Two Fatal Intoxications Involving Butyryl Fentanyl.

We present the case histories, autopsy findings and toxicology findings of two fatal intoxications involving the designer drug, butyryl fentanyl. The quantitative analysis of butyryl fentanyl in postmortem fluids and tissues was performed by an ultrahigh-performance liquid chromatography tandem mass spectrometry method. In the first case, butyryl fentanyl was the only drug detected with concentrations of 99 ng/mL in peripheral blood, 220 ng/mL in heart blood, 32 ng/mL in vitreous humor, 590 ng/mL in gastric contents, 93 ng/g in brain, 41 ng/g in liver, 260 ng/mL in bile and 64 ng/mL in urine. The cause of death was ruled fatal intoxication by butyryl fentanyl. In the second case, butyryl fentanyl was detected along with acetyl fentanyl, alprazolam and ethanol. The butyryl fentanyl concentrations were 3.7 ng/mL in peripheral blood, 9.2 ng/mL in heart blood, 9.8 ng/mL in vitreous humor, 4,000 ng/mL in gastric contents, 63 ng/g in brain, 39 ng/g in liver, 49 ng/mL in bile and 2 ng/mL in urine. The acetyl fentanyl concentrations were 21 ng/mL in peripheral blood, 95 ng/mL in heart blood, 68 ng/mL in vitreous humor, 28,000 ng/mL in gastric contents, 200 ng/g in brain, 160 ng/g in liver, 330 ng/mL in bile and 8 ng/mL in urine. In addition, the alprazolam concentration was 40 ng/mL and the ethanol concentration was 0.11 g/dL, both measured in peripheral blood. The cause of death in the second case was ruled a mixed drug intoxication. In both cases, the manner of death was accident.

Postmortem tissue distribution of acetyl fentanyl, fentanyl and their respective nor-metabolites analyzed by ultrahigh performance liquid chromatography with tandem mass spectrometry.

In the last two years, an epidemic of fatal narcotic overdose cases has occurred in the Tampa area of Florida. Fourteen of these deaths involved fentanyl and/or the new designer drug, acetyl fentanyl. Victim demographics, case histories, toxicology findings and causes and manners of death, as well as, disposition of fentanyl derivatives and their nor-metabolites in postmortem heart blood, peripheral blood, bile, brain, liver, urine and vitreous humor are presented. In the cases involving only acetyl fentanyl (without fentanyl, n=4), the average peripheral blood acetyl fentanyl concentration was 0.467 mg/L (range 0.31 to 0.60 mg/L) and average acetyl norfentanyl concentration was 0.053 mg/L (range 0.002 to 0.086 mg/L). In the cases involving fentanyl (without acetyl fentanyl, n=7), the average peripheral blood fentanyl concentration was 0.012 mg/L (range 0.004 to 0.027 mg/L) and average norfentanyl blood concentration was 0.001 mg/L (range 0.0002 to 0.003 mg/L). In the cases involving both acetyl fentanyl and fentanyl (n=3), the average peripheral blood acetyl fentanyl concentration was 0.008 mg/L (range 0.006 to 0.012 mg/L), the average peripheral blood acetyl norfentanyl concentration was 0.001 mg/L (range 0.001 to 0.002 mg/L), the average peripheral blood fentanyl concentration was 0.018 mg/L (range 0.015 to 0.021mg/L) and the average peripheral blood norfentanyl concentration was 0.002 mg/L (range 0.001 mg/L to 0.003 mg/L). Based on the toxicology results, it is evident that when fentanyl and/or acetyl fentanyl were present, they contributed to the cause of death. A novel ultrahigh performance liquid chromatography (UPLC) tandem mass spectrometry (MS/MS) method to identify and quantify acetyl fentanyl, acetyl norfentanyl, fentanyl and norfentanyl in postmortem fluids and tissues is also presented.

Postmortem Toxicology Findings of Acetyl Fentanyl, Fentanyl, and Morphine in Heroin Fatalities in Tampa, Florida.

