Altered hepatobiliary disposition of acetaminophen glucuronide in isolated perfused livers from multidrug resistance-associated protein 2-deficient TR(-) rats

Previous studies have demonstrated that phenobarbital treatment impairs the biliary excretion of acetaminophen glucuronide (AG), although the transport system(s) responsible for AG excretion into bile has not been identified. Initial studies in rat canalicular liver plasma membrane vesicles indicated that AG uptake was stimulated modestly by ATP, but not by membrane potential, HCO(3)(-), or pH gradients. To examine the role of the ATP-dependent canalicular transporter multidrug resistance-associated protein 2 (Mrp2)/canalicular multispecific organic anion transporter (cMOAT) in the biliary excretion of AG, the hepatobiliary disposition of acetaminophen, AG, and acetaminophen sulfate (AS) was examined in isolated perfused livers from control and TR(-) (Mrp2-deficient) Wistar rats. Mean bile flow in TR(-) livers was approximately 0.3 microl/min/g of liver ( approximately 4-fold lower than control). AG biliary excretion was decreased (>300-fold) to negligible levels in TR(-) rat livers, indicating that AG is an Mrp2 substrate. Similarly, AS biliary excretion in TR(-) livers was decreased ( approximately 5-fold); however, concentrations were still measurable, suggesting that multiple mechanisms, including Mrp2-mediated active transport, may be involved in AS biliary excretion. AG and AS perfusate concentrations were significantly higher in livers from TR(-) compared with control rats. Pharmacokinetic modeling of the data revealed that the rate constant for basolateral egress of AG increased significantly from 0.028 to 0.206 min(-1), consistent with up-regulation of a basolateral organic anion transporter in Mrp2-deficient rat livers. In conclusion, these data indicate that AG biliary excretion is mediated by Mrp2, and clearly demonstrate that substrate disposition may be influenced by alterations in complementary transport systems in transport-deficient animals.     

Glucuronidation and biliary excretion of acetaminophen in rats

Dose-dependent acetaminophen pharmacokinetics is thought to be due to saturation of sulfation and glucuronidation, although its glucuronidation has not been thoroughly examined. Because many drug-glucuronides are extensively excreted into bile, the excretion of acetaminophen metabolites in bile was examined in urethane-anesthetized rats which received 37.5, 75, 150, 300 or 600 mg/kg of acetaminophen i.v. Disappearance of acetaminophen from plasma exhibited clear dose-dependency as determined by prolongation of T 1/2 and decreases in total body clearance at 150 mg/kg or higher. Biliary excretion of the various metabolites of acetaminophen increased from 20 to 49% as the dosage was increased from 37.5 to 600 mg/kg. The glucuronide conjugate was the major form of acetaminophen in bile at all dosages. Biliary excretion of the glucuronide conjugate increased from 10.5 to 40.2% of the recovered dose as the amount administered was increased to 600 mg/kg, whereas urinary excretion of the glucuronide conjugate remained relatively constant at approximately 20% of that recovered. Although the fraction of acetaminophen excreted as the glucuronide conjugate increased to over 70% of that recovered at the highest dose, a significant decline in the rate constant for glucuronide formation was noted at 300 mg/kg and higher. Likewise, the rate for glutathione conjugation was also lower at 300 mg/kg, whereas the formation of the sulfate conjugate was decreased at lower dosages (75 mg/kg). The results of the present study show that glucuronidation is a high-capacity, high-dose saturable pathway of acetaminophen biotransformation whose product is preferentially eliminated in bile after high dosages.(ABSTRACT TRUNCATED AT 250 WORDS)     

