Role of sweat in accumulation of orally administered griseofulvin in skin.

Griseofulvin, an orally effective antimicrobial agent, appears in the stratum corneum within 4-8 h after oral administration. Griseofulvin distribution was found to be highest in the outermost layers of the stratum corneum (level I, 20.8+/-1.5 ng/mg) and lowest inside (level II, 10.0+/-1.5; level III, 7.5+/-2.2 ng/mg). In order to study the precise mechanism of griseofulvin transfer to stratum corneum, the role of sweat in the accumulation of griseofulvin was considered. Heat-induced total body sweating decreased the mean stratum corneum concentration of griseofulvin by 55%, and 200-300 ng of griseofulvin accumulated per ml of sweat. A silicone hydrophobic resin was used to differentiate between "wash-off" and carrier properties of sweat for griseofulvin. Prevention of transepidermal water and sweat loss by (a) topical application of formaldehyde-releasing cream to one palm, (b) occlusion by a 2 x 2-cm patch on one arm, and (c) wearing a rubber glove for 24 h, showed a lower griseofulvin concentration when compared to control areas in the same subjects. The results of the gloved hand experiment show that a complete equilibrium is established at all three levels of stratum corneum, thereby removing the reversed gradient. These results support the hypothesis that a "wick effect" is responsible for the observed reversed drug gradient within the stratum corneum. The results of the experiments suggest that sweat and transepidermal fluid loss play an important role in griseofulvin transfer in stratum corneum.

Nonlinear theophylline elimination.

Elimination kinetics of theophylline and its major metabolites were investigated in 14 healthy adults in single-dose studies and in a multiple-plateau study. The plasma concentrations of theophylline and the metabolites 3-methylxanthine (3-MX), 1-methyluric acid (1 MU), and 1,3-dimethyluric acid (13-MU) were monitored to about 0.020 mg/l and became convex descending at concentrations below 1 mg/l after single theophylline doses. Renal clearance values of 3-MX, 1-MU, and 13-MU were 12.0 +/- 1.3 l/hr, 22.5 +/-1.5 l/hr, and 22.6 +/- 1.6 l/hr. Metabolite formation of the three metabolites followed Michaelis-Menten kinetics and became capacity limited within the therapeutic range of theophylline. The apparent Michaelis-Menten parameters for each metabolite formation step were obtained by computer fitting. For the formation of 3-MX. 1-MU, and 13-MU, the approximate mean maximal rate of formation of metabolite (Vmax) values were 5 mg/hr, 13 mg/hr, and 34 mg/hr and the apparent concentration of theophylline at which metabolite formation rate is half of Vmax values were 2.7 mg/l, 9.3 mg/l, and 14.2 mg/l. The elimination of each of the metabolites was rate limited by the elimination of theophylline. Concomitant measurement of theophylline urinary excretion rate showed the renal clearance of the drug to be highly dependent on urine flow. The initial renal clearance, elevated due to diuresis, and the distribution phase tended to counterbalance the saturable metabolic formation clearance after a single therapeutic dose. Therefore, plasma theophylline concentration decayed roughly in a log-linear fashion and the convex-descending curve, characterized by capacity-limited elimination kinetics, was observed only at lower concentrations.

Influence of allopurinol on theophylline disposition in adults.

Theophylline kinetics after intravenous aminophylline were determined in 5 nonsmoking healthy males before and after allopurinol for 1 wk. There was no significant alteration in theophylline disposition.

Saturation kinetics of iodipamide.

To characterize the saturation kinetics of iodipamide, timed samples of blood, urine, and bile were taken from two unanesthetized dogs infused with iodipamide at increasing rates to achieve various steady state blood concentrations. Biliary excretion rate of iodipamide reached an asymptote with increasing blood concentration, indicating a biliary transport maximum (Tm) of 15.2 to 16.2 mgI/min. Urinary excretion was not a pure, first order process and urinary excretion rate was higher than the glomerular filtration rate corrected for plasma protein binding, suggesting that active tubular secretion may play a part. Extrarenal elimination followed Michaelis-Menten kinetics. Estimates of maximum rate (Vm) and Michaelis-Menten constant (Km) were obtained graphically. The estimated values of Vm were 4 to 6 times that of biliary Tm. In acute infusion experiments the iodipamide excreted in the bile and urine and that remaining in the organs analyzed accounted for only a fraction of the dose administered; no significant accumulation of iodipamide was found in the liver.

