Unbound Phenytoin Research Articles

Doxorubicin alters the disposition of phenytoin by reducing its metabolic elimination and binding affinity to serum albumin in rats.

We investigated the pharmacokinetic interaction of doxorubicin (DOX) with phenytoin (PHT) and the underlying mechanism in rats to clarify why the serum PHT concentration decreases despite the impaired PHT metabolic capacity in patients receiving DOX. Rats were administered 15 mg/kg of DOX or saline alone. The pharmacokinetic disposition of intravenously administered PHT was examined 4 days after DOX exposure. Enzyme kinetics of CYP2C-dependent PHT p-hydroxylation were analysed using hepatic microsomes. The unbound PHT concentration in serum was measured by the ultrafiltration method, and the relationship between the unbound fraction (fu) and serum albumin level was assessed. The total clearance (CLtot) of PHT was significantly increased by DOX, but the activity of PHT p-hydroxylation conversely decreased. The unbound serum PHT concentration and its fu were significantly higher in the DOX group than in the control group, and the CLtot/fu, a measure of intrinsic clearance, significantly decreased. An increase in the fu was observed even when using a serum sample with an albumin concentration equal to that in the control group. DOX treatment increases the unbound serum PHT concentration by depressing the metabolic capacity and alters the total PHT by reducing serum albumin and its affinity to PHT.

Response to “Characterization of Unbound Phenytoin Concentrations in Neurointensive Care Unit Patients Using a Revised Winter-Tozer Equation” by Sean P. Kane, Adam P. Bress, and Eljim P. Tesoro

Response to “Characterization of Unbound Phenytoin Concentrations in Neurointensive Care Unit Patients Using a Revised Winter-Tozer Equation” by Sean P. Kane, Adam P. Bress, and Eljim P. Tesoro

Phenytoin Removal by Continuous Venovenous Hemofiltration

To describe 2 cases of clinically significant phenytoin removal during continuous venovenous hemofiltration (CVVH) and review the relevant literature regarding phenytoin removal by renal replacement modalities. A 64-year-old female with chronic kidney disease and cirrhosis was admitted to the intensive care unit (ICU) with a traumatic subdural hematoma and seizures. The patient received a loading dose of intravenous phenytoin 1000 mg, followed by maintenance intravenous administration of phenytoin 100 mg and levetiracetam 250 mg every 12 hours. CVVH was initiated for acidosis. A 63-year-old male was admitted to the ICU after cardiac surgery complicated by hypotension. CVVH was initiated for fluid overload, and phenytoin was initiated 3 days later for seizures. A loading dose of intravenous phenytoin 2700 mg was administered, followed by maintenance dosing of intravenous phenytoin 150 mg every 8 hours. Concentrations of unbound phenytoin in serum and CVVH effluent samples were measured during concomitant treatment in each patient. In both patients, serum and effluent concentrations of unbound phenytoin fell steadily while they were on CVVH. Clearance of phenytoin by CVVH was calculated, as was the daily removal of phenytoin, as a percentage of total daily phenytoin dosage during each sampling period. Phenytoin clearance by CVVH ranged from 11 to 13 mL/min in these patients. The clearance of phenytoin with CVVH in these 2 patients was much higher than the renal clearance of phenytoin reported in healthy volunteers with normal renal function. Previous case reports have demonstrated that only small, clinically insignificant amounts of phenytoin are removed by hemodialysis, and the only published report of phenytoin removal by continuous renal replacement therapy used hemofiltration rates much lower than those used in the 2 cases described here. These cases demonstrate that a substantial amount-approximately 30%-of total daily phenytoin dose may be removed by CVVH, and patients may require higher than expected empiric doses. Phenytoin concentrations should be closely monitored in critically ill patients receiving CVVH.

The Quantitative Effect of Serum Albumin, Serum Urea, and Valproic Acid on Unbound Phenytoin Concentrations in Children

Dosing of phenytoin is difficult in children because of its variable pharmacokinetics and protein binding. Possible covariates for this protein binding have mostly been univariately investigated in small, and often adult, adult populations. We conducted a study to identify and quantify these covariates in children. We extracted data on serum phenytoin concentrations, albumin, triglycerides, urea, total bilirubin and creatinine concentrations and data on coadministration of valproic acid or carbamazepine in 186 children. Using nonlinear mixed effects modeling the effects of covariates on the unbound phenytoin fraction were investigated. Serum albumin, serum urea concentrations, and concomitant valproic acid use significantly influenced the unbound phenytoin fraction. For clinical practice, we recommend that unbound phenytoin concentrations are measured routinely. However, if this is impossible, we suggest to use our model to calculate the unbound concentration. In selected children, close treatment monitoring and dose reductions should be considered to prevent toxicity.

