Metabolism and hemoglobin adduct formation of acrylamide in humans.

Acrylamide (AM), used in the manufacture of polyacrylamide and grouting agents, is produced during the cooking of foods. Workplace exposure to AM can occur through the dermal and inhalation routes. The objectives of this study were to evaluate the metabolism of AM in humans following oral administration, to compare hemoglobin adduct formation on oral and dermal administration, and to measure hormone levels. The health of the people exposed under controlled conditions was continually monitored. Prior to conducting exposures in humans, a low-dose study was conducted in rats administered 3 mg/kg (1,2,3-13C3) AM by gavage. The study protocol was reviewed and approved by Institute Review Boards both at RTI, which performed the sample analysis, and the clinical research center conducting the study. (1,2,3-13C3) AM was administered in an aqueous solution orally (single dose of 0.5, 1.0, or 3.0 mg/kg) or dermally (three daily doses of 3.0 mg/kg) to sterile male volunteers. Urine samples (3 mg/kg oral dose) were analyzed for AM metabolites using 13C NMR spectroscopy. Approximately 86% of the urinary metabolites were derived from GSH conjugation and excreted as N-acetyl-S-(3-amino-3-oxopropyl)cysteine and its S-oxide. Glycidamide, glyceramide, and low levels of N-acetyl-S-(3-amino-2-hydroxy-3-oxopropyl)cysteine were detected in urine. On oral administration, a linear dose response was observed for N-(2-carbamoylethyl)valine (AAVal) and N-(2-carbamoyl-2-hydroxyethyl)valine (GAVal) in hemoglobin. Dermal administration resulted in lower levels of AAVal and GAVal. This study indicated that humans metabolize AM via glycidamide to a lesser extent than rodents, and dermal uptake was approximately 6.6% of that observed with oral uptake.     

Metabolism of CP-195,543, a leukotriene B4 receptor antagonist, in the Long-Evans rat and Cynomolgus monkey

1. The fate of [14C]CP-195,543, a novel leukotriene B4 receptor antagonist, was studied following oral administration to the Long-Evans rat and Cynomolgus monkey. 2. Most of the radioactivity was primarily excreted in the faeces, and urine was a minor route of excretion. 3. CP-195,543 was extensively metabolized in the two species, primarily by two metabolic pathways: glucuronidation of unchanged CP-195,543 and oxidative metabolism, presumably by cytochrome P450. 4. The sites of glucuronidation were the carboxylic acid moiety and the hydroxy group. The ester glucuronide was the predominant glucuronide conjugate detected in the rat, whereas the monkey generated the ether as well as the ester glucuronide. 5. The structures of oxidative metabolites were elucidated using mass spectrometry (in the positive- and negative-ion mode) and 1H-NMR. The sites of hydroxylation were the benzylic group and the 3-position of the benzopyran ring. 6. This study has indicated that CP-195,543 was mainly eliminated by Phase II metabolism in both species.     

Behavioral effects of AMI-193, a 5-HT(2A)- and dopamine D(2)-receptor antagonist, in the squirrel monkey

8-[3-(4-Fluorophenoxy) propyl]-1-phenyl-1,3,8-triazaspiro[4, 5]decan-4-one (AMI-193) was developed as a 5-HT(2A)-selective antagonist with in vivo activity suitable for behavioral studies. However, AMI-193 is a potent dopamine D(2)-receptor antagonist with low nanomolar affinity. Accordingly, D(2)-actions may contribute to its behavioral pharmacology. In the present study, the effects of AMI-193 on operant behavior were characterized in squirrel monkeys. In subjects trained under a fixed-interval (FI) schedule of stimulus termination, AMI-193 (0.003-0.01 mg/kg) dose-dependently decreased response rate. When administered in combination with cocaine (0.03-3. 0 mg/kg) or the selective dopamine uptake inhibitor, GBR 12909 (0. 03-3.0 mg/kg), the rate-decreasing effects of AMI-193 were reversed by both dopamine indirect agonists. In drug-discrimination experiments, AMI-193 (0.003 and 0.01 mg/kg) attenuated the discriminative-stimulus effects of cocaine. AMI-193 (0.003 and 0.01 mg/kg) also reduced response rate under a second-order schedule of i. v. self-administration of cocaine (0.1 mg/infusion). The profile of behavioral effects and drug interactions observed in the present study, in conjunction with the relatively high affinity of AMI-193 for dopamine D(2) receptors, suggests that its D(2)-antagonist effects play a prominent role in the behavioral pharmacology of AMI-193.     

