Effect Of Plasma Protein Binding Research Articles

Removal of Endotoxin and Cytokines by Adsorbents and the Effect of Plasma Protein Binding

High serum levels of endotoxin and cytokines, through which its activity is mediated, have been shown to be associated with disease severity in septic shock and in fulminant hepatic failure. In the present study, we have investigated the ability of activated charcoals (DHP-1 and Adsorba 150C) and uncharged resin (Amberlite XAD-7) to adsorb lipopolysaccharide (LPS) and various cytokines, namely tumour necrosis factor (TNF), interleukin-1 (IL-1), interleukin-6 (IL-6), interferon-alpha (IFN-alpha) and interferon-gamma (IFN-gamma). The capacities of the adsorbents were assessed by measurement of their equilibrium adsorption isotherms for these substances labelled with 125I. There was no single adsorbent that uniformly adsorbed LPS and the cytokines from phosphate buffered saline or human plasma. DHP-1 charcoal was superior to Adsorba 150C for all substances and was the most effective adsorbent for binding LPS, IL-1 alpha and IFN-gamma. Amberlite XAD-7 resin was most effective for TNF, IL-6 and IFN-alpha, but bound little LPS, particularly from human plasma. Ultrafiltration through a membrane which retains substances of molecular weight greater than 50 kD did not filter the cytokines from human plasma, although the molecular weight of the cytokines range from 17 to 22 kD. This demonstrated that, TNF, IL-1, IL-6, IFN-alpha and IFN-gamma readily bind to proteins and/or other large molecules in plasma.

Disposition of phenytoin in analbuminemic rats

The disposition characteristics of phenytoin were investigated in analbuminemic rats to study the effect of plasma protein binding on phenytoin disposition. Blood and plasma phenytoin concentration determinations, measurement of plasma protein phenytoin binding, and whole-body autoradiography were performed after intravenous bolus injection of 10 mg kg-1 of phenytoin. When plasma phenytoin was assayed, total body phenytoin clearance (CL) was faster and its apparent volume of distribution (Vd) greater in the analbuminemic rats in comparison to the controls. The plasma protein binding of phenytoin was significantly lower in the analbuminemic rats, suggesting the disposition characteristics of phenytoin were altered in the presence of low plasma albumin concentrations due to reduced plasma phenytoin protein binding. On the other hand, when blood phenytoin levels were analyzed, no difference in CL and a less pronounced difference in Vd were noted between the two groups of rats. The red blood cell-to-plasma phenytoin concentration ratios were greater in analbuminemic rats, suggesting that the distribution of phenytoin into red blood cells was greater in the mutant rats.

Disposition of warfarin in analbuminemic rats

The disposition characteristics of warfarin were studied in analbuminemic rats after intravenous bolus injection of 1 and 40 mg/kg of drug to investigate the effects of plasma protein binding on drug disposition. With both doses of warfarin, total body warfarin clearance (CL) was markedly faster and its apparent volume of distribution ( V d β ) significantly greater in the analbuminemic rats in comparison to controls. Further, the apparent elimination rate constant (β) was significantly greater and the corresponding elimination half-life (t 1 2β ) shorter in the rats with low plasma albumin. Whole body autoradiograms demonstrated that the distribution of warfarin to the liver, skeletal muscle and brain was greater in the analbuminemic rats which was in good agreement with the larger V d β observed in this group of rats. The results suggested that the disposition characteristics of warfarin were markedly altered in the rats with low plasma albumin concentrations due to reduced plasma warfarin protein binding.

