Formed and preformed metabolite excretion clearances in liver,a metabolite formation organ: Studies on enalapril and enalaprilat in the single-pass and recirculating perfused rat liver

Single-pass and recirculating rat liver perfusion studies were conducted with [¹⁴C]enalapril and [³H] enalaprilat, a precursor-product pair, and the data were modeled according to a physiological model to compare the different biliary clearances for the solely formed metabolite, [¹⁴C]enalaprilat, with that of preformed [³H]enalaprilat. With single-pass perfusion, the apparent extraction ratio (or biliary clearance) of formed [¹⁴C]enalaprilat was 15-fold the extraction ratio of preformed [³H] enalaprilat, an observation attributed to the presence of a barrier for cellular entry of the metabolite. Upon recirculation of bolus doses of [¹⁴C]enalapril and [³H]enalaprilat, the biliary clearance, estimated conventionally as metabolite excretion rate/midtime metabolite concentration, for formed [¹⁴C]enalaprilat was again 10-to 15-fold higher than the biliary clearance for preformed [³H]enalaprilat, but this decayed with perfusion time and gradually approached values for preformed [³H]enalaprilat. The decreasing biliary clearance of formed enalaprilat with recirculation was explained by the dual contribution of the circulating and intrahepatic metabolite (formed from circulating drug) to excretion. Physiological modeling predicted (i) an influx barrier (from blood to cell) at the sinusoidal membrane as the rate-limiting process in the overall removal of enalaprilat, (ii) a 15-fold greater extraction ratio or biliary clearance for formed [¹⁴C]enalaprilat over [³H]enalaprilat during single-pass perfusion, and (iii) the time-dependent and declining behaviour of the biliary clearance for formed [¹⁴C]enalaprilat during recirculation of the medium. In the absence of a direct knowledge of eliminating organs in vivo, this variable pattern for excretory clearance of the formed metabolite within the organ is indicative of a metabolite formation organ.Glossary C _R denotes the reservoir concentration - C _In andC _Out,L respectively, denote the input and output concentrations. - Q _L is the total hepatic plasma flow rate. - Q _Bile is the bile flow rate - f _p and f_L denote the unbound fractions in plasma and liver tissue, respectively - C_p is the concentration in renal plasma; C_L is the concentration in liver; - C _Bile is the concentration in bile. - v _R,V _p,V _L, andV _Bile denote the reservoir plasma, hepatic plasma, tissue, and bile volumes, respectively - CL _d ⁱⁿ andCL _d ^ef denote the influx and efflux clearances, respectively - CL _int,L ^m ,L represents the hepatic metabolic intrinsic clearance of the drug - CL _int,L ^b L denotes the biliary intrinsic clearance This work was supported by the Medical Research Council of Canada. I. A. M. de Lannoy was a recipient of the Ontario Graduate Fellowship from the Ontario Ministry of Health; K. S. Pang was a recipient of the Faculty Development Award, Medical Research Council, Canada.     

A physiological model for renal drug metabolism: Enalapril esterolysis to enalaprilat in the isolated perfused rat kidney

A physiologically based kidney model was developed to describe the metabolism of enalapril and explain the observed discrepancies between generated and preformed enalaprilat (metabolite) elimination in the constant flow single-pass and recirculating isolated perfused rat kidney preparations (IPKs) as a result of the differing points of origin of the metabolite within the kidney, subsequent to the simultaneous delivery of¹⁴Cenalapril and³H-enalaprilat. The model incorporated clearances for diffusion/transport of drug and metabolite across the basolateral and luminal membranes of the renal cells, an intrinsic clearance for renal drug metabolism, in addition to physiological variables such as perfusate flow rate, glomerular filtration rate, and urine flow rate. Nonlinear curve fitting of single-pass and recirculating data was performed to estimate the rate-limiting step in the renal elimination of enalaprilat. Through fitting and simulation procedures, we were able to predict metabolic and excretory events for enalapril (renal extraction ratio 0.25–0.3; fractional excretion, FE, was less than unity) and the relatively constant pattern of urinary excretion of preformed enalaprilat (extraction ratio 0.07; FE1). The extraction ratio of the intrarenally formed enalaprilat in single-pass IPK was about twofold that for the preformed metabolite, whereas the FEs of generated enalaprilat in recirculating IPKs were >1, and tended to increase, then decrease with perfusion time. These observations were explained by the optimized parameters which indicated that efflux from cell to lumen was rate-controlling in the excretion of enalaprilat, and another small transport barrier also existed at the basolateral membrane; the lower extraction ratio of preformed enalaprilat was due to its poor transmembrane clearance at the basolateral membrane. The variable FEs for generated enalaprilat vs. the relatively constant FE for preformed metabolite in the recirculating IPK was explained by the changing contributions of both circulating and intrarenal metabolite to metabolite excretion.This work was supported by the Medical Research Council of Canada. I. A. M. de Lannoy was a recipient of the Ontario Graduate Scholarship, and K. S. Pang is a recipient of the Faculty Development Award from MRC, Canada M5S 2S2.     

