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PubMedDate not supplied

Glucuronidation and the transport of the glucuronide metabolites in LLC-PK1 cells.

Formation and transport of glucuronide metabolites were studied in LLC-PK1 cells. Glucuronidation of 17beta-estradiol, 1-naphthol, mycophenolic acid, and 4-methylumbelliferone was examined in microsomes prepared from LLC-PK1 cells, human livers, human kidneys, and human intestines. The rate of glucuronide metabolite formation observed with LLC-PK1 microsomes was comparable to rates observed with various human tissue microsomes. The fate of the glucuronide metabolite formed in the LLC-PK1 cells was studied by examining its extracellular transport using mycophenolic acid as a model substrate. After administration of mycophenolic acid, the amount of the glucuronide metabolite exiting to the extracellular compartments significantly decreased in the presence of MK-571, an inhibitor for the multidrug resistance-associated protein (MRP) transporter. However, the intracellular levels of the glucuronide metabolite did not change, suggesting that MK-571 was probably blocking metabolite efflux. In summary, these results suggest that the glucuronidating enzyme(s) expressed in the LLC-PK1 cells are capable of sufficient glucuronidation activity and that endogenous transporter(s) in LLC-PK1 cells are active and determine the distribution of the formed metabolites. Since these cells have been previously used to study drug transport, they may be a useful tool in future studies to explore the effect of drug transporters on glucuronidation.

Animals↗
PubMed2000

A membrane-potential dependent ABC-like transporter mediates the vacuolar uptake of rye flavone glucuronides: regulation of glucuronide uptake by glutathione and its conjugates.

In this paper we present results on the vacuolar uptake mechanism for two flavone glucuronides present in rye mesophyll vacuoles. In contrast to barley flavone glucosides (Klein et al. (1996) J. Biol. Chem. 271, 29666-29671), the flavones luteolin 7-O-diglucuronyl-4'-O-glucuronide (R1) and luteolin 7-O-diglucuronide (R2) were taken up into vacuoles isolated from rye via a directly energized mechanism. Kinetic studies suggested that the vacuolar glucuronide transport system is constitutively expressed throughout rye primary leaf development. Competition experiments argued for the existence of a plant MRP-like transporter for plant-specific and non-plant glucuronides such as beta-estradiol 17-(beta-D-glucuronide) (E217G). The interaction of ATP-dependent vacuolar glucuronide uptake with glutathione and its conjugates turned out to be complex: R1 transport was stimulated by dinitrobenzene-GS and reduced glutathione but was inhibited by oxidized glutathione in a concentration-dependent manner. In contrast, R2 uptake was not increased in the presence of reduced glutathione. Thus, the transport system for plant-derived glucuronides differed from the characteristic stimulation of vacuolar E217G uptake by glutathione conjugates but not by reduced glutathione (Klein et al. (1998) J. Biol. Chem. 273, 262-270). Using tonoplast vesicles isolated with an artificial K+ gradient, we demonstrate for the first time for plant MRPs that the ATP-dependent uptake of R1 is membrane-potential dependent. We discuss the kinetic capacity of the ABC-type glucuronide transporter to explain net vacuolar flavone glucuronide accumulation in planta during rye primary leaf development and the possibility of an interaction of potential substrates at both the substrate binding and allosteric sites of the MRP transporter regulating the activity towards a certain substrate.

ATP-Binding Cassette Transporters↗
PubMed2006

UDP-glucuronosyltransferase 1A1 is the principal enzyme responsible for etoposide glucuronidation in human liver and intestinal microsomes: structural characterization of phenolic and alcoholic glucuronides of etoposide and estimation of enzyme kinetics.