The role of plasma protein binding on the metabolism and renal excretion of sulphadimethoxine and its metabolite N4‐acetylsulphadimethoxine in pigs

The effects of plasma protein binding on the elimination of sulphadimethoxine (SDM) were examined after intravenous administration of 6.25, 12.5, 25, 50, 100 and 150 mg/kg to pigs. At an early stage of the experiment, the animals were anaesthetised by inhalation of enflurane to obtain a more exact relationship between plasma concentration and the renal excretion. SDM and its acetylated conjugate, N4-acetylsulphadimethoxine (N4-SDM) were detected in plasma and urine of all animals, and the recovery of the doses was almost complete in two animals with negligible renal excretion of SDM. The percentages of plasma protein binding of SDM and N4-SDM were almost similar, and ranged from 30 to 95%, depending on the plasma concentration. The metabolic clearance of SDM by acetylation increased when the plasma protein binding decreased. These results suggested that the main elimination route of SDM in pigs is acetylation, and that the plasma protein binding can have a large effect on the elimination of SDM in pigs. The effect of plasma protein binding on the renal clearance of SDM was not so evident, because urine pH had a much greater effect on it. The deacetylation of N4-SDM was detected after 25 mg/kg intravenous administration of N4-SDM, which suggests that the metabolic clearance of SDM is part of an acetylation-deacetylation equilibrium. Saturation of the active tubular reabsorption of SDM and of the active tubular secretion of N4-SDM was also suggested after higher doses of SDM.

How chronically administered valproate increases chlordiazepoxide transfer through the blood‐brain barrier

Sodium valproate (VPA) is a drug widely used in the treatment of epileptics often in association with benzodiazepines. Recent animal studies have shown that the addition of valproate increases diazepam levels in the cortex and the cerebellum (Hariton et al, 1985). The aim of our study was to determine the effect of VPA on the transfer of benzodiazepines through the blood-brain barrier. They were investigated using the intracarotid injection technique in rats as described by Oldendorf (1971). Our results show that the 14C-chlordiazepoxide brain extraction is significantly higher in rats on prolonged valproate treatment than in controls. With regard to plasma protein binding effects on chlordiazepoxide transport, our data indicate that a fraction of the protein-bound chlordiazepoxide could transfer from the intracapillary space to the brain tissue space because of enhanced drug dissociation from albumin in the brain microcirculation (Kd in vitro = 74.1 microM; Kd in vivo = 793.7 microM). Two distinct mechanisms can be deduced from this study: 1) chlordiazepoxide is displaced from HSA by valproate, 2) in addition, this fatty acid could increase drug permeation through the blood brain barrier (PS/F (chlordiazepoxide) = 0.60 in controls, PS/F (chlordiazepoxide) = 0.97 in treated rats). On the contrary, the washout of the benzodiazepine from the rat brain does not seem to be modified by the addition of valproate.

Comparison of in vivo bioavailability of progestational agents into the rat uterus and liver and the effect of plasma protein binding

The in vivo bioavailabilities of three clinically available progestational agents were evaluated with a previously described rat model. With this model a single-injection, double-isotope technique was performed to calculate and compare the unidirectional extractions of progesterone, 19-nortestosterone, and medroxyprogesterone acetate into the rat uterus and liver. To assess the role of plasma protein binding in the extraction of these agents, steroids were presented in four different vehicles: (1) Ringer's solution, 0.1% bovine serum albumin; (2) Ringer's solution, 4% bovine serum albumin; (3) Ringer's solution, 0.8 mg/ml, alpha 1-acid glycoprotein; (4) human pregnancy serum. The results revealed that liver extraction always exceeded uterine extraction for each progestin, regardless of the vehicle, and that liver extraction was approximately 100% for all steroids. With regard to the uterus, when binding proteins were not present in the injectate, the extraction of the three steroids was similar; the uterine vasculature was relatively permeable to these progestins, and bovine serum albumin and alpha 1-acid glycoprotein had no major effect on the uptake into the uterus. However, when human pregnancy serum was in the injectate, the extractions of progesterone and 19-nortestosterone into the rat uterus were significantly diminished. In contrast, pregnancy serum had no effect on the uptake of medroxyprogesterone acetate into the uterus, with the extraction equal to that of Ringer's solution alone. This suggests that, regardless of potential binding proteins, medroxyprogesterone acetate displays greater bioavailability than that of the other progestogens studied.