A physiological model for renal drug metabolism: enalapril esterolysis to enalaprilat in the isolated perfused rat kidney

A physiologically based kidney model was developed to describe the metabolism of enalapril and explain the observed discrepancies between generated and preformed enalaprilat (metabolite) elimination in the constant flow single-pass and recirculating isolated perfused rat kidney preparations (IPKs) as a result of the differing points of origin of the metabolite within the kidney, subsequent to the simultaneous delivery of 14C-enalapril and 3H-enalaprilat. The model incorporated clearances for diffusion/transport of drug and metabolite across the basolateral and luminal membranes of the renal cells, an intrinsic clearance for renal drug metabolism, in addition to physiological variables such as perfusate flow rate, glomerular filtration rate, and urine flow rate. Nonlinear curve fitting of single-pass and recirculating data was performed to estimate the rate-limiting step in the renal elimination of enalaprilat. Through fitting and simulation procedures, we were able to predict metabolic and excretory events for enalapril (renal extraction ratio approximately equal to 0.25-0.3; fractional excretion, FE, was less than unity) and the relatively constant pattern of urinary excretion of preformed enalaprilat (extraction ratio approximately equal to 0.07; FE approximately equal to 1). The extraction ratio of the intrarenally formed enalaprilat in single-pass IPK was about twofold that for the preformed metabolite, whereas the FEs of generated enalaprilat in recirculating IPKs were greater than 1, and tended to increase, then decrease with perfusion time. These observations were explained by the optimized parameters which indicated that efflux from cell to lumen was rate-controlling in the excretion of enalaprilat, and another small transport barrier also existed at the basolateral membrane; the lower extraction ratio of preformed enalaprilat was due to its poor transmembrane clearance at the basolateral membrane. The variable FEs for generated enalaprilat vs. the relatively constant FE for preformed metabolite in the recirculating IPK was explained by the changing contributions of both circulating and intrarenal metabolite to metabolite excretion.     

Inhibition of esterolysis of enalapril by paraoxon increases the urinary clearance in isolated perfused rat kidney.

The effect of competing elimination pathways on the metabolic and excretory clearance estimates was examined with tracer concentrations of [(3)H]enalapril, which was both metabolized and excreted by the rat kidney. Perturbation was achieved with use of the carboxylesterase inhibitor paraoxon, which inhibited [(3)H]enalapril metabolism to [(3)H]enalaprilat in rat renal S9 fraction. At 0.1, 0.5, 1, and 10 microM paraoxon, esterolysis of enalapril was inhibited by 76 +/- 7, 93 +/- 5, 96 +/- 5, and 93 +/- 6%, respectively. The lowest concentration (0.1 microM) of paraoxon was chosen for single-pass isolated perfused kidney (IPK) studies because viability was least compromised, and the sodium and glucose reabsorptive functions of the IPK remained constant. After an equilibration period (15-20 min at constant pressure, 90-100 mm Hg), perfusion of the rat kidney with [(3)H]enalapril was carried out under constant flow (8 ml/min) for 30 min in the absence and presence of paraoxon (0.1 microM). The metabolic (from 1.83 +/- 0.52 to 1.48 +/- 0.47 ml/min/g) and total renal (from 1.87 +/- 0.46 to 1. 57 +/- 0.41 ml/min/g) clearances of [(3)H]enalapril in the IPKs were decreased significantly (p <.05) in the presence of paraoxon when compared with controls. Concomitantly, the urinary clearance (from 0. 04 +/- 0.07 to 0.09 +/- 0.09 ml/min/g) and the fractional excretion (from 0.23 +/- 0.18 to 0.52 +/- 0.25) of [(3)H]enalapril doubled (p <.05). The study illustrates that a reduction in cellular metabolism of the kidney brings forth a rise in the estimate of clearance of its complimentary pathway, estimate of the excretory (urinary) clearance.     