Etoposide, an important anticancer agent, undergoes glucuronidation both in vitro and in vivo. In this study, three isomeric glucuronides of etoposide, including one phenolic (EPG) and two alcoholic glucuronides (EAG1 and EAG2), were biosynthesized in vitro with human liver microsomes (HLMs), and identified by liquid chromatography-electrospray ionization-mass spectrometry and confirmed by beta-glucuronidase cleavage. In vitro UDP-glucuronosyltransferase (UGT) reaction screening with 12 recombinant human UGTs demonstrated that etoposide glucuronidation is mainly catalyzed by UGT1A1. Although UGT1A8 and 1A3 also catalyzed the glucuronidation of etoposide, their activities were approximately 10 and 1% of UGT1A1. Enzyme kinetic study indicated that the predominant form of etoposide glucuronide in HLMs and human intestinal microsomes (HIMs) was EPG, whereas EAG1 and EAG2 were the minor metabolites, with approximately an 8 to 10% glucuronidation rate of EPG. For the formation of EPG, the V(max) of HLMs (110 pmol/min/mg protein) was very similar to that of recombinant UGT1A1 (124 pmol/min/mg protein), whereas the V(max) of HIMs (54.4 pmol/min/mg protein) was 2-fold lower than those of HIMs and UGT1A1. The K(m) values of HLMs (530 microM) and HIMs (608 microM) were 2-fold higher than that of UGT1A1 (285 microM). The V(max)/K(m) values for the formation of EPG were 0.21 and 0.09 microl/min/mg protein for HLMs and HIMs, respectively. The data indicated that UGT1A1 is principally responsible for the formation of etoposide glucuronides, mainly in the form of phenolic glucuronide, suggesting that etoposide can be used as a highly selective probe substrate for human UGT1A1 in vitro.

Alcohols↗
PubMed1996

Bioactivation of a toxic metabolite of valproic acid, (E)-2-propyl-2,4-pentadienoic acid, via glucuronidation. LC/MS/MS characterization of the GSH-glucuronide diconjugates.

The hepatotoxicity of the anticonvulsant drug valproic acid may be associated with the formation of potentially reactive metabolites, one of which is (E)-2-propyl-2,4-pentadienoic acid ((E)-2,4-diene VPA). This report describes the characterization of new GSH-related conjugates of this diene. Bile samples collected from male Sprague-Dawley rats dosed ip with (E)-2,4-diene VPA (100 mg/kg) were analyzed by LC/MS/MS. Initial Q1 parent in scanning indicated that the daughter ions m/z 162 and 123 could be derived from the ions at m/z 624 and 480, respectively. Subsequent collision-induced dissociation (CID) of these parent ions revealed a common neutral loss of 176 Da which is diagnostic for glucuronides. A similar neutral loss of 176 Da was observed in daughter ion spectra of the biliary metabolites arising from [2H7]-4-ene VPA dosed ip to rats, where the ion fragments containing the VPA portion were 7 amu higher than those derived from the unlabeled drug. CID of the ion at m/z 624 also gave fragments characteristics for GSH conjugates such as the loss of glycine and glutamate moieties. Based on the MS data, the metabolites were assigned the diconjugate structures 1-O-(2-propyl-5-(glutathion-S-yl)-3-pentenoyl)-beta-D-glucur onide (5-GS-3-ene VPA-glucuronide I, MH+, 624) and the corresponding 5-NAC-3-ene VPA-glucuronide (MH+, 480). Further proof of structural identity was obtained from 1H NMR of HPLC-purified metabolites. The amount of biliary 5-GS-3-ene VPA-glucuronide I was 7-fold greater than the corresponding 5-GS-3-ene VPA, the sum of the two metabolites accounting for 6.6% of the dose. Incubation of 1-O-(2-propyl-2,4-pentadienoyl)-beta-D-glucuronide (2,4-diene VPA-glucuronide) with GSH in the presence or absence of GST enzyme led to the formation of 5-GS-3-ene VPA-glucuronide I which was readily detected by LC/MS/MS, suggesting that in vivo the diconjugate may arise from the reaction of GSH with 2,4-diene VPA-glucuronide. To our knowledge, this is the first recorded instance in which glucuronide formation activates a drug to further conjugate with GSH via a Michael addition reaction.