Effect of Plasma Protein Binding on Kinetics of PN 200‐110 in the Isolated Perfused Rat Heart

The myocardial accumulation and elimination pharmacokinetics of PN 200-110 (PN) were investigated in the single pass isolated perfused rat heart by two methods. A direct method, radioactivity measurement in myocardial tissue after various perfusion times, and an indirect method, concentration determination in coronary effluent, by fractionary collection of samples, during infusion and elimination periods. Both methods showed that the myocardium could be considered as a one-compartment model with regard to PN pharmacokinetics. The perfusion with a modified Krebs-Ringer (MKR) solution containing 1 nM of (+/-)PN 200-110 and [3H]-(+)PN 200-110 as radioactive tracer, led to an accumulation of about 61.4 fmol.mg-1 myocardial tissue at steady-state. The effect of protein binding on the uptake and pharmacokinetic parameters of PN has been investigated in this isolated perfused heart (IPH) model. binding of PN decreased as a function of increasing bovine serum albumin (BSA) levels in the perfusion solution. As a matter of fact, the mean steady state myocardial concentration of PN was decreased by 42.9, 56.2, 76.5, 83.9 and 95.5% for respectively, 1, 2.5, 6, 10 and 40 g.l-1 of BSA. In the same way, the free fraction, the apparent volume of distribution (Vd) and the distribution and elimination half-lives were decreased. On the contrary, the elimination rate constant was increased.

Further observations on the disposition characteristics of salicylic acid in analbuminemic rats.

The disposition characteristics of salicylic acid (SA) were investigated in analbuminemic rats after intravenous bolus injection of 10 and 173 mg kg-1 of SA to study the effects of plasma protein binding on drug disposition. Following the administration of 10 mg kg-1 of SA, total body SA clearance (CL) was markedly faster and its apparent volume of distribution (Vd) significantly greater in the analbuminemic rats in comparison to the controls. Further, the apparent elimination rate constant (kj) was two-fold greater and the corresponding elimination half-life (t 1/2) shorter in the rats with low plasma albumin. Whole body autoradiograms obtained following the administration of 14C-salicylic acid demonstrated that the tissue distribution of SA was greater in the analbuminemic rats which was in agreement with the larger Vd observed in this group of rats. After the administration of 173 mg kg-1 of SA, no differences in CL, Vd, kk or t 1/2 were noted between the analbuminemic and control rats. Dose-dependent SA disposition was observed in both the analbuminemic and control rats with the effects being more pronounced in the rats with low plasma albumin. The results suggested that the disposition characteristics of SA were markedly altered in the presence of low plasma albumin concentrations due to reduced plasma SA protein binding.

Effect of plasma protein binding on elimination of taurocholate by isolated perfused rat liver: comparison of venous equilibrium, undistributed and distributed sinusoidal, and dispersion models.

In the past, various models have been developed to allow better characterization of the hepatic elimination of substrates from plasma. In this study we investigated the applicability of the venous equilibrium, undistributed sinusoidal, several distributed sinusoidal, and dispersion models to the steady state elimination of sodium taurocholate by the isolated perfused rat liver. Rat livers were perfused with 24-14C-taurocholate (sodium salt) at a concentration of 25 microM (specific activity 500 microCi/mmole) in a single-pass design (n = 7) or at a rate of 0.5 mumol/min (specific activity 40 microCi/mmole) into the portal vein in a recirculating design (n = 5). In single-pass experiments, the changes in hepatic venous outflow concentration (C0) with changes in unbound fraction of taurocholate (fu) from 0.09 to 1.0 were fitted better by the venous equilibrium model, by the dispersion model, and by a distributed model in which heterogeneity in both hepatic blood flow (Q) and intrinsic clearance (CLint) was defined by separate density functions. The very large value of dispersion number (DN greater than 10(7] yielded by the dispersion model is consistent with a high degree of axial mixing of blood within sinusoids. The large coefficients of variation (0.7-232) for the density functions describing the transverse heterogeneity of Q and CLint obtained with the Q/CLint-distributed model were consistent with a large degree of heterogeneity in Q and CLint within the liver. In recirculation experiments, the steady state unbound concentration of taurocholate in the reservoir (Cuss) was independent of fu (range 0.05-0.9). This finding was not predicted by the undistributed sinusoidal model, but was in keeping with the venous equilibrium model, with the dispersion model, and with the Q/CLint-distributed model. Therefore, there is no need to invoke cell surface-mediated dissociation of albumin-ligand complexes in hepatic taurocholate uptake. As the dispersion and Q/CLint-distributed models are conceptually plausible and operationally accurate, it may be time to relinquish the venous equilibrium model, which, though operationally accurate, is conceptually flawed.

