Volatile substance abuse--post-mortem diagnosis

A substantial number of children and adolescents world-wide abuse volatile substances with the intention to experience an euphoric state of consciousness. Although the ratio of deaths to nonfatal inhalation escapades is low, it is an important and preventable cause of death in young people. In the analytical investigation of volatile substances proper sample collection, storage and handling are important in view of the volatile nature of the compounds. Volatile organic compounds in post-mortem matrices such as blood, urine and tissues are generally determined by gas chromatography after extracting the compounds with methods such as static and dynamic headspace or even with pulse-heating and solvent extraction. In post-mortem cases, metabolites in urine seem less relevant, however, trichloroethanol and trichloroacetic acid were determined in several cases. When interpreting qualitative and quantitative results, researchers should be aware of false conclusions. The main reason why scepticism is necessary is the occurrence of losses of analytes during sampling, sample handling and storage, which results in false quantitation.     

Urinary ortho-cresol concentrations as an indicator of toluene inhalation in glue-sniffers.

In a Japanese study of glue-sniffers, urinary ortho-cresol and expired air toluene concentrations were determined by gas chromatography, and urinary hippuric acid concentrations by high performance liquid chromatography. There were significant differences among three sample groups (30 male "glue-sniffers", 13 "non-glue-sniffers" and 32 industrial workers) in their expired air toluene, urinary hippuric acid and ortho-cresol concentrations. Moreover, there was a better correlation between expired air toluene and urinary ortho-cresol concentrations than between toluene and urinary ortho-cresol concentrations than between toluene and hippuric acid. These results suggest that urinary ortho-cresol is a good indicator of toluene inhalation in glue-sniffers.     

Inhalant abuse detection and evaluation in young Tunisians

Occupational exposure biological monitoring techniques were applied for the diagnosis of inhalation abuse and for the evaluation of the levels of exposure to benzene, toluene, ethylbenzene, xylenes, and n-hexane, in 44 Tunisian adolescents and children suspected for volatile substance addiction. Urinary trans,trans-muconic acid, hippuric acid (HA), mandelic acid, and methylhippuric acids determinations were performed by high performance liquid chromatography with a photodiode array detector, and urinary o-cresol (o-Cr) and 2,5-hexanedione (HD) were extracted simultaneously and measured using gas chromatography with a flame ionization detector. Given the high linearity ranges, HD and o-Cr occupational exposure monitoring techniques could be applied without modification. However, urinary sample dilution was necessary before HA analysis. Concentrations were compared with the maxima of normal values (MNVs) in the general population and to the biological exposure indices (BEIs) used in occupational toxicology. Values as high as 6610-fold the MNV and 68 times the BEI were registered. The subjects showed high exposure to toluene and hexane. Measured metabolites HA and/or o-Cr and HD enabled the easy detection and evaluation of exposure levels. The problem of inhalant abuse should be given more attention and treated through an effective prevention strategy.     

Identification and quantitation of xenobiotics by 1H NMR spectroscopy in poisoning cases

In order to analyse a wide range of xenobiotics and their metabolites present in biological fluids, NMR spectroscopy can be used. A large variety of xenobiotics (therapeutic agents, pesticides, solvents, alcohols) can be characterized and quantitated directly, without sample preparation. NMR investigations were applied to acute poisoning cases, involving drugs such as salicylates and valproic acid (VPA). In a salicylate poisoning case, the three major metabolites of acetylsalicylic acid have been detected in crude urine, and rapid identification of lysine revealed the origin of the intoxication, namely lysine acetylsalicylate (Aspegic). Valproic acid as its glucuronide was identified in urine samples from two poisoned patients. 1H NMR was also used to identify and quantitate paraquat (Gramoxone) in urine owing to its two aromatic signals at 8.49 and 9.02 ppm, in two acutely poisoned patients (183 and 93 mg/l). An intentional poisoning case with tetrahydrofuran (THF) was also investigated. Serum and urine samples were collected. THF was characterized by its resonances at 1.90 and 3.76 ppm, and quantified at 813 and 850 mg/l in the two biological fluids, respectively. Moreover, two other compounds were detected: lactate and gamma-hydroxybutyric acid (GHB). 1H NMR spectroscopic analysis of serum samples from three poisoned patients revealed methanol in one case and ethylene glycol in the two others. Moreover, in the same spectrum, the corresponding metabolites formate and glycolate were found. Compared with the reference chromatographic or spectrophotometric methods, requiring time-consuming extraction and/or derivatization steps, NMR spectroscopy allows the determination of many exogenous and endogenous compounds, without any pre-selection of the analytes.     

