Absorption of triazolam from pelleted drug-diet mixtures by the mouse: quantitation of alpha-hydroxytriazolam in urine.

The absorption of triazolam from pelleted drug-diet mixtures by mice under steady-state conditions was determined for doses up to 150 mg/kg/day by measuring alpha-hydroxytriazolam, the principal urinary metabolite of triazolam in the mouse, in urine samples collected over a 24-hour period. Following beta-glucuronide glucuronosohydrolase hydrolysis of the urine, quantitation of alpha-hydroxytriazolam was accomplished using a specific reverse-phase liquid chromatographic method which utilized UV detection at 214 nm. Assay precision was greater than 2.7% (CV) over the concentration range of interest. Statistical analysis of the excretion data indicated that the mathematical relationship between the triazolam dose and the quantity of alpha-hydroxytriazolam excreted was linear for female mice and nonlinear for male mice. Triazolam absorption, as reflected by alpha-hydroxytriazolam urinary excretion data, increased with triazolam dose.

Fast-LC determination of 1-methyl-1H-tetrazole-5-thiol in human plasma with column-switching sample clean-up.

A simple, fast and sensitive method for the quantitative determination of 1-methyl-1H-tetrazole-5-thiol using HPLC is reported. Samples are deproteinated by plasma water filtration and injected into a HPLC system capable of column switching and backflushing the analytical column. The limit of quantitation was determined to be 70 ng ml-1 and the limit of detection 22 ng ml-1. Between-day precision (expressed as percent coefficient of variation) of standard curve slopes was +/- 3.5% with a range of within-day percent coefficient of variations from 0.28 to 1.4%. Recovery and precision of spiked control samples was 96 +/- 7% over a concentration range of 630-6300 ng ml-1.

Is GABA involved in analgesia?

The effect of morphine and naloxone on gamma-aminobutyric acid (GABA) concentration in discrete areas of the rat brain has been studied. Neither morphine nor naloxone had a significant effect on regional steady-state concentrations of GABA. The results have been discussed with respect to the role of GABA in pain and analgesia.

Protein binding of tirilazad (U-74006) in human, Sprague-Dawley rat, beagle dog and cynomolgus monkey serum.

Determination of the serum protein binding of tirilazad across species was required to predict the pharmacokinetic behavior of this new drug. Equilibrium dialysis and ultrafiltration techniques, commonly used to study serum protein binding, were shown to be unsuitable for tirilazad due to high nonspecific binding and low aqueous solubility, resulting in unbound drug levels that were nondetectable with current analytical methodology. Ultracentrifugation appeared to offer a technique with which unbound tirilazad could be measured; however, after extensive studies, the apparent lipid partitioning behavior of tirilazad into low density and very low density lipoproteins showed that ultracentrifugation was also unsuitable for determination of the true unbound fraction of tirilazad. Fractions of tirilazad measured in the supernatant were highly correlated with total triglycerides and very low density lipoproteins in the sera of all species analyzed. Studies with delipidized human serum yielded a nonsaturable binding isotherm with free fractions of less than 0.6 +/- 0.02% (mean +/- S.D.) over a concentration range of 4.6 to 55.6 micrograms/ml (normal human in vivo range, 0.01-20 micrograms/ml). These data indicated that, as the triglyceride content of the sera increases, portions of tirilazad bound to serum proteins shift into the lipid phase of lipoproteins. What effect this has on the true unbound fraction is unknown and does not seem to be ascertainable with current technology.

Quantitative determination of itazigrel in rodent diet by reversed-phase HPLC and evaluation of its stability at 20 degrees C.

A simple, accurate and precise procedure for the quantitation of itazigrel (a potent lipophilic inhibitor of collagen and arachidonic acid-induced aggregation being studied for its effects on peripheral vascular disease) from granulated rodent diet is presented. The drug was extracted from rodent diet using methanol + water (80:20) following dissolution of the diet in water. Samples of the supernatant were injected into the HPLC and the eluent was monitored with a fluorescent detector (lambda ex = 320 and lambda em = 430) to achieve analytical specificity. Interday coefficients of variation of the calibration curve slope were +/- 6% on standards between 0 and 1000 micrograms/g. Potency and homogeneity of the drug spiked diet prepared over a 1 year interval at 70, 200 and 600 micrograms/g was 99.3 +/- 2.5%, 100 +/- 1.8%, and 101 +/- 1.9% of label, respectively. Samples prepared for chromatography were stable for 24 h at 20 degrees C, and drug in diet was stable for 102 days when protected from light and stored at 20 degrees C.

Effect of ranitidine on the disposition of orally and intravenously administered triazolam.

