CYP3A Polymorphism and Chronic Mercury Intoxication.

We analyzed the relationship between polymorphic loci of CYP3A genes (CYP3A4 (rs2740574), CYP3A5 (rs776746) and CYP3A7 (rs2257401)) with the development of chronic mercury intoxication. Of 170 men examined, 120 were workers chronically exposed to mercury vapors and 50 were carriers of GG-HSPA1B (+1267A/G) genotype associated with chronic mercury intoxication. Urinary content of 4-hydroxyantipyrine (4-HAP) generated in the reaction predominantly catalyzed by CYP3A4/CYP3A5 was studied in workers without chronic mercury intoxication (group 1, N=46) and patients in the delayed period of chronic mercury intoxication (group 2, N=74) depending on the genotypes of CYP3A4 and CYP3A5. For polymorphic loci CYP3A5 and CYP3A7, a tendency to an increase in the frequency of genotypes with rare alleles was found (p=0.071 and p=0.078) in the combined group (group 2 together with GGHSPA1B genotype carriers) relative to group 1. The high level of linkage disequilibrium was noted, especially for the pair rs776746 and rs2257401 (LD (r)=0.89). In group 2, a trend to 4-HAP decrease compared to group 1 (p=0.056 and p=0.065) was revealed for carriers of AA-CYP3A4 and GG-CYP3A5 genotypes. The involvement of CYP3A in the development of mercury neurotoxic effect remains unclear.

Impact of AhRR (565C > G) polymorphism on dioxin dependent CYP1A2 induction.

The current study uses the metabolic probe, antipyrine, and AhRR transcript expression (qRT-PCR) to examine the impact of the AhRR (565C > G or Pro185Ala, rs2292596) genetic polymorphism upon CYP1A2 inducibility in an established cohort of male firefighters with exposure to dioxin-like chemicals. The lipid adjusted concentrations of 29 dioxin and dioxin-like congeners were measured in serum. Possession of the G allele (CG and GG genotypes) was correlated with high expression AhRR transcript and lower CYP1A2 induction than found in individuals homozygous for CC. The induction of CYP1A2 was dioxin-dependent among carriers of the G allele. Multivariate models indicated that CYP1A2 activity, detected as urinary 3-hydroxymethylantipyrine, was significantly correlated with cotinine concentration and for those currently working as firefighters, dioxin body burden (ß = 0.54, p = 0.041). The efficacy of the AhRR in regulating the AhR signaling pathway is influenced by the AhRR (565C > G) polymorphism. Our study of firefighters using the induction of CYP1A2 as an indicator suggest that G allele proteins have variable AhR repressor activity which is manifested in a dioxin-dependent manner. These results provide evidence of metabolic differences that may affect susceptibility to dioxin-mediated health effects.

Monogenic control of variations in antipyrine metabolite formation. New polymorphism of hepatic drug oxidation.

To investigate mechanisms that control large variations among normal uninduced subjects in the elimination of the model compound antipyrine (AP) and other drugs, AP was administered to 144 subjects (83 unrelated adults and 61 members of 13 families). Thereafter, at regular intervals for 72 h, the urine of each subject was collected and concentrations of AP and its three main metabolites measured. From these urinary concentrations, rate constants for formation of each AP metabolite were calculated. Trimodal curves were observed when values for each AP rate constant were plotted in 83 unrelated subjects; probit plots of these values showed inflections at the two antimodes of each trimodal distribution. All members of our 13 families were assigned one of three phenotypes determined by where their AP metabolite rate constant placed them in the trimodal distributions derived from the 83 unrelated subjects. In each family, pedigree analysis to identify the mode of transmission of these three phenotypes was consistent with their monogenic control. These results provide evidence for a new polymorphism of drug oxidation in man.

Pharmacokinetics of drugs in patients with the nephrotic syndrome.