In the last two years, an epidemic of 40 fatal heroin overdose cases has occurred in the Tampa area of Florida. Of these cases, 14 involved fentanyl and acetyl fentanyl. Victim demographics, case histories, toxicology findings, and causes and manners of death for all 40 deaths are presented. In 26 deaths in which acetyl fentanyl or fentanyl were not involved, free and total peripheral blood morphine concentrations were consistent with fatal heroin intoxications, averaging 0.16 mg/L and 0.35 mg/L, respectively. In the heroin cases with fentanyl present (n=7), the average free morphine concentration was 0.040 mg/L, the average total morphine concentration was 0.080 mg/L, and the average fentanyl concentration was 0.012 mg/L. In the cases with heroin, fentanyl, and acetyl fentanyl (n=3), the average free morphine concentration was 0.010 mg/L, the average total morphine concentration was 0.030 mg/L, the average fentanyl concentration was 0.018 mg/L, and the average acetyl fentanyl concentration was 0.008 mg/L. In the cases involving only acetyl fentanyl (without heroin or fentanyl, n=4), the average acetyl fentanyl concentration was 0.47 mg/L and the average acetyl norfentanyl concentration was 0.053 mg/L. The presented cases, with associated drug concentrations, case histories, demographics, and causes and manners of death may help provide assistance with the interpretation of the postmortem findings. Based on case circumstances, autopsy results, and toxicology results, it is evident that fentanyl and/or acetyl fentanyl, when present, contributed to the cause of death.

5-HTTLPR genotype and associations with intoxication and intention to drive: results from a field study of bar patrons.

The serotonin transporter promoter polymorphism (5-HTTLPR) has been linked to a number of human behavioral traits and disorders. The variants of 5-HTTLPR are commonly reported in three forms, L/L, S/L and S/S, with the latter most often associated with emotional distress and/or behavioral dysfunction. Missing from the research literature are investigations that assess event-level associations between 5-HTTLPR genotype and specific incidents of risk behavior in natural drinking settings. This study reports associations between 5-HTTLPR, alcohol intoxication and intention to drive among young adult patrons exiting on-premise drinking establishments (i.e. bars) at night. Self-report measures, breath alcohol concentration (BrAC) readings and saliva samples for DNA analysis were collected from 477 bar patrons. Analyses were performed on 225 patrons likely to be near their peak intoxication level for the night. Results from a linear regression revealed that the 5-HTTLPR genotype was associated with exiting patron BrAC, after adjusting for random and fixed effects of other variables. An interaction effect involving 5-HTTLPR and bar-sponsored drink specials also had an independent association with BrAC, suggesting that selection of price-discounted alcoholic beverages increased intoxication in patrons with an L allele. In addition, results from logistic regression indicated that patrons with the S/S genotype were three times more likely to intend to drive a motor vehicle (after drinking on the night of study participation) compared with those with the L/L genotype. The 5-HTTLPR genotype may play an important role in the etiology of problems associated with on-premise drinking establishments.

Quantitation of benzodiazepines in blood and urine using gas chromatography-mass spectrometry (GC-MS).

The benzodiazepine assay utilizes gas chromatography-mass spectrometry (GC-MS) for the analysis of diazepam, nordiazepam, oxazepam, temazepam, lorazepam, alpha-hydroxyalprazolam, and alpha-hydroxytriazolam in blood and urine. A separate assay is employed for the analysis of alprazolam. Prior to solid phase extraction, urine specimens are subjected to enzyme hydrolysis. The specimens are fortified with deuterated internal standard and a five-point calibration curve is constructed. Specimens are extracted by mixed-mode solid phase extraction. The benzodiazepine extracts are derivatized with N-methyl-N-(tert-butyldimethylsilyl)trifluoroacetamide (MTBSFTA) producing tert-butyldimethyl silyl derivatives; the alprazolam extracts are reconstituted in methanol without derivatization. The final extracts are then analyzed using selected ion monitoring GC-MS.

Quantitation of fentanyl in blood and urine using gas chromatography-mass spectrometry (GC-MS).

Fentanyl is a potent, short-acting synthetic opioid analgesic. Fentanyl is measured in blood and urine following mixed-mode solid phase extraction. The specimens are fortified with deuterated internal standard and a five-point calibration curve is constructed. The final extracts are reconstituted in methanol and analyzed using selected ion monitoring gas chromatography-mass spectrometry.

Quantitation of opioids in blood and urine using gas chromatography-mass spectrometry (GC-MS).

