Eisai hyperbilirubinemic rats. Drug Metab. Dispos. 22:561â565. 18. Nakashima, E., R. Matsushita, M. Takeda, T. Nakanishi, and F. Ichimura. 1992. Comparative ...
ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Oct. 1995, p. 2258–2261 0066-4804/95/$04.0010 Copyright q 1995, American Society for Microbiology
Vol. 39, No. 10
Effect of a Bacterial Lipopolysaccharide on Biliary Excretion of a b-Lactam Antibiotic, Cefoperazone, in Rats SOHEILA HAGHGOO,1 TAKAAKI HASEGAWA,1* MASAYUKI NADAI,1 LI WANG,1 TOSHITAKA NABESHIMA,1 AND NOBUO KATO2 Department of Hospital Pharmacy1 and Department of Bacteriology,2 Nagoya University School of Medicine, Nagoya 466, Japan Received 13 June 1994/Returned for modification 8 December 1994/Accepted 5 August 1995
Klebsiella pneumoniae O3 lipopolysaccharide (LPS) has been found to dramatically modify the pharmacokinetics of the b-lactam antibiotic cefazolin in rats. This study investigated the effect of LPS on the biliary excretion of the b-lactam antibiotic cefoperazone (CPZ) in rats. CPZ is known to be actively secreted into the bile by a carrier-mediated transport system. LPS (250 mg/kg of body weight) was infused for 20 to 30 min 2 h before an intravenous administration of CPZ (20 mg/kg). The pharmacokinetic parameters of CPZ were estimated by a noncompartment model. LPS induced a significant decrease in the systemic clearance (by approximately 50%) and an increase in the mean residence time of CPZ. Significant decreases were also seen in the bile flow rate and in the biliary recovery of unchanged CPZ in the LPS-treated rats. LPS tended to increase the proportion of urinary excretion of CPZ. LPS significantly decreased the biliary clearance (by approximately 55%) and renal clearance (by approximately 35%) of CPZ. However, no changes in the volume of distribution at steady state for CPZ were observed between the treatment groups. Our findings suggest that LPS induces changes in the pharmacokinetics of CPZ as a result of changes occurring in the biliary secretory system.
foperazone (CPZ), an organic anion b-lactam antibiotic, was chosen as the model drug, since it is excreted into the bile in a primarily unchanged form more often than other well-known b-lactam antibiotics (19).
Lipopolysaccharide (LPS), which is an active component in the outer membrane of gram-negative bacteria, is known for its various biological and immunological activities. LPS has been found to induce various changes in the body: circulatory changes, disseminated intravascular coagulation, and damage to numerous organs such as the central nervous system, liver, kidneys, heart, gastrointestinal tract, and lungs (8). A number of studies have discussed the effects of LPS on the liver (1, 9, 10, 13, 20, 31), which is known to be vital in clearing circulatory LPS in rats (3, 37). Several investigators have also demonstrated that LPS is rapidly taken into the liver and localized in both the hepatocytes and Kupffer cells (2, 22, 23, 33). LPS has been reported to have a cholestatic effect by decreasing the bile flow (29) and decreasing the biliary clearance (CLBILE) of some organic anionic dyes such as sulfobromophthalein (BSP) and indocyanine green in isolated perfused rat livers (29, 30). From these findings, it is thought that the transport system of some organic anionic drugs, which are mostly excreted into the bile, may be affected by LPS-induced changes in the hepatofunctions and liver excretory system. Several investigators have examined the effects of hyperbilirubinemia and diabetes on the hepatofunctions and biliary excretion of certain organic anionic antibiotics (17, 18, 24, 35, 36). However, there is little data available regarding the effect of LPS on hepatic transport and the biliary secretion of drugs. A series of experiments was designed in our laboratories to establish a guideline for the safe use of antibiotics which are excreted mainly into the bile. Studies were intended to benefit patients with hepatobiliary infections by gram-negative bacteria. This study aimed at evaluating the effect of LPS on the hepatic elimination of b-lactam antibiotics, considering their importance in treating gram-negative bacterial infections. Ce-
