The Use of Miscellaneous Prescription Medications to Facilitate Sexual Assault.

Drugs used to facilitate sexual assaults are typically those that rapidly render the potential victim unconscious or sedated, and produce memory loss or amnesia. Many of these drugs are difficult to detect due to a delay in biological specimen collection. Detection is further hampered as the drugs are often administered in single low doses and are rapidly and extensively metabolized, resulting in low concentrations in biological specimens. Miscellaneous prescription drugs such as the barbiturates, antipsychotics, opioids, tricyclic antidepressants, ketamine, and chloral hydrate have the potential to produce varying degrees of sedation; however, they are not frequently detected in drug-facilitated sexual assault cases. A review of the literature shows that these drugs are often knowingly taken by the victim before or subsequent to the assault, and therefore may contribute to the sedation or unconsciousness experienced by the victim when ethanol or other central nervous system drugs are co-administered. Most barbiturates, opioids, and tricyclic antidepressants are routinely screened for in hospitals and forensic toxicology laboratories, and may be detectable in a urine specimen for several days. Antipsychotics, particularly the atypical class, ketamine, and chloral hydrate, generally require more targeted analyses. This review provides an overview of the pharmacodynamics, pharmacokinetics, and common analytical methods for the barbiturates, antipsychotics, opioids, tricyclic antidepressants, ketamine, and chloral hydrate.

Recommendations for toxicological investigations of drug-facilitated sexual assaults.

The recent increase in reports of drug-facilitated sexual assaults has caused alarm in the general public and prompted forensic toxicologists from across North America to address the toxicological issues surrounding this matter. The authors have developed recommendations and guidelines to inform law enforcement, medical, and scientific personnel of the requirements for performing successful toxicological examinations in cases of drug-facilitated rape.

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.

Determination of gabapentin in serum using solid-phase extraction and gas-liquid chromatography.

A gas-liquid chromatographic method for the determination of gabapentin (Neurontin) is described. The method involves extracting 0.5 mL of acidified sample by C18 solid-phase column, derivatization with MTBSTFA plus 1% tBDMCS, and analysis on an HP-1 column with a flame-ionization detector. Quantitation was performed with peak-height ratios of gabapentin to a gabapentin analogue [(1-aminomethyl-1-cycloheptyl) acetic acid] as the internal standard. The assay had a limit of detection of 0.2 mg/L and a linear range from 0.5 to 30.0 mg/L. Several compounds were analyzed for potential interference, and none interfered with the assay.

Perinatal methadone exposure produces physical dependence and altered behavioral development in the rat.

Pregnant rats were implanted with osmotic minipumps containing either methadone hydrochloride (9 mg/kg/day) or sterile water. Their offspring were cross-fostered so that the following prenatal/postnatal exposure groups were obtained: water/water, methadone/water, water/methadone and methadone/methadone. Methadone slightly reduced litter size, particularly the number of male offspring, and reduced litter birth weight. The induction or maintenance of physical dependence in the postnatal methadone exposure groups was confirmed by an experiment in which PD19 pups were challenged with naloxone (1 mg/kg, s.c.). Methadone concentrations were assayed in pup brain on postnatal days 4, 10 and 22. Postnatal exposure to methadone via maternal milk produced measurable levels of methadone which decreased with age. Neuromuscular and physical development were assessed. Exposure to methadone accelerated acquisition of the righting reflex, but tended to delay the acquisition of the negative geotaxic response. Postnatal exposure to methadone was associated with decreased somatic growth as measured through postnatal day 21. The older pups (postnatal day 21) exposed to methadone exhibited variations in activity levels: pups exposed to methadone both prenatally and postnatally exhibited the least amount of spontaneous locomotor activity and pups exposed only postnatally exhibited the most activity. Therefore, it is possible to induce and/or maintain physical dependence via lactation in rat pups fostered to methadone-treated dams. Perinatal exposure to methadone by this route produces several subtle disruptions of pup development in the absence of gross maternal or fetal toxicity.

Cocaine, ecgonine methyl ester, and benzoylecgonine plasma profiles in rhesus monkeys.