Disposition of acetaminophen in protein-calorie malnutrition

The influence of dietary protein deficiency on the pharmacokinetics, metabolism and disposition of acetaminophen was investigated in male Sprague-Dawley rats fed for 4 weeks on a 23% (control) or a 5% (low) protein diet ad libitum. Acetaminophen and its two major metabolites, acetaminophen glucuronide and acetaminophen sulfate in plasma and urine, were determined by a sensitive and specific high-performance liquid chromatography assay. After an i.v. dose of 100 mg/kg of acetaminophen, the average mean residence time was 40% higher in the protein-deficient rats, whereas the total plasma clearance per kilogram of body weight and elimination rate constant were both decreased by approximately 36% when compared to rats on a normal protein diet. No significant differences were found in the two groups of animals with respect to the apparent steady-state volume of distribution. Rats on a low protein diet excreted a larger percentage of the administered dose as the glucuronide conjugate (34.6 vs. 12.3%) and a smaller percentage as acetaminophen sulfate (41.0 vs. 70.1%). In addition, there was a reduction in the partial metabolic clearance to acetaminophen sulfate and a concomitant 2-fold increase in the partial metabolic clearance to acetaminophen glucuronide.     

Hepatic disposition of the acyl glucuronide 1-O-gemfibrozil-beta-D-glucuronide: effects of clofibric acid, acetaminophen, and acetaminophen glucuronide

Glucuronidation of carboxylic acid compounds results in the formation of electrophilic acyl glucuronides. Because of their polarity, carrier-mediated hepatic transport systems play an important role in determining both intra- and extrahepatic exposure to these reactive conjugates. We have previously shown that the hepatic membrane transport of 1-O-gemfibrozil-beta-D-glucuronide (GG) is carrier-mediated and inhibited by the organic anion dibromosulfophthalein. In this study, we examined the influence of 200 microM acetaminophen, acetaminophen glucuronide, and clofibric acid on the disposition of GG (3 microM) in the recirculating isolated perfused rat liver preparation. GG was taken up by the liver, excreted into bile, and hydrolyzed within the liver to gemfibrozil, which appeared in perfusate but not in bile. Mean +/- S. D. hepatic clearance, apparent intrinsic clearance, hepatic extraction ratio, and biliary excretion half-life of GG were 10.4 +/- 1.4 ml/min, 94.1 +/- 17.9 ml/min, 0.346 +/- 0.046, and 30.9 +/- 4.9 min, respectively, and approximately 73% of GG was excreted into bile. At the termination of the experiment (t = 90 min), the ratio of GG concentrations in perfusate, liver, and bile was 1:35:3136. Acetaminophen and acetaminophen glucuronide had no effect on the hepatic disposition of GG, suggesting relatively low affinities of acetaminophen conjugates for hepatic transport systems or the involvement of multiple transport systems for glucuronide conjugates. In contrast, clofibric acid increased the hepatic clearance, extraction ratio, and apparent intrinsic clearance of GG (P <.05) while decreasing its biliary excretion half-life (P <.05), suggesting an interaction between GG and hepatically generated clofibric acid glucuronide at the level of hepatic transport. However, the transporter protein(s) involved remains to be identified.     

Enterohepatic circulation of lorazepam and acetaminophen conjugates in ponies

Adult female ponies (130-225 kg) with chronically implanted external biliary fistulas (T-tubes) participated in three-way cross-over studies using either i.v. lorazepam (10 mg) or acetaminophen (2 g), two model drugs biotransformed mainly by hepatic conjugative reactions. The objectives were to determine the systemic pharmacokinetics, urinary and biliary excretion and degree of enterohepatic circulation (EHC) of these compounds. Trial conditions were: A: EHC intact, with blood and urine, but not bile, collected after i.v. drug administration; B: EHC interrupted, with blood, urine and bile collected after i.v. drug administration; and C: bile infused, EHC open, without i.v. drug administration, with bile collected from trial B (containing biliary excreted drug) infused into the duodenum via the T-tube, followed by collection of blood, urine and bile. At least 2 weeks elapsed between trials. Interruption of the EHC caused lorazepam plasma half-life to shorten (3.4 vs. 2.3 hr with the EHC intact, P less than .1), clearance to increase (9.2 vs. 12.3 ml/min/kg, P less than .1) and total area under the plasma concentration curve for lorazepam glucuronide to decrease (210 vs. 310 ng/ml X hr, P less than .06). Recovery of lorazepam as its glucuronide in bile was 24.5% of the i.v. injected dose. Urinary elimination of lorazepam glucuronide was reduced from 41 to 36% of the dose due to bile collection. Subsequent duodenal infusion of collected bile, containing an average of 2.45 mg of lorazepam as glucuronide, was followed by urinary excretion of 0.48 mg of lorazepam as glucuronide in urine and 0.36 mg re-excreted into bile.(ABSTRACT TRUNCATED AT 250 WORDS)     