Glutathione conjugation and mercapturic acid formation in the developing rat, in vivo and in vitro.

1. The levels of GSH-S-epoxidetransferase (GSH-S-transferase E, EC 2.5.1.18), gamma-glutamyl transpeptidase (EC 2.3.2.2) and S-substituted cysteine N-acetyltransferase have been measured in the liver and kidney of neonatal to adult rats. 2. GSH-S-epoxidetransferase and S-substituted cysteine N-acetyltransferase activities were less than 10% of the adult values in neonatal rats, rising gradually to reach adult values at about 40 days of age. Renal gamma-glutamyl transpeptidase activity was 27% of the adult value 2 days after birth and increased after 15 days reaching adult levels by 40 days. 3. The percentages of the doses of 1,2-epoxy-3-(p-nitrophenoxy)propane (ENPP) and of 1,2-epoxybutane, administered at the same dose level to rats aged 4 days to adult, excreted as the corresponding mercapturic acids in 24 h, were not significantly different. 4. Adult and 10 day old rats doses at the same dose level with ENPP excreted N-acetyl-S-[2-hydroxy-3-(p-nitrophenoxy)propyl]-L-cysteine (ENPP-MA) at the same rate. 5. In addition to ENPP-MA, dosed rats under 13 days of age excreted the corresponding substituted cysteine. 6. The correlation between results in vitro and in vivo is discussed.     

Metabolism of CP-195,543, a leukotriene B4 receptor antagonist,in the Long-Evans rat and Cynomolgus monkey

1. The fate of [¹⁴C]CP-195,543, a novel leukotriene B4 receptor antagonist, was studied following oral administration to the Long-Evans rat and Cynomolgus monkey. 2. Most of the radioactivity was primarily excreted in the faeces, and urine was a minor route of excretion. 3. CP-195,543 was extensively metabolized in the two species, primarily by two metabolic pathways: glucuronidation of unchanged CP-195,543 and oxidative metabolism, presumably by cytochrome P450. 4. The sites of glucuronidation were the carboxylic acid moiety and the hydroxy group. The ester glucuronide was the predominant glucuronide conjugate detected in the rat, whereas the monkey generated the ether as well as the ester glucuronide. 5. The structures of oxidative metabolites were elucidated using mass spectrometry (in the positive- and negative-ion mode) and ¹H-NMR. The sites of hydroxylation were the benzylic group and the 3-position of the benzopyran ring. 6. This study has indicated that CP-195,543 was mainly eliminated by Phase II metabolism in both species.     

Nonconserved residues in the second transmembrane-spanning domain of the D(4) dopamine receptor are molecular determinants of D(4)-selective pharmacology

The molecular determinants that govern selective ligand binding to the rat D(4) dopamine receptor were investigated by substituting D(2) dopamine receptor sequences into a D(4) dopamine receptor background. The resulting mutant D(4) dopamine receptors were then screened with a panel of 10 selective and nonselective ligands, which included two allosteric modulators as sensitive measures of protein conformational changes. Mutation of a phenylalanine at position 88 in the second transmembrane-spanning domain (TMS2) of the D(4) receptor to the corresponding valine in the D(2) receptor D(4)-F88V resulted in an approximately 100-fold decrease in the affinity of the highly D(4)-selective drug 3-([4-(4-iodophenyl) piperazin-1-yl]methyl)-1H-pyrrolo[2,3-b]pyridine (L-750,667) without substantially affecting the binding of the other ligands. Mutations at the extracellular side of D(4)-TMS3 produced moderate decreases in L-750,667 binding affinities with concomitant increases in binding affinity for the D(2)/D(3)-selective antagonist (-)-raclopride. However, the binding affinities of these same D(4)-TMS3 mutants for the allosteric modulator isomethylbutylamiloride also were an anomalous 6- to 20-fold higher than either wild-type receptor. In the combined D(4)-F88V/TMS3 mutants, L-750,667 binding affinity was further decreased, but this decrease was not additive. More importantly, the combined D(4)-F88V/TMS3 mutants had (-)-raclopride and isomethylbutylamiloride binding properties that reverted back to those of the wild-type D(4)-receptor. In contrast to the D(4)-F88V mutant, the adjacent D(4)-L87W mutant had an increased affinity for ligands with extended structures, but had essentially no effect on ligands with compact structures. These findings demonstrate that two residues near the extracellular side of D(4)-TMS2 are critical molecular determinants for the selective binding of L-750,667 and ligands with extended structures.     