Animals↗
PubMed1991

Urinary glucuronide excretion of fenofibric and clofibric acid glucuronides in man. Is it polymorphic?

The possible polymorphism of the glucuronidation reaction in man has been investigated using two hypolipidaemic compounds, fenofibrate and clofibrate, as the test probes. The formation of fenofibryl and clofibryl glucuronides was identified by their susceptibility to hydrolyses by beta-glucuronidase. The urinary excretion of the glucuronides was measured in 72 healthy volunteers after a single dose of fenofibrate, and in 104 subjects given a single dose of clofibric acid. Fenofibrate was excreted at a lower rate than clofibrate, since 13.94% and 26.55% of the doses of fenofibrate and clofibrate respectively, were recovered in urine in 8 h. Correlation analysis indicated that sex and body mass index significantly influenced the formation of fenofibryl glucuronide, whereas age and oral contraceptives affected the excretion of clofibryl acid glucuronide. The 8-hour urinary excretion patterns of clofibryl glucuronide and of clofibric acid presented a Gaussian distribution, whereas those of fenofibryl glucuronide and fenofibric acid showed 2 populations. When the metabolic ratio free fenofibric acid/glucuronide was considered, 84.7% of subjects presented the ratio 0.147, and 15.3% had the 3-fold higher ratio of 0.421. The study has shown, in the human population studied, that the glucuronidation of fenofibric acid but not that of clofibric acid may present a polymorphism.

Adult↗
PubMed1982

Kinetics of sulfation and glucuronidation of harmol in the perfused rat liver preparation. Disappearance of aberrances in glucuronidation kinetics by inhibition of sulfation.

Harmol is conjugated by glucuronidation and sulfation when it is given to the rat in vivo. In the once-through perfused rat liver preparation glucuronidation of harmol shows kinetic aberrances [Pang et al., J. Pharmac. exp. Ther. 219, 134 (1981)]. In order to further delineate the mechanism behind this, sulfation was inhibited to about 10% of control by 2,6-dichloro-4-nitrophenol. The loss of sulfation was compensated by an increase in the rate of glucuronidation, keeping the total clearance by the liver virtually constant in spite of the loss of sulfation. The inhibition of sulfation eliminated the previously observed lag-phase in the kinetics of glucuronidation; the rate of glucuronidation was now almost linear with the input concentration of the substrate harmol. The constant clearance of harmol in spite of inhibition of sulfation, the occurrence of the lag-phase in glucuronidation in the presence of sulfation, and the disappearance of this lag-phase in the absence of sulfation can be explained by either diffusion-limited metabolism of harmol or a heterogeneous sub-lobular distribution of the sulfating and glucuronidating systems. Activation of glucuronidation by harmol at high concentration can be excluded.

Alkaloids↗
PubMed1998

Inhibitory effect of phloridzin and phloretin on glucuronidation of p-nitrophenol, acetaminophen and 1-naphthol: kinetic demonstration of the influence of glucuronidation metabolism on intestinal absorption in rats.

Intestinal glucuronidation and absorption of p-nitrophenol (p-NP), acetaminophen (APAP) and 1-naphthol (alpha-NA) in the presence of phloridzin (inhibitor of Na+/glucose cotransporter) and phloretin (aglycone of phloridzin) were studied. Glucuronides of p-NP, APAP and alpha-NA appeared on both the serosal and mucosal sides. The amounts of glucuronides on the serosal side were decreased in the presence of phloridzin and phloretin. p-NP, APAP and alpha-NA appeared on the serosal side as well, and the amounts of p-NP, APAP and alpha-NA on the serosal side were increased by the presence of phloridzin and phloretin. Furthermore, the intestinal glucuronidation and absorption of alpha-NA at various concentrations were studied in the presence and absence of phloretin. Metabolic clearance was decreased in the presence of phloretin, and the absorption clearance was increased. The higher concentrations of alpha-NA caused higher absorption clearance. The lower the metabolic clearance, the higher the absorption clearance. The relationship between glucuronidation metabolism and absorption in intestine was kinetically analyzed by the metabolic inhibition model. Complete inhibition of glucuronidation improved the intestinal absorption of alpha-NA, and the absorption clearance increased to 7.17 microliter/min/cm. The formation of phloretin and an unknown metabolite from phloridzin were observed. An unknown metabolite from phloretin was observed, and was suppressed by the presence of alpha-NA. This suggests that phloridzin was hydrolyzed to phloretin, which was metabolized to glucuronide, and thereby inhibited glucuronidation of p-NP, APAP and alpha-NA.