Long-term diabetes alters the hepatobiliary clearance of acetaminophen, bilirubin and digoxin.

The effect of insulin-dependent diabetes on hepatobiliary clearance of acetaminophen, bilirubin and digoxin was determined using Sprague-Dawley rats that were treated with a 45 mg/kg dose of streptozotocin 28 days before experimentation. Urinary excretion of acetaminophen was increased 280%, whereas biliary excretion was decreased 65% and total clearance was unchanged. Both steady-state volume of distribution and elimination half-life of acetaminophen were decreased in diabetic rats by 23 and 25%, respectively. Biliary excretion of glucuronide and sulfate conjugates was decreased by 75 and 50%, respectively, whereas parent acetaminophen excretion was unchanged. The glutathione conjugate was only detected in normal and insulin-treated rats; however, comparable levels of a cysteine conjugate were detected in bile of diabetic rats. Administration of insulin reversed the hyperglycemia and the changes in volume of distribution, elimination half-life and glutathione excretion. Other diabetes-induced alterations were unchanged. In contrast, digoxin plasma disappearance, volume of distribution and total clearance were significantly increased in diabetic rats, whereas the elimination half-life was decreased. Administration of insulin reversed the changes in serum disappearance and partially reversed the increased biliary excretion of digoxin. No differences were observed for the serum disappearance, glucuronidation or biliary excretion of bilirubin in diabetic vs. normal rats. These data indicate that insulin deficiency for 1 month can alter hepatic excretory function and the pharmacokinetics of commonly used drugs.     

Abolition of valproate-derived choleresis in the Mrp2 transporter-deficient rat

Valproic acid (VPA) is a major therapeutic agent in the treatment of epilepsy and other neurological disorders. It is metabolized in humans and rats primarily along two pathways: direct glucuronidation to yield the acyl glucuronide (VPA-G) and beta-oxidation. We have shown much earlier in the Sprague-Dawley rat that i.v. administration of sodium valproate (NaVPA) caused a marked choleresis (mean of 3.3 times basal bile flow after doses of 150 mg/kg), ascribed to the passive osmotic flow of bile water following excretion of VPA-G across the canalicular membrane. Active biliary pumping of anionic drug conjugates across the canalicular membrane is now believed to be attributable to transporter proteins, in particular Mrp2, which is deficient in the TR(-) (a mutant Wistar) rat. In the present study, normal Wistar and Mrp2-deficient TR(-) rats were dosed i.v. with NaVPA at 150 mg/kg. In the Wistar rats, there was a peak choleretic effect of about 3.2 times basal bile flow, occurring at about 30 to 45 min postdose (as seen previously with Sprague-Dawley rats). In TR(-) rats given the same i.v. dose, there was no evidence of postdose choleresis. The choleresis was correlated with the excretion of VPA-G into bile. In Wistar rats, 62.8 +/- 7.7% of the NaVPA dose was excreted in bile as VPA-G, whereas in TR(-) rats, only 2.0 +/- 0.6% of the same dose was excreted as VPA-G in bile (with partial compensatory excretion of VPA-G in urine). This study underlines the functional (bile flow) consequences of biliary transport of xenobiotic conjugated metabolites.     