MATERIALS AND METHODS Chemicals. CPZ was donated by Toyama Chemical Industries (Tokyo, Japan). LPS was isolated from a cultured supernatant of Klebsiella pneumoniae LEN-1 (O3:K12) (6, 7), which is a decapsulated mutant strain derived from the K. pneumoniae strain of Kasuya (O3:K1) (21). The LPS used was identical to that used in previous studies (4, 5, 14–16, 34). All other reagents were commercially available and were used at analytical grade without the need for further purification. LPS and CPZ were dissolved in isotonic saline. Animal experiments. Eight- to nine-week-old male Wistar rats (Nippon SLC, Hamamatsu, Japan) were used in this study. One day before the experiments, the rats were anesthetized with sodium pentobarbital (25 mg/kg of body weight), were cannulated with polyethylene tubes in the right jugular vein for drug administration and blood sampling, and were then allowed to recover. On the following day, LPS was constantly infused over a period of 20 to 30 min (3.93 mg/min/kg), 2 h before administration of CPZ (LPS-treated group) under conditions previously described (4, 5, 14–16, 34). In the control group, rats were pretreated with isotonic saline instead of LPS in the same manner. Thirty minutes before the experiments were started, the bile ducts and urinary bladders were cannulated with polyethylene tubes for bile and urine sampling, respectively. All experiments were performed under light anesthesia, and body temperature was maintained at 378C with the assistance of a heat lamp. After the urine and bile flow were stabilized over a 10-min period, the rats received a bolus intravenous injection (within 5 s) of CPZ at a dose of 20 mg/kg. Blood samples of approximately 0.25 ml were collected at designated intervals of 2, 5, 10, 20, 30, 45, 60, 75, 90, and 120 min after CPZ administration, and plasma samples were immediately obtained by centrifugation at 6,000 3 g for 5 min. Bile samples were collected at three consecutive 10-min periods (0 to 10, 10 to 20, and 20 to 30 min) which were followed by six more consecutive 15-min periods (30 to 45, 45 to 60, 60 to 75, 75 to 90, 90 to 105, and 105 to 120 min). Urine samples were collected at 60-min intervals. The volumes of bile and urine were measured gravimetrically, with specific gravity assumed to be 1.0. Drug analysis. Concentrations of CPZ in the plasma, bile, and urine were measured by high-performance liquid chromatography (HPLC). Bile and urine samples were properly diluted in distilled water. Briefly, 50 ml of each sample, 50 ml of 10% perchloric acid, and 50 ml of phosphate buffer (pH 7.4) containing 3-butylxanthine as an internal standard were mixed and centrifuged at 15,000 3 g for 10 min. The resulting supernatant was injected into the HPLC apparatus. The HPLC apparatus was a Shimadzu LC-6A system (Shimadzu Company, Kyoto, Japan) consisting of an LC-6A liquid pump, an SPD-6A UV-VIS spec-
* Corresponding author. Mailing address: Department of Hospital Pharmacy, Nagaya University School of Medicine, 65 Tsuruma-cho, Showa-ku, Nagaya 466, Japan. Phone: 81-52-741-2111 (ext. 2552). Fax: 81-52-733-9415. 2258
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FIG. 1. Mean semilogarithmic plots of plasma concentration-time data for CPZ in control (E) and LPS-treated (F) rats after a single intravenous administration of CPZ (20 mg/kg). Plot points are means 6 standard deviations (n 5 5). Values marked by a are significantly different from the control values (P , 0.05).