Effect of mild therapeutic hypothermia on phenytoin pharmacokinetics.

The authors examined the effect of mild therapeutic hypothermia on phenytoin pharmacokinetics in 14 patients with brain damage. Each patient was given phenytoin during and after mild therapeutic hypothermia. Plasma concentrations of total phenytoin, unbound phenytoin, and 5-(p-hydroxyphenyl)-5-phenylhydantoin (5-p-HPPH), the major metabolite of phenytoin, were measured. Pharmacokinetic parameters during and after mild therapeutic hypothermia were compared. Plasma concentrations of total and unbound phenytoin were significantly higher during hypothermia than after hypothermia. The area under the plasma concentration-time curve (zero to infinity) was increased by 180% and mean residence time was prolonged by 180% during hypothermia compared with the corresponding values after hypothermia. Moreover, the elimination constant (ke) was decreased by 50% and elimination clearance of phenytoin was decreased by 67% during hypothermia compared with the corresponding values after hypothermia. The plasma concentration of 5-p-HPPH was significantly lower during hypothermia than after hypothermia. These findings suggest that phenytoin metabolism is inhibited by mild therapeutic hypothermia.

No gender effect on binding characteristics of phenytoin to serum proteins in monotherapy for adult patients with epilepsy.

The aim of the present study was to determine the gender-related binding characteristics of phenytoin (PHT) to serum proteins in adult patients with epilepsy. Serum samples examined in the study were obtained from 80 adult patients (40 men and 40 women) with epilepsy on PHT monotherapy. Their age ranged from 16 to 64 years (mean [SD], 36.0 [11.7] years). Protein binding of PHT was evaluated by ultrafiltration under current laboratory routine conditions (25 +/- 3 degrees C). The in vivo binding parameters of PHT to serum proteins were determined using a binding equation derived from the Scatchard equation for a one-site binding model. No significant differences were observed in age and serum concentrations of albumin between male and female patients (p > 0.05), but significant differences were observed in serum concentrations of total and unbound PHT between the two groups (p < 0.05). The mean association constant of PHT to serum proteins is the same value of 0.008 L micromol(-1) between male and female patients, whereas total concentration of binding sites seems to be similar between the two groups (1389 micromol L(-1) for men and 1345 micromol L(-1) for women). No significant differences were observed in binding characteristics of PHT to serum proteins between male and female patients (p > 0.05). Our results show that gender does not have a significant effect on the binding characteristics of PHT to serum proteins in adult patients receiving monotherapy under normal pathophysiologic conditions.

Effect of Temperature on Binding Characteristics of Phenytoin to Serum Proteins in Monotherapy Adult Patients With Epilepsy

The effects of temperature on binding characteristics of phenytoin (PHT) to serum proteins were determined in adult patients with epilepsy. Serum samples examined in the study were obtained from 47 adult patients (29 men, 18 women) with epilepsy on PHT monotherapy. Ages ranged from 18 to 64 years (mean [SD], 36.8 [12.1] years). Protein binding of PHT was evaluated by ultrafiltration under current laboratory routine conditions (25 +/- 3 degrees C) or at a temperature of 37 degrees C. The in vivo binding parameters of PHT to serum proteins were determined using a binding equation derived from the Scatchard equation for a one-site binding model. Significant differences were observed in serum concentrations of unbound PHT between paired data (P <.05). The mean association constants (K) of PHT to serum proteins are 0.009 L micromol(-1) at 25 +/- 3 degrees C and 0.003 L micromol(-1) at 37 degrees C, whereas mean total concentrations of binding sites [n(Pt)] are 1215 micromol L(-1) for 25 +/- 3 degrees C and 2263 micromol L(-1) for 37 degrees C. Significant differences were observed in binding characteristics of PHT to serum proteins between the data determined in different conditions of ultrafiltration (P <.05). Our study confirms that binding affinity for PHT-serum protein interaction is approximately 67% lower at 37 degrees C than at 25 +/- 3 degrees C, and, consequently, binding potential [K.n(Pt)] is approximately 38% lower at 37 degrees C than at 25 +/- 3 degrees C.