Saturation kinetics of iopanoate in the dog.

Experiments were carried out in dogs with a modified Thomas cannula in the duodenum through which the common bile duct could be catheterized. Constant intravenous infusion of sodium iopanoate at different infusion rates greater than the apparent excretion maximum revealed linearity of the blood concentration with time above a threshold concentration. When the slopes of the blood curves were plotted against the known infusion rates, a straight line relationship was obtained. The X axis intercept of this straight line represents an apparent transport maximum. This value obtained from the X axis intercept matched closely with the observed excretion maximum determined from bile and urine collection. The slope of this same line equals the inverse of the volume of distribution of the drug. Although previous workers have failed to appreciate an apparent transport maximum for iopanoate, the current studies clearly demonstrate that iopanoate is excreted by a saturable mechanism. Using this technique the apparent transport maximum for iopanoate was evaluated at high and low rates of taurocholate replacement to evaluate the quantitative effect of bile salt on the apparent transport maximum. A five-fold increase in taurocholate replacement led to an average 40% increase in the apparent transport maximum of iopanoate without effecting its volume of distribution significantly.

Gastrointestinal radiology. Pharmacokinetics of iopanoate and iodoxamate in rhesus monkeys.

The pharmacokinetics, biliary excretion, plasma protein binding, enterohepatic circulation, and biotransformation of iopanoic acid and iodoxamic acid in the rhesus monkey were evaluated by a dynamic infusion method. The dynamic method has the advantage that the pharmacokinetic parameters involved in the hepatic uptake and biliary excretion can be evaluated from a single infusion experiment. The percentage of iodoxamic acid not bound to plasma protein varied from 6.1-41.2% as iodoxamic acid plasma levels were from 42 microM to 912 microM. Using the Freundlich isotherm approach, more than one class of binding site for iodoxamic acid was found. A saturable biliary excretion mechanism or hepatic uptake mechanism was determined with a Vmax of 1.03 microM/kg/min. Less than 1% of iodoxamic acid injected into the duodenum was recovered in the bile in 12 hours. Iodoxamic acid was found to exist in blood as an unchanged species. Iopanoic acid was extremely highly bound to monkey plasma protein. As blood concentration increased from 18.9 to 464 microM, the percentage unbound in plasma protein varied from 0.1-2.8%. Biliary excretion rates of iopanoic acid were fitted by a computer to the Michaelis-Menten equation against unbound plasma concentration and the average Vmax value was found to be 0.85 microM/kg/min with an average Kmax value of 0.253. Iopanoic acid was found to exist in monkey blood as unchanged species and in the bile mainly as an ester glucuronide. Coadministration experiments revealed that the interaction of iodoxamic acid and iopanoic acid in the monkey is complex. The compounds appear to compete for plasma protein binding sites as well as for binding sites on intrahepatic protein. The biliary excretion data seem to fit the ligant exclusion model, in which iopanoic acid acts as an inhibitor and competes with iodoxamic acid for binding to either of two identical sites in the liver, which, presumably, is the rate-limiting step in the liver's overall elimination of these radiographic agents.

Pharmacokinetics of iopanoic acid in the rhesus monkey: biliary excretion, plasma protein binding and biotransformation.

A dynamic infusion method, originally developed for the pharmacokinetic studies of Iodoxamic acid, was applied to the kinetic studies of the biliary excretion of another cholecystographic agent, iopanoic acid. This dynamic method has an important advantage in that the pharmacokinetic parameters involved in the hepatic uptake or biliary excretion can be evaluated from a single infusion experiment. Using the equilibrium dialysis technique, iopanoic acid was found to be highly bound to the plasma proteins. A linear relationship was found when the logarithm of unbound plasma concentration of iopanoic acid was plotted vs. the logarithm of its blood concentration. When the biliary excretion rates of iopanoic acid were fitted by a computer to the Michaelis-Menten equation against its unbound plasma concentration, the average Vm value was found to be 0.85 micron/kg/min and the average Km value was found to be 0.253 micron. Iopanoic acid was found to exist in monkey blood as unchanged species and in the bile mainly as the ester glucuronide.