The effect of orally administered ranitidine on the pharmacokinetic properties of orally and intravenously administered triazolam was determined. Twelve healthy males with a mean age of 35 years were enrolled in this four-way, randomized, crossover study. Each subject received a total of four treatments, each separated by one week. The treatments consisted of (1) one orally administered 0.25-mg triazolam tablet after treatment with ranitidine; (2) one orally administered 0.25-mg triazolam tablet, with no ranitidine pretreatment; (3) a 0.25-mg intravenous dose of triazolam after treatment with ranitidine; and (4) a 0.25-mg intravenous dose of triazolam, with no ranitidine pretreatment. Ranitidine pretreatment consisted of five 150-mg oral doses (as the hydrochloride salt) given every 12 hours; the last dose was given two hours before triazolam was administered. Blood samples were taken at intervals up to 12 hours after triazolam treatment. Serum triazolam concentrations were measured by modified high-performance liquid chromatography, and pharmacokinetic values were calculated. Pretreatment with ranitidine had no effect on the disposition of intravenously administered triazolam but significantly increased the area under the serum drug concentration-time curve of oral triazolam. Ranitidine pretreatment had no effect on triazolam's terminal elimination rate constant or on the time to reach maximum serum triazolam concentration. Ranitidine pretreatment increased the systemic availability of triazolam by increasing its absorption.

A non-radioactive iothalamate and p-aminohippuric acid high-performance liquid chromatographic method for simultaneously measuring glomerular filtration rate and renal blood flow in the rat.

A high-performance liquid chromatographic (HPLC) assay method has been developed for the quantitative determination of iothalamate and p-aminohippuric acid (PAH) concentrations in serum and urine samples in the male rat. Glomerular filtration rate (GFR) was measured as clearance of iothalamate, while effective renal blood flow (ERBF) was measured as clearance of PAH. The method is simple, rapid and sensitive and detects iothalamate and PAH in rat serum and urine following administration of bolus doses and continuous infusions of iothalamate and PAH. Samples of serum and urine were deproteinized with two volumes of acetonitrile containing the internal standard, and an aliquot chromatographed on a C18 reversed-phase column. The mobile phase was comprised of 0.1 M sodium phosphate with 1.2 mM tetrabutylammonium phosphate: methanol, 85:15 (v/v), at a flow rate of 1.0 mL/min. The analytical column eluate was monitored with a UV detector at 254 nm with quantitation achieved using peak-height ratios. The precision of the method was 6.6 and 3.6% for iothalamate in serum and urine, and 5.6 and 4.9% for PAH in serum and urine, respectively. The lower limit of quantitation was 0.63 microgram/mL for iothalamate and 1.25 microgram/mL for PAH in serum, and 3.1 microgram/mL for iothalamate and 1.5 microgram/mL for PAH in urine. Recovery of iothalamate from serum and urine was 99.9 and 93.5%, respectively. Recovery of PAH from serum and urine was 99.8 and 92.6%, respectively. The present study demonstrated that non-radioactive iothalamate and PAH can be measured simultaneously using a HPLC assay to measure GFR and ERBF in the male rat.

Pharmacokinetics, metabolism, and excretion of irinotecan (CPT-11) following I.V. infusion of [(14)C]CPT-11 in cancer patients.

This study determined the disposition of irinotecan hydrochloride trihydrate (CPT-11) after i.v. infusion of 125 mg/m(2) (100 microCi) [(14)C]CPT-11 in eight patients with solid tumors. Mean +/- S.D. recovery of radioactivity in urine and feces was 95.8 +/- 2.7% (range 92.2-100.3%, n = 7) of dose. Radioactivity in blood, plasma, urine, and feces was determined for at least 168 h after dosing. Fecal excretion accounted for 63.7 +/- 6.8 (range 54.2-74.9%, n = 7) of dose, whereas urinary excretion accounted for 32.1 +/- 6.9% (range 21.7-43.8%; n = 7) of dose. One patient with a biliary T-tube excreted 30.1% of dose in bile, 14.2% in feces, and 48.2% in urine. Quantitative radiometric HPLC revealed that CPT-11 was the major excretion product in urine, bile, and feces. Aminopentane carboxylic acid (APC) and SN-38 glucuronide (SN-38G) were the most significant metabolites in urine and bile, whereas SN-38 and NPC, a primary amine metabolite, were relatively minor excretion products. SN-38 and APC were the most significant metabolites in feces. The relatively higher amount of SN-38 in feces compared with bile is presumably due to hydrolysis of SN-38G to SN-38 by enteric bacterial beta-glucuronidases. There was close correspondence between quantitative fluorescence HPLC and mass balance findings. CPT-11 was the major circulating component in plasma (55% of the mean radiochemical area under the curve), and CPT-11, SN-38, SN-38G, and APC accounted for 93% of the mean radiochemical AUC. These results show that the parent drug and its three major metabolites account for virtually all CPT-11 disposition, with fecal excretion representing the major elimination pathway.