Effect of plasma protein binding on drug disposition in muscle tissue: application of statistical moment analysis and network theory to in situ local single-pass perfusion system.

The local disposition characteristics of mitomycin C (MMC) and five lipophilic prodrugs in rabbit hind leg muscle were examined using an in situ single-pass perfusion experiment. Test compounds inputted into a perfusion line as a rectangular function (unit pulse) were perfused with or without albumin and their outflow patterns were analyzed by statistical moment analysis. In interpretation of statistical moment parameters, the well-stirred model was applied to the local perfusion system based on the plate theory of a chromatographic system and some general pharmacokinetic parameters (the disposition parameters) were derived from the moments. A new theory which elucidates the relationships among the moments for plasma protein binding, unbound (free), and total drug fraction was established based on network theory. Using this system, the following conclusions were made for mitomycin C and its five lipophilic derivatives: (i) In the absence of albumin, an increase in lipophilicity led to an increase in organ clearance and distribution volume; (ii) drug bound to albumin did not transfer to the extravascular space; (iii) in the presence of albumin, an increase in lipophilicity results in a decrease in clearance.

Kinetic analysis of cerebrovascular transport based on the multiple indicator dilution method.

The multiple indicator dilution method (MID) is one experimental technique for evaluating capillary transport parameters of highly permeable neuroreceptor ligands. This technique has been widely used for the determination of capillary permeability in various organs and was adapted for studies of the blood brain barrier in rat, dog and human. Recently, Hertz and Paulson found that the apparent extraction of various substances increases during the initial part of the dilution curves in human brain. This increase in the apparent extraction can be explained by heterogeneity of transit times through the cerebral microcirculation. It is also important to consider intravascular separation of the test and reference substances (the effect of plasma protein binding and red cell carriage) and the effect of recirculation. Due to the highly tortuous character of the brain microcirculation, a distributed model of vascular transit that assumes a well-mixed tissue compartment to analyze indicator dilution curves from brain was developed. MID experiments based on the heterogeneity of capillary transit time can be used to estimate the transcapillary fluxes (both influx and efflux processes) of a highly diffusable test compound in the brain of a single animal. This appoach is potentially usefull in the study of the transport processes of lipophilic drugs and receptor-ligands that rapidly permeate brain capillaries.

Blood-brain barrier transport of valproic acid.

Valproic acid distribution in brain is less than that of other anticonvulsants such as phenytoin or phenobarbital. Possible mechanisms for this decreased distribution space in brain include (a) increased plasma protein binding of valproate relative to the other anticonvulsants and (b) asymmetric blood-brain barrier (BBB) transport of valproate such that the brain-to-blood flux exceeds the blood-to-brain flux. These mechanisms are investigated in the present studies using the intracarotid injection technique in rats and rabbits. In the rat, the brain uptake index (BUI) of [14C]valproate relative to [3H]water is 51 +/- 6%, indicating the blood-to-brain transport of water is twofold greater than that of valproate. However, the BUI of [14C]valproate relative to [3H]water decreased with time after carotid injection during a 4-min washout period, which indicates that brain-to-blood transport of valproate is greater than that of water. This suggests that the permeability of the BBB to valproate is polarized, with antiluminal permeability being much greater than luminal permeability. In rabbits, the BUI of [14C]valproate is 47 +/- 7% in newborns and 17 +/- 6% in adult animals. However, the high drug extraction in newborns may be attributed to decreased cerebral blood flow in the neonate as the BBB permeability-surface area (PS) products are unchanged (e.g., PS = 0.13 and 0.11 ml min-1 X g-1 in the newborn and adult rabbit, respectively). With regard to plasma protein binding effects on valproate transport, brain valproate uptake was also measured in the presence of human, lamb, pig, rat, horse, goat, hamster, dog, and mouse sera. Higher brain uptakes were observed when the unbound fraction of drug increased. However, our data indicate that a fraction of the valproic acid entering the capillaries bound to plasma proteins had the capacity to equilibrate with brain because of enhanced drug dissociation from albumin in the brain microcirculation. Since plasma protein-bound valproate is available for uptake by brain, the major factor underlying the diminished distribution of the drug in brain appears to be the asymmetric transport properties of the BBB to valproic acid.