The opioid and 6-acetylmorphine assays utilize gas chromatography-mass spectrometry (GC-MS) for the analysis of morphine, codeine, hydromorphone, hydrocodone, and 6-acetylmorphine in blood and urine. The specimens are fortified with deuterated internal standard and a five-point calibration curve is constructed. Specimens are extracted by mixed-mode solid phase extraction. The morphine, codeine, hydromorphone, hydrocodone, and 6-acetylmorphine extracts are derivatized with N-methyl-bis(trifluoroacetamide) (MBTFA) producing trifluoroacetyl derivatives. The final extracts are then analyzed using selected ion monitoring GC-MS.

Quantitation of oxycodone in blood and urine using gas chromatography-mass spectrometry (GC-MS).

Oxycodone is a semisynthetic opioid analgesic used in pain management. In the following method, gas chromatography-mass spectrometry (GC-MS) is used to determine the presence and concentration of the drug in blood and urine. The specimens are fortified with deuterated internal standard and a five-point calibration curve is constructed. Specimens are extracted using mixed-mode solid phase extraction and derivatized with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) producing trimethylsilyl derivatives. The final extracts are then analyzed using selected ion monitoring GC-MS.

Event-level analyses of energy drink consumption and alcohol intoxication in bar patrons.

AIM: To assess event-level associations between energy drink consumption, alcohol intoxication, and intention to drive a motor vehicle in patrons exiting bars at night. METHOD: Alcohol field study. Data collected in a U.S. college bar district from 802 randomly selected and self-selected patrons. Anonymous interview and survey data were obtained as well as breath alcohol concentration (BrAC) readings. RESULTS: Results from logistic regression models revealed that patrons who had consumed alcohol mixed with energy drinks were at a 3-fold increased risk of leaving a bar highly intoxicated (BrAC> or =0.08g/210L), as well as a 4-fold increased risk of intending to drive upon leaving the bar district, compared to other drinking patrons who did not consume alcoholic beverages mixed with energy drinks. DISCUSSION: These event-level associations provide additional evidence that energy drink consumption by young adults at bars is a marker for elevated involvement in nighttime risk-taking behavior. Further field research is needed to develop sound regulatory policy on alcohol/energy drink sales practices of on-premise establishments.

Alcohol price and intoxication in college bars.

BACKGROUND: Many population studies find that alcohol prices are inversely related to alcohol consumption and alcohol-related problems, including among college students and young adults. Yet, little is known about the "micro-level" effects of alcohol price on the behavior of individual consumers in natural drinking settings such as college bars. Therefore, we assessed patron's cost per gram of ethanol consumed at on-premise drinking establishments and its association with intoxication upon leaving an establishment. METHODS: On 4 consecutive nights during April 2008, data were collected from 804 patrons exiting 7 on-premise establishments in a bar district located adjacent to a large university campus in the southeastern United States. Anonymous interview and survey data were collected as well as breath alcohol concentration (BrAC) readings. We calculated each patron's expenditures per unit of ethanol consumed based on self-reported information regarding the type, size, number, and cost of consumed drinks. RESULTS: A multivariable model revealed that a 10-cent increase in cost per gram of ethanol at on-premise establishments was associated with a 30% reduction in the risk of exiting an establishment intoxicated (i.e., BrAC > or = 0.08 g/210 l). CONCLUSIONS: The results are consistent with economic theory and population-level research regarding the price elasticity of alcoholic beverages, which show that increases in alcohol prices are accompanied by less alcohol consumption. These findings suggest that stricter regulation of the drink discounting practices of on-premise drinking establishments would be an effective strategy for reducing the intoxication levels of exiting patrons.

Event-specific analyses of poly-drug abuse and concomitant risk behavior in a college bar district in Florida.

OBJECTIVE: The authors describe the epidemiology of risk behavior associated with poly-drug use in a college bar district of a large campus community. PARTICIPANTS: A total of 469 bar patrons participated in the study. METHODS: The authors used self-report data and biological measures collected from patrons outside bars in July and August of 2007. RESULTS: The mean breath alcohol concentration of the exiting patrons was 0.09. Illicit and prescription drug use on the nights of data collection and in the recent past were significant features of the profile of patron risk behavior. About one-quarter of the patrons using only alcohol reported an intention to drive a vehicle within 60 minutes of leaving an establishment, compared with almost one-half of those using both alcohol and marijuana. CONCLUSIONS: A substantial amount of high-risk behavior was generated from the bar district on 4 typical nights. College bar districts should be a priority focus for prevention efforts.

