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K S Pang

Publications and source records attributed to K S Pang.

At least 73 records · Page 4Linked to original sources
PubMed1993

Kinetics of sequential metabolism. II. Formation and metabolism of nordiazepam and oxazepam from diazepam in the perfused murine liver.

Pharmacokinetic theory dictates that the extent of ensuing metabolism of a formed metabolite during drug transit through the liver is influenced by the number of consecutive reactions required for its genesis and the total intrinsic clearances of the precursors. This hypothesis was tested in the perfused murine liver by examining the successive conversion of the precursor diazepam (DZ) to its primary metabolite nordiazepam (NZ), and then the secondary metabolite oxazepam (OZ) and, finally, the tertiary metabolite, the oxazepam glucuronides. The concomitant C3-hydroxylation of DZ to temazepam, which can also be N-demethylated to form OZ, was minimal. The hepatic extraction ratios of NZ (E[NZ,DZ]) and OZ (E[OX,DZ]) after administration of [14C]DZ were compared to those obtained previously from [14C]NZ (E[NZ] and E[OZ,NZ]) and [3H]OZ (E[OZ]). The ability of three hepatic clearance models, the well-stirred, parallel-tube and dispersion models, to predict the experimental E[NZ,DZ] and E[OZ,DZ] was evaluated. DZ was highly extracted by the murine liver (E[DZ] = 0.95). The metabolism of NZ, generated in situ from DZ, was greater than that of preformed NZ (E[NZ,DZ] = 0.51; E[NZ] = 0.4), whereas E[OZ,DZ] (0.066) was similar to E[OZ,DZ] (0.056) and less than E[OZ] (0.0125). The unexpected observation of E[NZ,DZ] > E[NZ] may be explained by the coupling of N-demethylation and C3-hydroxylation/glucuronidation reactions or by a sequestration of hydrophobic substrates within the enzymic space, favoring sequential metabolism of products formed in situ. The atypical kinetic behavior of generated NZ may have also influenced the ensuing metabolic fate of its product, OZ, such that E[OZ,NZ] approximately E[OZ,DZ].(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Uptake of a protein-bound polar compound, acetaminophen sulfate, by perfused rat liver.

The hepatocytic entry of acetaminophen sulfate conjugate was examined in the rat liver, perfused with red cells with and without albumin, by use of the multiple-indicator dilution technique. [3H]acetaminophen sulfate was injected into the portal vein in a bolus of blood containing 51Cr-labeled red blood cells (a vascular reference), sucrose (a low-molecular-weight interstitial reference) or 125I-labeled albumin (a high-molecular-weight interstitial reference, included when albumin was present), and the time courses of their outflow into the hepatic venous blood were observed. The [3H]acetaminophen sulfate, which binds partially to albumin, emerged between albumin and sucrose in the presence of albumin, processed the upslope of the sucrose curve and showed a late low-in-magnitude tailing; the precession disappeared in the absence of albumin. Biliary excretion of [3H]acetaminophen sulfate was less than 1% of the dose. Quantitative evaluation with a barrier-limited, space-distributed variable transit time model (including rapidly equilibrating albumin binding) accounted for the albumin effect on [3H]acetaminophen sulfate behavior and demonstrated a low liver cell permeability for the acetaminophen sulfate and a small interstitial binding space for its nonalbumin-bound fraction in excess of that for sucrose, which in the absence of albumin was of similar dimensions.

Acetaminophen↗
PubMed1992

Dynamics of drug distribution. I. Role of the second and third curve moments.

