Species differences in the metabolism of suprofen in laboratory animals and man

The metabolism of the oral anti-inflammatory agent suprofen (S), 2-4-(2-thienylcarbonyl)phenyl)propionic acid, has been studied in mice, rats, guinea pigs, dogs, monkeys, and human volunteers. The major metabolites of S in the serum, urine, and feces of these species were determined by GC/MS and HPLC techniques. The metabolic pathways of S in these species involved reduction of the ketone group to an alcohol (S-OH), hydroxylation of the thiophene ring (T-OH), elimination of the thiophene ring to a dicarboxylic acid (S-COOH), and conjugation with glucuronic acid or taurine. In 72-hr urine and feces of these species after po dosing of 1.6 to 2 mg/kg of S, S and these metabolites accounted for 46 to 92% of the dose and were mainly excreted in the urine. S was present as a major product (excreted mainly in conjugated form) in all species. S-OH was a major component in guinea pig and dog but a minor one in other species. T-OH was identified as a major metabolite in monkey, rat, mouse, and man, but a minor one in guinea pig, and it was absent in the dog. S-COOH was present as the minor metabolite in mouse and rat, and present at trace levels in dog, monkey, and man. Conjugation of the propionic acid functionality with taurine was observed only in the dog; in the other species, conjugation with glucuronic acid was extensive. Absorption parameters of S in the rat and monkey were similar to those in man; however, other species were very different from man.     

Characterization of a new rat urinary metabolite of piperine by LC/NMR/MS studies

Potential of piperine, an active alkaloid of black and long peppers, to increase the bioavailability of drugs in humans is of great clinical significance owing to its omnipresence in food. In an attempt to further study the reported differences in its metabolism in rats and humans, a new major urinary metabolite was detected in rat urine and plasma using HPLC. The metabolite was partially purified using reverse phase column chromatography on Sephadex^®-LH 20 and characterized as 5-(3, 4-methylenedioxy phenyl)-2E,4E-pentadienoic acid-N-(3-yl propionic acid)-amide with the help of LC/NMR/positive ESI–MS studies. Complete mass fragmentation pattern could be assigned with MS/MS studies. The metabolite has a unique structure compared to the previously reported metabolites in that it retains methylenedioxy ring and conjugated double bonds while the piperidine ring is modified to form propionic acid group. Mechanism of formation of the metabolite by oxidation and cleavage of piperidine ring is proposed. Kidney appears to be the major excretion route for piperine metabolites in rats as no metabolite could be detected in feces.     

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.     

Biotransformation of styrene in mice. Stereochemical aspects

Biotransformation of styrene and its toxic metabolite, phenyloxirane (1), in mice in vivo was studied. Mice were treated with single intraperitoneal doses of styrene (400 mg/kg of body weight), and with (R)-, (S)-, or racemic styrene oxide (150 mg/kg of body weight). Profiles of neutral and acidic metabolites were determined by GC/MS. Mandelic acid (3) and two mercapturic acids, N-acetyl-S-(2-hydroxy-2-phenylethyl)cysteine (5) and N-acetyl-S-(2-hydroxy-1-phenylethyl)cysteine (6), were found to be major urinary metabolites of both styrene and phenyloxirane. 1-Phenylethane-1,2-diol (2) was the main neutral metabolite. The rate of excretion of this metabolite, as determined by GC, was 5-10 times lower than that of mandelic acid. Several minor acidic metabolites were also identified. Among them, novel phenolic metabolites, namely, 2-(4-hydroxyphenyl)ethanol (7), (4-hydroxyphenyl)acetic acid (11), and two isomeric hydroxymandelic acids (12), are of toxicological significance. Main stereogenic metabolites were isolated as methyl esters from extracts of pooled acidified urine treated with diazomethane. The mandelic acid that was obtained was converted to diastereomeric Mosher's derivatives prior to analysis by NMR. Mercapturic acids were analyzed directly by (13)C NMR. Pure enantiomers of 1 were metabolized predominantly but not exclusively to corresponding enantiomers of 3. Styrene yielded predominantly (S)-mandelic acid. Fractions of mercapturic acids 5 and 6 isolated from urine amounted to 12-15% of the dose for all compounds that were administered. Conversion to mercapturic acids was highly regio- and stereoselective, yielding predominantly regioisomer 5. Styrene, as compared to racemic phenyloxirane, yielded slightly more diastereomers arising from (S)-1 than from (R)-1. These data can be explained by formation of a moderate excess of the less mutagenic (S)-1 in the metabolic activation of styrene in mice in vivo.     