Organ Clearance Concepts: New Perspectives on Old Principles

The removal capacity of an eliminating organ by metabolism and/or excretion is often expressed as its clearance. Metabolic and excretory clearances are considered to be mutually independent, and the sum of these constitute the whole organ clearance. The influence of metabolism on estimates of the excretory clearance and vice versa was examined for the liver and kidney with physiologically based models. Mass transfer first-order rate equations describing transport and removal were derived. Upon inversion of the matrices originating from the coefficients of these equations, the area under the curve (AUC) and clearance (dose/AUC) were obtained with the liver or kidney as the eliminating organ. A more complex solution was found to exist for the kidney since glomerular filtration, secretion; reabsorption. and intrarenal metabolism were present. To ascertain the effect of excretion on estimates of the metabolic clearance as well as the effect of metabolism on estimates of the excretory clearance, intrinsic clearances for excretion or metabolism were set to zero. Clearance values were found to be altered when alternate pathways were present Whereas excretory clearance estimates were consistently reduced in the presence of metabolism, metabolic clearance estimates were affected differentially by excretion and varied according to the site of metabolism. Excretion reduced metabolic clearance estimates when metabolism occurred intracellularly. If metabolism occurred intraluminally (e.g., on the renal brush border or luminal membrane), the metabolic clearance estimate could become higher since the substrate was made available to the enzymes following its excretion. As expected, these changes depended on the relative magnitudes of the intrinsic clearances for metabolism and excretion. The above theory was applied to the elimination of enalapril which is both metabolized and excreted by the perfused rat liver and kidney preparations. Data obtained in these studies were consistent with a set of published physiologic parameters denoting transfer and intrinsic clearances. Perturbations on clearance estimates were studied by setting the metabolic/excretory intrinsic clearance to zero, then to some finite value. In liver, the avid hepatocellular metabolism of enalapril reduced biliary clearance by 73%. For the kidney, the fractional excretion (FE or unbound excretory clearance/glomerular filtration rate) was decreased modestly (from 0.64 to 0.44) with intracellular esterolysis, whereas if metabolism had occurred intraluminally. FE would have been significantly decreased (from 1.8 to 0.45). Simulation results show clearly that clearance estimates are affected by the presence of alternate removal pathways, and question the well-established principle that metabolic and excretory clearance estimates are independent of each other.     

Presence of a diffusional barrier on metabolite kinetics: enalaprilat as a generated versus preformed metabolite

Studies in the once-through perfused rat liver with the simultaneous delivery of 14 C-enalapril and its polar diacid metabolite, 3H-enalaprilat, revealed different extents of elimination (exclusively by biliary excretion) for the generated (14C-enalaprilat) and preformed (3H-enalaprilat) metabolite (18 and 5% dose) [Pang, Cherry, Terrell, and Ulm: Drug Metab. Dispos. 12, 309-313 (1984)]. The present re-examination of data provided an explanation for these discrepant observations: enalaprilat, being a polar dicarboxylic acid, encounters more of a diffusional barrier than its precursor, enalapril, an ethyl ester of enalaprilat. Programs written in Fortran 77 on mass balance relationships were employed to simulate data upon varying the diffusional clearances for drug (CLd) and metabolite [CLd(mi)] from 1 to 5000 ml/min. The metabolic and biliary intrinsic clearances for drug and metabolite were found by trial and error such that the combinations of all clearance parameters yielded data similar to enalaprilat, and 3H-enalaprilat. Our finding indicated that the diffusional clearance for enalaprilat was low (2 ml/min) compared to that of enalapril (75 ml/min). The presence of a diffusional barrier for enalaprilat retards entry of the preformed metabolite into hepatocytes but prevents efflux of the intracellularly formed generated metabolite into sinusoidal blood, thereby enhancing generated metabolite elimination.     

A comparative investigation of hepatic clearance models: Predictions of metabolite formation and elimination

Liver clearance models serve to improve our understanding of the relationships between the physiological determinants and hepatic clearance and predict changes in the disposition of substrates when homeostasis of the organ is perturbed. Their ability to describe metabolism was presently extended to the sequential formation and elimination of primary (M₁), secondary (M₂), and tertiary (M₃) metabolites during a single passage of drug (P) across the liver, under steady state and first-order conditions. The well-stirred model is distinct from other models in that metabolite formation and elimination is independent of enzymic distributions, the number of steps involved in metabolite formation, and the intrinsic clearances of the precursors. This model predicts that the extraction ratio of a formed primary metabolite derived from drug (E{M₁, P}) is identical to that for the preformed primary metabolite (E{M₁}), and that the extraction ratios of a secondary metabolite derived from drug (E{M₂, P}) and primary metabolite (E{M₂, M₁}) or preformed secondary metabolite (E{M₂}) are identical. For the more physiologically acceptable, parallel-tube and dispersion models, metabolite sequential elimination is highly influenced by the intrinsic clearances of the precursors and the enzymic distributions that mediate removal of precursor species and the metabolites. Furthermore, the extent of sequential metabolism recedes as the number of steps involved for metabolite formation increases. These models predict that E{M₁, P}1}, and E{M₂, P}2, M₁}2}, with the magnitude of the changes being less for the dispersion model than for the parallel-tube model. Competing pathways that divert substrate from entering the sequential pathway were found to exert only minimal influence on the sequential pathway.This work was supported by the Medical Research Council, Canada (DG-263, MA-9104, and MA-9765), and USHHS (NIH, GM-38250). M. V. St-Pierre was a recipient of the University Toronto Open Fellowship and the Ontario Ministry of Health Fellowship,. K. S. Pang and P. I. Lee are recipients of Faculty Development Awards, MRC, Canada.     