A field study of bar-sponsored drink specials and their associations with patron intoxication.

OBJECTIVE: The study examined associations between bar-sponsored drink specials and alcohol intoxication at the patron level. METHOD: Data were collected in a college bar district located in a large campus community in the southeastern United States. Random and self-selected samples of patrons were interviewed after exiting college bars at night on four different nights (N=383). Anonymous interview and questionnaire data were collected as well as breath alcohol concentration (BrAC) readings. RESULTS: Significant gender differences existed in patron drinking practices. Women were more likely to take advantage of drink specials, whereas men reported greater alcohol expenditures, consumed more drinks, and drank for longer periods of time. Gender differences in BrAC were very small and not meaningful. Patrons who did not take advantage of drink specials reported consuming more drinks before bar entry than patrons who did participate in these promotions. Participation in "all-you-can-drink" promotions was significantly associated with higher BrAC readings after adjusting for covariates and random effects attributable to drinking establishment. Other drink specials did not have significant associations with alcohol intoxication. CONCLUSIONS: The all-you-can-drink special may be the specific discounting practice with the greatest potential for boosting patron intoxication and thus may need to be a stronger focus of alcohol-control policies aimed at improving the beverage service of drinking establishments.

Caffeine content of brewed teas.

Caffeine is the world's most popular drug and can be found in many beverages including tea. It is a psychostimulant that is widely used to enhance alertness and improve performance. This study was conducted to determine the concentration of caffeine in 20 assorted commercial tea products. The teas were brewed under a variety of conditions including different serving sizes and steep-times. Caffeine was isolated from the teas with liquid-liquid extraction and quantitated by gas chromatography with nitrogen-phosphorus detection. Caffeine concentrations in white, green, and black teas ranged from 14 to 61 mg per serving (6 or 8 oz) with no observable trend in caffeine concentration due to the variety of tea. The decaffeinated teas contained less than 12 mg of caffeine per serving, and caffeine was not detected in the herbal tea varieties. In most instances, the 6- and 8-oz serving sizes contained similar caffeine concentrations per ounce, but the steep-time affected the caffeine concentration of the tea. These findings indicate that most brewed teas contain less caffeine per serving than brewed coffee.

Diminished nicotine withdrawal in adolescent rats: implications for vulnerability to addiction.

RATIONALE: Enhanced reinforcing effects of nicotine during adolescence appear to contribute to the rapid development of dependence in this age group. However, the contribution of nicotine withdrawal to dependence in adolescents is unclear. OBJECTIVE: We compared motivational and somatic signs of nicotine withdrawal in adolescent and adult rats. MATERIALS AND METHODS: In experiment 1, motivational signs of nicotine withdrawal were compared using intracranial self-stimulation procedures after administration of mecamylamine (1.5 mg/kg, i.p.) in adolescent and adult rats made dependent on nicotine (9 mg/kg/day). Somatic signs of withdrawal were compared in two experiments using various doses of nicotine (adolescent doses: 0, 1.6, 3.2, 4.7 mg/kg/day; adult doses: 0, 1, 2.1, 3.2 mg/kg/day, expressed as nicotine base) to produce dependence and one dose of mecamylamine (1.5 mg/kg, i.p.) to precipitate withdrawal (experiment 2) and in a subsequent experiment, using various doses of mecamylamine (0, 0.75, 1.5, 3.0 mg/kg, i.p.) to precipitate withdrawal and a dose of nicotine (adolescent dose: 4.7 mg/kg/day; adult dose: 3.2 mg/kg/day) that produced equivalent nicotine blood levels in these age groups (experiment 3). RESULTS: Adolescents did not display the decreases in brain reward function observed in adults experiencing withdrawal, and displayed fewer somatic signs of nicotine withdrawal relative to adults regardless of the dosing procedure used. CONCLUSION: The negative effects of nicotine withdrawal are lower during adolescence relative to later periods of development. Both the enhanced rewarding effects and the diminished nicotine withdrawal likely contribute to the rapid development of nicotine use during adolescence.

Second-hand exposure to aerosolized intravenous anesthetics propofol and fentanyl may cause sensitization and subsequent opiate addiction among anesthesiologists and surgeons.