Acetaminophen↗
PubMed2004

Effect of gonadectomy and hormones on sex differences in ketoprofen enantiomer glucuronidation and renal excretion of formed glucuronides in the rat.

PURPOSE: To investigate the sex hormone dependency of phase II metabolism using S-ketoprofen (S-KT) urinary excretion (sigmaXu) as a marker in the rat. METHODS: The effect of surgical gonadectomy, with or without concomitant estradiol or testosterone treatment, on the sigmaXu of glucuronidated S-KT was studied in male and female rats. Hepatic and renal glucuronidation of KT enantiomers was also determined using microsomal preparations from these animals. RESULTS: A controlling effect of testosterone was demonstrated by a rapid increase in sigmaXu of glucuronidated S-KT in castrated males (27.9 +/- 9.0%) compared to control males (7.2 +/- 3.9%). This approximated control female excretion (40.5 +/- 11.6%). Treatment of ovarectomized females with testosterone resulted in a steady reduction in sigmaXu of glucuronidated S-KT with time (13.4 +/- 5.4% at end point). Hepatic glucuronidation of S-KT by male rat liver microsomes was significantly higher than that of female, whereas renal glucuronidation of S-KT by female rat kidney microsomes was significantly higher than that of male. Significant correlations were found between hepatic (r = -0.78) or renal (r = 0.83) glucuronidation and sigmaXu of glucuronidated S-KT. CONCLUSIONS: Urinary excretion of S-KT-GC is sex hormone-dependent. This metabolite may have utility as a marker or probe for sex hormone-dependent studies of phase II metabolism.

Animals↗
PubMed2005

Kinetic studies on the intramolecular acyl migration of beta-1-O-acyl glucuronides: application to the glucuronides of (R)- and (S)-ketoprofen, (R)- and (S)-hydroxy-ketoprofen metabolites, and tolmetin by 1H-NMR spectroscopy.

Conjugation of carboxylate drugs with D-glucuronic acid is of considerable interest because of the inherent reactivity of the resulting beta-1-O-acyl glucuronides. These conjugates can degrade by spontaneous hydrolysis and internal acyl migration. beta-1-O-acyl glucuronides and their acyl migration products can also react covalently with macromolecules with potential toxicological consequences. The spontaneous degradation of the diastereoisomeric beta-1-O-acyl glucuronide metabolites of the racemic drug ketoprofen, two of its ring-hydroxylated metabolites and of tolmetin beta-1-O-acyl glucuronide was investigated by (1)H-NMR spectroscopy in buffer solutions, at pH 7.4 and 37 degrees C. A plot of the logarithm of the peak integrals against time revealed first-order kinetics. Degradation rates and half-lives were calculated for each glucuronide using first-order reaction equations. Tolmetin glucuronide had the fastest degradation rate, whilst all of the ketoprofen-related glucuronides had similar degradation rates. The degradation of the diastereoisomeric glucuronides was stereoselective, with the rate for the (S)-isomer always slower compared with the (R)-isomer by approximately a factor of 2.

Buffers↗
PubMed1993

Inhibition of beta-glucuronidase by natural glucuronides of kampo medicines using glucuronide of SN-38 (7-ethyl-10-hydroxycamptothecin) as a substrate.