Effect of diffusional barriers on drug and metabolite kinetics

Previous experimental and simulation studies have alluded to the presence of a diffusional barrier for enalaprilat, the polar, dicarboxylic acid metabolite of enalapril, entering hepatocytes. The present study examined the roles of diffusional clearances of drug and metabolite on the distribution and elimination characteristics in liver. The hepatic intrinsic clearances for enalapril (26.1 ml/min) and enalaprilat (0.7 ml/min), found in a previous study, were used for simulation because, along with their given diffusional clearances (75 and 2 ml/min, respectively), they yielded a high extraction ratio for drug (E = 0.86) and a poor extraction ratio for the preformed metabolite (E = 0.05). While maintaining the intrinsic clearances and hepatic blood flow rate (10 ml/min) constant, only drug and metabolite diffusional clearances were altered. The liver was modeled as three (blood, liver tissue, and bile) compartments, with blood flowing into sinusoids of uniform length L. Blood (sinusoidal) and tissue concentrations of drug and generated and preformed metabolites, at any point x along L and under linear kinetic conditions, were approximated numerically by computer simulations and expressed as the length-averaged or mean concentrations. The factors underlying drug and metabolite (preformed and generated) concentrations, hepatic clearances and elimination rates, and their interrelationships were illustrated graphically, emphasizing the roles of diffusional clearances for drug and metabolite on their spatial distributions and elimination in liver.     

Pharmacokinetic study of the phase III,randomized, double-blind,multicenter trial (TRIBUTE) of paclitaxel and carboplatin combined with erlotinib or placebo in patients with advanced Non-small Cell Lung Cancer (NSCLC)

Purpose To assess the pharmacokinetics and evaluate potential drug-drug interactions between erlotinib, paclitaxel and carboplatin. Experimental Design 1,079 previously untreated patients with advanced NSCLC were enrolled and randomized in a phase III trial (TRIBUTE) to receive either erlotinib or placebo in combination with paclitaxel 200 mg/m2 IV over 3 h and carboplatin at a calculated dose to achieve an AUC 6 mg∙min/mL. To determine possible drug-drug interaction with this combination, a subset of 24 (12 erlotinib, 12 placebo) patients were enrolled onto an intensive pharmacokinetic (IPK) substudy group at a single site. All IPK patients received either erlotinib 150 mg/day or placebo-controlled tablets. Analyses were completed using validated analytical methodologies. Non-compartmental modeling was utilized to estimate PK parameters. Results Complete blood sampling for pharmacokinetic analysis was obtained in 21 of 24 patients. Mean AUC_{0 - t} {\hbox{AU}}{{\hbox{C}}_{0 - \tau }} for erlotinib and the OSI-420 metabolite were 29,997 ng∙h/mL and 3,020 ng∙h/mL, respectively. Mean (SD) paclitaxel clearances (L/h/M²) were 11.7 (3.4) and 12.7 (6.7) in the placebo and erlotinib treatment groups, respectively. The resultant paclitaxel AUC_{0 - ￥} {\hbox{AU}}{{\hbox{C}}_{0 - \infty }} (ng∙h/mL) was 18,400 (5,300) for the placebo group and 17,800 (5,500) for the erlotinib group. For carboplatin, the mean (SD) clearances (L/h) were 16.8 (3.9) and 16.1 (4.4) for the placebo and erlotinib groups, respectively. The resultant carboplatin AUC_{0 - ￥} {\hbox{AU}}{{\hbox{C}}_{0 - \infty }} (ng/mL∙h) were 49,900 (9,700) for the placebo group and 48,400 (11,900) for the erlotinib group. No significant differences were observed in these paclitaxel or carboplatin pharmacokinetic group comparisons. Conclusions The addition of erlotinib to a standard chemotherapy regimen for NSCLC did not alter the systemic exposures (AUC_{0 - ￥} {\hbox{AU}}{{\hbox{C}}_{0 - \infty }} ) of paclitaxel (p = 0.80) and carboplatin (p = 0.756) when erlotinib-treated patients were compared to placebo-treated patients. The pharmacokinetics of erlotinib and its metabolite OSI-420 did not appear to be altered by the concomitant administration of paclitaxel and carboplatin.     