trophotometric detector, and an SIL-6A autoinjector. A Cosmosil 5C18 column (Nacalai Tesque, Kyoto, Japan) was used with a column oven (OTC-6A) heated to 508C. The UV detector was set at 266 nm. The mobile phase was 30 mM KH2PO4 buffer (pH 5.0)–methanol (80:20 [vol/vol]), and the flow rate was 1.5 ml/min. Calibration curves for measuring CPZ in the plasma, bile, and urine proved to be linear for concentrations ranging from 0.2 to 200 mg/ml. The inter- and intraday coefficients of variation for the assay were less than 6% at concentrations of 2 and 50 mg/ml. Blank plasma, urine, and bile samples did not interfere with the peak corresponding to CPZ. The detection limit of CPZ was 0.1 mg/ml. Data analysis. Plasma concentration-time data for CPZ were analyzed on the basis of a noncompartment model. The area under the plasma concentrationtime curve (AUC) and area under the first moment curve (AUMC) for CPZ were determined by the trapezoidal method with extrapolation to infinity by adding the following data. The value of the last measured plasma concentration was divided by the terminal elimination rate constant. The terminal elimination rate constant was calculated by determining the slope of the least-squares regression line from the terminal portion of the log concentration-time data (four to five points). Systemic clearance (CLSYS) was calculated by dividing the dose by the AUC. The mean residence time (MRT) and the volume of distribution at steady state (VSS) were calculated by the equations MRT 5 AUMC/AUC and VSS 5 CLSYS 3 MRT, respectively. The CLBILE and renal clearance (CLR) were calculated by dividing the total amount of CPZ excreted into the bile and urine, respectively, within the collection period (120 min) by the corresponding AUC (AUC120). All computer analyses were performed by using a nonlinear least-squares regression program with no weight function (MULTI; written by Yamaoka et al. [38]). Statistical analysis. The results are expressed as means 6 standard deviations for the indicated numbers of experiments. Statistical analyses were performed nonparametrically by the Mann-Whitney U test, with P , 0.05 taken as the limit of statistical significance.
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FIG. 2. Cumulative excretion of CPZ into the bile over the entire collection period (120 min) following an intravenous injection of CPZ (20 mg/kg) in control (E) and LPS-treated rats (F). Plot values are means 6 standard deviations (n 5 5). Values marked by a are significantly different from the control values (P , 0.05).
The corresponding pharmacokinetic parameters of CPZ are summarized in Table 1. LPS significantly decreased the CLSYS and increased the MRT for CPZ. However, there was no significant difference in the VSS for CPZ between the control rats and the LPS-treated rats. The cumulative biliary excretion of CPZ and bile flow rate within the collection period (120 min) after the administration are shown in Fig. 2 and 3, respectively. The bile flow rate remained nearly constant in both groups beginning 20 min after the administration of CPZ until the end of the experiments, although the rate was relatively high in the first sampling period. However, treatment with LPS led to a lower bile flow rate. LPS induced a significant decrease in the biliary excretion of CPZ. The percentages of the CPZ dose recovered in the bile and urine over the 120-min experimental period are summarized in Table 2. The urinary recovery of CPZ in an unchanged form in the LPS-treated rats tended to increase. Pretreatment with LPS induced significant decreases in the
RESULTS Mean semilogarithmic plasma concentration-time curves for CPZ in the control and LPS-treated rats following single intravenous administrations at a dose of 20 mg/kg are shown in Fig. 1. Pretreatment with LPS increased the level of CPZ in the plasma and delayed the disappearance of CPZ from plasma.
TABLE 1. Pharmacokinetic parameters of CPZ in control and LPS-treated ratsa Treatment group
Control LPS a b
CLSYS (liters/h/kg)
VSS (liters/h/kg)
MRT (h)
1.614 6 0.135 0.785 6 0.202b
0.452 6 0.059 0.385 6 0.093
0.273 6 0.036 0.483 6 0.039b
Values are means 6 standard deviations (n 5 5). Significantly different from control (P , 0.05).
FIG. 3. Effect of LPS on the bile flow rate during the collection period (120 min). Empty and shaded columns represent control and LPS-treated rat groups, respectively, after a single intravenous injection of CPZ (20 mg/kg). Columns show means 6 standard deviations (n 5 5). Values marked by a are significantly different from control values (P , 0.05).