Concentrations of toluene in the body killed by an injury to the head shortly after ingesting thinner

An autopsy was conducted on a male showing leather-like skin damage, revealing the cause of death to be an injury to the head. Thinner was found scattered around the scene of death, and stomach and intestine contents smelled strongly of solvent. Toxicological analysis was conducted to determine whether or not the solvent was of a lethal level. Using gas chromatography, peaks of toluene, xylene, and ethylbenzene were detected in the blood and gastric contents. No toluene was detected in the urine, and therefore it was concluded that the decedent died of a severe head injury shortly after solvent ingestion. In the literature, toluene concentrations in blood and lung samples were determined as both fatal and non-fatal but clear differences in the fatality of toluene in solid organ samples, namely, the brain, liver and kidneys were shown. The brain is especially useful in postmortem analysis. In this case, the concentration of toluene in the brain was 20.0 microl/g, which was considered as a non-lethal level.     

Hippuric acid and methyl hippuric acid in rat hair: possible monitoring of xylene and toluene exposure

Thinner is mainly composed of toluene and xylenes, and we studied the incorporation of the main metabolites of toluene and xylenes, hippuric acid (HA) and o-, m-, and p-methyl hippuric acids (o-, m-, p-MHA), in dark agouti rats' hair. Rat black hair was shaved before any exposure with an electric shaver designed for animals. Studies were performed in vivo with exposures of 30 min per day at three different concentrations (100, 300, and 1000 ppm) of toluene and o-, m-, and p-xylene for a total of 10 times over 2 weeks. Newly grown hair was tweezed out from the root with tweezers at seventh of the last exposure. Hair samples were then washed, extracted, derivatized, and analyzed by gas chromatography-mass spectrometry (GC-MS). HA and o-, m-, and p-MHA were not detected (ND) in the unexposed rat hair. After exposure, the metabolite concentration in the hair changed depending on the exposure concentration. Mean concentrations ranged from ND to 7.6 ng/mg, from ND to 13.8 ng/mg, from ND to 10.1 ng/mg, and from ND to 9.2 ng/ml hair for HA, o-, m-, and p-MHA, respectively. These results indicate that the metabolites concentrations in hair are effective indices of thinner exposure.     

Detection of cannabinoids in urine

Described in the paper are results of comparative examination of urine by the methods of immunochromatographic express analysis (ICA), fluorescence-polarization immune analysis (FIA) and of thin-layer chromatography (TLC) made for the purpose of detecting consumers of hemp products. A high specificity and a good sensitivity of the methods were demonstrated, which is a basis for using them as preliminary tests in detecting cannaboids and their metabolites in urine. The methods were evaluated quantitively and qualitatively versus the etalon method of chromato-mass-spectrometry. A possibility was pointed out to apply the methods of ICA and TLC as system analysis in the determination of 11-nordelta-9-tetrahydrocannabinol-9-carbonic acid in urine of marijuana users.     

The comparison of toluene determination between headspace-solid phase microextraction and headspace methods in glue-sniffer's blood and urine samples

An accurate and simple method was developed to determine the level of toluene in urine and blood quantitatively by using the gas chromatography/mass spectrometry (GC/MS) with headspace--solid phase microextraction (HS-SPME) technique. An assembly of SPME with a replaceable extraction fiber, coated with 100 microm polydimethylsiloxane, was used. The detection limit of toluene in blood and urine with HS-SPME technique was 10 times higher than that with headspace (HS) technique. To compare the HS-SPME with HS technique for the determination of toluene in biological fluids, blood and urine samples from glue sniffers were analyzed by both methods. The level of toluene by the two techniques was highly correlated: the correlation coefficient (r2) between the two sets of values were 0.98 and 0.96 in urine and blood, respectively.     

Comparison of extraction methods for methamphetamine and its metabolites in tissue

Five extraction methods were examined for analysis of methamphetamine and its major metabolites in tissue samples. The extraction methods studied were an acetone extraction method, an ethanol extraction method, an ammonium sulfate method, dialysis, and a direct solvent extraction. Acetone, ethanol, and dialysis methods showed no interference from endogenous components using thin-layer chromatography and gas chromatography, and gave satisfactory recovery of methamphetamine, amphetamine, and p-hydroxymethamphetamine when added to rabbit liver. These methods, however, proved time-consuming. The ammonium sulfate method and direct solvent extraction method were simple and more rapid, but recovery of the polar metabolite was poor.     

Determination of nimetazepam and 7-aminonimetazepam in human urine by using liquid chromatography–tandem mass spectrometry

We report determination of metabolites of popular drugs of abuse, including nimetazepam and nitrazepam, in urine by using liquid chromatography/mass spectrometry. Nimetazepam and its metabolites, 7-aminonimetazepam and nitrazepam, were extracted by solid-phase extraction using a DAU cartridge. An ammonium acetate buffer solution (pH 4) and a Luna polar-RP column were selected as the mobile and stationary phase, respectively, for liquid chromatography. Mass spectrometry was used for analysis and was optimized for operation in the positive mode for all analytes. The urine specimens were screened for the presence of nimetazepam and its metabolites nitrazepam and 7-aminonimetazepam at a concentration of 0.1 ng/mL. Presence of 7-aminonimetazepam in the urine was an indicator of the subject being a probable abuser of nimetazepam.     

