Multiple-drug toxicity caused by the coadministration of 4-methylmethcathinone (mephedrone) and heroin.

An accidental death caused by the combined use of a new designer drug, 4-methylmethcathinone (mephedrone), and heroin is reported. A 22-year-old Caucasian male was found unresponsive in his living quarters and was transported to the hospital where he died. During autopsy, needle marks were found along the decedent's lower legs and ankles. Investigators discovered the decedent and his roommate had been using "Black Tar" heroin and mephedrone. Routine toxicological analysis detected morphine in the decedent's blood at 0.06 mg/L. Additionally, 6-acetylmorphine, morphine, codeine, and doxylamine were detected in his urine. A designer drug screen, employing a basic liquid-liquid extraction followed by pentafluropropionic anhydride derivatization, was used to isolate mephedrone from both blood and urine specimens. The derivatized extracts were analyzed by gas chromatography- mass spectrometry (GC-MS) operating in full-scan mode. Quantitative analysis of mephedrone was performed by GC-MS operating in selective ion monitoring mode using methamphetamine-d(14) as an internal standard. Mephedrone was confirmed in the decedent's blood and urine at 0.50 and 198 mg/L, respectively. The physiological and pharmacological effects of mephedrone and any associated toxicity have not been reported. However, because of its structural similarities with methcathinone and the high concentration in the decedent's blood, the overall contribution of mephedrone to the death could not be minimized. Therefore, the medical examiner reported the cause of death as multiple-drug toxicity and the manner of death as accidental.

An overdose death involving the insufflation of extended-release oxymorphone tablets.

Two cases are reported involving the abuse of extended-release oxymorphone hydrochloride tablets (Opana® ER) in combination with alprazolam (Xanax®). Two juvenile females were discovered unresponsive and hypoxic by a male acquaintance. The trio had reportedly crushed and snorted Opana ER tablets and consumed Xanax for recreational purposes. Emergency personnel were able to stabilize one female. The second female was pronounced dead at the scene. Blood and urine samples from the surviving female were collected at the trauma center between 48 and 96 h post incident. Toxicology results showed the presence of oxymorphone, doxylamine, dextromethorphan, alprazolam, α-hydroxyalprazolam, oxazepam, and temazepam in her urine. No drugs were detected in her blood. Toxicology on the deceased female revealed the presence of 0.13 mg/L oxymorphone and 0.04 mg/L alprazolam in her blood. Gastric contents contained 0.25 and 0.93 mg/L of oxymorphone and alprazolam, respectively. Oxymorphone, alprazolam, and α-hydroxyalprazolam were present in her urine. Quantitative results were achieved by gas chromatography-mass spectrometry monitoring selected ions for the oxime-oxymorphone-trimethylsilyl derivative, alprazolam, and the α-hydroxyalprazolam tert-butyldimethylsilyl derivative. The established linearity ranges for the opiate and benzodiazepine methods were 0.050-3.000 and 0.025-1.000 mg/L, respectively. The cause of death was reported as multiple drug toxicity, and the manner of death was accidental.

Low concentrations of methamphetamine detectable in urine in the presence of high concentrations of amphetamine.

Twenty-two urine specimens reported by military drug-testing laboratories for the presence of high concentrations of amphetamine only were subject to further analysis for the presence of methamphetamine. The 22 urine specimens had concentrations of amphetamine in the range of 28,028 to 241,142 ng/mL. The specimens were also assayed for the respective isomeric ratio of d (S) and l (R) amphetamine and methamphetamine. The results suggest that urine specimens containing high concentrations of amphetamine in which the urine concentration ratio of methamphetamine to amphetamine is less than 0.5% with similar isomeric distribution of d-(S) and l-(R) amphetamine and methamphetamine, respectively, may not necessarily indicate polydrug use.

Concentration distribution of the marijuana metabolite Delta9-tetrahydrocannabinol-9-carboxylic acid and the cocaine metabolite benzoylecgonine in the department of defense urine drug-testing program.