Falsely Increased Immunoassay Measurements of Total and Unbound Phenytoin in Critically Ill Uremic Patients Receiving Fosphenytoin

Fosphenytoin, a phosphate ester prodrug of phenytoin, is metabolized to phenytoin in vivo. Phenytoin metabolites accumulate in renal insufficiency and cross-react in some phenytoin immunoassays. Our aim was to determine the accuracy of phenytoin immunoassays in renal patients treated with fosphenytoin. We measured phenytoin with HPLC and with the aca, ACS:180, TDx phenytoin II, Vitros, and AxSYM methods. Specimens were collected 2-120 h after fosphenytoin administration from 17 patients with renal insufficiency. The AxSYM, TDx phenytoin II, ACS:180, and Vitros assays displayed falsely increased phenytoin results up to 20 times higher than the HPLC results. The aca Star results for these specimens were comparable to the HPLC results. Although fosphenytoin can cross-react with phenytoin immunoassays, no fosphenytoin was detected by a sensitive HPLC method in any sample that was tested for its presence. These results are consistent with the formation of one or more novel metabolites or adducts of fosphenytoin that accumulate in some critically ill patients with renal insufficiency and that display significant cross-reactivity with some, but not all, phenytoin immunoassay methods.

Predicting Unbound Phenytoin Concentrations in the Critically Ill Neurosurgical Patient

To assess the correlation of measured unbound phenytoin concentration (dphF) to estimated unbound concentration (dphEF) using the Sheiner-Tozer equation in critically ill patients in the neurosurgical intensive care unit. The dphF and total phenytoin (dphT) trough serum concentrations were measured during the first week of therapy in 17 consecutive patients with albumin concentrations less than 3.5 g/dL. Serum albumin concentrations were measured within 24 hours of serum phenytoin concentration measurement. A university-affiliated urban teaching hospital. The study population consisted of 17 neurosurgical patients who were at least 18 years old. The predictability of the Sheiner-Tozer equation was tested by measuring dphF, dphT, and serum albumin concentrations. Measured phenytoin concentrations were compared with phenytoin concentrations calculated from the Sheiner-Tozer equation. To estimate correlation between variables linear regression was calculated. Mean absolute value of error and mean error were estimated to assess precision and bias between measures, respectively. The mean +/- SD dphT was 13.05 +/- 5.15 mu g/mL. The measured dphF was 1.89 +/- 0.80 compared with 2.00 +/- 0.8 mu g/mL for the dphEF (NS). Regression analysis for dphEF versus dphF revealed a significant correlation (r2 = 0.94, p = 0.001). The mean absolute value of error for the Sheiner-Tozer equation to predict dphEF was 0.167, which was 9% of the mean value of dphEF (1.89). These results indicate that, in this population, the Sheiner-Tozer estimate of dphEF provides an unbiased, precise clinical estimate of dphF in patients for whom measured dphF is unavailable or impractical.

Predicting Unbound Phenytoin Concentrations In Patients Receiving Valproic Acid: A Comparison of Two Prediction Methods