In vitro and in vivo murine metabolism of spirogermanium.

We report the identification of spirogermanium (SG) metabolites derived from incubation of the drug with a mouse liver microsomal preparation as well as those obtained from the urine of mice injected with the drug. GC/MS data using electron impact and chemical ionization indicate that the major metabolic products appearing in the urine of mice are hydroxylated metabolites resulting from oxidation of the ethyl substituents on germanium. Thermospray (TSP) LC/MS data suggest that these hydroxy metabolites are further oxidized to an acid and a deethylated metabolite that has undergone hydroxylation of the germanium atom. In a separate experiment, human urine from a subject undergoing therapy with SG was subjected to TSP-LC/MS analysis. The SG metabolite pattern observed in the urine from human was similar to that observed in the mouse urine. These results suggest that the metabolic fate of SG in human is qualitatively similar to that found in the mouse.     

Urinary metabolites of DX-8951, a novel camptothecin analog, in rats and humans

Urinary metabolites of DX-8951 ((1S,9S)-1-amino-9-ethyl-5-fluoro- 1,2,3,9,12,15-hexahydro-9-hydroxy-4-methyl-10H,13H- benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione, CAS 171335-80-1, exatecan) in rats and humans were identified. Rats were dosed with the drug, and two major metabolites (UM-1 and UM-2) in the urine were isolated and purified by using ion-exchange column and HPLC. From NMR and mass spectra, they are suggested to be 4-hydroxymethyl metabolite (UM-1) and 3-hydroxy metabolite (UM-2) of the drug. Their chemical structures were confirmed by comparing their NMR spectra with those of chemically synthesized metabolites. Two major metabolites were found in human urine obtained in phase I trial. They were also confirmed to be UM-1 and UM-2 by LC/MS/MS by comparing their mass fragment patterns with those of synthetic metabolites.     

Biotransformation of 3-(2′,4′,5′-Triethoxybenzoyl)propionic Acid,a New Biliary Smooth Muscle Relaxant with Choleretic Activity,in Rats

1. By the combined use of deuterium labelling and gas chromatographic-mass spectrometric analysis, the metabolite pattern of 3-(2′,4′,5′-triethoxybenzoy!)propionic, acid (triethoxybenzoylpropionic acid), a new biliary smooth muscle relaxant with choleretic activity, has been determined in the rat.

2. The metabolites excreted in urine and/or bile were isolated and characterized as follows: the parent drug, 3-(2′,5′-diethoxy-4′-hydroxybenzoyl)propionic acid (metabolite II), 3-(2′-ethoxy-4′-hydroxy-5′-methoxybenzoyl)propionic acid or 3-(2′-ethoxy-5′-hydroxy-4′-methoxybenzoyl)propionic acid (III), 3-(2′,4′-diethoxy-5′-hydroxybenzoyl)propionic acid (V) and 3-[2′,4′-diethoxy-5′-(2-hydroxyethoxy)benzoyl]propionic acid (VI).

3. The primary route of biotransformation of triethoxybenzoylpropionic acid in the rat was either O-de-ethylation at the C-4′ and C-5′ positions or β-hydroxylation of the ethoxy group at the C-5′ position in the parent molecule.

4. All these excreted products except metabolite VI were eliminated both unconjugated and in conjugates (probably glucuronides and/or sulphates).

5. The rat excreted mainly metabolites V and VI in the urine and parent drug, metabolites V and VI in the bile.     

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.     

