Serum Protein Binding Of Phenytoin Research Articles

Clinical Response in Epilepsy in Relation to Total and Free Serum Levels of Phenytoin

The relationships between total and free serum concentrations of phenytoin and the clinical control of seizures were investigated retrospectively in 114 patients. Total phenytoin levels were measured by enzyme-modified immunoassay (EMIT), and the free fraction by ultrafiltration at 37 degrees C using 14C-labelled phenytoin as a tracer. The median free fraction in 188 serum samples was 13.7% (range 8.9-27.0%). The free fraction was greater than 18% in 34 (18.1%) of the serum samples. In all but 5 samples, a likely reason for the elevated free fraction could be determined. The identifiable reasons were commonly hypoalbuminaemia and the presence of liver or renal disease. There was a significant negative correlation between serum albumin level and free fraction of phenytoin (n = 90, r = -0.68, p less than 0.001). The free phenytoin concentration was strongly correlated with the total phenytoin concentration in serum (n = 188, r = 0.94, p less than 0.001). The total phenytoin concentration provided as good an indication of clinical response as the free concentration in 91 patients (85.8% of the patients for whom response could be reliably determined). In the other 15 (14.2%) patients, free phenytoin concentrations were better related to clinical effect. These patients generally had significant reductions in the serum protein binding of phenytoin. The relationship between phenytoin toxicity and free serum concentrations was particularly strong--in 14 patients with toxicity, the total serum concentration of phenytoin was greater than 80 mumol/L in only 42.9% of cases, while the free phenytoin concentration was greater than 8 mumol/L in 85.7% of the cases (p less than 0.05 by chi-square test).(ABSTRACT TRUNCATED AT 250 WORDS)

Stereoselective inhibition of diphenylhydantoin metabolism by p‐hydroxyphenyl‐phenylhydantoin enantiomers in rats

Different doses of rac-p-HPPH (0.4 and 4 mg/h) were given repeatedly to rats infused with [14C]phenytoin. The serum levels of 14C-labeled and unlabeled p-HPPH, and [14C]phenytoin were measured by an HPLC method and radiometric analysis. The clearance of phenytoin and p-HPPH was determined by rate of dosing divided by the steady-state concentration. The phenytoin clearance was significantly lower in the high dose p-HPPH injection group than in the low dose group (87 versus 262 ml/h), whereas p-HPPH clearance showed no difference. The formation clearance of [14C]p-HPPH was also significantly lower in rats injected with high dose of p-HPPH (35 versus 169 ml/h). The clearance of other elimination pathways was also lower in rats with high dose of p-HPPH (53 versus 89 ml/h). The serum protein binding of phenytoin was lower in rats injected with high dose of p-HPPH. The result indicated that injections of rac-p-HPPH mainly inhibited on the formation of p-HPPH itself. The formation of (R)-p-HPPH and (S)-p-HPPH in microsomal preparation was measured by a ligand-exchange chromatographic method. The formation of (S)-p-HPPH or (R)-p-HPPH was not only inhibited by the enantiomer itself, but also cross-inhibited by the other enantiomer. To the formation of either (S)-p-HPPH or (R)-p-HPPH, (S)-p-HPPH showed a higher inhibitory activity. The use of rac-p-HPPH to inhibit phenytoin metabolism in vivo involved several mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Protein binding of phenytoin and lidocaine in pediatric patients with type I diabetes mellitus.

We evaluated serum protein binding of phenytoin and lidocaine, the extent of albumin and hemoglobin glycation, and alpha-1-acid glycoprotein concentrations in 47 children and adolescents (9-17 years) with type I diabetes mellitus (DM). Serum was incubated with phenytoin (n = 47) and lidocaine (n = 32) to yield total concentrations of 19.4 +/- 1.34 and 4.6 +/- 0.5 mg/l, respectively. A serum ultrafiltrate was prepared from an aliquot of each sample by membrane centrifugation. Total and free concentrations of phenytoin (free fraction 8.3 +/- 1.0%) and lidocaine (free fraction 25.8 +/- 11.6%) were determined using a fluorescence polarization immunoassay technique. A linear relationship (r = 0.63, p = 0.0001, y = 40.1 + [-0.2x]) was also found between the alpha-1-acid glycoprotein concentration (68.5 +/- 20.9 mg%, range 47.2-134.7 mg%) and lidocaine-free fraction (n = 32). No relationship existed between the extent of glycated albumin and the lidocaine-free fraction, in contrast to the linear correlation (r = 0.49, p = 0.0005, y = 3.7 + 0.4x) found between the extent of glycated albumin and the free fraction of phenytoin in serum (n = 47). A similar correlation (r = 0.49, p = 0.014, y = 3.7 + 0.3x) was found between glycated albumin (%) and the phenytoin-free fraction (3.9 +/- 0.9%) when examined in a separate (n = 17) set of patient samples spiked to contain a total serum phenytoin concentration of 32.6 +/- 1.4 (range 29.4-35.7) mg/l. Our data demonstrate a predictable increase in the free fraction of phenytoin as the extent of albumin glycation increases in pediatric patients with type I DM who are in poor glycemic control. This relationship was not found for lidocaine, a representative basic compound. These findings suggest that glycation of albumin alters the binding of phenytoin in children and adolescents with type I DM.