Since the binding of drugs to plasma proteins can significantly after the intensity of pharmacological and toxicological effects of drugs, we studied the pharmacokinetics of three drugs in patients with hypoalbuminemia secondary to the nephrotic syndrome, but with relatively normal renal function. No significant differences were seen in the pharmacokinetic parameters observed for antipyrine, a drug which is less than 10% bound to plasms proteins. The percentage of unbound diphenylhydantoin, a highly plasms protein-bound drug, was found in patients with the nephrotic syndrome to be twice that of healthy individuals (19,2 vs. 10.1%, P smaller than 0.001). However, there was also a lower steady-state plasma concentration of diphenylhydantoin (2.9 plus or minus 0.6 vs. 6.8 plus or minus 0.6 mug/ml, P smaller than 0.001) secondary to an increase in the plasms clearance (0.048 plus or minus 0.019 vs. 0.022 plus or minus 0.006 liter/kg.h, P smaller than 0.001) in the nephrotic patients. The net effect is no difference in the absolute concentration of unbound diphenylhydantoin in healthy individuals (0.69 plus or minus 0.05 mug/ml) and patients with the nephrotic syndrome (0.59 plus or minus 0.06 mug/ml). Qualitatively, similar differences were observed with clofibrate. The dose of these drugs need not be routinely reduced in patients with the nephrotic syndrome as long as they have reasonably normal renal function (creatinine clearance greater than 50 ml/min). With all highly bound acidic drugs, knowledge of the concentration of unbound drug is essential to the proper interpretation of total blood levels and subsequent treatment of the patient.

Impact of dioxins on antipyrine metabolism in firefighters.

Antipyrine (AP) metabolism was used to assess factors associated with the activity of hepatic oxidative enzymes in firefighters. Emphasis was placed on 3-hydroxymethylantipyrine (3HMAP), the metabolite with the greatest dependence on dioxin-inducible cytochrome P4501A2 (CYP1A2) activity. AP urinary metabolites were measured by HPLC in 38 male subjects from Eastern Siberia. Subjects were divided into three groups having similar ages and BMIs: current firefighters (n=11); former firefighters (n=17) and non-firefighters (n=10). Multiple regression models were constructed using the three major AP metabolites as a dependent variable to assess the influence of age, smoking as urinary cotinine concentration, dioxin exposure (as either WHO-TEQ or body burden), group, and CYP1A2*F (-163C>A) genotypes. Models for the proportion of dose excreted as the metabolite 3HMAP produced the best fit (adjusted R(2)=0.46, p<0.05). When the models were restricted to current firefighters, only those based on 3HMAP were statistically significant (adjusted R(2) of 0.80 (p<0.002)) due to contributions from urinary cotinine (ß=0.56, p<0.01) and dioxin expressed as body burden (ß=0.55, p=0.014). These results indicate that the antipyrine test can be used as metabolic probe of biological response to recent dioxin exposure provided the impact of smoking is carefully controlled.

[Markers of impact and effects of dioxines in firemen who participated in fire extinguishing at "Irkutskcabel" plant].

The article contains data tracing connection between CYP1A2 activity, dioxines level and dioxine-sensitive genes expression in firemen who participated in fire extinguishing at cabel plant.

The absence of significant biliary excretion of antipyrine or its metabolites in humans.

Three patients with complete bile duct obstructions requiring a percutaneous biliary fistuala were given an oral dose of antipyrine. Drug elimination was assessed through plasma t1/2 studies and urine and bile excretion of both antipyrine and its metabolites. Urine metabolite patterns were in agreement with reference standards, but analysis of bile revealed no antipyrine metabolites and minimal parent compound (mean of total administered dose excreted from the bile fistulas was 4%). This finding was not predicted from previous experiments in the bile-cannulated rat and suggests caution regarding interspecies extrapolation of data concerning the hepatic disposition of certain commonly used test drugs in clinical pharmacologic studies.

Interaction between metronidazole and drugs eliminated by oxidative metabolism.

Single-dose kinetics of 0.1 mg/kg intravenous diazepam, 10 mg/kg oral antipyrine, and 300 mg oral phenytoin were followed in healthy subjects before and after 400 mg metronidazole twice a day for 5 days. When data before metronidazole dosing were compared with those after metronidazole dosing, there were no changes in total plasma clearance of diazepam (0.53 and 0.65 ml/min/kg), antipyrine (39.0 and 38.0 ml/min/kg), or phenytoin (0.56 and 0.55 ml/min/kg). Plasma t 1/2s and volumes of distribution of the three drugs tested were not affected by metronidazole, but urinary excretion of 4-hydroxyantipyrine decreased after metronidazole dosing. There was no change in the elimination of phenytoin as its hydroxylated metabolite after metronidazole. It is concluded that, at therapeutic concentrations, metronidazole does not significantly inhibit oxidative drug metabolism.