Conventionally, the dynamics of distribution in the body is evaluated by the so-called distribution half-life (e.g., t1/2, alpha); but then the mean time of the distribution process is underestimated due to the influence of elimination. By contrast, information about the dynamics of distribution contained in drug disposition curves can be extracted by the second and third curve moments, parameters that are related to the variance (VDRT) and skewness (SDRT) of residence time distributions; whereas the equilibrium state characterized by the volume of distribution (Vss), is determined by the mean residence time (MDRT) or the first curve moment. The approach represents a general noncompartmental analysis that is independent of a detailed structural model or a particular disposition function. Two parameters are introduced to characterize the dynamics of drug distribution: (i) the degree of departure of the system from "well-mixed" behavior of instantaneous distribution equilibrium (related to VDRT) and (ii) the mean time until equilibration is achieved (mean equilibration time, MEQT), which additionally depends on SDRT. Both parameters are quantitative measures of the dynamics of distribution and display explicit physical significance in terms of distribution within the corresponding noneliminating system. It is further shown that the so-called "distribution phase" in biexponential disposition curves is related to a monoexponential mixing curve of its corresponding noneliminating system with an equilibration or mixing half-time, t1/2,M = t1/2,alpha (V beta/Vss*), where Vss* denotes the distribution volume of the noneliminating system. The results are applied to mixing and disposition curves measured for acetaminophen in liver-ligated and intact rats, respectively.

Acetaminophen↗
PubMed1992

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] less than E[M1], and E[M2, P] less than E[M2, M1] less than E[M2], 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.

Liver↗
PubMed1992

Enalaprilat handling by the kidney: barrier-limited cell entry.

The angiotensin-converting enzyme inhibitor enalaprilat is formed in vivo in liver and kidney by esterolysis of the antihypertensive drug enalapril. To gain insight into the renal elimination of enalaprilat, we carried out multiple-indicator dilution experiments in the isolated perfused rat kidney. Kidneys were perfused single pass with an amino acid-supplemented Krebs-Henseleit buffer containing 20% bovine red blood cells and 4% bovine serum albumin, at a flow rate of 0.11 +/- 0.02 (SD) ml.s-1 x g-1. A bolus of 51Cr-labeled red blood cells (vascular red blood cell indicator), 125I-labeled albumin (vascular plasma indicator), L-[14C]glucose (interstitial space indicator), and [3H]-enalaprilat was injected into the renal artery, and timed samples of venous blood (up to 1 min) and urine (up to 10 min) were collected. The data were analyzed using a variable-transit-time, space-distributed model with modifications accounting for glomerular filtration and the observed 14% protein binding of enalaprilat; the glomerular filtration rate (GFR) estimated from L-glucose clearance was 9.0 +/- 2.9% of total plasma flow. The ratio of renal clearance of unbound enalaprilat to GFR was 1.56 +/- 0.29, indicating both glomerular filtration and net tubular secretion of enalaprilat. Unidirectional influx from plasma to tubular cells exceeded tubular secretion by a factor of 2.2 +/- 0.5. Thus only about one-half of the enalaprilat taken up by the tubular cells was excreted into urine, with the remainder refluxing into the capillary blood stream, indicating bidirectional permeation of enalaprilat across the basolateral tubular membrane.

Algorithms↗
PubMed1991

High-performance liquid chromatographic method for the direct determination of 4-methylumbelliferone and its glucuronide and sulfate conjugates. Application to studies in the single-pass in situ perfused rat intestine-liver preparation.

A direct high-performance liquid chromatographic (HPLC) assay was developed for the separation and determination of 4-methylumbelliferone (4MU) and its glucuronide (MUG) and sulfate (MUS) conjugates in the cell-free perfusate ("plasma") from in situ perfused rat intestine-liver preparation. In addition, a procedure was developed to extract and determine 4MU in the whole blood perfusate. Perfusate plasma containing an internal standard (umbelliferone) was precipitated with methanol (1:4, v/v), and injected into a reversed-phase HPLC system with gradient elution. 4MU and the same internal standard were also extracted directly from the whole blood perfusate with ethyl acetate and injected into a reversed-phase HPLC system with isocratic elution. Inter- and intra-day precision studies (n = 5 for each) for both the plasma and whole blood procedures demonstrated relative standard deviation of less than 10% at all concentrations studied. The compounds were stable in either the plasma or blood extracts at room temperature for up to 72 h. The procedures were successfully used to analyze perfusate samples obtained from the single-pass in situ perfusion of rat intestine-liver system with either trace (0.95 nM) or 32.3 microM concentrations of 4MU. The intestine was responsible for the formation of most of the MUG formed by the intestine-liver preparation during steady-state perfusion with either input concentration of 4MU.