Topically applied griseofulvin in prevention and treatment of Trichophyton mentagrophytes.

Griseofulvin is not considered effective in treatment of superficial fungal infections. However, when applied topically in organic solvents, the drug appeared in very high concentration in all levels of stratum corneum, and it generally persisted there in measurable amounts for four or more days after a single application. Based on this observation, double-blind experimental prophylactic and therapeutic studies were conducted. Topically applied griseofulvin in alcohol solution was highly effective in preventing experimentally induced Trichophyton mentagraphytes infection, but it had no therapeutic effect once the infection was initiated. Topically administered griseofulvin may prove useful in prevention of recurrences or as an adjunct to therapy in selected cases of superficial fungal infection.

Topically applied antifungal agents. Percutaneous penetration and prophylactic activity against trichophyton mentagrophytes infection.

Penetration and prophylactic activity against Trichophyton mentagrophytes infection of topically applied antifungal agents were studied in human volunteers. The concentration and persistence of drug in the stratum corneum was determined after different treatment schedules and from different vehicles. Measurable levels of griseofulvin, clotrimazole, miconazole nitrate, and thiabendazole remained in the skin four days after discontinuation of topical applications. Griseofulvin, miconazole, and clotrimazole exhibited prophylactic activity against infection by T mentagrophytes spores in a standardized test. Griseofulvin was a significantly better prophylactic agent than either miconazole or clotrimazole.

Determination of quinidine and metabolites in urine by reverse-phase high-pressure liquid chromatography.

A new reverse-phase high-pressure liquid chromatography assay allowing simultaneous but separate quantitation of urinary levels of quinidine and its major metabolites, 2'-quinidinone, 3-OH-quinidine and a newly detected N-oxide, is described. The compounds were separated on a alkyl phenyl column using 0.05 M phosphate buffer pH 4.5/acetonitrile/tetrahydrofuran (80 : 15 : 5, v/v) as mobile phase and were detected by UV at lambda = 230 nm. The assay procedure includes extraction of the compounds from urine samples into a mixture of dichloromethane/isopropanol (4 : 1, v/v), evaporation of the organic extracts to dryness and reconstitution of the residue in acetonitrile. The new assay was compared to a modification of the Cramer and Isaksson fluorescence assay which has recently been recommended for analysis of quinidine in urine. The consistently higher quinidine levels observed in the fluorescence assay could be accounted for by the quinidine levels and metabolite carry-over as determined by HPLC.

GLC analysis of acetazolamide in blood, plasma, and saliva following oral administration to normal subjects.

An electron-capture GLC assay for acetazolamide in biological fluids was developed. Extraction efficiency was 69-104%. The minimum detectable amount of acetazolamide was 10 ng/sample. Concentrations of acetazolamide were determined by GLC in blood, plasma, plasma water, and saliva after oral administration of a single 250-mg dose to five volunteers. Erythrocyte levels were calculated from whole blood and plasma data. Concentration of free drug in the plasma was measured in samples of plasma water obtained by microultrafiltration. Peak plasma levels of 10-18 microng/ml were reached 1-3 hr after the dose. At least 1 hr later, erythrocyte levels reached peak concentrations of 13-29 microng/ml. Over 31 hr, plasma levels declined more rapidly than erythrocyte levels. Saliva concentrations averaged 1% of those in plasma and decreased at a rate equal to that of plasma. Saliva levels were proportional to, but not equal to, water concentration. Saliva to plasma ratios were consistent for any given individual and, therefore, offer a means of monitoring drug dosage without resorting to frequent blood sampling.

Iodipamide kinetics: capacity-limited biliary excretion with simultaneous pseudo-first-order renal excretion.