From a public health point of view, cocaine (COC) presents serious clinical problems and deaths from overdose and lifelong addiction patterns, not to mention its involvement in crime, in the United States. This study subjected rhesus monkeys to one intravenous administration of COC (1 mg/kg), which closely imitates the smoking of "crack" COC with regard to dose and effect. We monitored plasma concentrations over time, beginning when the primates were in a state of hyperarousal. Blood was sampled at 1, 6, 12, and 40 min after dosing. Plasma concentrations of COC decreased rapidly with a half life of 15.7 min. Mean COC concentrations in the drug-treated group (n = 7) for the four timepoints were 296, 225, 187, and 80 ng/mL, respectively. Ecgonine methyl ester (EME) concentrations ranged from 57 to 91 ng/mL. When compared with the 1-min COC concentrations, the mean EME concentration was 30.7%. Benzoylecgonine (BZE) ranged from 34 to 42 ng/mL, and the mean concentration was 11.5% of the mean COC concentration at 1 min. EME and BZE concentrations did not vary appreciably over the time course of the study. Plasma norcocaine concentrations were less than the limit of detection of 25 ng/mL. Because a rapid decline in plasma COC concentrations over time was observed along with a very small change in EME and BZE concentrations, we attribute tissue redistribution of COC, particularly to the brain, as significant and metabolism or hydrolysis of COC as minor.

Chromatographic procedures for the determination of felbamate in serum.

A gas-liquid chromatographic procedure and a high-performance liquid chromatographic (HPLC) procedure for the quantitative determination of the anticonvulsant drug felbamate in serum are presented. Both methods employ methyl felbamate as the internal standard. The efficiencies of felbamate extraction by both C18 solid-phase extraction (SPE) followed by HPLC and liquid-liquid extraction followed by gas chromatography (GC) were evaluated. The mean absolute recovery of felbamate over a serum concentration range of 10-100 mg/L for SPE and liquid-liquid extraction were 89.4 and 84.4%, respectively. Calibration curves for the GC and HPLC procedures were linear from 5 to 200 mg/L felbamate. The overall within-run precision of liquid-liquid extraction followed by GC analysis yielded coefficients of variation Os) of 7.3% at 16 mg/L (n = 10) and 5.8% at 100 mg/L (n = 10). The overall between-run precision for this method calculated by three determinations on a single day for three successive weeks yielded CVs of 7.8% at 16 mg/L (n = 45) and 6.3% at 100 mg/L (n = IO). The overall within-run precision of SPE followed by HPLC analysis yielded a CV of 2.9 % at 50 mg/L (n = 10), and the overall between-run precision for this method calculated by three determinations on a single day for three successive weeks yielded CVs of 3.3% at 25 mg/L (n = 40) and 3.5% at 50 mg/L (n = 37). The SPE-HPLC procedure was found to yield less variable results than the liquid-liquid-GC procedure. Limits of quantitation and detection of both methods were set at 5 mg/L felbamate. Popular anticonvulsant and acid or neutral extractable drugs did not interfere with either the GC or HPLC methods. The results of an interlaboratory, intermethod comparison study performed on patient serum specimens analyzed in one laboratory by the SPE-HPLC procedure and in another laboratory by the liquid-liquid extraction-GC procedure correlated well (r2 = 0.967; n = 41). Both methods provide clinically useful and comparable results.

Direct determination of benzoylecgonine in serum by EMIT d.a.u. cocaine metabolite immunoassay.

An EMIT d.a.u. immunoassay for urine testing was applied on the Syva ETS Plus analyzer for the detection of the cocaine metabolite, benzoylecgonine (BE), in human serum. Serum was analyzed without prior extraction, concentration, or matrix modification. Calibrators and serum controls were prepared from EMIT d.a.u. calibrators that were reconstituted and diluted with EMIT Tox serum calibrator. The assay cutoff concentration for BE was 50 ng/mL. The within-run and between-run precisions of the assay were both less than 5%. Analysis of 162 patient serums yielded 43 BE positive results. All EMIT positive serum BE results were confirmed by gas chromatography-mass spectrometry. All patients with positive BE serums also had BE positive urine samples. Serum bilirubin and triglycerides as high as 38 mg/dL and 319 mg/dL, respectively, did not interfere with the assay. Modification of the EMIT urine assay allowed for a simple, rapid, and reliable method for the detection of BE in serum.

Cocaine-induced rausch: overt behaviour and plasma concentrations in rhesus monkeys.