Urine drug testing has been employed for punitive purposes by the Department of Defense since December 1981 (Memorandum 62884, Deputy Secretary of Defense Frank C. Carlucci). Federal Workplace Drug Testing Programs were initiated in response to Executive Order 12564 issued on September 15, 1986, that required Drug-Free Federal Workplaces be established. In their respective programs, a positive urine drug test may be referred to a military court martial or to an administrative board. To address safety and insurance requirements, the testing of civilians has expanded beyond Federal Programs to include pre-employment and post-accident urine drug testing. During adjudication, an Expert Toxicologist may be asked to opine what can be discerned from the concentration of drug or drug metabolite found in the urine. Little can be opined with certainty from a positive urine drug test as to the amount of drug ingested, when the drug was ingested, and in most instances, whether the individual felt the effects of the drug, or was under the influence of the drug found in the urine. What may be useful to both the Expert and to the Trier-of-Facts is the frequency that a particular urine drug concentration is encountered in positive drug tests. The finding that 50% of all positive marijuana and cocaine urine metabolite concentrations in the military testing program over the three-year period of October 1, 2004 through September 30, 2007, are below a median value of 65 and 968 ng/mL, respectively, provide reference points. A median drug concentration combined with the percentile or frequency that a particular urine drug concentration occurs may provide evaluative information for a determination of the facts and the outcome of judicial or administrative proceedings. This may be especially useful to jurors when the concentration of marijuana or cocaine metabolite is perceptibly low. The information would also be applicable to medical review officers, medical examiners, drug treatment professionals, probation officers, and program analysts coordinating drug policy decisions.

The detection and quantitative analysis of the psychoactive component of Salvia divinorum, salvinorin A, in human biological fluids using liquid chromatography-mass spectrometry.

Salvia divinorum, a member of the mint plant family, has hallucinogenic properties that have become increasingly sought after by recreational drug users. The main psychoactive component, salvinorin A, has potency comparable to lysergic acid diethylamide. Though still legal to possess in most of the United States and much of Europe, little is known regarding the compound's long-term health effects, addiction liability, and pharmacokinetics. Limited data are available in the scientific literature, and few analytical methods are published for the detection in human biological fluids. These factors contribute to the unfamiliarity of the compound and complicate the method development process necessary to accommodate special requested testing for salvinorin A. A sensitive analytical method for the detection and quantitation of salvinorin A in human biological fluids was developed and validated to resolve analytical shortcomings. The method utilizes a solid-phase extraction technique coupled with liquid chromatography-electrospray ionization mass spectrometry operated in selected ion monitoring mode. The assay has a linear range of 5.0-100 ng/mL with a correlation coefficient of 0.997. The limit of detection and limit of quantitation were experimentally determined as 2.5 and 5.0 ng/mL, respectively. The method has been applied to blood and urine samples successfully and can be used to detect the presence of salvinorin A in forensic testing.

Delta9-tetrahydrocannabinol content of commercially available hemp products.

Delta9-Tetrahydrocannabinol (THC) is the main psychoactive compound present in marijuana. THC can also be found, as a contaminant, in some commercially available hemp products marketed in health food stores and on the internet as a good source of essential fatty acids. The products range from oil to alcoholic beverages to nutritional bars to candies, with oil being the most popular and commonly available. The analytical results are separated into two groups, products tested prior to and after publication of 21 CFR Part 1308, "clarification of listing of tetrahydrocannabinols." The data presented are a summary of 79 different hemp products tested for THC. THC was separated by a liquid-liquid or solid-liquid extraction, depending upon the product matrix. THC concentrations range from none detected to 117.5 microg THC/g material. Typical limits of detection for the assay (depending on matrix) are 1.0-2.5 microg THC/g material. Products that were of aqueous base (beer, tea) had much lower limits of detection (2.5 ng/mL). No THC was detected in 58% of the products from group 1 and 86% of the products from group 2. The amounts indicate that THC levels in currently marketed hemp products are significantly lower than in those products available before 2003 and reported in previous studies. The results reported here may be used as a general guideline for the THC content of hemp products recently found in the marketplace today.

Evaluation of four immunoassay screening kits for the detection of benzodiazepines in urine.

The identification of benzodiazepines (BZD) in biological fluids can be a challenging process. The large number of various BZD in pharmaceutical distribution, with similar core structures, and individual metabolic profiles all contribute to a complicated and time-consuming analysis. The purpose of the current study was to evaluate the performance of four commercially available immunoassay urine screening kits for use in a forensic urine analysis testing program. The four kits included the Roche Benzodiazepine Plus KIMS assay, Microgenics CEDIA Benzodiazepine assay, Microgenics CEDIA high sensitivity assay with beta-glucuronidase, and Microgenics DRI reagent ready Benzodiazepine assay. Each kit was evaluated for linearity, precision, accuracy, carryover, reagent specificity, and confirmation rates. BZD reagent specificity was compared by analysis of 55 structurally dissimilar compounds to BZD. Negative responses to all 55 compounds were elicited by all four reagent assays. Cross-reactivity for the assays was demonstrated by detecting 27 structurally similar BZD. In addition, greater than 10,000 randomly collected urine samples were screened at a 200 ng/mL cutoff for each assay. Positive samples were confirmed by gas chromatography-mass spectrometry using a panel of 13 BZD confirmation standards. The Microgenics CEDIA high sensitivity assay demonstrated the highest positive screening rate as well as the highest confirmation rate of the four assays.

