Biotransformation of diclofenac sodium (Voltaren) in animals and in man. I. Isolation and identification of principal metabolites.

1. The anti-inflammatory agent diclofenac sodium (o-[(2,6-dichlorophenyl)amino]phenylacetic acid sodium salt) is extensively metabolized by rat, dog, baboon and man. The main metabolites were isolated from the urine of all species and from the bile of rat and dog and identified by spectroscopy. 2. Metabolism involves direct conjugation of the unchanged drug, or oxidation of the aromatic rings usually followed by conjugation. Sites of oxidation are either position 3' or 4' of the dichlorophenyl ring or, alternatively, position 5 of the phenyl ring attached to the acetic acid moiety. 3. In the urine of rat, baboon and man conjugates of the hydroxylated metabolites predominate, but the major metabolite in dog urine is the taurine conjugate of unchanged diclofenac. 4. In the bile of rat and dog, the main metabolite is the ester glucuroniade of unchanged diclofenac.     

Disposition and metabolism of benoxaprofen in laboratory animals and man.

1. The absorption, distribution, metabolism and excretion of benoxaprofen, a novel anti-inflammatory compound, has been studied in the dog, mouse, rat, rabbit, rhesus monkey and man. 2. Benoxaprofen was well absorbed after oral administration of doses of 1 to 10 mg/kg in all six species. Only unchanged drug was detected in plasma. It was extensively bound to plasma proteins, the highest binding occurring in man (99.8%) and rhesus monkey (99.6%). 3. Species differences were observed in the plasma elimination half-life, the longest being in man (33 h). The rat and mouse also had high values (28 and 24 h respectively) whereas in the other species, values were less than 13 h. 4. After an oral dose of [14C]benoxaprofen (20 mg/kg) to female rats, tissue concn. was highest in liver, kidney, lungs, adrenals and ovaries. Tissue distribution in the pregnant rat was identical to the normal female. The compound was found in the foetus but at a concn. lower than in all maternal organs. 5. There was a marked species difference in the route of excretion. In man, rhesus monkey and rabbit, excretion in the urine was a major route, whilst biliary--faecal excretion was the only effective route in the rat and dog. 6. No major metabolic transformation of benoxaprofen was observed. Man and dog excreted the compound predominantly as the ester glucuronide whereas the rat, mouse, rabbit and rhesus monkey excreted a large proportion of the dose unchanged.     

Biotransformation of zeranol: disposition and metabolism in the female rat, rabbit, dog, monkey and man

The disposition of [3H]zeranol has been studied in the female Wistar rat, New Zealand rabbit, beagle dog, rhesus monkey and man. The blood elimination half-life of total radioactivity in rabbit was 26 h, monkey 18 h and man 22 h. In all species studied the drug was absorbed, oxidized and/or conjugated, and was extensively excreted via the bile in all species except rabbit and man, in which urinary excretion predominated. Blood total radioactivity in man probably consisted entirely of conjugates of zeranol and/or its metabolites. Urinary metabolites in all species included conjugates (beta-glucuronides and/or sulphates) of zeranol and the major metabolite zearalanone. A more polar minor metabolite was isolated from human urine and was shown to be hydroxy-zeranol. Taleranol (7 beta-zearalanol, the lower-melting diastereoisomer), a probable metabolite of zeranol (7 alpha-zearalanol, the higher-melting diastereoisomer) in animals and in man, was shown to be a urinary metabolite in a female New Zealand white rabbit which had received [3H]zeranol (8 mg/kg per day) for seven days. A reverse isotope dilution method was developed for the quantification of both diastereoisomers of zearalanol, and also zearalanone, in urine.     

Species differences in the metabolism and excretion of fenclofenac

1. The excretion and metabolism of radiolabelled fenclofenac (2-(2, 4-dichlorophenoxy)phenylacetic acid, Flenac^R) has been studied in five species.

2. In the rat, absorption of oral doses of fenclofenac was virtually complete and elimination occurred mainly by the bile and faeces. The guinea-pig excreted equal amounts of radioactivity in urine and faeces, while in rabbit, baboon and man renal excretion was the more important route.

