GC/MS analysis of m-hydroxybenzoylecgonine in urine. Forensic implication in cocaine use.

m-Hydroxybenzoylecgonine (m-OH-BE) and d3-m- hydroxybenzoylecgonine (d3-m-OH-BE) have been synthesized, and a GC/MS procedure with d3-m-OH-BE as internal standard has been developed. Among 24 human urine specimens that were positive for BE, all of them have shown detectable levels of m-OH-BE with 75% of the specimens exceeding the LoQ (5 ng/mL), compared with only 50% of the specimens containing detectable levels of EME. The presence of m-OH-BE in urine suggested that this metabolite may serve as a valuable marker of cocaine use in addition to BE and EME.

Military drug positive rates in the European Theater drug rates in Europe.

Urine samples were collected from Air Force and Army service members within the European Theater and analyzed for drugs of abuse employing radioimmunoassay and gas chromatography/mass spectroscopy (GC/MS). Data collected from January 1985 through December 1991 indicate that the total positive rate decreased from 4.67% to 0.69%. Of the drugs tested, tetrahydrocannabinol (THC) was the drug abused most in the European Theater during this time period.

HI-6 pharmacokinetics in rabbits after intravenous and intramuscular administration.

The pharmacokinetics of HI-6 ((4-carboxamidopyridinium (1) methyl)-(2'-hydroxyiminomethyl-pyridinium (1') methyl) ether dichloride) have been studied in rabbits receiving an intramuscular (50 micrograms kg-1) or intravenous (12.5 micrograms kg-1) dose. The plasma concentration-time profile for the intramuscular dose (n = 8) fits a one-compartment open model with first-order absorption and elimination. The absorption half-life was 2 min and maximum concentration (51 micrograms mL-1) was reached in 9 min. The pharmacokinetics for the intravenous dose (n = 8) was described by a two-compartment open model with first-order distribution and elimination. The apparent volume of distribution was 0.1 L kg-1. Half-lives of distribution and elimination were 5 and 38 min, respectively. The results indicate HI-6 is rapidly absorbed, distributed and eliminated in rabbits receiving an intramuscular dose.

The pharmacokinetics of atropine and diazepam in sheep: intramuscular co-administration.

The purpose of this study was to determine whether the co-administration of atropine and diazepam affect the rate and extent of absorption of either drug. A triple crossover pharmacokinetic study using adult sheep was conducted. Each of nine animals received single injections of atropine (2 mg), diazepam (10 mg), and a combination of the two compounds weekly over a 3-week period. The combination of the drugs was injected into a single intramuscular site through a specially designed tandem syringe. Blood samples were obtained from time 0 to 300 min post-injection. Serum samples were analyzed for atropine by radioimmunoassay and for diazepam by gas chromatography/mass spectrometry. Pharmacokinetic parameters were evaluated by non-compartmental analysis. The co-administration of atropine and diazepam intramuscularly in sheep caused a delay in the time to reach maximal concentration of atropine. However, at the time when a single injection of atropine reached its maximum serum concentration, 92 per cent of that concentration was reached by atropine in the presence of diazepam. Additionally, no difference was detected in the rate or extent of diazepam absorption when administered intramuscularly in combination with atropine at the same site.

Pharmacokinetics of diazepam intramuscularly administered to rhesus monkeys.

The purpose of this study was to define the pharmacokinetics of diazepam in monkeys following an im injection of 100 micrograms/kg (the minimum effective dose that prevents nerve agent-induced convulsions in pyridostigmine-pretreated, atropine- and 2-PAM-treated monkeys) in order to predict what im dose in humans is needed to prevent nerve agent-induced convulsions. Six rhesus monkeys were administered diazepam in the hind limb. Blood (3 mL) was collected via an indwelling saphenous catheter immediately prior to and 5, 10, 15, 25, 40, 60, 90, 120, 180, and 240 min after diazepam dosing. A contract laboratory, blind to the labeling code, analyzed diazepam serum concentrations by electron-capture gas chromatography and the percentage of unbound diazepam by equilibrium dialysis. The concentration-time data for total (unbound and bound) diazepam individually determined for each animal was best described by a one-compartment open model with first-order absorption and elimination. The average maximum serum concentration (50 ng/mL) was reached in 29 min. The volume of distribution and systemic clearance, assuming 100% bioavailability, were 1.5 L/kg and 19.4 mL/min/kg, respectively. The percentage of diazepam unbound to serum proteins was 4.6% and, therefore, the maximum concentration of free diazepam was 2.3 ng/mL. These results, when compared with human pharmacokinetic studies, allow a means of extrapolating effective monkey anticonvulsant doses to humans on a pharmacokinetic basis.

Pharmacokinetics of physostigmine intramuscularly administered to guinea pigs.