Iodipamide was infused into three dogs with bile fistulas to achieve various steady-state blood levels. When using ultracentrifugation techniques, iodipamide was found to be highly bound to plasma protein. The total blood clearance was low relative to hepatic blood flow. For either the whole blood concentration or the unbound concentration of iodipamide, the biliary excretion was shown to be capacity limited with a transport maximum, Tm, of approximately 1.0mumole/kg/min. The steady-state renal excretion rate, plotted against the whole blood concentration of iodipamide, resulted in a concave ascending curve, which could lead to the false conclusion that iodipamide was undergoing active renal tubular reabsorption. However, when corrected for plasma protein binding, a linear relationship was obtained, suggesting that the renal excretion of iodipamide is a pseudo-first-order process. The Michaelis-Menten parameters for the extrarenal elimination, when calculated using the whole blood concentration of iodipamide, led to a similar discrepancy compared to the parameter estimates obtained from biliary excretion rate data. This discrepancy can be eliminated when one uses the unbound concentration of iodipamide in the parameter estimates.

Kinetics of drug-drug interactions: biliary excretion of iodoxamic acid and iopanoic acid in rhesus monkeys.

The dynamic method originally developed for studying the capacity-limited kinetics of the cholecystographic agents iodoxamic acid and iopanoic acid was applied to study the in vivo interactions of these two compounds following coadminstration in the monkey. Results indicate that these interactions are complex. The compounds appear to compete for plasma protein binding sites as well as for binding sites on intrahepatic proteins. The biliary excretion data apparently fit the "ligand exclusion" model in which iopanoic acid acts as an inhibitor and competes with iodoxamic acid for binding to either of two identical sites within the liver. This competition probably is the rate-limiting step in the liver's overall elimination of these radiographic contrast agents.

Stereoselective disposition and glucuronidation of propranolol in humans.

Following oral dosing to steady state, the disposition of S(-)- and R(+)-propranolol and their corresponding glucuronide conjugates was studied in 4 healthy adults using doses from 40 to 320 mg/day of the racemate. Steady -state plasma concentrations of S(-)-propranolol and its corresponding glucuronide conjugate were greater than that for R(+)-propranolol and its corresponding conjugate. The average steady-state concentration of both enantiomers increased disproportionately to dose. There was a 52+/- 7 (mean +/- SD) % decrease in the intrinsic clearance (clint) of S(-)-propranolol and a 65 +/- 22% decrease in the Clint of R(+)-propranolol over the dosing range studied. The terminal elimination half-lives of S(-)-propranolol and its glucuronide conjugate were longer than for the R(+)-enantiomer at all doses. The formation of glucuzonide conjugates of S(-)- and R(+)-propranolol was best described by a saturable process in all subjects. Within individuals, the ratio of Vmax/Km for the glucuronide conjugate of S(-)-propranolol was from 2.1-to 4.9-fold greater than for the conjugate of the R(+)-enantiomer. These studies demonstrate for the first time, that propranolol undergoes stereoselective disposition in humans.

Determination of picogram nitroglycerin plasma concentrations using capillary gas chromatography with on-column injection.

A specific, sensitive, and precise capillary gas chromatographic (GC) assay capable of analyzing picogram concentrations of nitroglycerin in human plasma was developed. The analytical procedure involves a double extraction of 1 mL of plasma with pentane, after the addition of internal standard (1 ng of 2,6-dinitrotoluene), followed by evaporation and reconstitution in 50 microL of heptane. The extract (1 microL) was injected onto a capillary column using the on-column injection technique. The GC oven temperature was programmed from 120 degrees C to 180 degrees C at a rate of 5 degrees C/min. The oven temperature was then programmed to 250 degrees C and was maintained for 10 min. The nitroglycerin and internal standard retention times were 8.6 and 11.4 min, respectively. The position of the end of the capillary column inside the detector is a critical determinant of sensitivity: the column exit must be positioned such that nitroglycerin adsorption to the detector is minimized (i.e., sensitivity maximized). The assay limit of quantitation was 25 pg/mL (CV = 7.6%) using 1 mL of plasma. This GC assay, specific for nitroglycerin in the presence of its metabolites, isosorbide dinitrate, and several other drugs, may be used to quantitate plasma levels obtained after therapeutic nitroglycerin doses.