Predicting the hepatic clearance of xenobiotics in humans from in vitro data

Values for V_max and K_m determined during the in vitro metabolism of a xenobiotic to a known metabolite by a specific human isozyme of cytochrome P450 (P450) were used to predict the hepatic clearance (CL_H) of the xenobiotic to that metabolite. The calculated CL_H values were then compared to literature values of clearance (CL) to the same metabolite obtained during pharmacokinetic studies in humans. For the 6-hydroxylation of chlorzoxazone (P450 2E1) the predicted and actual clearances were 110±77 mL min^?1 and 110 mL min^?1, respectively. For the 6β-hydroxylation of cortisol, the deethylation of lidocaine (two studies), and the oxidation of nifedipine (all P450 3A3/4) the values were 13±15 mL min^?1 and 13 mL min^?1; 758±282 or 829±283 mL min^?1 and 875 mL min^?1; and 284±176 mL min^?1 and 294 mL min^?1, respectively. An increase to 72±25 mL min^?1 in the CL_H of cortisol to 6β-hydroxycortisol was calculated following rifampicin treatment. Finally, the polymorphic nature of the metabolism (P450 2D6) of mexiletine was confirmed. The usefulness of the method and its limitations are discussed.     

Radioimmunoassay for the quantitation of lisinopril and enalaprilat

A sensitive radioimmunoassay (RIA) capable of measuring either lisinopril (1-[N2-[(S)-1-carboxy-3-phenylpropyl]-L-lysyl] -L-proline) or enalaprilat (1-[N-[(S)-1-carboxy-3-phenylpropyl]-L-alanyl] -L-proline), the active metabolite of enalapril has been developed. A suitable antiserum was raised against an immunogen prepared from conjugation of lisinopril, the lysyl analogue of enalapril, with succinoylated keyhole limpet hemocyanin. A novel radiotracer was also prepared for use in the assay by acylation of the epsilon amine group on the lysyl side chain of lisinopril with N-succinimidyl [2,3-3H]propionate. The antiserum was used at a final dilution of 1:44,500 and the sensitivity of the assay for enalaprilat was estimated at 2 pmol/mL plasma sample and 0.4 pmol/mL for lisinopril. Enalapril, the ethyl ester of enalaprilat, exhibited little cross-reactivity (0.005%), and several other compounds (captopril, proline, lysine, tyrosine, hippuric acid, and tryptophan) were found not to crossreact. In rabbits given a 2.03 mumol/kg iv dose of enalapril, plasma concentrations of enalaprilat were determined by the RIA technique and compared with an estimation of the enalaprilat concentrations derived from the extent of inhibition of plasma angiotensin converting enzyme (ACE). The plasma levels estimated by ACE inhibition were less than those obtained by the RIA in the first 45 min but were always greater in the samples taken after this time. Both assay methods showed that the conversion of enalapril to enalaprilat was rapid, and also indicated that there was initial rapid clearance of enalaprilat from the plasma.     

Clinical pharmacokinetics of procainamide infusions in relation to acetylator phenotype

The pharmacokinetics of procainamide was determined in 21 lidocaine-resistant patients who received the drug according to a pharmacokinetically designed double-infusion technique. Thirteen patients were phenotyped as slow acetylators, seven as fast, and one as intermediate. The total body clearances (Cl_T) of PA in slow and fast acetylators were 22.6 and 34.8 liters/hr, respectively. The fraction of PA cleared by the formation of NAPA in the corresponding acetylator group was 0.2 and 0.4. Renal impairment affected the pharmacokinetics of PA more profoundly as the Cl_Ts of PA in patients with and without renal impairment were 17.9 and 31.2 liters/hr, respectively. None of the calculated volumes of distribution was affected by acetylator phenotype or renal impairment. These data identify the contribution of at least two of the major factors accounting for variability in PA disposition in patients undergoing therapy.Glossary PA Procainamide - NAP A N-Acetylprocainamide - LI Loading infusion - MI Maintenance infusion - V _p Volume of central compartment for PA - V _t Volume of tissue compartment for PA - V _ss ^d Volume of distribution of PA at steady state - V _d Volume of distribution of PA during postdistributive phase - Cl₁₂ Intercompartmental clearance of PA - Cl _T Total body clearance of PA - Cl _R Renal clearance of PA - Cl _A Acetylation clearance of PA - Cl_AP Apparent acetylation clearance of PA - Cl _M Metabolic (nonacetylation) clearance of PA - C _PA Serum concentration of PA - C _t Tissue concentration of PA - C _PA ^ss Steady-state serum concentration of PA - V _n Volume of distribution of NAPA - Cl _N Renal clearance of NAPA - Cl₀ Nonrenal clearance of NAPA - Cl_BN Total body clearance of NAPA - C _NAPA Serum concentration of NAPA - C _N ^ss Steady-state serum concentration of NAPA This work was supported by Grants 20852 and 24211 from the National Institutes of General Medical Sciences, National Institutes of Health.     