Factors causing age-dependent changes in phenytoin tissue and serum binding in rats.

The factors that cause age-dependent changes in phenytoin tissue and serum protein binding in rats were studied. It was confirmed that the age-dependent changes in the concentration of tissue constituents to which phenytoin bound mainly governed the change in phenytoin tissue binding. The concentration of the tissue constituents, protein and phospholipid, was changed by the water content in tissues in the growth process of rats. The increase in serum protein binding of phenytoin in the growth process of rats was caused by changes in both serum albumin concentration and binding parameters of phenytoin to serum albumin. The changes in binding parameters of phenytoin to serum albumin were led by those in the molar rations of free fatty acids to albumin.

Alteration of phenytoin binding by glycosylation of albumin in IDDM.

We measured glycosylated albumin and hemoglobin and serum protein binding of phenytoin in 57 children and adolescents with insulin-dependent diabetes mellitus (IDDM). Serum was incubated with phenytoin to yield concentrations of 15 and 25 mg/L, and a serum ultrafiltrate was prepared from an aliquot of each sample. We observed a linear correlation between glycosylated albumin and the free fraction of phenytoin at serum phenytoin concentrations of 15 mg/L (r = .35, P = .03) and 25 mg/L (r = .40, P = .003). A better correlation existed between the free fraction of phenytoin and total albumin concentrations for both serum concentrations (r = .45, P = .005 for 15 mg/L; r = .56, P = 10-5) for 25 mg/L), whereas the best linear correlation resided between the free fraction of phenytoin and the concentration of nonglycosylated albumin (r = .54, P = .0005 for 15 mg/L; r = .63, P less than 10(-6) for 25 mg/L). There was no correlation between the free fraction of phenytoin and the concentration of glycosylated albumin. Incubation of solutions of glycosylated and nonglycosylated albumin demonstrated significantly lower binding to the glycosylated fraction (P = 8.1 X 10(-6)). These results indicate that glycosylation of albumin diminishes the affinity of the phenytoin binding site on albumin. This alteration may have clinical significance in that it may alter the disposition of phenytoin in patients with IDDM and produce free phenytoin serum concentrations that are not accurately reflected by total serum phenytoin concentrations.

Hepatic extraction of endogenous inhibitors of drug binding to serum protein in the pregnant rat

Significant decreases in the serum protein binding of a fluorescent dye, 1-anilino-8-naphthalenesulfonate (ANS), and salicylic acid (SA) were observed in pregnant rats compared to that in nonpregnant (control) rats. A significant difference in the serum protein binding of ANS and SA between serum samples taken at the hepatic vein and portal vein or femoral artery was also observed in the pregnant rats, while such a sampling site difference in the serum protein binding was not observed in the control rats. In the pregnant rats the affinity of ANS binding to the primary binding site in the serum from the hepatic vein was approximately 70% higher than that in the case of serum from the portal vein. The hepatic extraction of nonesterified fatty acids (NEFA) was also determined, and the extraction ratios in the control and pregnant rats were 0.55 and 0.31 respectively. We concluded from these findings and other evidence that certain endogenous inhibitors of drug binding to serum protein (such as NEFA), which increase during pregnancy, were extracted efficiently by the liver and that the difference in the serum protein binding of ANS and SA between serum samples taken at the hepatic and portal veins or femoral artery in the pregnant rats may be explained by the hepatic extraction of endogenous inhibitors. Our present results support the previous finding by Chou et al. [ Int. J. Pharm. 18, 217 (1984)] that in pregnant rats the serum protein binding of phenytoin is greater in the hepatic vein than that in the femoral vein.

Serum protein binding of drugs and bilirubin in newborn infants and their mothers.