Evaluation of the absorption, excretion and metabolism of [14C] etoperidone in man.

1. The absorption, excretion and metabolism of 2-[3-[4-(3-chlorophenyl)-1-piperazinyl]propyl]-4,5-diethyl-2,4-dihydro-3H-1,2,4- triazole-3-one hydrochloride (etoperidone HCl) was investigated in six healthy men. Subjects were tasted overnight before receiving a single oral dose of a 100 mg solution [14C] etoperidone HCl. 2. Plasma (0-48 h), urine (0-120 h) and faecal (0-120 h) samples were collected. The terminal half-life of the total radioactivity from plasma was 21.7 +/- 2.8h with an apparent clearance of 1.01 +/- 0.08 ml min(-1). Recoveries of total radioactivity in urine and faeces were 78.8 +/- 3.6% and 9.6 +/- 4.1% of the dose, respectively. 3. Etoperidone and 21 metabolites were isolated and identified in the plasma, urine and faecal extracts. Unchanged etoperidone accounted for <0.01% of the dose in all excreta samples. Nine metabolites were identified in the plasma extracts and 21 urinary metabolites were identified. Seven faecal metabolites were identified. 4. Five proposed pathways were used to describe the formation of the metabolites: alkyl oxidation, piperazinyl oxidation, N-dealkylation, phenyl hydroxylation and conjugation. Alkyl oxidation of etoperidone resulted in the formation of 2-[3-[4-(3-chlorophenyl)-1-piperazinyl]propyl]-4-ethyl-2,4-dihydro-5- (1-hydroxyethyl)-3H-1,2,4-triazole-3-one. Piperazinyl oxidation of this metabolite leads to the formation of its N-oxide. N-dealkylation of the piperazinyl group led to the formation of 1-(3-chlorophenyl) piperazine and triazole propionic acid. Phenyl hydroxylation led to three important metabolites in the urine and faeces.     

Evaluation of the absorption,excretion and metabolism of [14 C] etoperidone in man

1. The absorption, excretion and metabolism of 2-{3-[4-(3-chlorophenyl)-1-piperazinyl]propyl}-4,5-diethyl-2,4-dihydro-3H-1,2,4 hydrochloride (etoperidone HCl) was investigated in six healthy men. Subjects were fasted overnight before receiving a single oral dose of a 100mg solution [14C] etoperidone HCl. 2. Plasma (0-48h), urine (0-120h) and faecal (0-120h) samples were collected. The terminal half-life of the total radioactivity from plasma was 21.7 ± 2.8?h with an apparent clearance of 1.01 ± 0.08 ml min-1. Recoveries of total radioactivity in urine and faeces were 78.8 ± 3.6% and 9.6 ± 4.1% of the dose, respectively. 3. Etoperidone and 21 metabolites were isolated and identified in the plasma, urine and faecal extracts. Unchanged etoperidone accounted for <0.01% of the dose in all excreta samples. Nine metabolites were identified in the plasma extracts and 21 urinary metabolites were identified. Seven faecal metabolites were identified. 4. Five proposed pathways were used to describe the formation of the metabolites: alkyl oxidation, piperazinyl oxidation, N -dealkylation, phenyl hydroxylation and conjugation. Alkyl oxidation of etoperidone resulted in the formation of 2-{3-[4-(3- chlorophenyl)-1-piperazinyl]propyl}-4-ethyl-2,4-dihydro-5-(1-hydroxyethyl)-3H-1 triazole-3-one. Piperazinyl oxidation of this metabolite leads to the formation of its N -oxide. N -dealkylation of the piperazinyl group led to the formation of 1-(3-chlorophenyl) piperazine and triazole propionic acid. Phenyl hydroxylation led to three important metabolites in the urine and faeces.     