Cimetidine-induced reduction in gastrointestinal absorption of antipyrine and rate constants for formation of its metabolites.

In 15 normal men, cimetidine taken orally in a dose of 300 mg twice a day for 3 days reduced to similar extents the rate constants for formation (ki) of the three principal metabolites of antipyrine (AP): 29.9% +/- 8.5% (mean +/- SD) for 4-hydroxyantipyrine (4-OH-AP); 28.3% +/- 6.3% for 3-hydroxymethylantipyrine (3-OHM-AP); and 22.4% +/- 5.6% for N-demethylantipyrine (NDM-AP). AP clearance declined 24.3%; AP salivary t 1/2 rose 33%; and corrected AP apparent volume of distribution was unchanged. In one apparently normal subject, however, kis for formation of 3-OHM-AP and NDM-AP rose after cimetidine even though AP clearance declined 19.7%. This surprising result, which suggests that cimetidine can exert an inductive effect on the hepatic mixed-function oxidases of some subjects, was checked by restudying the individual. Very similar values occurred on repetition. The average increase in kis for NDM-AP and 3-OHM-AP was 172.2% and 34.0%. These unusual results in this subject indicate that at least two distinguishable forms of cytochrome P-450 participate in AP metabolism in man. Cimetidine appeared to reduce the amount of AP absorbed from the gut in 10 of our 15 normal subjects.

Sources of interindividual variations in acetaminophen and antipyrine metabolism.

Our goal was to compare and contrast in the same normal twins the relative contribution of genetic and environmental factors to large interindividual variations in the metabolism of acetaminophen (APAP) and antipyrine. These drugs were selected because they are biotransformed by different mechanisms. A single oral dose of APAP (10 mg/kg) was given to six sets of monozygotic (MZ) and six sets of dizygotic (DZ) twins. All were normal, nonsmoking, nonmedicated, and male. Among these 24 subjects, there were 300% interindividual variations in rate constants for formation of the sulfate and glucuronide conjugates, as well as in the overall rate constant for APAP elimination. Intratwin variations for each measurement were as large within MZ as within DZ twinships, suggesting that predominantly environmental rather than genetic factors maintained interindividual variations. Two other observations support this conclusion: Intraindividual variations were frequently as large as interindividual variations, and regardless of zygosity for twins living together, intratwin correlation coefficients were almost twice those of twins living apart. Quite different results were obtained when these twins received antipyrine. After a single oral dose of antipyrine (18 mg/kg), 500% interindividual variations in rate constants for formation of the three main oxidative metabolites of antipyrine appeared to be mainly under genetic control. Also for antipyrine and its principal metabolites, intraindividual variations were much smaller than interindividual variations. In contrast to the results with APAP, regardless of zygosity, intratwin correlation coefficients for antipyrine were similar for twins living apart and twins living together. This comparison between APAP and antipyrine metabolism in the same carefully selected normal twins under apparently uniform environmental conditions reveals that interindividual variations in APAP metabolism arise from certain unidentified environmental factors, whereas genetic factors cause the large interindividual variations that occur in antipyrine disposition.

The effect of zileuton on antipyrine and indocyanine green disposition.

The effects of single and multiple oral doses of zileuton on the pharmacokinetics of antipyrine and indocyanine green were studied in 16 healthy, nonsmoking adult men by means of a double-blind, randomized, parallel placebo-controlled design. Indocyanine green disposition was not significantly altered by zileuton. Plasma antipyrine clearance declined by 20% (p < 0.0005) and 52% (p < 0.0005) after single and multiple dose zileuton exposure, respectively. Total urinary recovery of unchanged antipyrine and metabolites decreased with zileuton exposure. Selective declines from baseline of 16% (p = 0.007) and 20% (p = 0.003) after single-dose zileuton and 30% (p < 0.0005) and 43% (p < 0.0005) after multiple-dose zileuton were detected in recovery of 4-hydroxyantipyrine and 3-hydroxymethylantipyrine, respectively. Urinary recovery of the N-demethylantipyrine metabolite norantipyrine and percent of conjugation of 3-hydroxymethylantipyrine were unchanged by zileuton. In conclusion, zileuton therapy has no detectable effect on indocyanine green disposition but exerts marked effects on antipyrine plasma and urine metabolite disposition.