This study was designed to characterize the cocaine-induced rausch or hyperarousal syndrome in rhesus monkeys. This syndrome mimics the stage observed in human abusers bingeing on cocaine and is considered crucial in the progression from recreational use to compulsive abuse. However, little research has focused on this important aspect of cocaine use. Cocaine was administered i.v. at doses of 0.0, 0.5, 1.0 and 2.0 mg/kg. Plasma concentrations were determined by gas chromatograph mass spectrometry (GC/MS) using deuterated internal standards d3 cocaine and d3 benzoylecgonine (BE). Mean plasma concentrations of cocaine, were on samples collected 1 min after infusion, 46 +/- 31, 88 +/- 15 and 275 +/- 116 mg/microliters in the 0.5, 1.0 and 2.0 mg/kg dose groups, respectively. There were no detectable concentrations of BE in any of the specimens nor was cocaine detected in the saline controls. Analysis of the behavioural data revealed that the 0.5 and 1.0 mg/kg results were intermediate between the results obtained at doses of 0.0 and 2.0 mg/kg and that the 1.0 mg/kg dose produced a higher response than the 0.5 mg/kg dose up to the 12 min. Regarding individual behavioural signs, those designated escape attempts, checking, feinting, restlessness, searching, vocalizing, chewing, crouching and wide-eyed were noted most frequently. The results showed dose-response relationships for both plasma concentrations of cocaine and for the total number of overt behavioural signs. The plasma concentrations were in the range reported for human cocaine abusers.

Production of urinary ethanol after sample collection.

As the interest in urine drug testing grows, ethanol is frequently included in drug-abuse screening. Collection of urine for drug testing is less invasive than blood collection and is used to screen employees in a large cross-section of occupations. Because alcohol can be produced from carbohydrates via fermentation, our interest was to determine: (1) if ethanol could be produced in glucose-positive urine (2) under what microbiological conditions would this process occur, and (3) would the urine ethanol concentration be significant. Fourteen urine specimens were selected from the Urinalysis Laboratory of a large medical center. All specimens were tested for ethanol concentration on the day of voiding and were found to be negative (< 0.01 mg/100 mL). Urine glucose concentrations ranged from 0 to > or = 2000 mg/dL. Microbiological examinations were performed on all specimens. Storing the samples at room temperature, five of the specimens produced ethanol over the time course of the study (1 to 21 days) in concentrations ranging from 0.036 to 2.327 g/100 mL. Yeast was identified in the five glucose positive urine samples producing ethanol. Six glucose positive urine samples that did not produce ethanol were found to be yeast negative. Findings indicate that significant ethanol concentrations can develop from glucose and yeast positive urine, after the day of voiding.

EMIT-d.a.u. monoclonal amphetamine/methamphetamine assay. II. Detection of methylenedioxyamphetamine (MDA) and methylenedioxymethamphetamine (MDMA).

The cross-reactivity, stereoselectivity and clinical performance of the EMIT-d.a.u. monoclonal amphetamine/methamphetamine immunoassay (EM) for the detection of methylenedioxyamphetamine (MDA) and methylenedioxymethamphetamine (MDMA) in urine was evaluated. The cut-off concentrations of racemic MDA and MDMA were found to be approximately 800 ng/ml and 3000 ng/ml, respectively. The EM assay demonstrated a high selectivity for the S(+) isomer of both MDA and MDMA. Urines collected over a 24-h period from rats administered 20 mg i.v. racemic MDMA were all positive when analyzed by the EM assay. The EM was found vastly superior to the EMIT d.a.u. polyclonal amphetamine/methamphetamine assay for the detection of MDA and MDMA. The EM assay displayed sufficient sensitivity for detection of these drugs following clinical intoxication.

EMIT-d.a.u. monoclonal amphetamine/methamphetamine assay. I. Stereoselectivity and clinical evaluation.

The stereoselectivity, cross-reactivity and clinical performance of the EMIT-d.a.u. monoclonal amphetamine(A)/methamphetamine (MA) immunoassay (EM) were evaluated. The cut-off calibrator of the assay was 1000 ng/ml S(+)MA. Analysis of drug-added urines and 72 clinical specimens demonstrated a cut-off for S(+)-amphetamine of approximately 400 ng/ml. The stereoisomeric selectivity of the assay was determined in a concentration vs. response manner by adding pure S(+) or R(-)isomers of A and MA, to drug free urine. The EM assay demonstrated a high selectivity for S(+)-isomers with only one of 16 urine specimens collected following excessive use of nasal inhalers yielding a positive result. This specimen contained 6000 ng/ml R(-)MA. Five-hundred clinical urine specimens were simultaneously analyzed for A or MA by the EM and EMIT-d.a.u. polyclonal (EP) amphetamine assay with 131 positive results confirmed by GC/MS. In five specimens negative by EM while positive by EP, MA was present at concentrations below the 1000 ng/ml cut-off. Two ME false positive results were apparently caused by chlorpromazine (CPZ) metabolites. A study of other phenothiazines or their metabolites gave no false positive results. The possible cross reactivity of the EM assay was further studied for phenyl-isopropylamine analogs or drugs previously reported to react with the EP assay. The EM assay showed much less cross-reactivity than EP to all drugs tested.