High dose methotrexate population pharmacokinetics and Bayesian estimation in patients with lymphoid malignancy

The purpose of present study was to develop a population pharmacokinetic model of high dose methotrexate (HD‐MTX) infusion in patients with lymphoid malignancy, to investigate the biological and clinical covariates related to the drug distribution and elimination. It is also the purpose to propose a limited sampling strategy (LSS) for the estimation of the time above the threshold (0.2 µmol·L^?1). A total 82 patients with lymphoid malignancy were involved in the study. A pharmacokinetic model was developed using nonlinear mixed‐effect model. The influence of demographic characteristics, biological factors, and concurrent administration were investigated. The final predictive performance was validated by bootstrap and cross‐validation. Bayesian estimation was evaluated. The pharmacokinetics of HD‐MTX was described by a two‐compartment model. The pharmacokinetic parameters and the inter‐individual variability were as follows: the clearance CL, 7.45 L·h^?1 (inter‐individual variability 50.6%), the volume of the central and peripheral compartment V₁, 25.9 L (22.5%), V₂, 9.23 L (97.8%), respectively, and the intercompartmental clearance Q, 0.333 L·h^?1 (70.4%). The influence of serum creatinine on CL and weight on V₁ was retained in the final model. The protocol involved one sampling time at 44 h after the start of the infusion, allowing one to predict the time at which the MTX concentration reached the expected threshold (0.2 µmol·L^?1). Serum creatinine and weight showed significant influence on methotrexate CL and V₁, respectively. Furthermore, a Bayesian estimation based on the covariates and 44 h sample was developed, allowing prediction of the individual methotrexate pharmacokinetic parameters and the time to 0.2 µmol·L^?1. Copyright © 2009 John Wiley & Sons, Ltd.     

Renal handling of oxalate a micropuncture study in the rat

Summary Clearance and micropuncture experiments were performed in rats to study the renal handling of oxalate. The ¹⁴C-oxalate to ³H-inulin clearance ratio (C _ox /C _in ) was 1.36±0.04 and was lowered by probenecid (200 mg/kg) to 1.11±0.03 (±S.E., n=6, P<0.005). an attempt was made to localize the assumed secretion of oxalate in three different micropuncture protocols. In free flow micropuncture experiments single nephron clearance of oxalate was not different when obtained from proximal or distal tubular puncture sites. The fractional delivery of oxalate averaged 0.84 ±0.03 regardless of the puncture site from midproximal to late distal. This finding indicates a net outflux of oxalate in an early proximal loop since oxalate is freely ultrafilterable. In microperfusion experiments the mean recovery of oxalate ranged from 79–90%. The outflux of oxalate correlated linearly with the tubular load (r=0.95). The results suggest that no net secretion occurs in superficial nephron segments accessible for micropuncture. Since whole kidney clearances of oxalate always exceeded glomerular filtration rate, it is concluded that net addition of oxalate into the tubular fluid can occur at sites beyond the superficial late distal tubules or is due to higher delivery of oxalate by deep cortical nephrons.This work was generously supported by the Martin Brinkmann Foundation, Bremen, West Germany     

中国成年患者万古霉素群体药动学研究

目的:利用万古霉素治疗药物监测(TDM)数据建立群体药动学(PPK)模型,用于估算个体化药动学参数。方法:选择使用万古霉素成年患者,详细记录用药、TDM数据以及病理生理资料。采用非线性混合效应模型(NONMEM)法建立万古霉素群体药动学模型。结果:169例患者数据来源于血液科及重症监护(ICU)病房等9个科室,共获得385个血药浓度数据,其中峰浓度39个,谷浓度346个。根据文献资料及TDM数据建立二室PPK模型,万古霉素清除率(CL)、中央室(V1)及外周室(V2)分布容积、室间清除率分别为4.08 L·h-1、21.7 L、65.3 L、5.95 L·h-1,患者肌酐清除率及体重分别对CL及V2具有显著影响。根据模型预测169位患者AUC0-24h为(450.1±231.8)mg·L-1·h。结论:本研究建立的万古霉素PPK模型可以用于中国成年患者个体化药动学参数估算。     