Animals↗
PubMed1991

D2O as a substitute for 3H2O, as a reference indicator in liver multiple-indicator dilution studies.

The hepatic space of distribution and the processes underlying uptake of tracer substrate may be appraised by the multiple-indicator dilution technique after simultaneous injection of noneliminated vascular (51Cr-labeled red blood cells), extracellular 125I-labeled albumin and [14C]sucrose (or [58Co]EDTA) as high- and low-molecular-weight interstitial references, respectively], and cellular (3H2O or [14C]urea) indicators, together with the tracer-labeled substrate. The use of 3H2O or [14C]urea, with [14C]- or [3H]sucrose, however, precludes the simultaneous introduction and analysis of the behavior of 3H- and 14C-labeled substrate and metabolite. An assay for the quantitation of D2O in plasma by Fourier transform infrared spectrometry was therefore developed such that D2O could be used in lieu of 3H2O in multiple-indicator dilution studies in the blood-perfused rat liver. In experiments performed with an injection dose containing 51Cr-labeled red blood cells, 125I-labeled albumin, [14C]sucrose, 3H2O, and D2O, D2O was found to behave virtually identical to 3H2O in blood and liver; the accessible cellular water spaces were 0.625 and 0.621 ml/g liver for 3H2O and D2O, respectively, and the corresponding ratios of the sum of the cellular water plus the interstitial water space to the sinusoidal water space were 3.87 and 3.89. D2O was found to be an ideal substitute and is much superior to [14C]urea, which exhibits a small red blood cell carriage effect and which is slightly less dispersed than 3H2O.

Animals↗
PubMed1991

Esterases for enalapril hydrolysis are concentrated in the perihepatic venous region of the rat liver.

Perfusion of substrate via only the hepatic artery with simultaneous substrate-free perfusion of portal vein or hepatic vein [hepatic artery portal vein-hepatic artery hepatic vein] was used to examine the enzymic distribution of carboxylesterases towards the hydrolysis of enalapril to enalaprilat in the perfused rat liver preparation. In this single-pass method, [14C]enalapril was delivered by the hepatic artery at 2 ml/min into the liver, whereas drug-free perfusate entered the portal vein or hepatic vein at 10 ml/min for HAPV and HAHV perfusions, respectively. During steady state, a multiple indicator dose of 51Cr-labeled red blood cells (vascular marker), [58Co]EDTA (interstitial space marker, which behaves similarly to labeled tracer sucrose), and 3H2O was given into the hepatic artery. Labeled enalapril and the reference indicators entering via the hepatic artery will reach virtually all sinusoidal spaces during HAPV, and will be confined to the peripheral region during HAHV. By defining the steady-state extraction ratios of enalapril (Etot) and segregating the components of biliary excretion and metabolism, and by assessing the intracellular water spaces and the corresponding transit times during HPAV and HAHV, the metabolic sequestration rate constant (metabolic intrinsic clearance per unit accessible cell water space) for the periportal region/whole liver (HAHV/HAPV) was 0.344. The data suggest that the carboxylesterase activity for enalapril conversion to enalaprilat is primarily localized in the perihepatic venous region of the rat liver.

Animals↗
PubMed1990

Physiological modeling of drug and metabolite: disposition of oxazepam and oxazepam glucuronides in the recirculating perfused mouse liver preparation.