Physostigmine pharmacokinetics was determined in guinea pigs following im administration of 5-146 micrograms/kg. Eighteen male guinea pigs were divided into three equal groups and given dosages of 5, 27, and 146 micrograms/kg, respectively. Physostigmine was given in the right hind limb and blood samples were collected at various times up to 300 min postinjection via an indwelling carotid catheter. Unbound physostigmine plasma concentrations were analyzed by HPLC. The concentration-time profile for each animal was fitted to standard pharmacokinetic models. A one-compartment open model with first-order absorption and elimination provided the best fit. For all dosage groups, physostigmine concentrations peaked in approximately 30 min. Apparent volumes of distribution (assuming 100% bioavailability) ranged from 1.9 to 2.2 L/kg. Systemic clearances and elimination half-lives were 30-36 mL/min/kg and 40-50 min, respectively. The area under the concentration-time curve and the Cmax were linearly related to the dose, indicating pharmacokinetic linearity. In conclusion, physostigmine, intramuscularly administered to the guinea pig, is absorbed, distributed, and eliminated rapidly, and the pharmacokinetics behave linearly within the 5-146-micrograms/kg dosage range.

A high-performance liquid chromatographic assay for HI-6 oxime in plasma.

A high-performance liquid chromatographic (HPLC) assay was developed to determine HI-6 (1-(2-hydroxyiminomethyl-1-pyridinio-3-(4-carbamoyl-1-py ridiniol-2-oxapropane dichloride)) concentrations in small volumes of plasma. A 100-microL plasma sample added to 900 microL of distilled water was microfiltered. Filtrate (200 microL) was injected onto an HPLC instrument containing a 100-microL sample loop, a C18 column, and an ultraviolet (UV) wavelength detector. Limit of sensitivity for HI-6 was 2.5 micrograms/mL. Extraction of efficiency (n = 12) at 10 and 100 micrograms HI-6/mL plasma was 69.4 +/- 6.6% (SD) and 81.5 +/- 2.0% (SD), respectively. Protein-plasma binding of HI-6 did not occur. HI-6 was stable when frozen at -20 degrees C for up to 10 days (0.025 less than p less than 0.05). Correlation coefficients representing standard curve linearity ranged from 0.9986 to 0.9999 (n = 6). Within-day and between-day coefficients of variation (n = 6) for unknown samples ranged from 4.4 to 8.3% and 5.8 to 17.1%, respectively. Bias of unknown samples ranged from -10.5 to 5.7%. The method's sensitivity, accuracy, and precision indicate that it can be used to accurately measure HI-6 concentrations in 100 microL of plasma.

Intramuscular administration of atropine in the rat: jet spray versus conventional needle injection.

The characteristics of atropine plasma levels after jet spray injection were compared to those after conventional needle injection (i.m.) in 12 male rats, six per group. Blood samples were sequentially collected from the tip of the tail over a 7h period. Injection of atropine sulfate (8.0 mg kg-1) using the jet spray resulted in mean peak plasma levels of 650 ng ml-1 (95 per cent C.I. = 90) compared to 488 ng mg-1 (95 per cent C.I. = 64) using a conventional needle. Times to reach maximum concentration were 30 min (95 per cent C.I. = 12) and 58 min (95 per cent C.I. = 6) for the jet spray and needle, respectively. Histopathologic examination (5 days post-injection) of target muscle showed that minimal fiber damage resulted from using the low pressure setting on the jet spray. The results suggest that the jet spray may offer a means of increasing the antidotal benefit over that achieved with conventional techniques using presently available therapeutic formulations for acetylcholinesterase poisoning.

Development and application of a radioimmunoassay for physostigmine.

Antiphysostigmine antibodies were produced in rabbits using a physostigmine analog, 1,3-dimethyl-3-[2- [N-methyl-N-(7-carboxyheptanoyl)] aminoethyl]-5-(N-methyl-carbamoyloxy)-2,3-dihydroindole hydrochloride, conjugated to keyhole limpet hemocyanin. These antibodies were used to develop a radioimmunoassay ranging from 0.2 to 15.0 ng/ml of physostigmine in a 0.1-ml plasma sample. A typical standard curve gave an r2 value of 0.992. This assay measures physostigmine in plasma with better sensitivity and much greater through-put than do current state-of-the-art, high-performance liquid chromatography techniques. In addition, only small volumes (100 microliters) of the plasma samples are required. Precision represented by within and among day coefficients of variance was less than 20% for 1.0 to 50.0 ng/ml and less than 22% for 0.2 ng/ml. Accuracy for the 1.0 to 50.0 ng/ml range varied less than 15% and was 22% for 0.2 ng/ml. Plasma levels of physostigmine were determined in the rat after i.m. administration of physostigmine salicylate to give a free base equivalency of 27 micrograms/kg. Estimates of the various pharmacokinetic parameters were calculated using the computer program PCNONLIN. The results were as follows: apparent volume of distribution = 5.9 liters/kg, absorption rate half-life = 2.7 min. elimination rate half-life = 17.4 min, area under the curve = 118 ng x min/ml, maximal plasma concentration = 3.5 ng/ml and time to maximal plasma concentration = 7.7 min.