Nonlinear elimination and hepatic concentration of conjugation- metabolite of valproate in guinea-pigs

The plasma clearance and metabolic rate characteristics of valproic acid (VPA) were studied using guinea-pigs placed on various (0.08-9 μmol ml^?1 = 11–1303 μg ml^?1) steady-state plasma concentrations (C_ss) by constant intravenous (i.v.) infusion. The total clearance (CL) was significantly decreased at plasma concentration of 0.61 μmol ml^?1 (88 μg ml^?1). The metabolic clearance of VPA was apparently biphasic. The maximum metabolic rate (V_max) and the Michaelis-Menten constant (K_m) for the primary (V_maxl, K_ml) and the secondary (V_max2, K_m2) pathways were V_maxl = 1.52 μmol min ^?1kg^?1, K_ml = 0.15 μmol ml^?1, V_max2 = 24.98 μmol min ^?1 kg^?1 and K_m2 = 11.70 μmol ml^?1, respectively. The K_ml value was within clinical therapeutic concentration range. The formation of conjugated VPA (cjVPA) metabolite in liver was shown to be saturable. Plasma protein binding of VPA was also nonlinear. The dose-dependent decrease in metabolic clearance was counterbalanced by the increased unbound fraction (f_u), resulting in a relatively constant apparent clearance of VPA over a wide concentration range. The hepatic concentration of VPA was not significantly different from the plasma unbound concentration, again over a wide concentration range. The biliary and hepatic concentrations of VPA were not significantly different; but the concentration ratio of cjVPA in bile compared with that of VPA in liver decreased against hepatic concentration of VPA, which suggests a saturable conjugation rate. The K_m value estimated from hepatic cjVPA production as a function of plasma VPA concentration was comparable with the K_ml value. These results implied that the primary metabolic parameters may describe the conjugation pathway which is nonlinear within the clinical therapeutic concentration range.     

Age and the pharmacokinetics of angiotensin converting enzyme inhibitors enalapril and enalaprilat.

The pharmacokinetics of angiotension converting enzyme (ACE) inhibitors enalapril (10 mg orally) and its active metabolite, enalaprilat (10 mg intravenously) were studied in nine young healthy volunteers aged 22-30 years and nine sex matched elderly subjects aged 65-73 years. After both drugs, a biexponential curve was fitted to the decline in plasma enalaprilat concentration. Area under the plasma concentration-time curve (AUC) was greater in the elderly for both drugs. Clearance (CL) and clearance/bioavailability (CL/F) were less in the elderly for enalaprilat and enalapril, respectively. There was no difference in F between young (0.62 +/- 0.16) and elderly subjects (0.61 +/- 0.15). Enalaprilat CL and enalapril CL/F were significantly and positively correlated to endogenous creatinine clearance. There was a significant difference in the weight corrected volume of distribution at steady state after enalaprilat between the young and elderly (P less than 0.02). The relationship between plasma enalaprilat concentrations and percentage ACE inhibition, using the Hill equation, showed no difference in the sensitivity to ACE inhibition between the young and the elderly group. The pharmacokinetic differences observed are likely to be related to an age dependent decline in renal function as well as changes in body composition. Kinetic differences partly explain the greater pharmacodynamic response in the elderly.     