Hepatic drug clearance in patients with mild cystic fibrosis.

The plasma disposition of three model substrates (lorazepam, indocyanine green, and antipyrine) and the formation clearance of antipyrine metabolites (3-hydroxymethylantipyrine, norantipyrine, and 4-hydroxyantipyrine) were evaluated in 15 subjects with mild cystic fibrosis and in 15 healthy control subjects. Plasma clearance was significantly greater in patients with cystic fibrosis for both lorazepam (1.7 +/- 0.4 versus 1.2 +/- 0.5 ml/min/kg) and indocyanine green (14.2 +/- 6.1 versus 9.1 +/- 3.0 ml/min/kg). In contrast, the clearance of antipyrine was not significantly different (1.0 +/- 0.7 versus 0.8 +/- 0.3 ml/min/kg), but the formation clearance for 3-hydroxymethylantipyrine was significantly greater in patients with cystic fibrosis. Lorazepam and antipyrine apparent steady-state volume of distribution were not different between groups. These results suggest that clearance of drugs that undergo conjugation (e.g., lorazepam) or biliary excretion (e.g., indocyanine green) is increased in patients with mild cystic fibrosis. In contrast, the increased formation clearance of only one antipyrine metabolite suggests that alterations in clearance of drugs metabolized by cytochrome P450 enzymes are substrate specific and isoform specific in patients with cystic fibrosis.

Lack of effect of influenza vaccine on the pharmacokinetics of antipyrine, alprazolam, paracetamol (acetaminophen) and lorazepam.

31 healthy male (n = 17) and female (n = 14) volunteers, aged 20 to 45 years, were divided into 4 groups and received on 3 separate occasions either: paracetamol (acetaminophen) 650mg intravenously (n = 9); alprazolam 1mg orally (n = 7); antipyrine (phenazone) 1g intravenously (n = 8); or lorazepam 2mg intravenously (n = 7). Doses were administered prior to influenza vaccine (0.5ml, intramuscularly) and at 7 and 21 days post-vaccination. The overall differences among the 3 trials in clearance of antipyrine were of borderline significance (p less than 0.0611), with a trend towards reduced clearance in both of the post-vaccination trials. There were no overall differences observed in the elimination half-life of antipyrine, nor were there significant differences between trials in cumulative urinary excretion or fractional recovery of intact antipyrine, 4-hydroxyantipyrine, norantipyrine, or 3-hydroxymethyl antipyrine. For paracetamol and alprazolam, there were no significant differences among the 3 trials in any of the kinetic variables. The elimination half-life of lorazepam varied significantly among trials, but differences were small and not systematic. Lorazepam clearance did not vary significantly among trials. Thus, clearance of drugs which undergo hepatic conjugative reactions such as glucuronidation and sulphation are unlikely to be affected by the coadministration of influenza vaccine. Furthermore, not all drugs which are biotransformed by hepatic microsomal oxidation necessarily have impaired clearance due to coadministration of influenza vaccine.

The relationship between phenazone (antipyrine) metabolite formation and theophylline metabolism in healthy and frail elderly women.

The influence of aging on the metabolism of phenazone (antipyrine), and the relationship between the formation of 3 phenazone metabolites and the metabolic clearance of theophylline in healthy and frail elderly women, were examined. Whereas the elimination half-life did not change, clearance of phenazone decreased by about 50% with age in healthy women receiving phenazone without theophylline. However, the summation of the urinary recovery of phenazone and the measured metabolites, expressed as percentage of the phenazone dose, was lower in the healthy elderly (37 +/- 9% vs 74 +/- 15%). In both healthy and frail females the clearance of formation of 4-hydroxy-phenazone and the metabolic clearance of theophylline correlated strongly (r = 0.93 and 0.90, respectively). In non-healthy elderly females, strong correlations were also observed between the other metabolic pathways of phenazone and the metabolic clearance of theophylline. Coadministration of theophylline in the elderly increased the percentage of the phenazone dose excreted as the measured metabolites. A considerably higher interindividual variability in the disposition of phenazone and theophylline was observed in the frail elderly women. This high degree of variability in drug metabolism may be one of the explanations for the problems often occurring after drug prescription in the elderly.