Effect of aging on drug-metabolizing enzymes important in acetaminophen elimination

The effects of aging on selected drug-metabolizing enzyme activities, the pattern of phenol and bile salt sulfotransferase isoenzymes and the pharmacokinetics of acetaminophen were examined in male Fischer 344 rats at ages 5, 14 and 25 months. Aging decreased sulfotransferase activity toward acetaminophen while activity toward glycolithocholate increased with age. Glucuronosyltransferase activity toward estrone increased with age, while activity toward testosterone, morphine and naphthol remained constant. Glutathione-S-transferase (1-chloro-2,4-dinitrobenzene) activity was also unchanged through the various age groups. Cytochrome P-450 content and monooxygenase activity (p-nitroanisole demethylation) activity decreased with advancing age. Overall, the age-related in vitro changes in enzyme activities approached or equaled values measured in 5-month-old female Fischer 344 rats. Moreover, age-related alterations in total phenol sulfotransferase activity and the isozyme pattern paralleled changes in the in vivo elimination kinetics and metabolic fate of acetaminophen. The fraction of drug excreted as the sulfate conjugate and the partial clearance to acetaminophen sulfate decreased with increasing age. Conversely, the fraction excreted as the glucuronide and the partial clearance to acetaminophen glucuronide increased with increasing age. There was no effect of aging on the total clearance of acetaminophen. The gender-related differences in the pattern of sulfotransferase isozyme activity toward phenolic and bile salt acceptors disappeared with age. Age-related changes in sulfation and perhaps glucuronidation in male rats appear to feminize hepatic biotransformation and may arise due to altered gonadal hormone status.     

Cyclosporin A, but not tacrolimus, inhibits the biliary excretion of mycophenolic acid glucuronide possibly mediated by multidrug resistance-associated protein 2 in rats

The onset of diarrhea after the administration of mycophenolate mofetil (MMF) is possibly associated with the biliary excretion of its metabolite, mycophenolic acid glucuronide (MPAG). This study was undertaken to clarify the mechanism underlying the biliary excretion of MPAG. Intravenously administered mycophenolic acid (MPA, 5 mg/kg) rapidly disappeared from plasma and was efficiently excreted as MPAG in the bile of Wistar (26% of dose) and Sprague-Dawley rats (21% of dose) over 1 h. On the other hand, in spite of the rapid disappearance of MPA from plasma, the biliary excretion of MPAG was very limited in Eisai hyperbilirubinemic rats (EHBRs), which display mutations in multidrug resistance-associated protein 2 (Mrp2)/canalicular multispecific organic anion transporter, and constituted only 0.5% of dose. Instead, high levels of MPA were noted in the plasma of EHBRs. Intravenous administration of CsA (5 mg/kg) to Wistar rats significantly lowered the biliary excretion of MPAG. However, intravenously administered tacrolimus (0.1 mg/kg) failed to produce such effect. In conclusion, it is suggested that there is an efficient MPAG transport mediated by Mrp2 on the bile canalicular membrane of rat hepatocytes and that the therapeutic range of CsA potentially interferes with Mrp2. However, the therapeutic range of tacrolimus does not inhibit the transporter. Thus, it should be noted that MMF coadministered with tacrolimus instead of CsA might increase the occurrence of diarrhea related to the biliary excretion of MPAG in transplant recipients.     