Application of a new radiometric high-performance liquid chromatographic assay to define physostigmine pharmacokinetics in guinea pigs.

A sensitive high-performance liquid chromatographic method was developed to determine pharmacokinetic parameters of [3H]physostigmine from serial plasma samples from guinea pigs. Physostigmine was totally resolved from its metabolite, eseroline. The limit of sensitivity was 0.05 ng/ml from 0.2 ml plasma. Extraction efficiency was 99.6%. Within-run and among-run coefficients of variation (n = 6) for 0.2, 0.75, 1.5 and 2.5 ng/ml [3H]physostigmine ranged from 0.7 to 20% and 16 to 32%, respectively. Physostigmine (5 micrograms/kg) intramuscularly administered to the guinea pig (n = 6) reached maximum serum concentration (1.5 ng/ml) in 26 min. The apparent volume of distribution and systemic clearance were 1.4 l/kg and 26 ml/min/kg, respectively. This method was successful in defining physostigmine pharmacokinetic parameters in guinea pigs and can be employed for other small animal pharmacokinetic studies.

Relationships among nortriptyline, 10-OH(E)nortriptyline, and 10-OH(Z)nortriptyline steady-state plasma levels and nortriptyline dosage.

The postulated therapeutic activity of nortriptyline metabolites has prompted investigation of dosage adjustments based on plasma levels of nortriptyline (NT) and its metabolites. The method assumes that metabolite concentrations vary independently of nortriptyline concentrations among patients. This study tests that assumption and investigates different means of obtaining metabolite concentrations. Forty-two psychiatric inpatients were divided into three maintenance dose groups: 50, 100, and 150 mg NT/day. After a 28-day study for each inpatient, steady-state plasma concentrations for days 14, 18, 21, 25, and 28 were determined. Concentrations were averaged for each patient. Nortriptyline concentrations did not correlate well with corresponding 10-OH(E)nortriptyline (p greater than 0.05) or 10-OH(Z)nortriptyline concentrations (r2 = 0.31, p less than 0.05). Concentrations of 10-OH(E)nortriptyline did not correlate well with corresponding 10-OH(Z)nortriptyline concentrations (r2 = 0.236, p less than 0.05). Neither dose/body weight nor obesity were good predictors of individual concentrations of nortriptyline, 10-OH(E)nortriptyline, or 10-OH(Z)nortriptyline or even the sum of the three. In conclusion, optimal drug therapy may involve dosage adjustments according to the combined plasma levels of nortriptyline and metabolites. Assurance of obtaining certain plasma concentrations requires plasma level monitoring.

A radioimmunoassay for pyridostigmine.

Antipyridostigmine antibodies were produced in rabbits using a pyridostigmine analog conjugated to keyhole limpet hemocyanin. These antibodies were used for development of a radioimmunoassay that has a linear standard curve (r2 = 0.986) ranging from 0.5 to 10.0 ng/ml of pyridostigmine bromide in a 0.1-ml plasma sample. This assay measures pyridostigmine in plasma with better sensitivity and much greater through-put than do current state-of-the-art high-performance liquid chromatography techniques. In addition, only small volumes (100 microliter) of the plasma samples are required. Plasma levels of pyridostigmine were determined in the rat after intramuscular administration (0.056 mg/kg) of pyridostigmine bromide. Estimates of the various pharmacokinetic parameters were calculated using the computer program NONLIN84. The results were as follows: apparent volume of distribution = 1.97 l/kg, absorption rate constant = 0.277 min-1, elimination rate constant = 0.0273 min-1, area under the curve = 1010 ng x min/ml, absorption rate half-life = 2.41 min, elimination rate half-life = 24.8 min, maximal plasma concentration (Cmax) = 21.3 ng/ml and time to Cmax = 9.02 min.

Quantification of amitriptyline, nortriptyline, and 10-hydroxy metabolite isomers in plasma by capillary gas chromatography with nitrogen-sensitive detection.

A selective, sensitive method for the determination of amitriptyline and its metabolites is described. This method involves liquid-liquid extraction and capillary gas chromatography with nitrogen-sensitive detection. The detection limits of amitriptyline, nortriptyline, 10-hydroxy(E)amitriptyline, 10-hydroxy(E)nortriptyline, and 10-hydroxy(Z)nortriptyline were slightly less than 0.5 ng/ml in 1.0-ml plasma samples. The coefficients of variation for within-run and between-run analyses of samples containing 100 ng/ml were less than 12% and 9%, respectively. The method offers rapid analysis of individual isomers, increased sensitivity over high-performance liquid chromatographic methodology and the conveniences of the gas chromatographic technique.