1. 7-Ethyl-10-[4-(piperidino)-1-piperidino] carbonyloxycamptothecin (CPT-11), a potent anticancer agent currently under development for clinical use, is metabolized in vivo to 7-ethyl-10-hydroxycamptothecin (SN-38), which is subsequently conjugated to 7-ethyl-10-hydroxycamptothecin glucuronide (SN-38-glucuronide). The SN-38-glucuronide was hydrolysed by beta-glucuronidase from E. coli to aglycones and glucuronic acid. 2. Four purified natural glucuronides including baicalin, wogonoside, luteolin-3'-glucuronide, and glycyrrhizin, inhibited beta-glucuronidase using SN-38-glucuronide as substrate. The inhibition potencies of these natural glucuronides toward beta-glucuronidase were similar to that of saccharic acid 1,4-lactone. 3. These results indicate that plant materials of Kampo (Japanese herbal) medicines containing these glucuronides could be used in vivo to decrease the enterohepatic circulation of SN-38 and possibly that of other drugs.

Antineoplastic Agents, Phytogenic↗
PubMed1999

Glucuronidation of benzidine and its metabolites by cDNA-expressed human UDP-glucuronosyltransferases and pH stability of glucuronides.

Although glucuronidation is considered a necessary step in aromatic amine-induced bladder cancer, the specific enzymes involved are not known. This study assessed the capacity of five different human recombinant UDP-glucuronosyltransferases expressed in COS-1 cells to glucuronidate benzidine, its metabolites and 4-aminobiphenyl. [(14)C]UDP-glucuronic acid was used as co-substrate. UGT1A1, UGT1A4 and UGT1A9 each metabolized all of the aromatic amines. UGT1A9 exhibited the highest relative rates of metabolism with preference for the two hydroxamic acids, N-hydroxy-N-acetylbenzidine and N-hydroxy-N,N'-diacetylbenzidine. UGT1A9 metabolized 4-aminobiphenyl approximately 50% faster than benzidine or N-acetylbenzidine. UGT1A4 N-glucuronidated N'-hydroxy- N-acetylbenzidine at the highest relative rate compared with the other transferases. UGT1A6 was effective in metabolizing only four of the eight aromatic amines tested. UGT1A1 demonstrated more extensive metabolism of the hydroxamic acid, N-hydroxy-N,N'-diacetylbenzidine, and the ring oxidation product, 3-OH-N,N'-diacetylbenzidine, than it did for the other six amines. UGT2B7 was the only product of the UGT2 gene family examined and it metabolized all the aromatic amines at similar low relative levels compared with a preferred substrate, 4-OH-estrone. The K(m) values for N-acetylbenzidine metabolism by UGT1A1 and UGT1A4 were 0.37 +/- 0.14 and 1.8 +/- 0.4 mM, respectively. The O-glucuronide of 3-OH-N,N'-diacetylbenzidine was not hydrolyzed during a 24 h 37 degrees C incubation at either pH 5. 5 or 7.4. Likewise, the O-glucuronide of 3-OH-benzidine was stable at pH 7.4, with 52% remaining at pH 5.5 after 24 h. These results suggest the following relative ranking of transferase metabolism: UGT1A9 > UGT1A4 > > UGT2B7 > UGT1A6 approximately UGT1A1. The relative pH stability of O-glucuronides is consistent with a role in detoxification and excretion of aromatic amines, while the acid lability of N-glucuronides is consistent with delivery of these amines to the bladder epithelium for activation, resulting in DNA adducts which may lead to mutations.

Base Sequence↗
PubMed2006

Biosynthesis of imipramine glucuronide and characterization of imipramine glucuronidation catalyzed by recombinant UGT1A4.