Primary,secondary, and tertiary metabolite kinetics

Because of the propensity of nascently formed metabolites towards sequential metabolism within formation organs, theoretical and experimental treatments that achieve mass conservation must recognize the various sources contributing to primary, secondary, and tertiary metabolite formation. A simple one-compartment open model, with first-order conditions and the liver as the only organ of drug disappearance and metabolite formation, was used to illustrate the metabolism of a drug to its primary, secondary, and tertiary metabolites, encompassing the cascading effects of sequential metabolism. The concentration-time profiles of the drug and metabolites were examined for two routes of drug administration, oral and intravenous. Formation of the primary metabolite from drug in the gut lumen, with or without further absorption, and metabolite formation arising from first-pass metabolism of the drug and the primary metabolite during oral absorption were considered. Mass balance equations, incorporating modifications of the various absorption and conversion rate constants, were integrated to provide the explicit solutions. Simulations, with and without consideration of the sources of metabolite formation other than from its immediate precursor, were used to illustrate the expected differences in circulating metabolite concentrations. However, a simple relationship between the area under the curve of any metabolite, M,or [AUC{m}],its clearance [CL{m}],and route of drug administration was found. The drug dose, route, fraction absorbed into the portal circulation, F_abc,fraction available of drug from the liver, F,availabilities of the metabolites F{m}from formation organs, and CL{m}are determinants of the AUC{m}'s.After iv drug dosing, the area of any intermediary metabolites is determined by the iv drug dose divided by the (CL{m}/F{m})of that metabolite. When a terminal metabolite is not metabolized,its area under the curve becomes the iv dose of drug divided by the clearance of the terminal metabolite since the available fraction for this metabolite is unity. Similarly, after oral drug administration, when loss of drug in the gut lumen does not contribute to the appearance of metabolites systemically, the general solution for AUC{m} isthe product of F_abc and oral drug dose divided by [CL{m}/F{m}].A comparison of the area ratios of any metabolite after po and iv drug dosing, therefore, furnishes F_abc.When this fraction is divided into the overall systemic availability or F_sys,the drug availability from the first-pass organs, F,may be found. The potential application of these relationships to other schemes, namely, drugs that have competing metabolic pathways within the liver and/or intestine as well as reversible metabolism is briefly discussed.In view of the various contributing sources of metabolite formation, and the modulation of circulating metabolite concentrations by sequential first-pass metabolism of the metabolite, caution is given against the use of area ratios of metabolite after iv drug and metabolite administration for estimations of metabolite formation clearances.This work was supported by the Medical Research Council of Canada (MA-9104 and MA-9765) and the NIH (GM-38250). KSP is a recipient of the Faculty Development Award from MRC, Canada.     

Effect of chronic oral furosemide administration on the 24-hour cycle of lithium clearance and electrolyte excretion in humans

Summary The effect of chronic furosemide treatment on the circadian cycle of lithium clearance (CL_Li) and electrolyte excretion has been examined in 8 young, male volunteers, by performing two 24 h clearance experiments, before and after one week of treatment with furosemide 80 mg once daily.After 8 days on furosemide there was a significant decrease in creatinine clearance (–21%), plasma Na (–8.4 mM) and plasma K (–0.58 mM). At that time, however, there were no changes in 24 h-values of CL_Li or Na excretion, although the magnitude of the circadian variation in CL_Li and other renal parameters was increased. Both CL_Li and CL_Na were increased in the first 3 h following furosemide administration and thereafter they fell below the control level in the remaining hours of the experiment.From the absolute and fractional changes in CL_Li it is suggested that compensatory Na conservation in response to chronic furosemide treatment occurs between doses, and that it involves decreased output from the proximal tubules combined with increased fractional Na reabsorption in the distal nephron.Abbreviations CL renal clearance - C_u urinary concentration - C_p plasma concentration - V_ur urinary volume - f_e fractional excretion - GFR glomerular filtration rate - CR creatinine     

Differential disposition of intra-renal generated and preformed glucuronides: studies with 4-methylumbelliferone and 4-methylumbelliferyl glucuronide in the filtering and nonfiltering isolated perfused rat kidney

Objectives This study was designed to investigate the renal disposition of 4‐methylumbelliferone (4MU) and 4‐methylumbelliferyl glucuronide (4MUG) to characterise the contribution of excretion and metabolic clearance to total clearance in the kidney. Methods The isolated perfused kidney (IPK) from the male Sprague–Dawley rat was used in filtering and non‐filtering mode to study the renal disposition of 4MU, renally generated 4MUG and preformed 4MUG. Perfusate and urine (filtering IPK only) was collected for up to 120 min and 4MU and 4MUG in perfusate and urine were determined by HPLC. Analytes were also measured in kidney tissue collected at 120 min. Non‐compartmental analysis was used to derive pharmacokinetic parameters. Key findings The concentration of 4MU in perfusate declined with a terminal half‐life of approximately 120 min following administration to the filtering IPK and nonfiltering IPK. There was a corresponding increase in the concentration of 4MUG. Metabolic clearance of 4MU accounted for 92% of total renal clearance. After bolus dosing of preformed 4MUG in the perfusion reservoir of the filtering IPK, the perfusate concentration declined with the terminal half‐life of approximately 260 min. The renal excretory clearance of preformed 4MUG accounted for 96% of total renal clearance. 4MU was extensively metabolized by glucuronidation in the filtering and nonfiltering IPK, and the total renal clearance of 4MU was far greater than its renal excretory clearance. This indicated that glucuronidation was the major elimination pathway for 4MU in the kidney. Conclusions The data confirmed an important role for the kidney in the metabolic clearance of xenobiotics via glucuronidation and signalled the lack of impact of impaired glomerular filtration on renal drug metabolism.