Isolation and identification of the polar metabolites of chlorpheniramine in the dog

The metabolites of chlorpheniramine were isolated from dog urine. After daily repeated dosing with chlorpheniramine, [methylene-14C]chlorpheniramine maleate was given as a tracer and urine was collected until less than 1% of the labeled dose was excreted daily. An average of 54% of the oral radioactive dose was recovered in the urine. In addition to the N-demethylated metabolites, one very polar metabolite accounting for about 18% and two less polar metabolites accounting for a total of about 30% of the total urine radioactivity were isolated. Hydrolysis studies of the most polar metabolite indicated that it was a conjugate, though not a glucuronide or sulfate. The metabolite identified after hydrolysis was 3-(p-chlorobenzyl)-3-(2-pyridyl)propionic acid. One of the two less polar metabolites was identified as the corresponding alcohol. The least abundant metabolite could not be identified.     

Disposition and metabolism of letosteine in rats

The metabolism and disposition of letosteine, labeled either with 14C or 35S, has been investigated in Sprague-Dawley rats. In separate experiments, rats received 20 mg/kg, iv or orally, [14C]letosteine or [35S]letosteine. Radioactivity was rapidly excreted, mainly in urine, after iv and oral administration. Recovery of radioactivity from 0-72-hr excreta averaged 95% after both routes of [14C]letosteine administration, whereas only 50% was recovered when [35S]letosteine was administered. 14CO2 accounted for about 7.3% (iv) and 5.1% (po) of the dose of [14C]letosteine. Comparison of the iv and oral areas under the plasma 14C radioactivity concentration-time curves suggested that oral absorption of letosteine was complete. Analysis of the radioactivity content of urine showed that letosteine undergoes rapid and extensive metabolism. Several metabolites were identified by TLC, HPLC, and MS. The findings are consistent with a splitting of the ester group of letosteine and subsequent cleavage of the thiazolidinyl ring, yielding cysteine, hypotaurine, taurine, and inorganic sulfate. The metabolite derived from the side chain was identified in the urine as 3-(hydroxycarbonylmethylthio)propanoic acid. It undergoes further oxidation into sulfoxide and sulfone derivatives, which are also present in the urine.     

Absorption and excretion of suprofen in rats

DL-2-(4-(2-Thienylcarbonyl)phenyl)propionic acid (suprofen, S) was rapidly absorbed in rats after oral administration. Blood levels after a single oral dose of 2, 10, 50, or 100 mg/kg of 3H-S reached maxima within 30 min and were dose-dependent. The major portion of the drug was shown to be absorbed from the upper part of the small intestine and a portion from the stomach. The radioactivity in rat plasma was extensively bound to the plasma protein in vivo; this was found to be unchanged S and four metabolites. Elimination of S and its metabolites from blood was rapid; 3H was mostly excreted in the urine and feces within 24 hr after oral administration of 3H-S. No significant amounts of 14CO2 were excreted in expired air after administration of 14C-S. Rat urine contained S and four metabolites found in rat plasma, accounting for about 60% of the urinary radioactivity. After rats with biliary fistulas were given an oral dose of 2 mg/kg of 3H-S, 41% of the dose was excreted in the bile during 48 hr; there was significant enterohepatic circulation. When single or 21 consecutive daily doses of 3H-S were administered to rats, the blood levels after the multiple doses were higher than those after a single dose but no significant difference was found in excretion of 3H.     

Metabolism of a bisphosphonate ester (PNU-91638) in rat

1. Metabolites of the cyclic bisphosphonate ester, 4-[2,2′-bis-(5,5-dimethyl-1,3,2-dioxaphosphorinan-2-yl)] butanoyl-3-fluoro-benzene (PNU-91638) in bile or urine of the male Sprague-Dawley rat were characterized by mass spectrometry. The chronically bile duct/duodenal-cannulated male rats received a single oral dose of 100?mg/kg [¹³C] [¹³C]PNU-91638. Bile and urine samples were analysed for radioactivity and profiled by hplc with radiometric and UV detection. 2. The 0-28-h bile and urine accounted for 46.0 and 19.7% of dose respectively. The structures of radioactive peaks were investigated by ionspray and liquid secondary ion mass spectrometry (LSIMS) and LSIMS/MS analysis. 3. Major metabolites in urine included two regioisomeric phenol glucuronides, a gem-methyl hydroxylated metabolite of the bisphosphonate heterocycle, a phenol metabolite, parent drug and a benzylic alcohol metabolite. Additional metabolites in bile included an unstable phenol glutathione adduct (from a putative epoxide intermediate) with several minor isobaric regioisomers, and a carboxylic acid derived from the gem methyl hydroxylated bisphosphonate ring. 4. The structures proposed have not been confirmed by nmr due to discontinuation of the development of PNU-91638.     