3. In all species the majority of excreted radioactivity was present as fenclofenac ester glucuronide. Amino acid conjunction with fenclofenac was minimal in all species studied.

4. Mono- and di-hydroxylated metabolites have been detected in urine from guineapig, baboon and man. The major hydroxylated metabolite in baboon urine has been identified as 2-(2,4-dichlorophenoxy)-5′-hydroxyphenylacetic acid.     

Study on metabolism of pyridonecarboxylic acid II. Determination of metabolites of (+-)-7-(3-amino-1-pyrrolidinyl)-6-fluoro-1-(2,4-difluorophenyl)- 1,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic acid p-toluenesulfonate hydrate (T-3262) in blood, urine, bile, and feces

The fate of (+-)-7-(3-amino-1-pyrrolidinyl)-6-fluoro-1-(2,4-difluorophenyl-1,4- dihyro-4-oxo-1,8-naphthyridine-3-carboxylic acid p-toluenesulfonate hydrate (T-3262) was studied using T-3262 and 14C-T-3262 in various animals. 1. Metabolites in serum and urine were assayed for mouse, rat, rabbit, dog and monkey following oral administration of T-3262. In serum, besides unchanged T-3262 base, T-3262A (N-acetylated) was detected in rat, rabbit and monkey; T-3262B (deamino-hydroxylated) was detected in monkey. In urine, unchanged T-3262 base was excreted mainly. But a few of metabolites (T-3262A, T-3262B, T-3262 glucuronide, T-3262A glucuronide, T-3262B glucuronide, and unknown compound M-1) were detected, and species difference existed in types of metabolites. 2. Metabolites in bile and feces were assayed for mouse and rat following oral administration of T-3262 and 14C-T-3262. Metabolites in bile were similar to the urine, but the volume of T-3262A and T-3262A glucuronide was larger than in urine. In feces, the excreted compounds mainly consisted of unchanged T-3262 base. 3. p-Toluenesulfonic acid, which is the counter acid for T-3262 base, was absorbed following the oral administration of T-3262, and excreted in urine in the unchanged form.     

Pharmacokinetics of nisoldipine. III. Biotransformation of nisoldipine in rat, dog, monkey, and man

After intraduodenal administration of 14C-labelled (+/-) 3-isobutyl-5-methyl 1,4-dihydro-2,6-dimethyl-4-(2-nitrophenyl)-pyridine-3,5-dicarboxylate (nisoldipine, Bay k 5552) to rats approx. 68% of the dose was excreted in the bile in the first 6 h. In an isolated perfused rat liver model the excretion with the bile was 56% of the total dose within 3 h. The recovery of radioactivity from orally administered [14C] nisoldipine was approx. 32% (rat), 23% (dog), 73% (monkey) and 74% (man), resp., in the urine. The unchanged drug was neither detected in the urine nor in the bile, but nisoldipine was present in plasma of the rat 30 min after dosing and up to 24 h in man. The drug was extensively metabolized: 18 biotransformation products were identified by comparison with synthetic reference compounds using combined GC-MS, 1 NMR-spectroscopy, mass spectrometry, gas chromatography/radio-gas chromatography and two-dimensional thin layer chromatography, 6 of them being quantitatively important (about 80% of the radioactivity excreted in urine). The metabolites identified accounted for approx. 82% (rat: bile and urine), 19% (dog, due to the low renal excretion), 58% (monkey: urine) and 64% (man: urine) of the excreted dose, resp. The following biotransformation steps occurred: hydroxylation of the isobutyl moiety, dehydrogenation of the 1,4-dihydropyridine system, oxidative ester cleavage, hydroxylation of one of the methyl groups in 2- or 6-position and subsequent oxidation to the carboxylic acid, oxidation of one of the methyl groups of the isobutyl moiety to the carboxyl group reduction of the aromatic nitro group (minor biotransformation reaction) and glucuronidation as phase II reaction.     

The disposition of [14C]iprindole in man, dog, miniature swine, rhesus monkey and rat.