Metabolism of 2,3,4',6-tetrachlorobiphenyl: formation and tissue localization of mercapturic acid pathway metabolites in mice

2,3,4',6-Tetrachlorobiphenyl (tetraCB) and the corresponding 14C-labelled compound (14C-tetraCB) were synthesized. Two reference compounds, 4-methylthio- and 4-methylsulphonyl-2,3,4',6-tetrachlorobiphenyl were also prepared and characterized. TetraCB and 35S-cysteine were given to groups of female mice. Formation of methyl[35S]sulphonyl-tetraCB was indicated by the presence of extractable sulphuric acid-soluble radioactivity in lung, liver, kidney and fat of the tetraCB-treated mice. As demonstrated by gel permeation chromatography followed by gas chromatography-mass spectrometry, the tissues of the tetraCB-treated mice contained mainly methylsulphonyl-tetraCB, minor amounts of tetraCB and traces of methylthiotetraCB. The major compound present in lung was 4-methylsulphonyl-tetraCB, indicating the presence of specific binding sites for this metabolite in lung tissue. According to autoradiography of mice injected with 14C-tetraCB, these binding sites were present mainly in the tracheo-bronchial mucosa.     

R(-)2-fluoro-N-n-propylnorapomorphine: a very potent and D2-selective dopamine agonist.

A series of 2-substituted N-n-propylnorapomorphine (NPA) derivatives were synthesized and compared with other DA agonists for affinity to D1 and D2 dopamine (DA) receptors in rat brain corpus striatum tissue. The 2-substituents tested reduced D1 affinity similarly, but enhanced D2 affinity in the rank order: F greater than OH greater than Br greater than OCH3 greater than H greater than or equal to NH2. The extraordinarily high D2 affinity (Ki = 12 pM) and D2 vs. D1 selectivity (57,500) of 2-F-NPA far-exceeded that of all other DA agonists tested, and it was about 10-times more potent than NPA in vivo.     

1,2-Dichloropropane: investigation of the mechanism of mercapturic acid formation in the rat

1. Three mercapturic acid metabolites were identified in the urine of male and female Fischer 344 rats given 1,2-dichloropropane (DCP) orally (100 mg/kg) or by inhalation exposure (100 ppm, 6 h). 2. These compounds (N-acetyl-S-(2-hydroxypropyl)-L-cysteine, N-acetyl-S-(2-oxopropyl)-L-cysteine and N-acetyl-S-(1-carboxyethyl)-L-cysteine) were isolated from the urine following acidification and extraction with ethyl acetate. The extracts were derivatized with diazomethane and N,O-bis(trimethylsilyl)trifluoroacetamide and analysed by chemical ionization g.l.c.-mass spectrometry. 3. Further mechanistic studies were carried out with the stable isotope-labelled analogue, D6-DCP (105 mg/kg, orally). Analysis of the resulting mass spectra indicated retention of primarily three deuterium atoms in the 2-hydroxypropyl-mercapturic acid formed from D6-DCP. Similar isotope retention was observed for the 2-oxopropyl-mercapturic acid metabolite. 4. These results do not support a sulphonium ion intermediate in the formation of the 2-hydroxypropyl-mercapturic acid metabolite of DCP. Instead, this metabolite is thought to arise via direct oxidation of DCP, either prior to or following conjugation with glutathione.     

A novel leukotriene D4/E4 antagonist, SR2640 (2-[3-(2-quinolylmethoxy)phenylamino]benzoic acid)