The disposition of tracer doses of 3H-oxazepam was studied in the recirculating perfused mouse liver preparation. 3H-Oxazepam was biotransformed primarily to the diastereomeric 3H-oxazepam glucuronides, which either effluxed into the circulation or underwent biliary excretion. Three additional, unknown metabolites constituted a small fraction (5-10%) of the total radioactivity recovered in bile (7% of dose); no other metabolite was detected in perfusate. A physiologically based model, comprising the reservoir, liver blood and tissue, and bile, was fitted to reservoir concentrations of 3H-oxazepam and 3H-oxazepam glucuronides, and the cumulative amount excreted into bile. The model allowed for consideration of elimination pathways other than glucuronidation and the presence of a transport barrier for the oxazepam glucuronides across the hepatocyte membrane. The fitted results suggest a slight barrier existing for the transport of metabolites across the sinusoidal membrane, inasmuch as the transmembrane clearance was comparable to liver blood flow rate. Upon further comparison of estimates of formation, biliary, and transmembrane clearances for the oxazepam glucuronides, the rate-limiting step in the overall (biliary) clearance appears to be a poor capacity for biliary excretion. The influence of the cumulative volume loss that a recirculating perfused organ system incurs upon repeated sampling was discussed, and a compartmental method of correcting the observed concentrations of drug and generated metabolite was presented.

Animals↗
PubMed1990

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.

Animals↗
PubMed1990

Transfer of enalaprilat across rat liver cell membranes is barrier limited.

The liver cell entry of enalaprilat, the polar, pharmacologically active dicarboxylic acid metabolite arising from esterolysis of enalapril, a new angiotensin-converting enzyme inhibitor, was examined in the perfused rat liver by use of the multiple-indicator dilution technique. [Phenylpropyl-2,3-3H]enalaprilat was injected into the portal vein in a bolus of blood containing 51Cr-labeled red blood cells (a vascular reference) and 125I-labeled albumin and [14C]sucrose (interstitial references that do not enter cells), with observation of the time courses of their outflow into the hepatic venous blood. A quantitative evaluation of the data was carried out by use of the barrier-limited, space-distributed variable transit time model (C.A. Goresky, G.G. Bach, and B. E. Nadeau, J. Clin. Invest. 52:991-1009, 1973). For data up to 60 s after injection, the transfer coefficients for influx, efflux, and sequestration were 0.018 +/- 0.004 (means +/- SD), 0.044 +/- 0.017, and 0.033 +/- 0.007 s-1, respectively. The influx permeability surface area product (influx clearance) per gram was 0.0057 +/- 0.0013 ml.min-1.g-1, and the rapidly accessible cellular equilibrium distribution space for enalaprilat was 0.137 +/- 0.022 ml water/g wet wt. At times beyond 60 s, the fitted data deviated systematically from the experimental data, suggesting the presence of an additional intracellular pool. With this addition, the coefficients for transfer between the intracellular pools were 0.0150 +/- 0.0045 s-1 for the direction cytoplasmic pool (pool 1)----additional pool (pool 2) and 0.0234 +/- 0.0069 s-1 for the opposite direction, and the fitted volumes of pools 1 and 2 became 0.126 +/- 0.021 and 0.082 +/- 0.018 ml/g, the total accessible cellular distribution space became 0.208 +/- 0.036 ml/g wet wt, and the sequestration transfer coefficient became 0.027 +/- 0.007 s-1. The data indicate that, as previously postulated (I. A. M. de Lannoy and K. S. Pang, Drug Metab. Dispos. 14: 513-520, 1986), the enalaprilat flux across liver cell membranes is retarded by a diffusional barrier. The results also indicate that enalaprilat is excluded from part of the volume of hepatocytes, as expected for an anionic compound crossing the membrane in charged rather than protonated form, given the negative electrical potential of the cytosol vs. the extracellular space.

Animals↗
PubMed1990

[14C]urea and 58Co-EDTA as reference indicators in hepatic multiple indicator dilution studies.