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TABLE 2. Renal and biliary clearance data of CPZ in control and LPS-treated ratsa Treatment group
CLBILE (liters/h/kg)
CLR (liters/h/kg)
% of dose excreted in: Bile
Urine
Control 1.318 6 0.110 0.329 6 0.079 81.41 6 8.33 20.04 6 3.88 LPS 0.574 6 0.188b 0.215 6 0.066b 69.50 6 6.25b 29.57 6 11.41 a b
Values are means 6 standard deviations (n 5 5). Significantly different from control (P , 0.05).
CLBILE (56%) and CLR (35%) for CPZ compared with those for the control rats (Table 2). DISCUSSION In our earlier studies, we found that K. pneumoniae LPS dramatically modifies the pharmacokinetics and renal handling of drugs, including the b-lactam antibiotic cefazolin, which are excreted primarily into the urine (4, 5, 14–16, 34). However, direct evidence is limited regarding whether LPS contributes to changes in the biliary excretion of drugs, including that of b-lactam antibiotics. In the present study with the b-lactam antibiotic CPZ, we demonstrated that K. pneumoniae LPS dramatically modifies the pharmacokinetics of CPZ, since LPS induced a significant decrease in the CLSYS for CPZ as a result of a decrease in the CLBILE. Recent studies in our laboratory have illustrated that K. pneumoniae LPS does not change the volume of distribution for the aminoglycoside antibiotic tobramycin or the extracellular fluid volume of rats (14), although it does increase the distribution volume of drugs which strongly bind to plasma protein because of decreases in the extent of protein binding (15, 16, 34). In the present study, the K. pneumoniae LPS had no effect on the VSS of CPZ. This supports earlier evidence showing that the protein binding potency of CPZ is low in rats (18). A preliminary experiment also showed that the protein binding potency was very low (,30%) in a concentrationindependent manner (data not shown). Tsuji et al. (26, 28) have reported that b-lactam antibiotics localize in the extracellular water space and bind to plasma proteins in the intravascular and interstitial fluids in nonclearing organs. Such experimental outcomes may serve to confirm that LPS has no effect on the VSS for CPZ. In the present study, approximately 80% of the administered dose of CPZ was excreted into the bile in the control rats, which is consistent with values reported previously (18). As shown in Table 2, LPS decreased the amount of biliary excretion of CPZ to approximately 85% of the control and tended to increase the urinary excretion of CPZ. In addition, CPZ was recovered almost completely in the bile and urine in an unchanged form in both groups. On the other hand, LPS reduced the CLBILE and CLR of CPZ to approximately 44 and 65% of those of the control, respectively, although the renal excretion of CPZ contributed little to eliminating CPZ from the body (approximately 20% of CLSYS). The decrease in the CLSYS and increase in the MRT for CPZ as observed in the LPStreated rats may be caused by the reduction in the CLBILE. The increased urinary excretion of CPZ in the LPS-treated rats may likewise compensate for a reduction in biliary excretion ability. These findings suggest that LPS inhibits the biliary excretion pathway of CPZ in addition to decreasing the CLR. Similar outcomes have been seen in our previous studies with other drugs as well (4, 5, 14–16, 34). However, the precise mechanism for this decrease remains unclear at this stage.