16(R)-hydroxy-5,8,11,14-eicosatetraenoic acid, a new arachidonate metabolite in human polymorphonuclear leukocytes

Intact human polymorphonuclear leukocytes (PMNL) incubated with substimulatory amounts of arachidonic acid in the absence of a calcium ionophore formed four metabolites that were isolated by reverse-phase HPLC and characterized structurally by GC/MS. A major metabolite eluting as the most abundant peak of radioactivity lacked UV chromophores above 215 nm, and its formation was sensitive to 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride (SKF525A) but not 3-amino-1-[m(trifluoromethyl)phenyl]-2-pyrazoline (BW755C), suggesting that it was likely to be a product of cytochrome P450. The GC/MS analysis revealed the presence of two components: 20-hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE) and 16-hydroxy-5,8,11,14-eicosatetraenoic acid (16-HETE) in an approximate ratio of 4:1. The minor metabolites were identified as 15-HETE and 5-HETE. Although 20-HETE has been observed previously as a product of arachidonic acid metabolism in PMNL, the occurrence of 16-HETE was a novel finding. The stereochemistry of the hydroxyl group in PMNL-derived 16-HETE was established by analysis of 1-pentafluorobenzyl-16-naphthoyl derivatives on a chiral-phase chromatographic column and comparison with authentic synthetic stereoisomers. The PMNL-derived radioactive metabolite co-eluted with the synthetic 16(R)-HETE stereoisomer. Analysis of the total lipid extracts from intact PMNL followed by mild alkaline hydrolysis resulted in detectable amounts of 16-HETE (108+/-26 pg/10(8) cells) and 20-HETE (341+/-69 pg/10(8) cells), which suggested that these HETEs were formed from endogenous arachidonic acid and esterified within PMNL lipids. Thus, in contrast to calcium ionophore-stimulated neutrophils that generate large amounts of 5-lipoxygenase products, the intact PMNL generate 20-HETE and 16(R)-HETE via a cytochrome P450 omega- and omega-4 oxygenase(s).     

Identification of some benproperine metabolites in humans and investigation of their antitussive effect

AIM: To identify 4 unknown metabolites of benproperine (BPP, 1) in human urine after a po dose, and to investigate the antitussive effect of monohydroxylate metabolites. METHODS: The putative metabolite references were prepared using chemical synthesis. Their structures were identified using 1H and 13C nuclear magnetic resonance, and mass spectrometry. The metabolites in human urine were separated and assayed using liquid chromatography-ion trap mass spectrometry (LC/MS/MS), and further confirmed by comparison of their mass spectra and chromatographic retention times with those of synthesized reference substances. The antitussive effects of metabolites were evaluated on coughs induced by 7.5% citric acid in conscious guinea pigs. RESULTS: 1-[1-Methyl-2-[2-(phenylmethyl)phenoxy]-ethyl]-4-piperidinol (2), 1-[1-methyl-2-[2-(phenylmethyl)phenoxy] ethyl]-3-piperidinol (3) and their glucuronides 4 and 5 were obtained from chemical synthesis. Four urinary metabolites in human urine showed peaks with the same chromatographic retention times and mass spectra in LC/MS/MS as synthetic substances 2, 3, 4 and 5. Phosphates of compounds 2 and 3 prolonged the latency of cough and reduced the number of coughs during the 3 min test using citric acid, but did not reduce the number of coughs during the 5 min immediately after the test in conscious guinea pigs. CONCLUSION: Compounds 2, 3, 4, and 5 were identified as the metabolites of BPP in human urine. Among them, compounds 2 and 3 are inactive in the antitussive effect.     