Differential permeability of uterine and liver vascular beds to estrogens and estrogen conjugates.

The role of capillary membrane permeability and the effect of plasma protein binding on the influx of unconjugated and conjugated estrogens into a target organ, the uterus, and a metabolic organ, the liver, were studied in anesthetized rats. In the absence of plasma proteins, estrone (E1) and estradiol (E2) were freely diffusible through the uterine capillaries, but influx was significantly reduced for estriol (E3) and estetrol. In the uterus, the influx of the conjugated estrogens was markedly restricted and approximated the influx of dextran, a vascular space marker. The polarity of the compound (based on the number of hydrogen bond-forming functional groups and the presence of charged groups) appeared to predict uterine endothelial membrane permeability better than the octanol/Ringer's partition coefficient. In contrast to the selective permeability properties of the uterine endothelial barrier, the limiting membrane lining the hepatic microcirculation, the hepatocyte cell membrane, was highly permeable to all unconjugated and conjugated estrogens. The addition of 4% albumin to the injection solution led to a significant inhibition of uterine influx of E2, but not E1 or E3. In the liver, only the influx of E1 sulfate was slightly diminished by 4% albumin. In all cases, the influx of estrogens greatly exceeded the rate that would be expected if only the fraction that was free (dialyzable) in vitro was diffusible in vivo. Human sera containing sex hormone-binding globulin and albumin caused inhibition of influx of E1 and E2 through the uterine capillary barriers, whereas in the liver, the influx of E2 sulfate, and E3 glucuronide were diminished. The results are compatible with a difference in permeability of the microvasculature of the two organs and a differential availability of protein-bound estrogen for influx into liver and uterus. With the exception of E1, which is nearly completely diffusible into both organs, the influx of estrogens and estrogen conjugates into liver is greatly amplified compared to that into a peripheral organ such as the uterus.

Experimental Evidence for Concentration-Dependent Plasma Protein Binding Effects on the Apparent Half-Lives of Restrictively Cleared Drugs

Experimental Evidence for Concentration-Dependent Plasma Protein Binding Effects on the Apparent Half-Lives of Restrictively Cleared Drugs

Effects of human cirrhotic serum on estradiol and testosterone transport into rat brain.

It is known that sex hormone-binding globulin (SHBG), which binds both testosterone and estradiol in human serum, is markedly elevated in cirrhosis. In addition, it is known that the net metabolic clearance of testosterone is decreased in cirrhosis, while no change in the metabolic clearance of estradiol is observed. Since net clearance estimates are not direct measures of plasma protein binding effects, the present studies were designed to examine the effects of cirrhotic human serum on the unidirectional clearance of sex steroids using an in vivo rat brain paradigm previously described. Cirrhotic serum was characterized by the following changes from control serum: 2.6-fold increase in SHBG, 40% decrease in albumin, and 30% and 22% decreases in the dialyzable fractions of testosterone and estradiol, respectively. In addition, the non-SHBG-bound fraction of estradiol was decreased 41% in cirrhosis, but no significant change in the brain extraction of estradiol was observed using cirrhotic serum. In contrast, the unidirectional testosterone clearance was decreased 33% by cirrhotic serum. These studies indicate that changes in the net metabolic clearance of sex steroids closely parallel the changes in unidirectional clearance caused by alterations in plasma proteins. The absence of a decrease in estradiol clearance in cirrhosis in association with the substantial decrease in the non-SHBG-bound estradiol fraction is an unexpected finding, since previous studies have shown that a close parallel exists between these two parameters. A possible explanation is that estradiol bound to the SHBG in human cirrhotic serum is partially available for transport into peripheral tissues such as the brain.