The mechanism for transporting the b-lactam antibiotics from blood to bile involves various processes, including active uptake through a sinusoidal membrane into the hepatocytes, intracellular translocation, and excretion into the bile through the canalicular membrane by an active transport system (27). Tsuji and colleagues (25, 27) showed that b-lactam antibiotics, including CPZ, are taken into the hepatocytes by a carriermediated transport system and are actively excreted into the bile through the bile canalicular membrane. Utili et al. (30) demonstrated that the biliary excretion rate and maximum transport of the organic anion dye BSP, which has the same transport system as CPZ, were decreased by Escherichia coli LPS in experiments with isolated perfused rat liver, although the removal rate of BSP from the perfusing medium remained unchanged. They also reported that LPS induced an increase in the amount of BSP stored in the liver at the end of the experiments (30). Information from such studies indicates that LPS impairs the transport process of BSP from intracellular storage to the bile via the canalicular membrane rather than through sinusoidal membrane transport from blood to the hepatocytes. Our current study also showed that LPS decreases the bile flow rate, a finding which is comparable to results reported by Utili et al. (29, 30) and other investigators (33). In another study, Utili et al. (32) reported that LPS decreases the Na1, K1-ATPase activity in the canalicular membrane and leads to a decrease in the bile flow rate because of an inhibition of the bile salt-independent fraction in the bile formation, which is regulated by Na1, K1-ATPase activity (12). Kitamura et al. (11) recently reported that BSP was transported into canalicular membrane vesicles that were derived from the plasma membrane of hepatocytes by an ATP- and temperature-dependent, saturable transport process. From these reports and the results of this study, it may be speculated that LPS decreases the ATP-dependent transport of CPZ in the canalicular membrane as a result of a decrease in Na1, K1-ATPase activity. It is further known that LPS stimulates the production of superoxide and nitric oxide in the liver, which in turn induces organ injury, and it also induces increases in the release of several cytokines, such as tumor necrosis factor, interleukin-1, and interleukin-6, and induces abnormalities in arachidonic acid metabolism (8). From this we may hypothesize that the LPS-induced decrease in the biliary excretion of CPZ may be caused by such endogenous mediators; however, further studies are needed to confirm this. In summary, K. pneumoniae O3 LPS dramatically decreased the biliary excretion of CPZ. The LPS-induced decrease in the biliary excretion of CPZ may be caused by an inhibition of the anion transport system across the sinusoidal and/or bile canalicular membrane. Further studies with the sinusoidal and bile canalicular membranes from rats treated with LPS are needed to clarify the precise mechanism for the LPS-induced decrease in the biliary excretion of CPZ. The results reported here, at least, should provide further evidence regarding the biliary excretion of anionic drugs in endotoxemia and useful information for designing drug regimens for patients with gram-negative bacterial infections. REFERENCES 1. Durham, S. K., A. Brouwer, R. J. Barelds, M. A. Horan, and D. L. Knook. 1990. Comparative endotoxin-induced hepatic injury in young and aged rats. J. Pathol. 162:341–349. 2. Freudenberg, M. A., N. Freudenberg, and C. Galanos. 1982. Time course of cellular distribution of endotoxin in liver, lung and kidney of rat. Br. J. Exp. Pathol. 63:56–65. 3. Freudenberg, M. A., K. Kleine, N. Freudenberg, and C. Galanos. 1984. The fate of lipopolysaccharide in rats: evidence for chemical alteration in the molecule. Rev. Infect. Dis. 6:483–487.