Regiospecific and stereospecific triangulation of the structures of metabolites formed by sequential metabolism at multiple prochiral centers

Structures of in vivo secondary metabolites of a norbornane-containing drug candidate with multiple prochiral centers were triangulated, in a regio- and stereospecific fashion, using in vitro metabolism data from synthetic primary metabolites and in vivo metabolism data from the separate administration of a radiolabeled primary metabolite, [(14)C]-(S)-2-((1R,2S,4R,5S)-5-hydroxybicyclo[2.2.1]heptan-2-ylamino)-5-isopropyl-5-methylthiazol-4(5H)-one (M1). A mass balance study on the 11β hydroxysteroid dehydrogenase type 1 enzyme inhibitor [(14)C]-(S)-2-((1S,2S,4R)-bicyclo[2.2.1]heptan-2-ylamino)-5-isopropyl-5-methylthiazol-4(5H)-one (AMG 221) in rats was dosed at 2 mg/kg. Radioactivity was excreted mainly in urine. Metabolites of AMG 221 were quantified by high-performance liquid chromatography with radiometric detection and characterized by liquid chromatography-tandem mass spectrometry (LC-MS/MS). LC-MS/MS revealed at least 38 metabolites. Seven monohydroxylated metabolites mediated formation of the other 31 metabolites. Twenty-eight metabolites were identified regio- and stereo-specifically. Little parent drug was observed in urine or feces. Monohydroxy metabolite M1 was the major metabolite comprising 17 to 24% of excreted dose, and seven monohydroxy metabolites comprised 29 (male) and 37% (female) of dose. Of 11 quantifiable isobaric dihydroxy metabolites that comprised 8.3 (male) and 24% (female) of dose, 10 were identified regio- and stereospecifically by triangulation. A single trihydroxy metabolite comprised approximately 10% of dose. Complex secondary metabolism of drugs with multiple prochiral centers can be elucidated in a regio- and stereospecific fashion without NMR through synthesis and in vitro and in vivo studies on the metabolism of chiral primary oxidation products.     

Hopantenic acid beta-glucoside as a new urinary metabolite of calcium hopantenate in dogs

The metabolism of calcium hopantenate (HOPA) was studied in beagle dogs. After oral administration of 14C-labeled HOPA, 25.5% of the administered radioactivity was excreted in the urine within 24 hr, mostly in the form of unchanged drug. The only metabolite, accounting for 4.2% of the radioactivity in the urine, was isolated by HPLC. The metabolite was hydrolyzed by the treatment of beta-glucuronidase (Helix pomatia), acid phosphatase, or beta-glucosidase. These enzyme activities were not inhibited by treatment with D-glucaric acid 1,4-lactone or PO4(3-), but with D-gluconic acid 1,5-lactone, demonstrating that the metabolite is a glucose conjugate. The compound was identified as HOPA-glucoside, 4'-O-(beta-D-glucopyranosyl)-D-hopantenic acid, by GC/MS analyses after derivatization of the metabolite and the synthetic compound. This is the first reported instance of glucose conjugation to a non-acidic hydroxyl group in the metabolism of xenobiotics in mammals.     

Urinary metabolites of busulfan in the rat

After ip administration of 15 mg/kg [1,4-14C]busulfan to rat, the urinary excretion was 70% of the total radioactivity after 72 hr. Three major metabolites were isolated and quantified by HPLC. Of the total radioactivity in the urine, unchanged busulfan was excreted as a minor amount (6%) and the following metabolites were identified as: 3-hydroxysulfolane (39%), tetrahydrothiophene 1-oxide (20%), and sulfolane (13%) using GC/MS and NMR spectroscopy. The cytotoxicity of busulfan and its major metabolites was examined using a V79 Chinese hamster cell line.