甲基苯丙胺滥用与司来吉兰服用的区分:尿液中甲基苯丙胺和苯丙胺的手性分析

目的考察司来吉兰及其代谢物在尿液中的含量变化,并结合实际案例探讨手性分析区分甲基苯丙胺滥用与司来吉兰服用的可行性。方法采用CHIROBIOTICTM V2手性液相色谱柱对尿液样品进行手性分离和液相色谱-串联质谱(LC-MS/MS)法测定,并对司来吉兰服药志愿者尿样、疑服用司来吉兰的涉毒人员尿样进行甲基苯丙胺和苯丙胺的手性分析。结果服用5 mg司来吉兰后,尿液中司来吉兰的检出时限仅为7h。尿液中R(-)-甲基苯丙胺和R(-)-苯丙胺约在7h质量浓度最高,分别为0.86μg/m L和0.18μg/m L,并在80 h和168 h后无法检出。应用该方法成功分析了疑服用司来吉兰的涉毒人员尿液中甲基苯丙胺和苯丙胺的来源。结论甲基苯丙胺和苯丙胺的手性分析以及司来吉兰代谢物检测可区分甲基苯丙胺滥用与司来吉兰服用。     

尿中氯胺酮及其代谢物检测的研究

目的建立氯胺酮滥用者尿中氯胺酮及其代谢物检测方法。方法尿液用有机溶剂液-液萃取,气相色谱/氮磷检测器、电子捕获检测器、氢火焰检测器和气-质联用仪测定。结果确认了尿液中氯胺酮的主要代谢物,尿液中氯胺酮及去甲氯胺酮的最小检测限均为2ng/mL,脱氢去甲氯胺酮的最小检测限为5ng/mL。结论所建方法快速、灵敏、准确,能够满足氯胺酮滥用者尿液检测的需要。     

Synthesis and Analysis of Glucuronic Acid‐Conjugated Metabolites of 4‐Bromo‐2,5‐Dimethoxyphenethylamine

In the study reported here, two glucuronic acid‐conjugated metabolites of 4‐bromo‐2,5‐dimethoxyphenethylamine (2C‐B)—a ring‐substituted psychoactive phenethylamine—were chemically synthesized for the first time and a method for analyzing them in urine was developed. β‐D‐Glucuronide of 4‐bromo‐2,5‐dimethoxyphenylethylalcohol was successfully synthesized using methyl 2,3,4‐tri‐Ο‐acetyl‐1‐O‐(trichloroacetimidoyl)‐α‐D‐glucuronate as a glucuronyl donor and boron trifluoride diethylether complex as a Lewis acid catalyst. β‐D‐Glucuronide of 4‐bromo‐2,5‐dimethoxyphenylacetic acid was synthesized by condensing 4‐bromo‐2,5‐dimethoxyphenylacetic acid and benzyl D‐glucuronate followed by benzyl group deprotection based on catalytic hydrogenation. Two glucuronic acid‐conjugated metabolites of 2C‐B in urine were qualitatively and semiquantitatively evaluated via direct liquid chromatography/mass spectrometry (LC/MS) analysis of a diluted urine sample. The simple method proposed is expected to be useful for studying the metabolic fate of 2C‐B.     

Occurrence of contamination by controlled substances in Euro banknotes from the Spanish archipelago of the Canary Islands

The social problems of drug abuse are a matter of increasing global problem. Nowadays, international agencies need fresh methods to monitor trends of the use of illicit drugs. In this sense, small amounts of drugs are transferred to banknotes and they could be detected and quantified. An analytical procedure based upon extraction with organic solvent, liquid chromatography separation, and mass spectrometric detection allowed the identification of 21 drugs and metabolites in 120 used Euro banknotes collected in the Canary Islands (Spain). Most of the banknotes analyzed showed detectable drug residues (92.5%). Cocaine was the most frequently detected drug, present in approximately 90% of the samples. In addition, 75%, 35%, and 15% of the banknotes showed residues of amphetamine derivatives, opiates, and benzodiazepines, respectively. An average of three drug residues per banknote was detected. In summary, the presence of drug residues in banknotes could be useful as tracer for drugs prevalence.     

Hair and urine analysis: relative distribution of drugs and their metabolites

This work studies the distribution of cocaine and heroin metabolites in hair and urine of living polidrug abusers. Cocaine, benzoylecgonine (BEG), ecgonine methyl ester (EME), morphine, codeine and 6-monoacetylmorphine (6-MAM) were simultaneously extracted and analyzed by GC/MS in SIM mode. The results obtained show a different distribution of heroin and cocaine metabolites in urine and hair. In urine, we generally find BEG and EME for cocaine abuse, and morphine for heroin abuse. In hair, we detect cocaine and MAM as major metabolites for cocaine and heroin abuse, respectively.     