1. Absorption of a single oral dose of [14C]iprindole was rapid in rats, rhesus monkeys, miniature swine, dogs and human volunteers. In all species except the rat, most of the radioactivity in the blood resided in the plasma. Small amounts of unchanged iprindole were detected in the plasma of rats and rhesus monkeys but not in man and miniature swine. 2. Radioactivity was excreted mainly in the urine of man, miniature swine and rhesus monkey, but in the faeces of rat and dog. 3. Urinary radioactivity was associated with basic (free and conjugated), acidic and highly polar, water soluble metabolites. At least 20 metabolites as well as small amounts of unchanged drug were detected in the basic fractions of each species' urine. 4. Many of these metabolites were common to all species; however, qualitative as well as quantitative differences were apparent. Mass-spectrometric analysis of several metabolites indicated N-demethylation and oxidation of the alicylic ring or a combination of both pathways.     

Dydrogesterone: metabolism in animals

The metabolic pattern of dydrogesterone was investigated in the rat, dog, mouse, rabbit and rhesus monkey. The drug was administered orally in 3H-labelled form. Following enzymatic hydrolysis of conjugates the radioactive metabolites were extracted from the urine, and in rat and dog also from bile. The separation method used for the development of the metabolite patterns was reversed-phase high performance liquid chromatography. Dydrogesterone and 4 derivatives, known or suspected to be metabolites, were used as marker substances. In all the species a substantial portion of the urinary or biliary radioactivity was too polar to be extracted, or it was not resolved in the chromatographic system used. The radioactivity which did develop into a pattern coincided with two or more of the marker substances. Only in the monkey, the pattern contained a peak of some substance which did not coincide with any marker. The urinary patterns of rat, dog, and mouse differed substantially, from each other as well as from those of rabbit and monkey. The patterns for the latter two animals showed certain similarities, both to each other and to the human urinary pattern as reconstructed from previous studies. It is concluded that with regard to the metabolic fate of dydrogesterone, the rabbit resembles man more than does any other species.     

The absorption, metabolism and excretion of disodium cromoglycate in nine animal species

The plasma concentrations and excretion of disodium cromoglycate after iv administration were studied in 9 animal species: mouse, rat, rabbit, dog, silky marmoset, squirrel monkey, Java (cynomolgus) monkey, stump-tailed macaque and baboon. In each species the decline of plasma concentration was rapid and the compound was quickly excreted in the bile and urine. Bile: urine ratios of excreted compound varied, the squirrel monkey excreted most of the dose in the bile (78–88%) whereas the rabbit excreted most of the dose in the urine (74–84%). Other species were intermediate between these two extremes. Tissue residues were low. Poor absorption (1–5%) after oral administration was found. No metabolites were detected in any species.     

Pharmacokinetics,metabolism and excretion of the glycine antagonist GV150526A in rat and dog

1. The pharmacokinetics, metabolic fate and excretion of 3-[-2(phenylcarbamoyl) ethenyl-4,6-dichloroindole-2-carboxylic acid (GV150526), a novel glycine antagonist for stroke, in rat and dog following intravenous administration of [C14]-GV150526A were investigated. 2. Studies were also performed in bile duct-cannulated animals to confirm the route of elimination and to obtain more information on metabolite identity. 3. Metabolites in plasma, urine and bile were identified by HPLC-MS/MS and NMR spectroscopy. 4. GV150526A was predominantly excreted in the faeces via the bile, with only trace metabolites of radioactivity in urine (< 5%). Radioactivity in rat bile was predominantly due to metabolites, whereas approximately 50% of the radioactivity in dog bile was due to parent GV150526. 5. The principal metabolites in bile were identified as glucuronide conjugates of the carboxylic acid, whereas in rat urine the main metabolite was a sulphate conjugate of an aromatic oxidation metabolite. Multiple glucuronide peaks were observed and identified as isomeric glucuronides and their anomers arising from acyl migration and muta-rotation.     