We hypothesize that aerosolization of anesthetics administered intravenously to patients in the operating room may be an unintended source of exposure to physicians. This may lead to inadvertent sensitization, which is associated with an increased risk for developing addiction. This may contribute to the over-representation of certain specialties among physicians with addiction. We retrospectively reviewed the de-identified demographic information of all licensed physicians treated for substance abuse in the State of Florida since 1980, to determine if medical specialty was associated with addiction in this group of individuals. Then, to identify the potential for exposure, two mass spectrometry assays were developed to detect two intravenously administered drugs, fentanyl and propofol, in air. Since 1980, 7.6% of licensed Florida physicians underwent treatment for addiction. Addiction in anesthesiologists was higher than expected. Opiate abuse was greater in anesthesiologists and surgeons compared to other specialties. Aerosolized fentanyl was detected in the air of the cardiothoracic operating room, in patients' expiratory circuits, and in the headspace above sharps boxes, but not in adjoining hallways. Aerosolized propofol was detected in the expirations of a patient undergoing transurethral prostatectomy. While access and stress may place anesthesiologists and surgeons at greater risk for substance abuse, an additional risk factor may be unintended occupational exposure to addictive drugs. This report provides preliminary evidence of detection of aerosolized intravenous anesthetics using two newly developed analytical methods. We conclude that the potential exists for chronic exposure to low levels of airborne intravenously administered drugs. Further studies are under way to determine the significance of this exposure.

Putting an Ecstasy test kit to the test: harm reduction or harm induction?

OBJECTIVE: To evaluate the DanceSafe Complete Adulterant Screening Kit for Ecstasy with regard to its accuracy in identifying 3,4-methylenedioxymeth-amphetamine (MDMA) and methylenedioxyamphetamine (MDA) derivatives and its ability to detect certain contaminants. METHODS: In part 1, 39 street-grade tablets purported to be MDMA were tested with the Marquis, Mecke, and Simon's reagents provided by the DanceSafe testing kit. The tablets then were submitted to gas chromatography-mass spectrometry for identification of active ingredients. In part II, seven known drugs of abuse were tested with the Marquis, Mecke, and Simon's reagents. These drugs were codeine, dextromethorphan, dihydrocodeine, ketamine, MDMA, morphine, and d-norpropoxyphene. RESULTS: The Marquis, Mecke, and Simon's reagents did not differentiate pure MDMA from adulterated forms. They lacked both sensitivity and specificity for the purpose of MDMA identification when tested by persons unfamiliar with these reagents. Also, experienced toxicologists using this unfamiliar procedure generated false-positive results. CONCLUSIONS: Neither the Marquis, Mecke, nor Simon's reagents should be used by the public for harm reduction purposes. These agents do not help identify pure MDMA tablets. Gas chromatography-mass spectrometry remains the most sensitive and specific testing method for identifying MDMA and its contaminants.

Expression of aquaporin 1 and 5 in the developing mouse inner ear and audiovestibular assessment of an Aqp5 null mutant.

To examine the potential roles of aquaporins 1 and 5 (AQP1 and AQP5, respectively) in inner ear development and function, we defined their spatial and temporal expression patterns in the developing mouse inner ear and examined the morphologic and physiologic effects of loss of Aqp5 function. Standard in situ hybridization (ISH) and immunohistochemical (IHC) assays were used for expression studies with routine morphologic, behavioral, and physiologic assessments of hearing and balance in Aqp5 null mutant mice. AQP1 was first detected at embryonic day 10.5 (E10.5) in the otocyst but eventually localized to specific nonsensory portions of the inner ear and connective tissue cells surrounding the membranous labyrinth. AQP5 displayed specific cochlear expression, first detectable at E15.5 in the nonsensory epithelium and later restricted to the lateral wall of the cochlear duct near the spiral prominence. AQP5 expression continued through postnatal periods with a change of expression domain to the stria vascularis between postnatal day 7 (P7) and P14. By in situ hybridization and immunohistochemical techniques, subtle differences between transcript and protein expression patterns were noted for both AQP1 and 5. Although AQP5 is dynamically expressed in the developing mouse inner ear, adult Aqp5 knockout mice show normal hearing when tested and normal inner ear structural development. These results suggest redundant or alternative mechanisms that likely regulate water homeostasis in the developing and mature inner ear.