尿中地西泮及其代谢物的气相色谱电子捕获检测法分析

目的　建立尿中地西泮及其代谢物替马西泮、去甲西泮和奥沙西泮的GC/ECD分析方法。方法　尿样经β 葡萄糖醛酸酶水解后 ,用有机溶剂提取 ,再以N ,O 双三甲硅烷基三氟乙酰胺进行衍生化 ,衍生物用GC/ECD法分析。结果　检材中分析物的回收率 70 %以上 ,检出限 5ng/ml以下。结论　本法可进行口服地西泮 10mg人体 48h内尿液中地西泮代谢物的分析。     

Measurement and analysis of diastereomer ratios for forensic characterization of brodifacoum

The highly toxic anticoagulant rodenticide brodifacoum is an organic compound that has two diastereomeric forms. In this paper, we consider the hypothesis that the relative population of the diastereomers is a characteristic of forensic value for the association or source attribution of specimens of brodifacoum. In general, the stereoisomer distribution in an organic compound depends on the reagents, conditions, and methods used for synthesis and purification, and may vary over time due to differential stabilities of the stereoisomers. The stereoisomer distribution may thus serve as an identifier of the production methods and history of samples and provide a basis for comparing recovered specimens. We refer to this novel approach for signature detection as stereoisomer distribution analysis or SDA. If the stereoisomers are diastereomers, quantitative determination of the diastereomer ratio in a specimen can be performed by a number of techniques, notably gas or liquid chromatography or nuclear magnetic resonance (NMR) spectroscopy. This paper describes an NMR spectroscopic analysis of ten commercial technical grade brodifacoum samples from distinct batches originating from three different sources. The results reveal detectable source-to-source and batch-to-batch variations in diastereomer ratios.     

生物检材中氯胺酮及其代谢物的检测

近年来氯胺酮的滥用越来越普遍,建立快速、准确的检测方法越来越重要。氯胺酮在生物体内的代谢物主要有去甲氯胺酮、脱氢去甲氯胺酮等。目前,常用的生物检材有血液、尿液、毛发等。常用的检测方法有气相色谱法、气相色谱-质谱联用法、高效液相色谱法、液相色谱-质谱联用法、高效毛细管电泳法等。本文参考近年来的相关文献对生物检材中氯胺酮及其代谢物的检测方法作一综述,为法医毒物分析等相关领域提供参考。     

Application of solvent microextraction to the analysis of amphetamines and phencyclidine in urine

A fast and simple method to detect some commonly abused illicit drugs, amphetamine, methamphetamine, 3,4-methylendioxy-amphetamine (MDA), 3,4-methylendioxy-methamphetamine (MDMA), 3,4-methylendioxy-N-ethylamphetamine (MDEA) and phencyclidine (PCP) in urine using solvent microextraction (SME) combined with gas chromatography (GC) analysis has been developed. The extraction is conducted by suspending a 2 microl drop of chloroform in a 2 ml urine sample. Following 8 min of extraction, the organic solvent is withdrawn into the syringe and injected into a GC with a pulsed discharge helium ionization detector (PDHID). The effects of different extraction solvents and times, pH and sample preparation were studied. The optimized method was capable of detecting drugs in urine at concentrations below Substance Abuse and Mental Health Services Administration (SAMHSA) established cut-off values for preliminary testing. Good linearity and reproducibility of extraction were obtained. The limits of detection were 0.5 microg/ml for amphetamine, 0.1 microg/ml for methamphetamine and MDA, 0.05 microg/ml for MDMA, 0.025 microg/ml for MDEA and 0.015 microg/ml for PCP. Relative standard deviation (R.S.D.) values ranged between 5 and 20% for the studied drugs.     

Use of SPE and LC/TIS/MS/MS for rapid detection and quantitation of ketamine and its metabolite, norketamine, in urine

Ketamine (K) has become more and more popular for drug abuse in recent years. A lot of pre-treatment work such as extraction and derivatizing increase difficulties in the tests for ketamine in biological specimens. A rapid method to detect and quantitate ketamine and its metabolite norketamine in urine used deuterated dilution followed by solid phase extraction and liquid chromatography/TurboIonSpray/tandem mass spectrometry (LC/TIS/MS/MS) is described. Control recovery for both low and high concentrations can reach to 90%. Ten ketamine positive urines were examinated by this method. Concentrations ranged from 114 to 2925 ng/mL and from 453 to 9805 ng/mL for norketamine. The method was sensitive, specific, accurate and provided easy operation to detect and quantitate ketamine and its metabolites in urine.