Immediate decrease by hydrocortisone of the plasma half-life of antipyrine.

Infusion of hydrocortisone in man caused an immediate shortening of the plasma half-life of antipyrine. There was no change in the 'apparent' volume of distribution of antipyrine and the plasma concentrations of hydrocortisone during the infusion remained within physiological limits. Similar changes in plasma half-lives of antipyrine were observed in the dog, but in vitro studies of drug oxidation with dog liver failed to show any difference between biopsy samples taken before and during steroid infusion.Many drugs and chemicals are known to stimulate rates of drug oxidation in both man and animals. Administration of these inducing agents results in an increase in the activity of enzymes catalyzing drug oxidation, most of which are located with-in the endoplasmic reticulum of liver cells. In man there is usually a delay of 4-8 days before an appreciable change in rates of drug metabolism is seen when such an agent is administered (Breckenridge, Orme, Thorgeirsson, Davies & Brooks, 1971); in rats, the administration of enzyme-inducing agents causes increased rates of drug metabolism only 24-48 h after their administration.We wish to report in this paper an immediate and hitherto undescribed effect of hydrocortisone on rates of antipyrine elimination in man.

Metabolism of antipyrine in vivo in two rat models of liver cirrhosis. Its relationship to intrinsic clearance in vitro and microsomal membrane lipid composition.

Antipyrine metabolism depends on at least three isoenzymes of cytochrome P450 forming the main metabolites 3-OH-, 4-OH- and norantipyrine. We investigated to what extent antipyrine clearance and metabolite formation are impaired in two models of liver cirrhosis in the rat, namely micronodular cirrhosis induced by chronic exposure to phenobarbital/CCl4 and biliary cirrhosis induced by bile duct ligation. Salivary antipyrine clearance was decreased to a similar extent in cirrhosis induced by CCl4 and bile duct ligation (-35%). Clearance for production of 3-OH-antipyrine was decreased in both models, while 4-hydroxylation was maintained. Metabolic clearance of both 3-OH-antipyrine and 4-OH-antipyrine in vivo correlated with their clearance in vitro (r = 0.658 and r = 0.583) but not with that of norantipyrine. The microsomal cholesterol content was increased by 16% and 90% in CCl4 and bile duct-ligated cirrhotic rats (P < 0.001), respectively. Membrane fluidity, expressed as the ratio of phospholipids to cholesterol, correlated with the in vivo clearance for production of norantipyrine (r = 0.841) but not of 3-OH- or 4-OH-antipyrine, while clearance in vitro was not related to altered lipid composition. Our results demonstrate that the cytochrome P450 isoenzymes responsible for the different pathways of antipyrine metabolism are affected to different extents by cirrhosis. Alterations in intrinsic clearance explain only part of the loss of hepatocellular function. Altered lipid composition contributes to this loss of function but other factors, among them loss of hepatocytes and changes in microcirculation, could be more important determinants of the decrease in xenobiotic metabolism in cirrhosis.

Lack of effect of halofantrine on hepatic drug metabolism in the rat in vivo and in vitro.

The effect of the antimalarial drug halofantrine (Hf) on hepatic drug metabolism in the rat has been studied in vivo and in vitro using different model drug substrates. Hf in vitro produced no significant effect on the values of Km and Vmax for aminopyrine N-demethylation or 7-ethoxycoumarin O-dealkylation in microsomes incubated with Hf (0.01-0.1 mM) or on the rate of N-demethylation of aminopyrine or O-dealkylation of Ec in microsomes produced from rats dosed chronically with Hf (200 mg/kg) for 4 days. The disposition of antipyrine (Ap) was investigated in the isolated perfused rat liver preparation (IPRL). Following the administration of bolus doses of Hf (0.5, 2.5 and 5.0 mg) no significant changes were observed in the half-life (t1/2), clearance (Cl) or apparent volume of distribution (Vd) for Ap compared with controls. Pentobarbitone induced sleeping time was also assessed in mice. No significant difference was determined in time to recovery of the righting reflex for mice receiving Hf as single oral doses or chronically over 4 days when compared with appropriate controls. The potential for selective isoenzyme effects was studied in vivo. The three principal urinary metabolites of Ap, norantipyrine (Np), 3-OH and 4-OH Ap were measured in rat urine, with no significant change in urinary recovery of Ap or any of the metabolites in the presence of Hf (1.25 mg/kg i.p.) compared with controls. These results suggest that Hf is not, in contrast to many commonly used quinoline antimalarials, a potent or specific inhibitor of drug metabolism in vitro or in vivo.