Evaluation of a surgical procedure to measure drug biliary excretion of rats in regulatory safety studies

A surgical procedure was evaluated to allow bile collection from the freely moving male Sprague-Dawley rats for the assessment of drug biliary excretion during regulatory safety studies. A catheter was implanted into the bile duct to divert the bile flow via an exteriorized loop. Following recovery from the surgery and verification of normal hepatic function, the exteriorized catheter was sectioned to allow collection of the bile and replacement with a commercial bile salt solution. Approximately 80% of the catheterized animals (10 females and 10 males) had normal serum liver enzyme levels 2 days after surgery. Then, the effect of acute or repeated administrations of the immunosuppressant tacrolimus on the biliary excretion of 14C diazepam was studied to validate the technique. A first group of 12 rats received an intravenous injection of 10 mg/kg 14C-diazepam and the total and sequential amounts of diazepam excreted in the bile were measured over 72 h. Biliary excretion accounted for 80% of diazepam elimination. These rats were then given an oral administration of 3 mg/kg tacrolimus on days 7 and 8 followed by the same intravenous dose of 14C-diazepam. Another group of 10 catheterized rats was given 21 daily oral doses of 3 mg/kg tacrolimus followed by a single intravenous administration of 14C-diazepam. No significant changes in diazepam biliary excretion were observed following either acute or repeated administration of tacrolimus. This study demonstrates the feasibility of drug biliary excretion investigations under Good Laboratory Practices conditions as a complement to regulatory acute or repeated dose safety studies.     

Disposition of acetaminophen at 4, 6, and 8 g/day for 3 days in healthy young adults

The objective of this study was to determine the disposition and tolerability of 1, 1.5, and 2 g acetaminophen every 6 h for 3 days. Group I healthy adults received acetaminophen (4 then 6 g/day) or placebo; Group II received acetaminophen (4 then 8 g/day) or placebo. Acetaminophen and metabolites were measured in plasma and urine. Hepatic aminotransferases were measured daily. At steady state, acetaminophen concentrations were surprisingly lower than predicted from single-dose data, although sulfate formation clearance (fCL) was lower as expected, indicating cofactor depletion with possible sulfotransferase saturation. In contrast, glucuronide fCL was unexpectedly higher, strongly suggesting glucuronosyltransferase induction. This is the first evidence that acetaminophen induces its own glucuronidation. No dose-dependent differences were detected in fCL of thiol metabolites formed via cytochrome P4502E1. Hepatic aminotransferases stayed within reference ranges, and the incidence and frequency of adverse events were similar for acetaminophen and placebo. Although dose-dependence of acetaminophen disposition was reported previously, this study shows a novel finding of time-dependent disposition during repeated dosing. Unexpected increases in glucuronide fCL more than offset decreases in sulfate fCL, thus increasing acetaminophen clearance overall. Thiol metabolite fCL remained constant up to 8 g/day. These findings have important implications in short-term (3 day) tolerability of supratherapeutic acetaminophen doses in healthy adults.     

Species variation in toxication and detoxication of acetaminophen in vivo: a comparative study of biliary and urinary excretion of acetaminophen metabolites

Acetaminophen (AA) is converted to a toxic electrophile that may subsequently form a glutathione conjugate (AA-GS). In addition to the toxication pathway metabolites, which consist of AA-GS and its hydrolysis products (AA-cysteinylglycine, AA-cysteine and AA-mercapturate), detoxication pathway metabolites, such as AA-glucuronide and AA-sulfate, are also formed. In order to evaluate the role of these opposing pathways in the reported species variations in susceptibility to AA-induced liver injury, AA was administered to hamsters and mice, species which are susceptible to AA-induced liver injury, and to rats, rabbits and guinea pigs, species which are relatively resistant to AA-induced liver injury, and the biliary and urinary excretion of AA metabolites were measured simultaneously for 2 hr after administration of AA (1 mmol/kg i.v.). The AA-susceptible species excreted 27 to 42% of the dose as toxication pathway metabolites, whereas the resistant species excreted only 5 to 7% of the dose as toxication pathway metabolites. Most of the toxication pathway metabolites appeared in bile, where their composition reflected hepatic gamma-glutamyltranspeptidase activity; hamsters and mice (low gamma-glutamyltranspeptidase activity) excreted mainly AA-GS, whereas bile from rabbits and guinea pigs (high gamma-glutamyltranspeptidase activity) contained significant amounts of AA-GS hydrolysis products. Thus, the biliary excretion of AA-GS and its hydrolysis products may be used as an index of toxic activation of AA. The excretion of the detoxication pathway metabolites (AA-glucuronide and AA-sulfate) was 74, 62, 41, 27 and 12% of the dose in guinea pigs, rats, mice, rabbits and hamsters respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of prevention of inorganic sulfate depletion on the pharmacokinetics of acetaminophen in rats