We have studied the leukotriene antagonistic activity of a novel compound, SR2640 (2-[3-(2-quinolylmethoxy)phenylamino]benzoic acid), in vitro and in vivo. SR2640 inhibited LTD4- but not histamine-induced contractions of guinea-pig ileum and trachea in a concentration-dependent manner. Schild plot analysis of tracheal LTD4 antagonism yielded a pA2 value of 8.7 and a slope not different from unity. SR2640 concentration dependently inhibited the binding of 0.4 nM [3H]LTD4 to guinea-pig lung membranes with an IC50 value of 23 nM. The curve was parallel to that of unlabelled LTD4 (IC50 = 2.2 nM). SR2640 was equally effective in antagonizing LTD4 and LTE4, but was much less potent in reducing LTC4-induced ileum contractions. In vivo, SR2640 in the dose range 0.03-1.00 mg/kg shifted the dose-response curve for guinea-pig bronchoconstriction induced by intravenous LTD4 administration to the right at a rate proportional to the dose of SR2640, without reducing the maximum attainable obstruction: the slope of the Schild plot was 0.99. SR2640 (1 mg/kg) also caused a significant inhibition of antigen-induced bronchoconstriction in anaesthetized guinea-pigs pretreated with pyrilamine, indomethacin, propranolol and suxamethonium. In conclusion, SR2640 appears to be a potent and selective competitive LTD4/LTE4 antagonist, and may be useful in elucidating the role of leukotrienes in human asthma.     

Dopamine turnover and glutamate decarboxylase activity in the rat brain after acute and chronic treatment with raclopride, a dopamine D2-selective antagonist

The effect of acute and chronic (94, 141 and/or 199 days) oral treatment of rats with the dopamine D2-selective antagonist raclopride (0, 5, 15, 45 and 135 mumol/kg) upon the turnover of dopamine in the striatum and limbic system and upon the activity of glutamate decarboxylase (GAD) in the s. nigra, striatum and frontal cortex has been investigated. A dose-dependent tolerance to the effect of raclopride on the turnover of dopamine was observed after chronic treatment, although the degree of tolerance was marginal at the 5 and 15 mumol/kg doses. Acute treatment with raclopride was without effect on the activity of GAD in the s. nigra, striatum or frontal cortex, whereas chronic (199 days) treatment with 45 mumol/kg of raclopride produced an increase in the activity of GAD in the s. nigra and striatum.     

1,2-Dichloropropane: investigation of the mechanism of mercapturic acid formation in the rat

1. Three mercapturic acid metabolites were identified in the urine of male and female Fischer 344 rats given 1,2-dichloropropane (DCP) orally (100mg/kg) or by inhalation exposure (100 ppm, 6h).

2. These compounds (N-acetyl-S-(2-hydroxypropyl)-L-cysteine, N-acetyl-S-(2-oxopropyl)-L-cysteine and N-acetyl-S-(1-carboxyethyl)-L-cysteine) were isolated from the urine following acidification and extraction with ethyl acetate. The extracts were derivatized with diazomethane and N,O-bis(trimethylsilyl)trifluoroacetamide and analysed by chemical ionization g.l.c.-mass spectrometry.

3. Further mechanistic studies were carried out with the stable isotope-labelled analogue, D₆-DCP (105mg/kg, orally). Analysis of the resulting mass spectra indicated retention of primarily three deuterium atoms in the 2-hydroxypropyl-mercapturic acid formed from D₆-DCP. Similar isotope retention was observed for the 2-oxopropyl-mercapturic acid metabolite.

4. These results do not support a sulphonium ion intermediate in the formation of the 2-hydroxypropyl-mercapturic acid metabolite of DCP. Instead, this metabolite is thought to arise via direct oxidation of DCP, either prior to or following conjugation with glutathione.     

The formation of a novel mercapturic acid during the metabolism of an N-methyl aromatic amine, 4-cyano-N,N-dimethylaniline

As part of a collaborative study on the genotoxicity of aromatic amines¹ we have recently investigated the metabolism of 4-cyano-N,N-dimethylaniline (CDA) in rats and mice. We now wish to report the formation of a sulphur containing metabolite, which indicates an important role of glutathione in the metabolism of an aromatic amine N-methyl group.     

Metabolism of 2,3,4′,6-tetrachlorobiphenyl: formation and tissue localization of mercapturic acid pathway metabolites in mice

1. 2,3,4′,6-Tetrachlorobiphenyl (tetraCB) and the corresponding ¹⁴C-labelled compound (¹⁴C-TetraCB) were synthesized. Two reference compounds, 4-methylthio- and 4-methylsulphonyl-2,3,4′,6-tetrachlorobiphenyl were also prepared and characterized.

2. TetraCB and ³⁵S-cysteine were given to groups of female mice. Formation of methyl[³⁵S]sulphonyl-tetraCB was indicated by the presence of extractable sulphuric acid-soluble radioactivity in lung, liver, kidney and fat of the tetraCB-treated mice.