Induction of cytochrome P-450 and related drug-oxidizing activities in muscone (3-methylcyclopentadecanone)-treated rats.

In the present study, we investigated the effects of muscone on both in vitro and in vivo parameters of the hepatic microsomal drug-metabolizing enzyme system and other enzyme activities in rats. In the in vivo study, the serum dimethadione (DMO)/trimethadione (TMO) ratios at 2 hr after oral administration of TMO (100 mg/kg) were significantly increased in both male and female rats treated with 75 and 150 but not 40 mg muscone/kg. Antipyrine metabolite profile in 24 hr urine of rats pretreated with muscone (150 mg/kg) was examined. The results showed that the excretion of norantipyrine was significantly increased as compared to the control group. In the in vitro study, we found that the content of cytochrome P-450, and activities of aminopyrine, N-demethylase, aniline hydroxylase and delta-aminolevulinic acid (ALA) synthetase were significantly increased as compared to the controls in both male and female rats treated with muscone (75 and 150 mg/kg). This type of induction of the hepatic metabolizing enzymes was similar to that seen after treatment with a prototype drug, phenobarbital.

The effect of age and sex on metabolism and urinary excretion of antipyrine.

BACKGROUND: Drug-metabolizing capacity is generally reduced in the elderly. The purpose of this investigation was to study antipyrine clearance and metabolite excretion in old subjects of both sexes. METHODS: Saliva clearance of antipyrine and the production clearances of antipyrine metabolites were studied in young and elderly volunteers of both sexes. Seventy-six elderly subjects (mean age 81 years) were compared with a group of 24 young subjects (mean age 29 years). RESULTS: After oral administration, salivary antipyrine clearance declined with age in both males and females, whether or not this variable was corrected for weight, and antipyrine half-life was significantly prolonged in elderly groups of either sex. The percentage urinary excretion of the antipyrine metabolites (hydroxymethylantipyrine, HMA; norantipyrine, NORA; and 4-hydroxyantipyrine, OHA) was reduced at 48 h in the elderly compared to young subjects by 23%, 31%, and 10%, respectively, in males, and by 41%, 41%, and 24%, respectively, in females. The formation clearance of HMA was reduced by 47% in males and by 52% in females. NORA clearance declined by 42 and 56%, respectively, in males and females. A decrease of 30% in males and 44% in females was observed in OHA clearance. CONCLUSIONS: The findings suggest that aging leads to altered disposition of antipyrine in both males and females and that the main metabolic pathways of the compound are not different in the elderly.

The effect of diltiazem on hepatic drug oxidation assessed by antipyrine and trimethadione.

The effect of pretreatment for 3 days with diltiazem 60 mg three times a day on the pharmacokinetics of 500-mg antipyrine and 250-mg trimethadione was studied in six healthy male subjects. Diltiazem decreased the total body clearance from 34.0 +/- 8.0 to 28.6 +/- 6.1 mL/min (P less than .01), and prolonged the elimination half-life from 12.6 +/- 3.0 to 14.3 +/- 2.5 hours (P less than .01) of antipyrine without any changes in volume of distribution. The cumulative renal excretion (% dose) of antipyrine was significantly increased from 2.23 +/- 0.73 to 2.78 +/- 0.83% (P less than .05). Clearances of production for three major antipyrine metabolites, norantipyrine (4.31 +/- 1.64 to 3.50 +/- 1.28 mL/min, P less than .01), 3-hydroxymethylantipyrine (4.67 +/- 1.63 to 3.82 +/- 1.34 mL/min, P less than .01) and 4-hydroxyantipyrine (10.47 +/- 3.41 to 8.16 +/- 2.82 mL/min, P less than .01) were reduced significantly by diltiazem. On the other hand, diltiazem did not produce any significant changes in pharmacokinetic parameters of trimethadione and plasma concentration ratio, oxidative major metabolite of trimethadione to trimethadione itself. These results suggest that other drugs metabolizing the same hepatic oxidative pathways as antipyrine, may be influenced by diltiazem.