The elimination of large doses of acetaminophen is associated with substantial depletion of endogenous inorganic sulfate which is utilized for the formation of acetaminophen sulfate. This depletion has pronounced dose- and time-dependent effects on the pharmacokinetics of acetaminophen. The purposes of this investigation were to determine the pharmacokinetics of acetaminophen in rats when endogenous sulfate depletion is prevented by administration of inorganic sulfate and to develop a simple, physiologically based pharmacokinetic model for the elimination of acetaminophen under these conditions. Adult Sprague-Dawley rats received an i.v. injection and a continuous infusion of sodium sulfate as well as an i.v. injection of acetaminophen, either 15, 30, 150 or 300 mg/kg. Serum inorganic sulfate concentrations remained at or above the physiologic level at all times. Plasma concentrations of acetaminophen declined exponentially with time after the two lower doses but exhibited initial downward curvature in log-linear plots after the two larger doses. The time-averaged plasma clearance of acetaminophen decreased with increasing dose whereas the terminal half-life was dose-independent. Most of the drug was eliminated in the urine as acetaminophen sulfate but the dose fractions of acetaminophen glucuronide and unmetabolized drug excreted in the urine increased with increasing dose. The renal clearance of acetaminophen did not exhibit dose-dependence but the apparent formation clearance of acetaminophen glucuronide tended to decrease with increasing dose. The formation of acetaminophen sulfate is describable by Michaelis-Menten kinetics, with a Vmax of about 6.5 mumol/min/kg and an in vivo KM (referenced to plasma) of about 100 microM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of aging on hepatic biotransformation in female Fischer 344 rats: changes in sulfotransferase activities are consistent with known gender-related changes in pituitary growth hormone secretion in aging animals

The effect of aging on hepatic drug conjugation in 5- to 6-, 12- to 13- and 22- to 23-month-old female Fischer 344 rats was examined. The overall disposition of acetaminophen including the formation and elimination kinetics of its sulfate and glucuronide metabolites were used as in vivo probes. The effects of aging on selected in vitro drug metabolizing enzyme activities and on the pattern of phenol and bile salt sulfotransferase isoenzymes were also determined. Aging decreased the total clearance of acetaminophen and the partial clearance of acetaminophen to acetaminophen sulfate by 36 and 47%, respectively. Increasing age also resulted in a reduced partial clearance of acetaminophen to the glucuronide- (24%) and to the glutathione-derived conjugates (29%). UDP glucuronosyltransferase activity toward 1-naphthol, morphine and testosterone was unaffected by advanced age, whereas there was a significant correlation between increased age and increased UDP glucuronosyltransferase activity toward estrone. Cytochrome P-450 concentration and glutathione-S-transferase activity toward 1-chloro-2,4-dinitrobenzene were unchanged by aging. Oxidative demethylase activity toward p-nitroanisole was decreased 18% and sulfotransferase activities toward p-nitrophenol, acetaminophen and glycolithocholate were decreased 27, 12 and 12%, respectively, in the 22- to 23-month-old rats, compared to the 5- to 6-month-old animals. In contrast to the age-related feminization in the pattern of sulfotransferase isoenzyme activities that occurs in male rats, there was no effect of aging on the pattern of phenol and bile salt sulfotransferase isoenzyme activities in female rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lithocholate glucuronide is a cholestatic agent