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4. Hasegawa, T., M. Nadai, L. Wang, S. Haghgoo, T. Nabeshima, and N. Kato. 1994. Influence of endotoxin and lipid A on the renal handling and accumulation of gentamicin in rats. Biol. Pharm. Bull. 17:1651–1655. 5. Hasegawa, T., M. Nadai, L. Wang, Y. Takayama, K. Kato, T. Nabeshima, and N. Kato. 1994. Renal excretion of famotidine and role of adenosine in renal failure induced by bacterial lipopolysaccharide in rats. Drug Metab. Dispos. 22:8–13. 6. Hasegawa, T., M. Ohta, M. Mori, I. Nakashima, and N. Kato. 1983. The Klebsiella O3 lipopolysaccharide isolated from culture fluid: structure of polysaccharide moiety. Microbiol. Immunol. 27:683–694. 7. Hasegawa, T., M. Ohta, I. Nakashima, N. Kato, K. Morikawa, T. Hanada, and T. Okuyama. 1985. Structure of polysaccharide moiety of the Klebsiella O3 lipopolysaccharide isolated from culture supernatant of decapsulated mutant (Klebsiella O3:K12). Chem. Pharm. Bull. 33:333–339. 8. Hewett, J. A., and R. A. Roth. 1993. Hepatic and extrahepatic pathobiology of bacterial lipopolysaccharides. Pharmacol. Rev. 45:381–411. 9. Hinshaw, L. B., D. A. Reins, and R. J. Hill. 1966. Response of isolated liver to endotoxin. Can. J. Physiol. Pharmacol. 44:529–541. 10. Hirata, K., A. Kanako, K. Ogawa, H. Hayasaka, and T. Onoe. 1980. Effect of endotoxin on rat liver. Analysis of acid phosphatase isozymes in the liver of normal and endotoxin-treated rats. Lab. Invest. 43:165–171. 11. Kitamura, T., P. Jansen, C. Hardenbrook, Y. Kamimoto, and Z. Gatmaitan. 1990. Defective ATP-dependent bile canalicular transport of organic anions in mutant (TR2) rats with conjugated hyperbilirubinemia. Proc. Natl. Acad. Sci. USA 87:3557–3561. 12. Klaassen, C. D., and J. B. Watkins. 1984. Mechanisms of bile formation, hepatic uptake and biliary excretion. Pharmacol. Rev. 36:1–67. 13. Levy, E., F. C. Path, and B. H. Ruebner. 1967. Hepatic changes produced by a single dose of endotoxin in the mouse. Light microscopy and histochemistry. Am. J. Pathol. 51:269–285. 14. Nadai, M., T. Hasegawa, K. Kato, L. Wang, T. Nabeshima, and N. Kato. 1993. Influence of a bacterial lipopolysaccharide on the pharmacokinetics of tobramycin in rats. J. Pharm. Pharmacol. 45:971–974. 15. Nadai, M., T. Hasegawa, K. Kato, L. Wang, T. Nabeshima, and N. Kato. 1993. Alterations in pharmacokinetics and protein binding behavior of cefazolin in endotoxemic rats. Antimicrob. Agents Chemother. 37:1781–1785. 16. Nadai, M., T. Hasegawa, K. Kato, L. Wang, T. Nabeshima, and N. Kato. 1993. The disposition and renal handling of enprofylline in endotoxemic rats by bacterial lipopolysaccharide (LPS). Drug Metab. Dispos. 21:16–21. 17. Nadai, M., T. Hasegawa, L. Wang, O. Tagaya, and T. Nabeshima. 1994. Alterations in the pharmacokinetics and protein binding of enprofylline in Eisai hyperbilirubinemic rats. Drug Metab. Dispos. 22:561–565. 18. Nakashima, E., R. Matsushita, M. Takeda, T. Nakanishi, and F. Ichimura. 1992. Comparative pharmacokinetics of cefoperazone and cephradine in untreated streptozotocin diabetic rats. Drug Metab. Dispos. 20:730–735. 19. Neu, H. C. 1981. A review and summary of the pharmacokinetics of cefoperazone: a new, extended-spectrum b-lactam antibiotic. Ther. Drug Monit. 3:121–128. 20. Nolan, J. P., and C. J. O’Connell. 1965. Vascular response in the isolated rat liver. I. Endotoxin, direct effects. J. Exp. Med. 122:1063–1073. 21. Ohta, M., M. Mori, T. Hasegawa, F. Nagase, I. Nakashima, S. Naito, and N. Kato. 1981. Further studies of polysaccharide of Klebsiella pneumoniae possessing strong adjuvanticity. I. Production of the adjuvant polysaccharide by noncapsulated mutant. Microbiol. Immunol. 25:939–948.