Detection of 1-benzylpiperazine and 1-(3-trifluoromethylphenyl)-piperazine in urine analysis specimens using GC-MS and LC-ESI-MS.

Designer piperazines, such as 1-benzylpiperazine (BZP) and 1-(3-trifluoromethylphenyl)-piperazine (TFMPP), are widely available and have become popular party drugs throughout the world. Used in many countries as legal alternatives to methamphetamine and ecstasy, these designer piperazines exhibit several of the same stimulant and psychoactive properties of their illicit counterparts. Presented is a case study of seven urine analysis specimens analyzed for designer piperazines. A full scan gas chromatography-mass spectrometry screen detected the presence of BZP and TFMPP in all seven specimens. Confirmation using liquid chromatography-electrospray ionization-mass spectrometry operating in selected ion monitoring mode (SIM) yielded urinary concentrations ranging from 13.0 to 429.1 mg/L and 0.79 to 25.4 mg/L for BZP and TFMPP, respectively.

Isomerization of delta-9-THC to delta-8-THC when tested as trifluoroacetyl-, pentafluoropropionyl-, or heptafluorobutyryl- derivatives.

For GC-MS analysis of delta-9-tetrahydrocannabinol (delta-9-THC), perfluoroacid anhydrides in combination with perfluoroalcohols are commonly used for derivatization. This reagent mixture is preferred because it allows simultaneous derivatization of delta-9-THC and its acid metabolite, 11-nor-delta-9-THC-9-carboxylic acid present in biological samples. When delta-9-THC was derivatized by trifluoroacetic anhydride/hexafluoroisopropanol (TFAA/HFIPOH) and analyzed by GC-MS using full scan mode (50-550 amu), two peaks (P1 and P2) with an identical molecular mass of 410 amu were observed. On the basis of the total ion chromatogram (TIC), P1 with a shorter retention time (RT) was the major peak (TIC 84%). To identify the peaks, delta-8-THC was also tested under the same conditions. The RT and spectra of the major peak (TIC 95%) were identical with that of P1 for delta-9-THC. A minor peak (5%) present also correlated well with the latter peak (P2) for the delta-9-THC derivative. The fragmentation pathway of P1 was primarily demethylation followed by retro Diels-Alder fragmentation (M - 15-68, base peak 100%) indicating P1 as a delta-8-THC-trifluoroacetyl compound. This indicated that delta-9-THC isomerized to delta-8-THC during derivatization with TFAA/HFIPOH. Similar results were also observed when delta-9-THC was derivatized with pentafluoropropionic anhydride/pentafluoropropanol or heptafluorobutyric anhydride/heptafluorobutanol. No isomerization was observed when chloroform was used in derivatization with TFAA. In this reaction, the peaks of delta-8-THC-TFA and delta-9-THC-TFA had retention times and mass spectra matching with P1 and P2, respectively. Because of isomerization, perfluoroacid anhydrides/perfluoroalcohols are not suitable derivatizing agents for analysis of delta-9-THC; whereas the TFAA in chloroform is suitable for the analysis.

Stable isotopes as valid components for identification of drugs in biological specimens.

Selected ion monitoring in mass spectrometric methods (SIM-MS) is generally used to confirm the presence of a drug in biological samples. Criteria for identification of a compound by MS are based on some specific guidelines. However, some disparities exist between the guidelines as to how many and what type of ions to monitor. Although European guidelines allow the monitoring of isotopic ions, such monitoring is not valid by SOFT/AAFS guidelines. The feasibility of monitoring a stable isotopic ion as an alternate to the fragment ion was examined in our study. Area ratios of stable isotopic ion m/z 275 and its parent ion m/z 274 of optical isomers of methamphetamine as (R)-(-)-alpha-methoxy-alpha-(trifluoromethyl)phenylacetyl derivative were found to be within +/- 4% of theoretical value (14.969) calculated from fragment formula C(13)H(15)F(3)NO(2) and isotopic abundances (C(13) = 1.1%, H(2) = 0.015%, F = 0%, N(15) = 0.37%, and O(17) = 0.037%). In another example, the area ratios of a stable isotopic ion m/z 283 and the parent ion m/z 282 of 6-pentafluoropropionyl codeine was also within +/- 4% of theoretical value (20.542) calculated from fragment formula C(18)H(20)NO(2). This relationship between the isotopic abundance and fragment composition was also useful in assigning structures to many fragment ions of methamphetamine, LSD, morphine, and 6-acetylmorphine derivatives, whereas structural assignment to the ions based on mass alone was difficult. The predictability of the relative abundance of the examined isotopic ions has proven reliable in our studies. The use of an isotope was found to be an important additional tool for compound identification by MS.