Postinjury scopolamine administration in experimental traumatic brain injury.

A single bolus dose of scopolamine (1.0 mg/kg) or saline (equal volume) was injected (i.p.) at 15, 30 or 60 min after fluid percussion traumatic brain injury in the rat. Scopolamine administered at 15 min postinjury significantly reduced beam walking deficits and body weight loss assessed for 5 days after injury. Scopolamine treatment at 30 or 60 min postinjury had no effect on behavioral outcome assessed for 5 days after injury. Plasma concentrations of scopolamine were measured with a radioreceptor assay. The plasma half-life for scopolamine was 21.6 min in injured rats and 17.3 min in normal rats (P less than 0.05). These results, along with evidence from previous studies, suggest that a brief period of excessive neuronal excitation can produce relatively long-lasting behavioral deficits. The temporal effectiveness of receptor antagonist intervention in this process appears to be brief.

Demonstration of physical dependence following chronic continuous methadone delivery via osmotic minipumps in pregnant rats.

The effects of a 14-day (gestation days 7-20) chronic methadone (6.3-9.0 mg/kg/day) infusion via osmotic minipumps were studied on the induction of physical dependence in both pregnant and nonpregnant female rats. Following continued methadone exposure, an acute injection of naloxone (2.0 mg/kg, SC) produced the following symptoms of withdrawal in both pregnant and nonpregnant methadone-exposed rats: increased frequency of head shakes, teeth-chattering and face-rubbing episodes, as well as the induction of burrowing, diarrhea, facial tremor, squeaking and vaginal sniffing. Increased fetal movement in the maternal abdomen was also observed in the pregnant rats. In the saline-exposed pregnant controls, naloxone failed to induce a significant effect. In addition, brain and plasma methadone levels during the various stages of pregnancy (gestation days 8-20) were determined. The methadone levels in plasma were initially variable (gestation days 8-12) but became more constant (approximately 50 ng/ml) from gestation day 14 to 20. Methadone brain levels also followed a similar pattern, except that the brain methadone content was at least 20-fold greater than plasma concentrations at any given time. Thus, relative to the high brain levels, the present data suggest that acute changes in methadone plasma concentration may not be a good index of pharmacological effect.

Evidence for arsenic as the immunosuppressive component of gallium arsenide.

Gallium arsenide (GaAs) has been shown previously to suppress the in vivo antibody-forming cell (AFC) response to sheep erythrocytes (SRBC) when administered intratracheally at concentrations between 50 and 200 mg/kg. In the present studies, direct addition of GaAs to in vitro-generated antibody cultures resulted in dose-dependent suppression of the primary antibody response, and was only seen when GaAs was added within 36 hr following immunization. Using atomic absorption spectrophotometry on tissue samples from mice exposed to 200 mg/kg GaAs, arsenic concentrations were found to peak in the spleen at 24 hr and decline, whereas gallium concentrations continue to rise through 14 days. Concentrations of each metal in the spleen at 24 hr are comparable to the concentrations achieved for each metal when GaAs is added at 25 microM to the in vitro model system. The 24 hr time point was chosen for comparison because all in vivo-in vitro studies were conducted using spleens from mice 24 hr after GaAs exposure. NaAsO2 and Ga(NO3)3 suppressed the AFC response dose-dependently, and in a time-dependent manner similar to GaAs when added to the in vitro system. However, based on IC50 values for each salt, the role of the gallium component in the immunosuppression appears weak. Oxalic acid (OA) and meso-2,3-dimercaptosuccinic acid (DMSA), chelators of gallium and arsenic respectively, were added to cultures with GaAs to confirm that arsenic was the primary immunosuppressive component. DMSA dose-dependently blocked GaAs-induced immunosuppression in vitro, while OA had no effect. The metal-binding compounds were determined to be specific for the metals used in these studies and did not cross-react with one another. DMSA was evaluated for its ability to prevent suppression of the AFC response in splenocytes from GaAs-exposed mice and was able to block GaAs-induced suppression of the AFC response when given sc every 4 hr beginning 1 hr prior to GaAs exposure. These data indicate that the arsenic component of GaAs is the major contributor to the GaAs-induced immunosuppression and that this effect occurs within the first 36 hr of the 5-day culture period in a concentration-dependent manner.