To compare the predictive performance of 2 equations that estimate unbound (free) phenytoin plasma concentrations when valproic acid (VPA) and phenytoin are administered concurrently. Eighty-eight adults receiving VPA and phenytoin concurrently were included in the study. Steady-state plasma concentration measurements of total phenytoin, total VPA, and unbound phenytoin were collected prospectively in the inpatient group (group 1) and retrospectively in the outpatient group (group 2). Using the equations developed by Haidukewych and May, unbound phenytoin concentrations were calculated. The mean predicted unbound phenytoin concentrations then were compared with mean actual unbound phenytoin concentrations measured in the laboratory. Identical assays were performed to measure unbound phenytoin, total phenytoin, and VPA from each patient group. Antiepileptic drug concentration measurements were collected from 43 inpatients (mean age 34.8 y) from the epilepsy unit at Abbott Northwestern Hospital and 45 outpatients (mean age 37.3 y) at the MINCEP Epilepsy Care clinic, Minneapolis, MN. Mean prediction error (MPE) and mean squared error (MSE) were calculated to determine which equation was the least biased and most precise in predicting unbound phenytoin when total VPA and phenytoin concentrations are known. Linear regression of predicted unbound phenytoin on measured unbound phenytoin values determined the correlation coefficients (r). A paired Student's t-test also was performed comparing mean predicted unbound phenytoin concentration with mean actual unbound phenytoin concentrations in both groups. The MPE from May's equation was -0.49 and -0.45 for groups 1 and 2, respectively; using the Haidukewych equation, MPE was -0.02 and 0.08 for groups 1 and 2, respectively. The MSE using May's equation was 0.34 for both groups. Using the Haidukewych equation, group 1 MSE was 0.07, and for group 2, 0.12. Correlation coefficients were more than 0.91 (p < 0.001) from each equation in both patient groups. In group 1, mean actual unbound phenytoin concentration was 2.02 micrograms/mL; May's equation predicted 1.52 micrograms/mL (p < 0.001) and the Haidukewych equation predicted 2.00 micrograms/mL (p = 0.64). In group 2, mean actual unbound phenytoin concentration was 2.10 micrograms/mL; May's equation predicted 1.65 micrograms/mL (p < 0.001) and the Haidukewych equation predicted 2.18 micrograms/mL (p = 0.11). Haidukewych's equation predicts unbound phenytoin concentrations with the least bias and most precision with statistical significance. May's equation consistently underpredicted unbound phenytoin concentrations. Because unbound phenytoin fraction is not constant (and usually more than the expected 10%) in patients comedicated with VPA, unbound phenytoin concentrations cannot be predicted even though total VPA and phenytoin concentrations are known. If unbound phenytoin concentrations cannot be readily measured, Haidukewych's equation is a reliable predictor of unbound phenytoin concentrations.

Effect of barbiturate therapy on phenytoin pharmacokinetics.

To evaluate the effect of high-dose pentobarbital therapy on phenytoin pharmacokinetics. A prospective, clinical study. The intensive care unit of a university hospital. Ten adult patients with cerebral lesions requiring anticonvulsants and control of intracranial pressure. Each patient received phenytoin sufficient to maintain a plasma concentration at 15 micrograms/mL (60 mumol/L) both before and after barbiturate therapy. Plasma concentrations of total phenytoin, unbound phenytoin, and the major metabolite of phenytoin, 5-(p-hydroxyphenyl)-5-phenylhydantoin, were measured, and pharmacokinetic variables obtained before and after barbiturate therapy were compared. Plasma concentrations of total phenytoin remained within the therapeutic range during the 12-hr period preceding barbiturate therapy. After barbiturate therapy, plasma concentrations of both total and unbound phenytoin were significantly less than those concentrations before barbiturate therapy. For total phenytoin, maximum metabolic velocity was increased by 62% (1.09 +/- 0.62 to 1.77 +/- 0.52 mg/L/hr, 1.20 +/- 0.68 to 1.95 +/- 0.57 nmol/L/sec, p < .05), and area under the plasma concentration-time curve (0 to infinity) and mean residence time were each decreased by 73% (32.5 +/- 20.0 to 8.7 +/- 3.1 min.mg/mL, 2.14 +/- 1.25 to 0.57 +/- 0.19 sec.mmol/L, p < .01, and 135,000 +/- 69,000 to 37,000 +/- 11,000 secs, p < .005, respectively) after barbiturate therapy. The plasma concentration of the principal metabolite of phenytoin, 5-(p-hydroxyphenyl)-5-phenylhydantoin, was significantly increased after barbiturate therapy. Phenytoin metabolism is increased by barbiturate therapy, and supplemental doses of phenytoin and frequent drug monitoring may be required after barbiturate therapy.

Unbound Phenytoin and Valproate Monitoring

All antiepileptic drug blood level determinations at the University of Virginia Medical Center were analyzed over a 13 month period.

