alcohol was induced in rats and the rates of hepatic oxidation of 7-ethoxycoumarin to 7-hydroxycoumarin and of the subsequent conjugation of 7-hydroxycoumarin were studied using isolated perfused livers. Seven to 11.2 g of ethanol per kg body weight per day were given to rats in 2 divided doses at 10 a.m. and at 8 p.m. for 7, 9, 11, and 13 consecutive days. All rats surviving these courses of ethanol administration have demonstrated moderate degrees of withdrawal signs. In the isolated perfused livers obtained from these alcohol dependent rats, 100 microM 7-ethoxycoumarin was infused. The rate of 7-hydroxycoumarin formation was increased from 36 to 50 nmoles per min per g of liver tissue. This increase has occurred without significant changes of the liver weights. With such increase in the rate of mixed function oxidation, greater amounts of the 7-hydroxycoumarin metabolite were conjugated not only to glucuronide (from 4.9 to 6.7 nmoles), but also to sulfate ester (from 27.6 to 42.1 nmoles). Furthermore, a greater percentage of the produced metabolite was conjugated in the ethanol treated rat liver. A high concentration of 7-hydroxycoumarin (100 microM) was also infused in order to determine the effect of chronic alcohol administration on the maximal available rates of conjugation. Results indicated that the maximal rates of conjugation were significantly decreased. However, these decreased conjugation rates were still much greater than the maximal production rate for 7-hydroxycoumarin from the infused 7-ethoxycoumarin, for which the rate of oxidation has been elevated by the chronic ethanol treatment. Thus, the livers obtained from rats chronically treated with alcohol have increased rates for both the initial drug oxidation and the subsequent conjugation, even though the maximal rates of conjugation, per se, are reduced.