Urinary excretion of benzoylecgonine following ingestion of Health Inca Tea.

Four males ingested one cup of Health Inca Tea which contained 1.87 mg of cocaine. Urine specimens collected for 36 h post-ingestion were analyzed for benzoylecgonine (BE) by EMIT-d.a.u., TDx and gas chromatography/mass spectrometry (GC/MS). Positive immunoassay results were obtained for 21-26 h post tea ingestion. Discrepant immunoassay results occurred with only one specimen: EMIT positive; TDx negative, 0.25 mg/l; GC/MS, 0.273 mg/l. Quantitative TDx results were well correlated with GC/MS results, r2 = 0.963, n = 45. Maximum urinary BE concentrations ranged from 1.4-2.8 mg/l, occurring from 4-11 h, post ingestion. Total BE excretion in 36 h ranged from 1.05 to 1.45 mg, 59-90% of the ingested cocaine dose. Urinary excretion rate constant (Km) ranged from -0.073 to 0.111/h. Health Inca Tea ingestion should be considered when interpreting urinary BE concentrations.

Arsenic poisoning: acute or chronic? Suicide or murder?

The case of the death by arsenic poisoning of a 62-year-old white man is presented. One year prior to death, he developed intermittent bouts of severe gastroenteritis with vomiting and diarrhea, hyperpigmentation and keratosis of the skin, neutropenia, and Guillain-Barré-like neuropathy for which he was hospitalized several times. Urine test results 6 months prior to death indicating 36 mg/L arsenic were believed to be in error. At the patient's last admission, he appeared in the emergency room with severe gastroenteritis, hypotension, and dehydration. He died 3 days later. Antemortem as well as autopsy specimens revealed elevated arsenic concentrations. Arsenic micrograms/g analysis by neutron activation of hair pulled from the man's head revealed by centimeter segmental analysis proximal to distal: 226, 104, 28, 56, 41, 40, and 74. The wife of the decedent was charged with murder by arsenic poisoning of this, her fifth, husband. The defense alleged that the decedent had committed suicide. The judge awarded a directed verdict of "not guilty." Particulars of the medical, toxicological, and investigative findings are presented.

Determination of chlorinated hydrocarbon pesticides by solid-phase extraction and capillary GC with electron capture detection.

Pesticides and their metabolites are extracted and concentrated from serum using C18 solid-phase extraction cartridges. The internal standard aldrin is added to 4 mL of serum or plasma, treated with 2 mL of methanol, and the resultant supernatant applied to the C18 cartridge. After several washes, pesticides are eluted from the column with isoctane and the eluate quantified with capillary gas chromatography using electron capture detection. Extraction efficiency is from 70 to 75% and the method detection limit ranges from 0.1-0.7 ng/mL depending on the analyte. Precision studies demonstrate that CVs range from 3.5 to 25.2%. Standard curves are linear to at least 7 ng/mL for lindane and chlordane isomers, heptachlor, heptachlor epoxide, oxychlordane, trans-nonachlor, dieldrin, dichlorodiphenyldichloroethylene (p,p'-DDE), dichlorodiphenyldichloroethane (p,p'-DDD), and dichlorodiphenyltrichloroethylene (p,p'-DDT).

Evaluation of the Abbott ADx total serum tricyclic immunoassay.

The ADx total serum tricyclic antidepressant (TCA) fluorescence polarization immunoassay (Abbott Diagnostics) for the semi-quantitation of imipramine or amitriptyline and their respective N-demethylated metabolites in cases of TCA overdose was evaluated. The assay is linear from 75-1000 ng/mL total TCA in serum, and flaggs as "HI" all results exceeding 300 ng/mL. The within and between run precision of the assay for patient serum containing imipramine or amitriptyline and their metabolites gave CV's of less than 5.5% and 8.9%, respectively. A good correlation between the results of patient serum containing imipramine and desipramine simultaneously analyzed by ADx and gas liquid chromatography (GC) was observed, r2 = 0.964, n = 32. Results of patient serum containing amitriptyline and nortriptyline or doxepin and desmethyldoxepin analyzed by ADx and GC or GC-mass spectrometry were not well correlated; r2 = 0.738, n = 44 and r2 = 0.695, n = 21, respectively. The assay consistently flagged as "HI" serum with total imipramine and desipramine concentrations above 300 ng/mL by GC, and serum with greater than 360 ng/mL of amitriptyline and nortriptyline by GC/MS. No significant cross-reactivity was observed for drugs other than the TCA.