Pharmacokinetic Pitfalls of Present Antiepileptic Medications

Some patients fail to respond to antiepileptic drugs (AEDs) or their response varies over time. Unexpected fluctuations in AED concentrations can cause loss of seizure control or side effects. Recent advances in AED clinical pharmacology have characterized a number of factors that alter AED concentrations. Storage of carbamazepine (CBZ) and phenytoin (PHT) formulations in hot, humid conditions alters the dosage form and reduces bioavailability up to 50%. Diurnal changes in gastrointestinal physiology affect disintegration of valproate (VPA) enteric-coated tablets, reducing nighttime drug concentrations 30 to 40% compared with daytime values. Drug-drug interactions or pathophysiologic conditions (e.g., hypoalbuminemia) may displace AEDs from protein-binding sites reducing total but not unbound drug concentrations decrease. VPA not only displaces PHT from protein-binding sites but inhibits its metabolism, causing a decrease in total concentration and an increase in the unbound concentration, occasionally necessitating a reduction in PHT dosage. Alterations in drug metabolism can cause fluctuations in the concentrations of AEDs and active metabolites. Enzyme inhibitors such as cimetidine or VPA can increase concentrations of both CBZ and CBZ epoxide (CBZE). Enzyme inducers such as ethanol, PHT, CBZ, and phenobarbital accelerate the metabolism of other AEDs. Some forms of physiologic stress increase binding to alpha 1-acid glycoprotein (AAG), which is stimulated within hours of a myocardial infarction or major surgery. Total CBZ and CBZE, both of which bind to AAG, increase over the same period. Stress may also activate hepatic drug metabolism. Unbound PHT clearance increases 7 to 21 days following head trauma, necessitating larger maintenance doses. Age greater than 65 years is associated with decreases in protein binding and drug clearance and longer elimination half-lives. In elderly patients, AED dosage may need to be reduced and the dosing interval extended. Knowledge of these factors permits prospective assessment of risk and the design of treatment plans that minimize fluctuations in response.

Nomogram for the Prediction of Unbound Phenytoin Concentrations in Patients on a Combined Treatment of Phenytoin and Valproic Acid

The interpretation of the total phenytoin (PT) concentration can be problematic if valproic acid (VPA) is given as a comedication, because VPA displaces PT from the protein binding sites and can increase the free fraction of PT. In order to estimate the free or unbound PT concentration (PTf) from the total PT concentration (PTt) and VPA concentration, a nomogram was constructed and evaluated. Data of 84 patients on combined therapy with PT and VPA were used in drawing up the regression equation (PTf = 0.0792.PTt + 0.000636.PTt.VPA) from which the nomogram was constructed. The predictions were checked using another 33 patients whose serum concentrations were determined in the morning and in addition several times during the day. The results show that using this method the PTf concentrations can be accurately estimated from the PTt and VPA concentrations.

Fluctuations of Unbound and Total Phenytoin Concentrations During the Day in Epileptic Patients on Valproic Acid Comedication

The influence of daily fluctuations in the concentration of valproic acid (VPA) on the unbound and total concentration of phenytoin (PT) was examined in a prospective study. The serum concentrations of 28 patients with epilepsy (group PT + VPA) who were treated with PT and concurrently with VPA and 15 patients (group PT) who were treated with PT but without VPA comedication were determined at 8.00, 11.00, 14.00, 17.00, and in part at 20.00 h. The results show that there are significantly greater fluctuations in the total PT concentration among patients on VPA than those not on VPA. The fluctuations in the total PT concentration during the day correlated with the fluctuations in the VPA concentration. On the other hand, the fluctuations in the concentrations of unbound PT of patients on VPA were comparable with those not on VPA. In the absence of VPA, the diurnal fluctuations of the total PT concentration correlated highly significantly with the fluctuations of the unbound PT concentration. This is not the case in patients on VPA. Our data are a further indication of the smaller significance of the total PT concentration as compared with the unbound PT concentration in the combined treatment of PT and VPA.

Brain interstitial fluid and intracellular distribution of phenytoin.

After intravenous (i.v.) administration (10 mg/kg), the biodisposition of phenytoin (PHT) in serum (total and free concentration), cerebrospinal fluid (CSF), brain, and the interstitial fluid (IF) of the normal brain were determined in dogs. A sufficient volume of IF was obtained through a multiperforated polypropylene ball implanted into the left parietotemporal region for 4-5 weeks. PHT brain distribution coefficient values ranged between 1.9 and 3.75, while the ratios of IF to free serum PHT concentrations ranged between 0.19 and 1.04; thus, our data indicate that most of the free unbound PHT which enters the brain parenchyma accumulates in the cellular compartment. Furthermore, at 60 and 90 min the peak CSF and IF concentrations are delayed; thus, for PHT, an apparent diffusion front from the CSF into the extracellular space of the brain seems to occur.

Phenytoin binding in healthy volunteers.