Lithocholic acid and its taurine, glycine, and sulfate derivatives are potent cholestatic agents. Lithocholate glucuronide is present in the plasma and urine of patients with cholestatic syndromes, but little is known of its metabolism, excretion, and cholestatic potential. [3 beta-3H]lithocholate 3-O-beta-D-glucuronide was synthesized, and chemical and radiochemical purity were established. The aqueous solubility of lithocholate glucuronide was determined and found to be greater than that of lithocholic acid or several of its derivatives. In the range of concentrations examined, calcium ions precipitated lithocholate glucuronide stoichiometrically. The material was administered to rats prepared with an external biliary fistula. When 17-25 micrograms quantities were administered, 89.1 +/- 4.5% (mean +/- SEM) of the radiolabel was secreted in bile within the first 20 h after administration, the major fraction being secreted in less than 20 min. Four-fifths of the radiolabeled material in bile was the administered unaltered parent compound, while a minor fraction consisted of a more polar derivative(s). We showed that increasing biliary concentrations of more polar derivatives were observed with milligram doses of [3H]lithocholate glucuronide, and with time after the administration of these loading doses. Milligram doses of [3H]lithocholate glucuronide resulted in partial or complete cholestasis. When induced cholestasis was partial, secretion in bile remained the primary excretory route (82.5-105.6% recovery in bile), while, when complete cholestasis was induced, wide tissue distribution of radiolabel was observed. Cholestasis developed rapidly during infusion of [3H]lithocholate glucuronide. Bile flow was diminished within 10-20 min of the start of an infusion of 0.05 mumol, 100 g-1 body weight, minute-1, administered concomitantly with an equimolar infusion of taurocholate. The results establish that lithocholate glucuronide exerts cholestatic effects comparable to those exerted by unconjugated lithocholic acid.     

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

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

Shift from biliary to urinary elimination of acetaminophen-glucuronide in acetaminophen-pretreated rats

Despite its toxicity, acetaminophen (APAP) is used increasingly as an analgesic, antipyretic, and anti-inflammatory agent. We examined the effect of prior exposure to APAP on its biliary and urinary elimination. The biliary and urinary elimination of a test dose of APAP (150 mg/kg i.v.) was determined in male Wistar rats 24 h after pretreatment with vehicle, a single dose (1.0 g/kg i.p.), or increasing daily doses (0.2, 0.3, 0.6, and 1.0 g/kg/day i.p.) of APAP. Although elimination of the parent APAP was minimally affected, biliary excretion of APAP glucuronide was significantly decreased 70 and 80%, whereas urinary excretion was significantly increased 90 and 100% in the groups pretreated with single and repeated doses of APAP, respectively, relative to vehicle controls. Western analysis and confocal immunofluorescent microscopy indicated a marked increase in hepatic expression of multidrug resistance-associated protein 3 (Mrp3) in both groups pretreated with APAP, relative to expression of Mrp2. ATP-dependent transport of [3H]taurocholate, an Mrp3 substrate, was significantly increased in basolateral liver plasma membrane vesicles from rats pretreated with repeated doses of APAP relative to controls. Enterohepatic recirculation of APAP glucuronide after administration of the same test dose of the drug was significantly decreased in rats pretreated with repeated doses of APAP. These data indicate that APAP pretreatment induced a shift from biliary to urinary elimination of APAP glucuronide, consistent with the increased expression of Mrp3 in the basolateral domain of the hepatocyte. We postulate that decreased enterohepatic recirculation contributes to decreased APAP hepatotoxicity by reducing liver exposure.     