3. As demonstrated by gel permeation chromatography followed by gas chromatography-mass spectrometry, the tissues of the tetraCB-treated mice contained mainly methylsulphonyl-tetraCB, minor amounts of TetraCB and traces of methylthio-TetraCB.

4. The major compound present in lung was 4-methylsulphonyl-tetraCB, indicating the presence of specific binding sites for this metabolite in lung tissue. According to autoradiography of mice injected with ¹⁴C-tetraCB, these binding sites were present mainly in the tracheo-bronchial mucosa.     

Nigrostriatal dopaminergic denervation enhances dopamine D(4) receptor binding in rat caudate-putamen.

Radioligand binding to dopamine (DA) D(4) receptors was examined in adult rat forebrain 5 weeks after unilateral 6-hydroxydopamine (6-OHDA) lesioning of substantia nigra to remove ascending nigrostriatal dopaminergic projections. D(4) receptor binding was increased by up to 47% in denervated caudate-putamen (CPu) in rats that rotated away from the lesioned side with apomorphine challenge, with lesser changes in rats that failed to rotate with apomorphine. Functional significance of D(4) receptor upregulation induced by the lesions was investigated by examining behavioral effects of the highly selective D(4) agonist CP-226,269 and antagonist CP-293,019. Neither agent induced rotation at doses as high as 30 mg/kg ip. Pretreatment with the D(4) antagonist CP-293,019 did not affect rotation induced by either a D(1)-like (SKF-38393) or D(2)-like receptor (quinpirole) agonist. These findings provide the first evidence that D(4) receptors can be upregulated by nigrostriatal dopaminergic denervation. They also suggest that, unlike D(1) and D(2) receptors, D(4) receptors do not play a pivotal role in rotational behavior in rats with unilateral dopaminergic lesions.     

Zuclopenthixol, a combined dopamine D1/D2 antagonist, versus haloperidol, a dopamine D2 antagonist, in tardive dyskinesia.

Animal data suggest that a D₁ antagonistic component in neuroleptic drugs counteracts development of dopamine supersensitivity and of tolerance to cataleptic effect. This has led to the hypothesis that neuroleptics with D₁ antagonistic activity should cause a better suppression of tardive dyskinesia (TD) and less rebound aggravation after withdrawal than pure D₂ antagonists. In this study the effect of zuclopenthixol (mixed D₁/D₂ antagonist) and haloperidol (D₂ antagonist) was evaluated in chronic psychotic patients with TD. Fifteen patients completed a randomized crossover study with blind evaluation of TD and parkinsonism. The test medications, haloperidol and zuclopenthixol, caused a significant suppression of TD and a significant increase of parkinsonism. No significant differences between haloperidol and zuclopenthixol were observed. No TD aggravation was seen. The lack of differences between the mixed D₁/D₂ antagonist and a D₂ antagonist suggest that tolerance and DA supersensitivity play no or a minor role for development of TD.     

Pharmacokinetics of the novel,high-affinity and selective dopamine D3 receptor antagonist SB-277011 in rat,dog and monkey: in vitro/in vivo correlation and the role of aldehyde oxidase

1. In vitro studies with the selective dopamine D3 receptor antagonist SB-277011 were conducted in liver microsomes and homogenates from rat, dog, cynomolgus monkey and human to correlate the rate of metabolism with the in vivo pharmacokinetics of the compound in rat, dog and cynomolgus monkey. 2. In the presence of NADPH, SB-277011 was relatively stable in the presence of liver microsomes from rat, dog, cynomolgus monkey and human with an intrinsic clearance (CL_i) of <2ml min^-1 g^-1 liver for all species. In total liver homogenates, SB-277011 was metabolized at a similar rate in rat and dog (CL_i <2mlmin^-1 g^-1 liver) to that in liver microsomes but in cynomolgus monkey and human (CL_i = 9.9 and 45 mlmin ^-1 g-^-1 liver, respectively) the intrinsic clearance was ~6- and 35-fold higher, respectively, than that in liver microsomes. 3. In the absence of NADPH, SB-277011 was rapidly cleared in liver homogenates from cynomolgus monkey and human (CL_i = 7.4 and 27ml min^-1 g^-1 liver, respectively) demonstrating that a significant pathway of metabolism of this compound was via an NADPH-independent non-microsomal oxidative route. This pathway was sensitive to inhibition with isovanillin suggesting that the enzyme responsible was aldehyde oxidase. 4. The in vivo pharmacokinetics showed that the plasma clearance of SB-277011 was low in rat (20 mlmin^-1 kg^-1), moderate in dog (14 mlmin^-1 kg^-1) and high in cynomolgus monkey (58 mlmin^-1 kg^-1), which is consistent with the in vitro findings and demonstrated a greater capacity for the monkey to metabolize this compound. The oral bioavailability of SB-277011 in rat, dog and cynomolgus monkey was 35, 43 and 2%, respectively. Given the high clearance of this compound in cynomolgus monkey, the low oral bioavailability is probably as a result of high first-pass elimination, specifically by aldehyde oxidase, rather than poor absorption. 5. The high in vitro clearance of SB-277011 in human liver homogenates and the involvement of aldehyde oxidase in the metabolism of SB-277011 indicates that the bioavailability of the compound is likely to be low in human.     