AIM: To study the profile of imipramine N+-glucuronidation using homogenates of recombinant uridine-5'-diphosphoglucuronosyltransferase 1A4 (UGT1A4) from baculovirus-infected sf9 cells. METHODS: Recombinant UGT1A4 was obtained from sf9 cells infected with recombinant baculovirus. Imipramine N(+)-glucuronide was biosynthesized by incubating imipramine with recombinant UGT1A4 and then purified with solid-phase cartridges. A reversed phase-high pressure liquid chromatography (RP-HPLC) assay method was used to directly measure the concentration of imipramine and its metabolite, imipramine N(+)-glucuronide, with p-nitrophenol as the internal standard. The validated method was used to characterize the activity of recombinant UGT1A4 and carry out kinetic studies on imipramine glucuronidation in vitro. RESULTS: The high concentration of imipramine inhibited glucuronide conjugation, so the formula V=V(max).S/(Km+S+S(2)/K(i)) was used to calculate the parameters, using MATLAB software. The values of apparent K(m), K(i), and V(max) for imipramine glucuronidation via UGT1A4 were 1.39+/-0.09 mmol/L, 6.24+/-0.45 mmol/L and 453.81+/-32.12 pmol/min per mg cell homogenate (n=3), respectively. CONCLUSION: As a specific substrate of UGT1A4, imipramine was used as a convenient method to characterize the activity of recombinant UGT1A4 by using HPLC. Furthermore, the profile of imipramine glucuronidation was evaluated by using recombinant UGT1A4 in vitro.

Animals↗
PubMed1993

Intrathecal morphine-3-glucuronide does not antagonize spinal antinociception by morphine or morphine-6-glucuronide in rats.

Morphine or morphine-6-glucuronide either alone or in combination with morphine-3-glucuronide was administered intrathecally to rats. Antinociceptive effects were evaluated with the tail flick and the hot plate tests. Motor function was tested using the rotarod test. Estimated ED50 from the dose-response curves for morphine and morphine-6-glucuronide showed about a 30 times more potent antinociceptive effect of morphine-6-glucuronide compared with morphine. Morphine-3-glucuronide had no antinociceptive effect. Simultaneous administration of morphine-3-glucuronide 5.0 micrograms did not show any significant effect on antinociception induced by morphine 1.0 microgram or morphine-6-glucuronide 0.05 microgram.

Analgesics↗
PubMed2003

Intestinal absorption and metabolism of retinoyl beta-glucuronide in humans, and of 15-[14C]-retinoyl beta-glucuronide in rats of different vitamin A status.

In order to prove the hypothesis that humans and animals with adequate vitamin A status do not absorb and metabolize orally administered all-trans retinoyl beta-glucuronide, unlabeled retinoyl glucuronide (0.1 mmol) was orally dosed to fasting well-nourished young men. Neither retinoyl glucuronide nor retinoic acid, a possible metabolite, appeared in the blood within 12 h after ingestion. Next, radiolabeled all-trans 15-[14C]-retinoyl beta-glucuronide was chemically synthesized by a new procedure, and fed orally to rats of different vitamin A status. Analysis of blood and other tissues 5 or 24 h after the dose, showed the presence of radioactivity ( approximately 0.5%) in the blood of vitamin A deficient rats, but not in sufficient rats. Livers of all rats contained small, but detectable amounts (0.3 to 1.1% of the dose) of radioactivity. The accumulation of radioactivity in the liver was highest in deficient rats. Analysis of the retinoids showed that the radioactivity in serum and liver was due to retinoic acid formed from retinoyl glucuronide. Within 24 h after the dose, 31 to 40% of the administered radioactivity was excreted in the feces, and 2 to 4.7% of the dose was excreted in the urine. Results of the present studies show that oral administration of retinoyl beta-glucuronide did not give rise to detectable changes in blood retinoyl glucuronide and/or retinoic acid concentrations in humans or rats with adequate vitamin A status.

Animals↗
PubMed1975

The radioimmunoassay of steroid glucuronides. The oestrogen C-3 glucuronides as haptens.