The serum protein binding of dexamethasone, diazepam, phenytoin, sulfisoxazole, and bilirubin was determined in full-term newborn infants and their mothers immediately after delivery. Diazepam and sulfisoxazole were bound more extensively in infant serum than in maternal serum, dexamethasone was bound more extensively in the maternal serum, and the serum protein binding of phenytoin and bilirubin was much the same in the infants and their mothers. These results are consistent with and explain clinical observations that diazepam concentrations are higher in cord than in maternal serum and that phenytoin levels in newborn and maternal sera are in the same range when the mothers had taken one of these drugs for some time before delivery. There were negative correlations between free fraction of drug and albumin concentration in serum only for phenytoin (infants and mothers), diazepam (infants only), and sulfisoxazole (mothers only). Except for dexamethasone (infants and mothers) and phenytoin (mothers), there were positive correlations between the free fraction values of the various compounds and those of salicylic acid. It was previously determined that salicylic acid is more extensively bound in newborn than in maternal serum. Induction or augmentation of labor with oxytocin was associated with decreased serum protein binding of diazepam and possibly of bilirubin in mothers. There was a significant correlation between serum free fraction values in newborns and their mothers only in the case of phenytoin. Clinical Pharmacology and Therapeutics (1980) 28, 58–63; doi:10.1038/clpt.1980.131

Effects of sodium valproate on the serum protein binding of phenytoin, and on liver enzyme activity.

The effect of a single dose of sodium valproate on the serum protein binding of phenytoin was studied in 6 epileptic patients receiving phenytoin maintenance therapy. Phenytoin was significantly displaced by valproic acid, but the free concentration of phenytoin was unchanged because of redistribution to tissues. As a result of this effect, total serum phenytoin concentration was significantly lowered. Induction of metabolism probably does not play a part in this interaction because antipyrine half-lives and urinary D-glucaric acid excretion were not altered by chronic administration of sodium valproate to 8 patients receiving this drug alone.

Pharmacokinetics of drugs in rabbits with experimental acute renal failure.

Serum protein binding and serum levels of antipyrine, phenytoin and phenylbutazone were measured in rabbits with acute renal failure induced by uranyl nitrate. The kinetics of antipyrine are not altered in the uraemic rabbit. Serum protein binding of phenytoin and phenylbutazone is decreased and the volume of distribution of total drug is increased. The volume of distribution of free phenytoin is unchanged, that of free phenylbutazone is decreased in acute renal failure. The half-lives of phenytoin and phenylbutazone are unchanged, but the serum clearance is increased in experimental acute renal failure. The intrinsic clearance of free drug, i.e. the ability of the liver enzymes to metabolize the drug, is not altered for antipyrine, is increased for phenytoin and is decreased for phenylbutazone. It is difficult to extrapolate these data obtained in rabbits with acute renal failure to humans in chronic renal failure.

Intraindividual Relationships between Serum Protein Binding of Drugs in Normal Human Subjects, Patients with Impaired Renal Function, and Rats

The serum protein binding of phenytoin, salicylic acid, sulfisoxazole, and warfarin was determined in normal human adults, in patients with impaired renal function (kidney donor and recipient), and in adult male Sprague-Dawley rats. The free fraction values for salicylate and sulfisoxazole were significantly correlated in all three groups. The other correlations were statistically significant in only one or two of these groups. There was a statistically significant negative correlation between albumin concentration and the free fraction values of salicylic acid and sulfisoxazole (but not of phenytoin and only under special circumstances with warfarin) in normal human subjects and of phenytoin, salicylic acid, and sulfisoxazole (but not warfarin) in rats. No such correlation was observed for any of the drugs in patients with impaired renal function. These observations show that no single weakly acidic drug can serve as an index for quantitatively determining the effect of disease or species differences on the serum protein binding of other weakly acidic drugs.

Frequency distribution of free warfarin and free phenytoin fraction values in serum of healthy human adults.

The protein binding of racemic warfarin was determined in serum from 57 normal adults (27 men and 30 women). The free fraction of warfarin (total concentration, 0.8 mug/ml) ranged from 0.0050 to 0.0186 and was log-normally distributed. The frequency distribution differs from that in rats in which the serum free fraction values of warfarin are trimodally distributed. The protein binding of phenytoin was determined in serum from 39 of the subjects. The free fraction of phenytoin (total concentration, 15 mug/ml) ranged from 0.111 to 0.155 and was unimodally distributed. There was no apparent correlation between the extent of protein binding of warfarin and phenytoin in individual serum samples. The pronounced intersubject variation in serum free fraction of warfarin observed in consistent with our previous finding that serum protein binding is an important determinant of interindividual differences in the total clearance of warfarin in man. On the other hand, the relatively narrow distribution of free phenytoin fraction values suggests that differences in serum protein binding of phenytoin are not an important cause of the prounounced interindividual differences in the total clearance of phenytoin by subjects with normal renal function.