The space distribution and the processes underlying uptake of tracer substrate may be appraised by the multiple indicator dilution technique, after the simultaneous injection of noneliminated vascular (51Cr-labeled red blood cells), extracellular (125I-labeled albumin and [14C]sucrose for high and low molecular weight interstitial space, respectively), and cellular (3H2O) indicators and tracer substrate. When tracer substrates and/or their metabolites containing 14C or 3H labels are being studied, it becomes necessary to find substitutions for the similarly labeled noneliminated indicators. In red blood cell-perfused rat livers, 58Co-EDTA is found to be a good replacement for the low molecular weight interstitial reference; it has a space distribution indistinguishable from that for [14C]sucrose. [14C]urea and 3H2O, which distribute similarly in red blood cells, have similar outflow dilution profiles. With corrections for minor deviations in recovery [a ratio of 0.936 +/- 0.024 (SD)] and transit time [a ratio of 0.962 +/- 0.008 (SD)], the cellular water space can be closely approximated from the [14C]-urea curve. 58Co-EDTA and [14C]urea are reasonable substitutes for [14C]sucrose and 3H2O, respectively, in multiple indicator dilution liver perfusion studies for investigating transfer and removal characteristics of tracer substrates.

Animals↗
PubMed1990

Sequential metabolism of salicylamide exclusively to gentisamide 5-glucuronide and not gentisamide sulfate conjugates in single-pass in situ perfused rat liver.

Gentisamide (GAM), the hydroxylated metabolite of salicylamide (SAM), underwent sequential metabolism, and GAM-5-glucuronide was the only metabolite detected in the single-pass in situ rat liver preparation (10 ml/min/liver) perfused at varying SAM steady-state input concentrations (CIn). The exclusive formation of GAM-5-glucuronide was unexpected, because GAM, when administered to the rat liver, formed predominantly monosulfate conjugates at the 5- and 2-positions (Morris et al., J. Pharmacol. Exp. Ther. 245: 614-624, 1988). Kinetic parameters obtained from single-pass studies for SAM sulfation (CIn increasing from 31 to 347 microM) and glucuronidation and hydroxylation (CIn decreasing from 1383 to 130 microM, with 0.85 mM SO4(2-), and CIn increasing from 32 to 800 microM in an absence of SO4(2-), with 2,6-dichloro-4-nitrophenol, a sulfation inhibitor) revealed sulfation as a high-affinity, high-capacity pathway, glucuronidation as a lower-affinity, high-capacity pathway and hydroxylation as a low-affinity, low-capacity pathway. However, these parameters would not explain the exclusive glucuronidation of GAM when generated from SAM. Rather the zonal localization of metabolizing activities [a periportal sulfation, evenly distributed glucuronidation, and perivenous hydroxylation system (Xu and Pang, J. Pharmacokinet. Biopharm. 17: 645-671, 1989; Morris et al., J. Pharmacokinet. Biopharm. 16: 633-656, 1988)] could explain the complete lack of sulfation in SAM sequential metabolism. The different metabolite patterns arising from the administrations of a metabolite precursor (SAM) versus a preformed metabolite (GAM), due to the proximity of enzymes for formation and sequential metabolism, may serve to explain the differential toxic or pharmacologic effects observed in other precursor-metabolite pairs.

Animals↗
PubMed1990

Effects of retrograde flow on measured blood volume, Disse space, intracellular water space and drug extraction in the perfused rat liver: characterization by the multiple indicator dilution technique.