Pharmacokinetics of iodoxamic acid in rhesus monkey: biliary excretion, plasma protein binding, and enterophepatic circulation.

The previously reported steady-state method allowed estimation of the capacity-limited pharmacokinetics of the cholangiographic agent, iodipamide. To circumvent the long time period required to establish each steady-state level, a dynamic method was applied to the study of the rate processes involved in the hepatic uptake and biliary excretion of a new cholangiographic agent, iodoxamic acid, in rhesus monkeys. The dynamic method has the advantage that the pharmacokinetic parameters involved in capacity-limited hepatic uptake or biliary excretion can be obtained from a single infusion experiment. The V max was 1.03 +/- 0.25 mumoles/kg/min (mean +/- SD); Km varied from animal to animal and ranged from 1.5 to 16.4 micrometer. Protein binding was estimated using equilibrium dialysis. The Freundlich isotherm yielded a linear plot when the natural logarithm of unbound iodoxamic acid concentration in plasma was plotted against the natural logarithm of its blood concentration. The plasma protein binding data also could be fitted to the Langmuir isotherm, presuming two independent classes of binding.

Dependence of renal clearance on urine flow: a mathematical model and its application.

A mathematical model is developed to explain the dependence of renal clearance on urine flow rate. The model is tested using human data from the literature on compounds that are neither secreted nor reabsorbed by active or pH-sensitive mechanisms. The physiologically derived model explains and predicts the relationship between renal clearance and urine flow for a broad spectrum of compounds (i.e., butabarbital, chloramphenicol, creatinine, ethanol, theophylline, and urea) for which appropriate data are available.

Negligible excretion of unchanged ketoprofen, naproxen, and probenecid in urine.

On the average, 0.6% of a dose of ketoprofen or naproxen or 1.2% of a dose of probenecid was found in the urine of normal male volunteers assayed immediately after its collection. Between approximately 60 and 85% of the dose of these drugs can be excreted in the urine as conjugates, which rapidly hydrolyze at body temperature, at room temperature, and even during frozen storage, thereby regenerating the parent drug. Since urine collections involved sample retention in the bladder at 37 degrees for collection intervals as long as 2--3 hr, the given percentages excreted unchanged probably are overestimates. It is possible that no unchanged ketoprofen, naproxen, or probenecid is excreted in urine. This study contrasts with previous reports of up to 50% of a dose of ketoprofen and 15--17% of doses of naproxen and probenecid being excreted in urine as the parent compound. Those reports probably reflect primarily the duration of frozen sample storage between collection and assay along with the urine collection schedules employed the speed of the clinical procedures, and the analytical procedures used. Attention should be given to potential conjugate hydrolysis whenever the pharmacokinetics of carboxylic acids are studied.

An automated HPLC assay for simultaneous quantitation of methylated xanthines and uric acids in urine.

To investigate the elimination kinetics of caffeine and its metabolites, as well as the interaction between them, an automated HPLC method is described. This method involves a single extraction procedure, followed by a gradient elution. Fourteen methylated xanthines and uric acids are well separated with an assay sensitivity of 1 microgram/ml when one-half ml of urine is used. The assay is highly selective, from endogenous compounds, and reproducible. This method is recommended for accurate pharmacokinetic studies.

An automated HPLC method for the assay of propranolol and its basic metabolites in plasma and urine.

An automated HPLC method is described for the simultaneous determination of propranolol, 4-hydroxypropranolol, and N-desisopropylpropranolol in plasma and urine before and after beta-glucuronidase/aryl sulfatase treatment. It involves extraction with ether at pH 10 in the presence of ascorbic acid, added to prevent oxidation of 4-hydroxypropranolol. The compounds are then back extracted into dilute acid and assayed on an HPLC using a fluorescence detector. Three HPLC columns have been used (a phenyl, an octyl, and an octadecyl column). The last column was found to be most reproducible with minimal intercolumn variation. The solvent system includes a combination of acetonitrile, methanol, and phosphoric acid. Concentrations as low as 0.2, 1.0, and 0.2 ng/ml of propranolol, 4-hydroxypropranolol, and N-desisopropylpropranolol, respectively, can be measured using 1 ml of plasma.