Pharmacokinetics,metabolism and excretion of the glycine antagonist GV150526A in rat and dog

1. The pharmacokinetics, metabolic fate and excretion of 3-[-2(phenylcarbamoyl) ethenyl-4,6-dichloroindole-2-carboxylic acid (GV150526), a novel glycine antagonist for stroke, in rat and dog following intravenous administration of [C14]-GV150526A were investigated. 2. Studies were also performed in bile duct-cannulated animals to confirm the route of elimination and to obtain more information on metabolite identity. 3. Metabolites in plasma, urine and bile were identified by HPLC-MS/MS and NMR spectroscopy. 4. GV150526A was predominantly excreted in the faeces via the bile, with only trace metabolites of radioactivity in urine (< 5%). Radioactivity in rat bile was predominantly due to metabolites, whereas approximately 50% of the radioactivity in dog bile was due to parent GV150526. 5. The principal metabolites in bile were identified as glucuronide conjugates of the carboxylic acid, whereas in rat urine the main metabolite was a sulphate conjugate of an aromatic oxidation metabolite. Multiple glucuronide peaks were observed and identified as isomeric glucuronides and their anomers arising from acyl migration and muta-rotation.     

Excretion and metabolism of recainam, a new anti-arrhythmic drug, in laboratory animals and humans

The metabolic disposition of recainam, an antiarrhythmic drug, was compared in mice, rats, dogs, rhesus monkeys, and humans. Following oral administration of [14C]recainam-HCl, radioactivity was excreted predominantly in the urine of all species except the rat. Metabolite profiles were determined in excreta by HPLC comparisons with synthetic standards. In rodents and rhesus monkeys, urinary excretion of unchanged recainam accounted for 23-36% of the iv dose and 3-7% of the oral dose. Aside from quantitative differences attributable to presystemic biotransformation, metabolite profiles were qualitatively similar following oral or iv administration to rodents and rhesus monkeys. Recainam was extensively metabolized in all species except humans. In human subjects, 84% of the urinary radioactivity corresponded to parent drug. The major metabolites in mouse and rat urine and rat feces were m- and p-hydroxyrecainam. Desisopropylrecainam and dimethylphenylaminocarboxylamino propionic acid were the predominant metabolites in dog and rhesus monkey urine. Small amounts of desisopropylrecainam and p-hydroxyrecainam were excreted in human urine. Selective enzymatic hydrolysis revealed that the hydroxylated metabolites were conjugated to varying degrees among species. Conjugated metabolites were not present in rat urine or feces, while conjugates were detected in mouse, dog, and monkey urine. Structural confirmation of the dog urinary metabolites was accomplished by mass spectral analysis. The low extent of metabolism of recainam in humans suggests that there will not be wide variations between dose and plasma concentrations.     

Metabolism and disposition of 14C-granisetron in rat,dog and man after intravenous and oral dosing

1. The disposition and metabolic fate of ¹⁴C-granisetron, a novel 5-HT₃ antagonist, was studied in rat, dog, and male human volunteers after intravenous and oral administration.

2. Complete absorption occurred from the gastrointestinal tract following oral dosing, but bioavailability was reduced by first-pass metabolism in all three species.

3. There were no sex-specific differences observed in radiometabolite patterns in rat or dog and there was no appreciable change in disposition with dose between 0·25 and 5 mg/kg in rat and 0·25 and 10mg/kg in dog. Additionally, there were no large differences in disposition associated with route of administration in rat, dog and man.

4. In rat and dog, 35–41% of the dose was excreted in urine and 52–62% in faeces, via the bile. Metabolites were largely present as glucuronide and sulphate conjugates, together with numerous minor polar metabolites. In man, about 60% of dosed radioactivity was excreted in urine and 36% in faeces after both intravenous and oral dosing. Unchanged granisetron was only excreted in urine (5–25% of dose).

5. The major metabolites were isolated and identified by MS spectroscopy and nmr. In rat, the dominant routes of biotransformation after both intravenous and oral dosing were 5-hydroxylation and N1-demethylation, followed by the formation of conjugates which were the major metabolites in urine, bile and plasma. In dog and man the major metabolite was 7-hydroxy-granisetron, with lesser quantities of the 6,7-dihydrodiol and/or their conjugates.     

Biotransformation of the antidepressant D,L-rolipram. II. Metabolite patterns in man,rat, rabbit,rhesus and cynomolgus monkey

1. The metabolism of D,L-rolipram in man was studied by comparison of?h.p.l.c. radiochromatograms of plasma and urine obtained at various times following oral and/or s.c. administration to man, rat, rabbit, and rhesus and cynomolgus monkey.