Reduced elimination clearance of micafungin in rats with cholestatic hyperbilirubinemia

We examined whether the pharmacokinetic disposition of micafungin (MCFG), an echinocandin class antifungal agent, is altered in hyperbilirubinemia using a rat model prepared by bile duct ligation (BDL). Serum bilirubin levels were increased depending upon the duration of BDL. The elimination rate constant and total body clearance (CL_tot) of MCFG were reduced by 24% and 16%, respectively, after BDL for 1 h, but there was no significant change in the apparent volume of distribution at steady‐state. The degree of reduction in the CL_tot was much greater 7 days after BDL as compared with that 1 h after BDL (44% vs. 16%). However, the proportion of the biliary clearance in the CL_tot was about 10%. This is similar to the extent of decrease in the CL_tot by occlusion of the bile duct, demonstrating that decreased biliary excretion of MCFG makes only a minor contribution to its pharmacokinetic change. These findings suggest that the metabolic capacity of MCFG is markedly impaired in hepatic hypofunction secondary to hyperbilirubinemia, providing a fundamental explanation for the previous clinical report that there is a significant correlation between dose‐adjusted plasma MCFG concentration and serum bilirubin levels.     

Metabolism of Vitamin D3-3H in Human Subjects: Distribution in Blood, Bile, Feces, and Urine

Vitamin D(3)-(3)H has been administered intravenously to seven normal subjects, three patients with biliary fistulas, and four patients with cirrhosis. Plasma D(3)-(3)H half-times normally ranged from 20 to 30 hours. in vivo evidence that a metabolic transformation of vitamin D occurs was obtained, and a polar biologically active vitamin D metabolite was isolated from plasma.Urinary radioactivity averaged 2.4% of the administered dose for the 48-hour period after infusion, and all the excreted radioactivity represented chemically altered metabolites of vitamin D. The metabolites in urine were mainly water-soluble, with 26% in conjugated form.From 3 to 6% of the injected radioactivity was excreted in the bile of subjects with T-tube drainage and 5% in the feces of patients having no T-tube. The pattern of fecal and biliary radioactivity suggested that the passage of vitamin D and its metabolites from bile into the intestine represents an essential stage for the fecal excretion of vitamin D metabolites in man.Abnormally slow plasma disappearance of vitamin D(3)-(3)H in patients with cirrhosis was associated with a significant decrease in the quantity and rate of glucuronide metabolite excretion in the urine.     

Disposition of beta-glucuronidase-resistant "glucuronides" of valproic acid after intrabiliary administration in the rat: intact absorption, fecal excretion and intestinal hydrolysis

The major metabolite of valproic acid (VPA) is its beta-glucuronidase-susceptible glucuronide conjugate (VPA-G). At slightly alkaline pH such as in bile, VPA-G undergoes intramolecular rearrangement into at least six beta-glucuronidase-resistant isomers (VPA-G-R). The in vivo disposition of VPA-G-R was compared with those of VPA-G and VPA, each at 100 mg of VPA per kg, after intrabiliary administration to surgically prepared rats fasted during the experiments. Administered VPA was rapidly and completely absorbed into blood (peak 30 micrograms of VPA per ml at 0-2 hr). Administered VPA-G was predominantly hydrolyzed (beta-glucuronidase) in the intestine and liberated VPA absorbed into blood (peak 5 micrograms of VPA per ml at 6-9 hr). Administered VPA-G-R was disposed along at least three pathways: (1) part excretion, mainly unchanged, in feces (12% of dose); (2) part absorption (intact) from gut to blood and excretion in urine as VPA-G-R (3.6% of dose); and (3) part hydrolysis in the intestine (most likely by nonspecific esterases) with absorption of liberated VPA into blood (peak 2 micrograms of VPA per ml at 12-24 hr). The VPA/VPA-G/VPA-G-R composition of recovered dose in bile and urine was determined after all doses. In fed, nontraumatized rats given VPA-G-R p.o. at 100 mg of VPA per kg, 50% of the dose was recovered (mainly unchanged) in feces, a portion was absorbed intact into blood (2.5% of dose VPA-G-R excreted in urine) and the remainder hydrolyzed in the intestine with absorption of liberated VPA into blood.(ABSTRACT TRUNCATED AT 250 WORDS)