The pharmacologic effect of phenytoin is directly related to the unbound concentration in the serum, which previously has been reported in the literature to be approximately 10%. The results of 13 out of 14 20-35 year-old healthy male volunteers studied indicate that less than 10% unbound phenytoin is present in the majority of subjects taking two different doses of phenytoin.

Effect of plasma exchange on phenytoin plasma concentration.

The effect of plasma exchange on the concentrations of total and unbound phenytoin (DPH) was determined in one normal subject and four patients undergoing therapeutic plasma exchange (PE). The average percentage of plasma exchanged ranged from 57 to 70% among the five patients studied. The reduction in DPH concentration was more variable and ranged from 10 to 16%. However, the extent of reduction within each patient for both total DPH and "free" DPH serum concentrations was minimal; in addition, the ratio of free to total DPH remained virtually unchanged by plasma exchange. Thus, the replacement of endogenous albumin with exogenous albumin, which is an integral part of therapeutic plasma exchange, does not appear to alter DPH binding. Our findings indicate that rapid equilibration of DPH occurs between the intra- and extravascular compartments and that for the occasional plasma exchange procedure, dose adjustment does not appear indicated.

A New Approach to Establishing Individualized Maintenance Doses of Phenytoin

A new method for arriving at individualized doses of phenytoin (PHT) is described. This method, designated the eta method, operates on two measurements of unbound plasma PHT. One plasma sample is collected 48 h after a single, oral PHT dose (approximately 4.5 mg/kg). From the single dose, single sample measurements of plasma unbound PHT individual estimates of intrinsic unbound PHT clearance (CLuint) can be made. With the use of CLuint and weight-adjusted mean population values of Vmax predicted plasma concentrations of unbound PHT for 11 subjects exhibited a mean prediction error (mpe) of 0.07 mg/L with 95% confidence limits of -0.11 to 0.26 mg/L and an error coefficient of variation [root mean squared error (rmse)/mean unbound concentration at steady state (Cuss)] of 0.4. The second plasma unbound PHT is collected at steady state. From CLuint and one Cuss measurement, Vmax and Km can be directly calculated. Doses calculated by the eta method to produce plasma unbound PHT concentrations of approximately 1.0 mg/L were identical to those calculated by the Bayesian method to yield plasma total PHT concentrations of approximately 10 mg/L. When those doses were administered to six subjects, the plasma unbound PHT concentrations predicted by the eta method exhibited a mpe of -0.13 mg/L, with 95% confidence limits ranging from -0.33 to 0.06 mg/L and an error coefficient of variation of 0.2. The plasma PHT concentrations predicted by the Bayesian method exhibited an mpe of -1.30 mg/L, with 95% confidence limits of -4.02 to 1.42 mg/L and an error coefficient of variation of 0.3. Values of Vmax and doses of PHT calculated to produce a Cuss of 1.0 mg/L calculated by the eta method were nearly identical to Vmax values and doses of PHT calculated by Bayesian double feedback (based on two different Css values) to produce a Css of 10.0 mg/L.

Phenytoin as a probe of drug metabolism. Predicting clearance with a salivary sample.

Phenytoin (PHT) was administered in single 300-mg doses to each of 12 healthy, male subjects. Serial blood samples and salivary samples were collected for the next 48 h, and concentrations of plasma total PHT, unbound plasma PHT (plasma ultrafiltrates), and salivary PHT (salivary ultrafiltrates) were measured by immunofluorescence polarization. The following parameters were estimated: CLint/F, CL'int/F, V/F, and total and unbound PHT plasma disappearance half-lives. Estimates of CL'int/F (CL'int/F) were calculated from single 48-hour salivary PHT measurements. Mean (+/- SD) values were 0.026 +/- 0.009 1 . h-1 . kg-1, 0.385 +/- 0.148 1 . h-1 . kg-1, 12.6 +/- 3.5 l/kg (referenced to unbound drug), 28.0 +/- 12.1 h, and 25.9 +/- 13.5 h, respectively. When V/F referenced to unbound PHT was set at 14 l/kg, CL'int/F estimates were good predictors of the actual CL'int/F values demonstrating a mean prediction error of -9.012 1.h-1.kg-1. These data demonstrate that under specified conditions, intrinsic unbound PHT clearance can be estimated from a single PHT measurement in saliva, thereby permitting PHT to be used safely and sampled simply and noninvasively as a probe of hepatic mixed function oxygenase activity in humans.