2. Seven metabolites isolated previously from man, rat, and rhesus monkey urine by preparative?h.p.l.c. and identified by mass spectrometry and n.m.r. analysis were used for metabolite identification.

3. In plasma, 3-5 metabolites were found in addition to the unchanged drug. In urine, which was the main excretion route for rolipram metabolites, >10 metabolites were detected but not the parent compound.

4. Between 40 and 70% of the compounds eliminated renally, were identified by reference to isolated metabolites.

5. Biotransformation proceeded by ether cleavage at the methoxy and pentyloxy groups, and by hydroxylation in positions 2 or 3 of the pentyloxy ring followed by sulphation.

6. In man, not in the other species studied, the 5-position of the pyrrolidone ring was also hydroxylated. This compound cross-reacted with the antibody raised against rolipram which was used formerly for the determination of rolipram in biological fluids.     

Regioselective glucuronidation of denopamine: marked species differences and identification of human udp-glucuronosyltransferase isoform.

Denopamine is one of the oral beta(1)-adrenoceptor-selective partial agonists. Denopamine glucuronide is the most abundant metabolite in human, rat, and dog urine when administered orally. Species differences in denopamine glucuronidation were investigated with liver microsomes obtained from humans and experimental animals. In rat and rabbit, only the phenolic glucuronide was detected, whereas in dog and monkey, not only the phenolic glucuronide but also the alcoholic glucuronide was found. In contrast, in humans, the alcoholic glucuronide was detected exclusively. The kinetics of denopamine glucuronidation in human liver microsomes showed a typical Michaelis-Menten plot. The K(m) and V(max) values accounted for 2.87 +/- 0.17 mM and 7.29 +/- 0.23 nmol/min/mg protein, respectively. With the assessment of denopamine glucuronide formation across a panel of recombinant UDP-glucuronosyltransferase (UGT) isoforms (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A8, UGT1A9, UGT1A10, UGT2B4, UGT2B7, UGT2B15, and UGT2B17), only UGT2B7 exhibited high denopamine glucuronosyltransferase activity. The K(m) value of denopamine glucuronidation in recombinant UGT2B7 microsomes was close to those in human liver and jejunum microsomes. The formation of denopamine glucuronidation by human liver, jejunum, and recombinant UGT2B7 microsomes was effectively inhibited by diclofenac, a known substrate for UGT2B7. The denopamine glucuronidation activities in seven human liver microsomes were significantly correlated with diclofenac glucuronidation activities (r(2) = 0.685, p < 0.05). These results demonstrate that the denopamine glucuronidation in human liver and intestine is mainly catalyzed by UGT2B7 and that glucuronidation of the alcoholic hydroxyl group, but not the phenolic hydroxyl group, occurs regioselectively in humans.     

The disposition and metabolism of flurbiprofen in several species including man.

Flurbiprofen was rapidly absorbed in all species studied. 2. Half-lives of elimination measured 0 to 12 h after a single dose were: mouse 3.4 h, rat 2.5 h, dog 10.1 h, baboon 3.1 h and man 3.9 h. A second phase of elimination was seen in the dog. Flurbiprofen accumulated in the circulation of the dog on repeated dosing. 3. After dosing with [14C]flurbiprofen, tissue levels of radioactivity in dog and baboon were similar to that in plasma. In the rat, levels were slightly elevated in liver, kidney, large intestine and thyroid after repeated dosing. 4. The dog excreted equal amounts of radioactivity in urine and faeces. In other species renal excretion was the more important route. 5. Six metabolites have been detected, the most important being: 2-(2-fluoro-4'-hydroxy-4-biphenylyl)propionic acid (metabolite 1), 2-(i-fluoro-3',4'-dihydroxy-4-biphenylyl)propionic acid (metabolite 2) and 2-(2-fluoro-3'-hydroxy-4'-methoxy-4-biphenylyl)propionic acid (metabolite 3). The proportions of the metabolites and the extents of their conjugation varied among the species. 6. Metabolites were detected in the circulation of rat, mouse and baboon but not in dog and man. 7. Flurbiprofen did not affect the hepatic drug-metabolizing enzyme system of rat. 8. Flurbiprofen was extensively bound to serum protein of rat, dog, baboon and man.     