Antisera were prepared against three related oestrogen ring-A glucuronides, oestrone 3-glucuronide, oestradiol 3-glucuronide and oestriol 3-glucuronide. The corresponding 6,7-3H-labelled conjugates were synthesized as radioligands and the cross-reactions of the antisera against ring-A oestrogen glucuronides and other steroid conjugates were examined. The specificity of the antiserum against oestriol 3-glucuronide was compared with that raised against oestriol 16alpha-glucuronide, and the measurement of the former conjugate in late-pregnancy urine is discussed.

Antibody Specificity↗
PubMed1981

Significance of 17-hydroxycorticosteroid glucuronide in urine as an index of salicylamide glucuronidation in guinea pigs.

The glucuronidation of 17-hydroxycorticosteroids (17-OHCS) and its effectiveness as an index of salicylamide (SAM) glucuronidation were examined in guinea pigs, the same cortisol type animals as man, treated with carbon tetrachloride (CCl4), D-galactosamine, phenobarbital, D,L-ethionine and alloxan. There was not a statistically significant correlation between the plasma clearance of SAM glucuronidation (CLglu) and the urinary excretion ratio of 17-OHCS glucuronide to total 17-OHCS in the intact, and CCl4, D-galactosamine, alloxan and D,L-ethionine treated guinea pigs (r = 0.158). However, there was a statistically significant correlation between the CLglu and the clearance of 17-OHCS glucuronidation obtained as urinary 17-OHCS glucuronide/plasma cortisol concentration (r = 0.738, p less than 0.1). It was proposed that the clearance of 17-OHCS glucuronide metabolized from endogenous cortisol could be used as an index to assess the drug glucuronidation activity.

17-Hydroxycorticosteroids↗
PubMed1989

Enzymic synthesis of two glucuronides of the hydroxyisoxazole GABA-agonist, THIP, and the in vivo glucuronidation of THIP in rat.

1. A method for preparative enzymic synthesis of two glucuronides of THIP (3-hydroxy-4,5,6,7-tetrahydro-isoxazolo[5,4-c]pyridine) is described. 2. Using FAB mass spectrometry, u.v. and 1H- and 13C-n.m.r. spectroscopy, the two glucuronides were identified as N- and O-glucuronides respectively. 3. An h.p.l.c. method for determination of THIP and the two intact glucuronides in urine has been developed. 4. The glucuronidation pattern of THIP in rats has been examined; THIP was excreted as a THIP-O-glucuronide but not as a THIP-N-glucuronide.

Animals↗
PubMed2000

The glucuronidation of morphine by dog liver microsomes: identification of morphine-6-O-glucuronide.

Canines are used extensively in the pharmaceutical industry for the preclinical screening of novel therapeutics, yet comparatively little is known about the phase 2 metabolism in this species. In humans, morphine is known to undergo extensive metabolism by glucuronidation, and the UDP-glucuronosyltransferase isoform, which catalyzes the formation of morphine-3-O-glucuronide and morphine-6-O-glucuronide is UGT2B7. This study was designed to investigate the glucuronidation of morphine using dog liver microsomes. Liver microsomes from beagle dogs catalyzed the glucuronidation of morphine-3(and 6)-O-glucuronide at rates 4 to 10 times that of rhesus monkey and human liver microsomes. The K(m) of morphine using beagle dog liver microsomes was approximately 270 microM, which is similar to that found for expressed human UGT2B7. The V(max) for morphine, using dog liver microsomes, was 27 nmol/min/mg of protein. Flunitrazepam inhibited the glucuronidation of morphine in dog liver microsomes, and the K(i) was 40 microM, which is similar to human UGT2B7 for other substrates. The effects of detergents were also investigated with dog liver microsomes, and Brij 35 and Brij 58 were found to be the best detergents to use for maximal activation of the dog liver morphine UGT. These studies suggest that dog has a UGT2B isoform similar to human UGT2B7.

Animals↗