Pharmacokinetics of the novel, high-affinity and selective dopamine D3 receptor antagonist SB-277011 in rat, dog and monkey: in vitro/in vivo correlation and the role of aldehyde oxidase

1. In vitro studies with the selective dopamine D3 receptor antagonist SB-277011 were conducted in liver microsomes and homogenates from rat, dog, cynomolgus monkey and human to correlate the rate of metabolism with the in vivo pharmacokinetics of the compound in rat, dog and cynomolgus monkey. 2. In the presence of NADPH, SB-277011 was relatively stable in the presence of liver microsomes from rat, dog, cynomolgus monkey and human with an intrinsic clearance (CLi) of < 2 ml min(-1) g(-1) liver for all species. In total liver homogenates, SB-277011 was metabolized at a similar rate in rat and dog (CLi < 2 ml min(-1) g(-1) liver) to that in liver microsomes but in cynomolgus monkey and human (CLi = 9.9 and 45 ml min(-1) g(-1) liver, respectively) the intrinsic clearance was approximately 6- and 35-fold higher, respectively, than that in liver microsomes. 3. In the absence of NADPH, SR-277011 was rapidly cleared in liver homogenates from cynomolgus monkey and human (CLi = 7.4 and 27 ml min(-1) g(-1) liver, respectively) demonstrating that a significant pathway of metabolism of this compound was via an NADPH-independent non-microsomal oxidative route. This pathway was sensitive to inhibition with isovanillin suggesting that the enzyme responsible was aldehyde oxidase. 4. The in vivo pharmacokinetics showed that the plasma clearance of SB-277011 was low in rat (20 ml min(-1) kg(-1)), moderate in dog (14 ml min(-1) kg(-1)) and high in cynomolgus monkey (58 ml min(-1)kg(-1)), which is consistent with the in vitro findings and demonstrated a greater capacity for the monkey to metabolize this compound. The oral bioavailability of SB-277011 in rat, dog and cynomolgus monkey was 35, 43 and 2%, respectively. Given the high clearance of this compound in cynomolgus monkey, the low oral bioavailability is probably as a result of high first-pass elimination, specifically by aldehyde oxidase, rather than poor absorption. 5. The high in vitro clearance of SB-277011 in human liver homogenates and the involvement of aldehyde oxidase in the metabolism of SB-277011 indicates that the bioavailability of the compound is likely to be low in human.     

Metabolism of a novel CCK-B antagonist in rat,dog and human liver microsomes

1. The in vitro metabolism of a novel CCK-B antagonist ((+)-N-[1-adamantane-1- methyl)-2,4-dioxo-5-phenyl-2,3,4,5-tetrahydro-1H-1,5-benzodiazepin-3-yl]N′-phenylurea; GV150013X) was investigated using rat, dog and human liver microsomes. 2. Four monohydroxy and four dihydroxy metabolites of GV150013X in rat and man were identified by comparison with authentic standards using HPLC and mass spectrometry. 3. The dihydroxy metabolite M1 was not detected in dog liver microsomes mixtures. 4. The formation of dihydroxylated metabolites proceeds via monohydroxylated metabolites M5 and M8 and not directly from GV150013X. 5. A monohydroxy metabolite M5 was the major metabolite in rat and dog, with M5 and dihydroxy metabolites M2 and M3 major metabolites in man.