Previous rat liver perfusion studies had shown a preferential derecruitment of downstream (phenacetin O-de-ethylation) in relation to upstream (acetaminophen sulfation) hepatocyte activities with reductions in prograde (normal) flow, with substrates entering from the portal vein and exiting the hepatic vein. The reduction in metabolic activities was found associated with a reduction in accessible intracellular water space (Pang et al., J. Pharmacokinet. Biopharm. 16: 595, 1988). In the present studies, we investigated the changes in metabolic activities of the liver and its accessible spaces during retrograde flow (substrates entering from the hepatic vein and exiting the portal vein), an approach used to probe zonal heterogeneity in the liver. The multiple indicator dilution technique was used to gain information concerning the accessible sinusoidal blood volume, Disse space, and the intracellular water space; once-through perfusion with tracer [3H]acetaminophen and [14C]phenacetin was performed simultaneously within the same liver, to estimate metabolic activities at the periportal and perihepatic venous regions of the liver, respectively, at the designated flows. The estimated sinusoidal blood volume, total albumin and sucrose distribution spaces, the Disse space and the accessible albumin and sucrose Disse spaces showed weak but positive correlations with retrograde perfusate flow rate. The values were substantially greater than those for rat livers perfused with normal (prograde) flows, suggesting an increase in the vascular space and accessible Disse space of the liver during retrograde flow. The total water and accessible intracellular water spaces also showed weak but positive correlations with retrograde perfusate flow, and the average intracellular water space (which is also accessed by the substrates) at both retrograde flows of 8 and 12 ml/min was 0.52 +/- 0.08 and 0.55 +/- 0.08 ml/g liver, respectively, values that were not statistically different. When the intracellular water space was plotted vs. water flow rate (ml water/g liver), a weakly positive correlation was again found, suggesting that over the flow range, the accessible intracellular water space decreased in a very gradual and linear fashion with a reduction in water flow rate. When the estimated intracellular water space for retrograde flow was compared with values obtained for the same flow rate in a prograde fashion, slightly lower values were obtained. After consideration of the effect of the distended vascular and interstitial spaces on the weights of the liver, the values obtained for retrograde and prograde flow rates were closely similar. In both cases, it appeared that with reduction in flow, the accessible intracellular water spaces were reduced, regardless of the flow direction.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetaminophen↗
PubMed1989

The multiple-indicator dilution technique for characterization of normal and retrograde flow in once-through rat liver perfusions.

The technique of normal and retrograde rat liver perfusion has been widely used to probe zonal differences in drug-metabolizing activities. The validity of this approach mandates the same tissue spaces being accessed by substrates during both normal and retrograde perfusions. Using the multiple-indicator dilution technique, we presently examine the extent to which retrograde perfusion alters the spaces accessible to noneliminated references. A bolus dose of 51Cr-labeled red blood cells, 125I-albumin, 14C-sucrose and 3H2O was injected into the portal (normal) or hepatic (retrograde) vein of rat livers perfused at 10 ml per min per liver. The outflow perfusate was serially collected over 220 sec to characterize the transit times and the distribution spaces of the labels. During retrograde perfusion, red blood cells, albumin and sucrose profiles peaked later and lower than during normal perfusion, whereas the water curves were similar. The transit times of red blood cells, albumin and sucrose were longer (p less than 0.005), whereas those for water did not change. Consequently, retrograde flow resulted in significantly larger sinusoidal blood volumes (45%), albumin Disse space (42%) and sucrose Disse space (25%) than during normal flow, whereas the distribution spaces for total and intracellular water remained unaltered. The distension of the vascular tree was confirmed by electron microscopy, by which occasional isolated foci of widened intercellular recesses and spaces of Disse were observed. Cellular ultrastructure was otherwise unchanged, and there was no difference found between normal and retrograde perfusion for bile flow rates, AST release, perfusion pressure, oxygen consumption and metabolic removal of ethanol, a substrate with flow-limited distribution, which equilibrates rapidly with cell water (hepatic extraction ratios were virtually identical: normal vs. retrograde, 0.50 vs. 0.48 at 6 to 7.4 mM input concentration). These findings suggest that the functional and metabolic capacities of the liver remain unperturbed during retrograde perfusion, rendering the technique suitable for the investigation of zonal differences in drug-metabolizing enzymes.

Albumins↗