2261
22. Praaning-van Dalen, D. P., A. Brouwer, and D. L. Knook. 1981. Clearance capacity of rat liver Kupffer, endothelial and parenchymal cells. Gastroenterology 81:1036–1044. 23. Ruiter, D. J., J. Van Der Meulen, A. Broywer, W. J. R. Hummel, B. J. Mauw, J. C. M. Van Der Ploeg, and E. Wisse. 1981. Uptake by liver cells of endotoxin following its intravenous injection. Lab. Invest. 45:38–45. 24. Sathirakul, K., H. Suzuki, K. Yasuda, M. Hanano, O. Tagaya, T. Horie, and Y. Sugiyama. 1993. Kinetic analysis of hepatobiliary transport of organic anions in Eisai hyperbilirubinemic mutant rats. J. Pharmacol. Exp. Ther. 265:1301–1312. 25. Tamai, I., T. Maekawa, and A. Tsuji. 1990. Membrane potential-dependent and carrier-mediated transport of cefpiramide, a cephalosporin antibiotic, in canalicular rat liver plasma membrane vesicles. J. Pharmacol. Exp. Ther. 253:537–544. 26. Tsuji, A., T. Terasaki, N. Imaeda, and E. Nakashima. 1985. Effect of extracellular water volume on the distribution kinetics of b-lactam antibiotics as a function of age. J. Pharmacobio-Dyn. 8:167–174. 27. Tsuji, A., T. Terasaki, K. Takanosu, I. Tamai, and E. Nakashima. 1986. Uptake of benzylpenicillin, cefpiramide and cefazolin by freshly prepared rat hepatocytes. Evidence for a carrier-mediated transport system. Biochem. Pharmacol. 35:151–158. 28. Tsuji, A., T. Yoshikawa, K. Nishide, H. Minami, M. Kimura, E. Nakashima, T. Terasaki, E. Miyamoto, C. H. Nightingale, and T. Yamana. 1983. Physiologically based pharmacokinetic model for b-lactam antibiotics. I. Tissue distribution and elimination in rats. J. Pharm. Sci. 72:1239–1252. 29. Utili, R., C. O. Abernathy, and H. J. Zimmerman. 1976. Cholestatic effect of Escherichia coli endotoxin on the isolated perfused rat liver. Gastroenterology 70:248–253. 30. Utili, R., C. O. Abernathy, and H. J. Zimmerman. 1977. Studies on the effects of E. coli endotoxin on canalicular bile formation in the isolated perfused rat liver. J. Lab. Clin. Med. 89:471–482. 31. Utili, R., C. O. Abernathy, and H. J. Zimmerman. 1977. Endotoxin effects on the liver. Life Sci. 20:553–568. 32. Utili, R., C. O. Abernathy, and H. J. Zimmerman. 1977. Inhibition of Na1, K1-adenosinetriphosphatase by endotoxin: a possible mechanism for endotoxin-induced cholestasis. J. Infect. Dis. 136:583–587. 33. Van Bossuyt, H., R. B. De Zanger, and E. Wisse. 1988. Cellular and subcellular distribution of injected lipopolysaccharide in rat liver and its inactivation by bile salts. J. Hepatol. 7:325–337. 34. Wang, L., T. Hasegawa, M. Nadai, I. Muraoka, T. Nabeshima, and N. Kato. 1993. The effect of lipopolysaccharide on the disposition of xanthines in rats. J. Pharm. Pharmacol. 45:34–38. 35. Watkins, J. B., and T. P. Dykstra. 1987. Alteration in biliary excretory function by streptozotocin-induced diabetes. Drug Metab. Dispos. 15:177– 183. 36. Watkins, J. B., and H. Noda. 1986. Biliary excretion of organic anions in diabetic rats. J. Pharmacol. Exp. Ther. 239:467–473. 37. Yamaguchi, Y., K. Yamaguchi, J. L. Babb, and H. Gans. 1982. In vivo quantitation of the rat liver’s ability to eliminate endotoxin from portal vein blood. J. Reticuloendothel. Soc. 32:409–422. 38. Yamaoka, K., Y. Tanigawara, T. Nakagawa, and T. Uno. 1981. A pharmacokinetic analysis program (MULTI) for microcomputer. J. PharmacobioDyn. 4:879–885.
