Urinary excretion of d-amphetamine following oral doses in humans: implications for urine drug testing.

Seven healthy male volunteers received a single oral dose of 5 mg, 10 mg, or 20 mg of d-amphetamine. Urine was collected at 2, 4, 8, 12, 18, and 24 h post-dose. Total urine volume was measured, and pH and creatinine were determined. All specimens were analyzed by TDx Amphetamine/Methamphetamine II (TDx), Emit-d.a.u. Monoclonal Amphetamine/Methamphetamine (EM), and Emit II Amphetamine/Methamphetamine (EII) immunoassays at a cutoff value of 1000-ng/mL amphetamine. Quantitation of urinary amphetamine in all specimens was performed by gas chromatography-mass spectrometry. All urine testing results by the three immunoassays, EM, EII, and TDx, were in agreement; there were no discordant findings. Of the 42 total urine specimens collected following a 5-mg dose of amphetamine, only 8 (19%) screened positive by immunoassay. Twenty-four of 36 (67%) urine specimens yielded positive responses following a 10-mg dose, and 37 of 42 (88%) were positive by immunoassay following a 20-mg dose. These data demonstrate the present guideline for regulated forensic urine drug testing (FUDT) for amphetamine with a screening cutoff of 1000 ng/mL is too high to consistently detect the administration of a single 5-mg oral dose of d-amphetamine. There was considerable overlap of amphetamine concentrations in individual specimens following the various doses. Peak urinary amphetamine ranged from 620 to 3160 ng/mL following 5-mg doses. The time to peak concentration also varied widely at each dose, occurring in urines collected 2 to 18 h post-administration. The mean percent of dose excreted as unchanged amphetamine over 24 h at each dose ranged from 35 to 44%. The data demonstrated that amphetamine excretion increases with increasing urine flow and decreasing urine pH. Thus, a positive FUDT result for amphetamine means only that the individual was administered or self-administered amphetamine at some time prior to collection of the specimen.

Comparison of methods for the assessment of central nervous system stimulant response after dextroamphetamine administration to healthy male volunteers.

The objective of this investigation was to evaluate a series of potential pharmacodynamic measures of central nervous system stimulation, including quantitative electroencephalography (EEG) and neuroendocrine, mood, and psychomotor performance measures. The reproducibility and sensitivity of the measures were compared. The study was conducted in two parts. The first part investigated the interindividual and intraindividual variability associated with a series of potential pharmacologic response measures under baseline (i.e., drug-free) conditions. It was an open-label, three-period pilot study in which healthy male volunteers underwent a series of tests (EEG, a visual continuous performance task, a finger tapping task, and self-rated mood scales) repeatedly during each study period. The second part evaluated the sensitivity of a series of potential response measures to detect the effects of dextroamphetamine, and was a double-blind, placebo-controlled, four-period crossover study in nine healthy male volunteers. Subjects received 5 mg, 10 mg, or 20 mg of dextroamphetamine or placebo orally and underwent the same series of tests as in Part I in addition to blood collection for determination of serum prolactin and dextroamphetamine levels. Peripheral response to dextroamphetamine was assessed by heart rate and blood pressure measurement. The greatest variability among days, within days, and among participants was associated with the quantitative electroencephalographic parameters studied. First-session effects were apparent for several of the tests, including EEG. Consistent response on EEG (increased alpha power) to dextroamphetamine was observed only in the three subjects who had a baseline alpha activity greater than 35%. Serum prolactin levels were inversely associated with the amount of dextroamphetamine administered, with the largest decrease in serum prolactin levels observed after the 5-mg dose, and this finding was statistically significant. Mood scales showed that three of nine participants experienced dysphoria after at least one dose level of dextroamphetamine. The effect on mood was generally greater as the dose increased. Doses could not be distinguished based on the results of the performance tests. Serum prolactin concentration was the most sensitive measure of central nervous system stimulation on EEG produced by dextroamphetamine under these study conditions. Cardiovascular measures were more sensitive measures of dextroamphetamine effects than the central nervous system measures.

Age-related changes in valproic acid binding to rat serum proteins in vitro.

The effect of age on the in vitro binding of valproic acid (VPA) to serum proteins was investigated in rats ranging in age from 14 days (preweaning) to 24 months (senescent). The influence of free fatty acid (FFA) and total protein (TP) concentrations on age-related changes in binding was examined. The protein binding of VPA was altered during development and aging. The VPA fraction unbound (fu) at low VPA concentrations was significantly higher in older age groups (12 and 24 months old; fu = 0.26-0.30) than in younger animals (14, 20, and 40 days old; fu = 0.16-0.18). Binding was best described by a model incorporating a saturable and a nonsaturable binding site. Binding affinity at the saturable binding site was lowest at the extremes of age. Changes in binding at either the saturable or the nonsaturable site were not predicted by changes in TP or FFA with age. Changes in nonsaturable binding were marginally associated with age (p = 0.0952). A 3-fold increase in FFA concentrations was necessary to produce a 1.5-fold increase in VPA fu. There was less than a 2-fold difference in FFA concentrations between the age groups (range 0.219-0.379 mmol/L). Thus, the difference in FFA concentrations between the age groups may not have been large enough to cause measurable differences in displacement of VPA from binding sites. Changes in protein binding may contribute to age-related changes in disposition of VPA observed in the rat. Changes in the serum concentrations of specific FFA or proteins may play a role in the altered VPA binding with age, but changes in total FFA or protein concentrations do not account for the age-related differences observed. Further investigation is required to identify the mechanism(s) responsible for age-related changes in binding of VPA to serum proteins measured in vitro.

Age-dependent intestinal hydrolysis of valproate glucuronide in rat.

1. Age-dependent differences in the intestinal hydrolysis of the glucuronide conjugate of valproic acid were evaluated in the Fisher-344 rat at 14 and 40 days, and 24 months of age. 2. Hydrolysis occurred more quickly when incubations were conducted under anaerobic as compared with aerobic conditions. 3. The rate of hydrolysis of valproate glucuronide was most rapid in the contents of the large intestine (caecum and colon); no difference in rate was noted between age groups during incubations with large intestinal contents. 4. Hydrolysis in the tissues of the large and small intestines, and the contents of the small intestine, was more rapid in the 14-day-old rat than in the older age groups. Differences in the rates and sites of hydrolysis in the 14-day-old animal may be due to regional differences in the number and types of microorganisms or mammalian beta-glucuronidase present in the gastrointestinal tract. 5. Differences in intestinal hydrolysis of valproate glucuronide may account in part for age-related changes in enterohepatic recirculation of valproate in young animals; other mechanisms apparently are responsible for altered valproate disposition in senescent animals.