The electrochemical interactions of heparin and the aminoglycoside antibiotics

Various aminoglycoside antibiotics—tobramycin "analytical standard"; tobramycin, gentamycin, kanamycin, and amikacin, all containing preservatives added in pharmaceutical preparations; and streptomycin—without preservatives—yield in aqueous solutions a conductance peak when titrated against a heparin aqueous solution, indicating the formation of a charge transfer complex, which subsequently dissociates. At the volume ratio corresponding to the peak, a colloidal suspensoid forms which is possibly a micellar complex stabilized by the ions produced by the dissociation of the complex. The clinical significance of this interaction is suggested by preliminary microbiological tests. The possible effects of plasma protein binding and dissociation of the complexes here reported in tissues, however, were not studied.

Effect of Plasma Protein Binding on Clearance of Drugs Metabolized by Michaelis-Menten Kinetics

Effect of Plasma Protein Binding on Clearance of Drugs Metabolized by Michaelis-Menten Kinetics

Effect of Plasma Protein Binding on Renal Clearance of Drugs

Effect of Plasma Protein Binding on Renal Clearance of Drugs

Studies Comparing the Metabolic Clearance Rate of 11β,17,21-Trihydroxypregn-1,4-Diene-3,20-Dione (Prednisolone) after Oral 17,21-Dihydroxypregn-l,4-Diene-3,11,20-Trione and Intravenous Prednisolone

The MCR of prednisolone (11 beta,17,21-trihydroxypregn-1,4-diene-3,20-dione) and the absorption of prednisone (17,21-dihydroxypregn 1,4-diene-3,11,20-trione) were studied in five normal subjects and four patients. Plasma and urinary prednisolone were measured by a competitive radioassay. The MCR was determined after iv administration of 30 mg nonisotopic prednisolone using one-compartment (MCR1) and two-compartment (MCR2) analyses. These values were compared with the MCR determined after oral administration of nonisotopic prednisone (MCR0). MCR1 and MCR2 were closely correlated, indicating the applicability of first order kinetics to the study of prednisolone metabolism. In subjects with normal gastrointestinal function, MCR0 was consistently lower than MCR2 (mean MCR2 = 0.175 liters/h.kg; MCR0 = 0.145 liters/h.kg). In two patients with steroid malabsorption, the MCR0 was significantly greater than MCR2. Knowledge of the expected relationship between MCR0 and MCR2 allowed quantitation of the degree of malabsorption. With or without impaired absorption, the absorptive process was essentially complete by the time of the peak plasma concentration. Estrogen therapy lowered the MCR0, and high prednisone dose increased the MCR0. Those effects and the effects of iv prednisolone administration on the MCR are explained by the effects of plasma protein binding of prednisolone. These studies demonstrate the usefulness of the oral MCR determination in the evaluation of steroid absorption and metabolism.

Comparative Pharmacokinetics of Coumarin Anticoagulants XXXI: Effect of Plasma Protein Binding on Distribution Kinetics of Dicumarol in Rats

The purpose of this investigation was to determine, with respect to dicumarol, the effect of plasma protein binding on the pharmacokinetic parameters used conventionally to describe the distribution kinetics of a drug on the basis of the time course of its plasma concentration. After rapid intravenous injection, plasma dicumarol concentrations in adult male Sprague-Dawley rats declined triexponentially, with the terminal exponential phase starting at about 4hr. The free fraction, f, of dicumarol in the serum of individual animals ranged from 0.000150 to 0.000790. The parameters of the equation Ct=Pe−πt+Ae−αt+Be−βt for plasma concentration Ct at time t were obtained by nonlinear least-squares computer fitting of the experimental data and varied appreciably between animals. Of these parameters, only β showed a significant correlation with f. These observations indicate that the distribution kinetics of this very extensively plasma protein-bound drug, as reflected by the time course of its plasma concentration after intravenous injection, are apparently not affected by intersubject differences in plasma protein binding. There is a remarkable similarity in the values of P, A, B, π, and α for dicumarol and warfarin, even though the serum free fraction of these drugs differs considerably.