Species differences in the metabolic conjugation of clofibric acid and clofibrate in laboratory animals and man

The urinary metabolites of single doses of clofibric acid (p-chlorophenoxyisobutyric acid), and its ethyl ester, clofibrate, have been investigated in rat, guinea pig, rabbit, dog, cat, ferret, and human volunteers. Human volunteers, rodents, and rabbits given clofibric acid excreted 60-90% of the 14C dose in the urine in 24 hr, and the only metabolite found was the ester glucuronide of clofibric acid, together with small amounts of the unchanged acid. In the dog, cat, and ferret, however, urinary excretion of 14C was much slower (23-39% of dose in 24 hr) and these species all formed the taurine conjugate of clofibric acid, excreted together with the unchanged acid. The ester glucuronide was found in the urine of dog and ferret but not cat. The fate of clofibrate, the ethyl ester of clofibric acid, in rat, guinea pig, rabbit, and man was similar to that of the parent acid. The characterization of the glucuronic acid and taurine conjugates of clofibric acid is described.     

Carnitine and glucuronic acid conjugates of pivalic acid

The [1-14C]pivaloyloxyethyl ester of methyldopa administered to man and cynomolgus monkeys resulted in the elimination in the urine of 14C-pivalic acid metabolites. Pivaloyl glucuronide and pivaloyl carnitine were identified as the major radioactive urinary metabolites in monkey urine and human urine, respectively. N.m.r. analysis indicated that pivaloyl carnitine had a cyclic structure. Although the role of carnitine is in the transport of fatty acids across mitochondrial membranes, it may also function in the conjugation of carboxylic acid xenobiotics in humans.     

Metabolism and clearance of proxicromil--studies in rat, hamster, rabbit, dog, squirrel monkey, cynomolgus monkey, baboon and man

Proxicromil was extensively metabolized and eliminated as metabolites in urine and faeces by the rat, hamster, rabbit, squirrel monkey, cynomolgus monkey, baboon and man after oral administration. The pathway of metabolism in these species was by hydroxylation of the alicyclic ring principally to yield monohydroxylated metabolites with trace amounts of a dihydroxylated product. Elimination of proxicromil by the dog, however, was essentially as the unchanged drug. The lack of metabolism of the drug by the dog resulted in the dog having a dependence on biliary excretion of the unchanged drug for clearance. These differences in clearance routes between species were reflected in the plasma clearance of the drug. The value for rat, a species capable of metabolism, was approximately 20 fold (4.1 ml min-1 kg-1) greater than the corresponding value for dog (0.2 ml min-1 kg-1). Inhibiting the metabolism of proxicromil in the rat with SKF-525A lowered plasma clearance of proxicromil (0.6 ml min-1 kg-1) and elevated the proportion of unchanged drug cleared by biliary excretion.     

Species-related differences in the stereoselective glucuronidation of oxazepam

The concentrations of (R)-(-)- and (S)-(+)-oxazepam glucuronides in plasma and urine of several species have been measured. The relative amounts of these diastereoisomers vary among species. Thus, in the plasma and urine of rhesus monkeys the concentrations of the R-isomer are higher, whereas in man and dog more of the S-isomer is present. In plasma and urine of miniature swine the amounts of the two diastereoisomers are about equal. In the urine of rabbits the S-isomer prevails. Similar species-related differences are observed in the in vitro formation of the isomeric oxazepam glucuronides. Homogenates of dog, miniature swine, rabbit, and rat liver produce more of the S-isomer, whereas with monkey liver the formation of (R)-oxazepam glucuronide is favored. The agreement between in vivo and in vitro data is fairly good for rhesus monkey, miniature swine, and rabbit. However, for the dog the ratio of S- to R-isomers in the liver homogenate is much higher than in plasma and urine. This species-dependent stereoselective glucuronidation of oxazepam is not related to the phylogenetic or dietary grouping of these species.