Headspace solid phase microextraction for the gas chromatographic analysis of methyl-parathion in post-mortem human samples. Application in a suicide case by intravenous injection

A simple and rapid procedure for the determination of methyl-parathion (m-p) in post-mortem biological samples was developed using headspace solid phase microextraction (SPME) and gas chromatography (GC) with nitrogen-phosphorous detection (NPD). Methyl-parathion was extracted on 85 microm polyacrylate SPME fiber. Salt addition, extraction temperature, and extraction time were optimized to enhance the sensitivity of the method. The linearity (y = 0.0473x - 0.0113, R2 = 0.9992) and the dynamic range (0.1-40 microg/ml) were found very satisfactory. The recoveries of methyl-parathion were found to be 46% in spiked human whole blood, 53% in spiked homogenized liver tissue, and 54% in spiked homogenized kidney tissue compared with samples prepared in water. The coefficients of variations for 2, 4, and 20 microg/ml of methyl-parathion in blood ranged from 0.9 to 5.1%, whereas the detection limit of the method was satisfactory (1 ng/ml in aqueous samples, 50 ng/ml in whole blood). The developed procedure was applied to post-mortem biological samples from a 21-year-old woman fatally poisoned (suicide) by intravenous injection of methyl-parathion. The intact insecticide was found in the post-mortem blood at a concentration of 24 microg/ml. No methyl-parathion was detected in the liver, kidneys, and gastric contents.     

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.     

固相微萃取结合气质联用测定血浆中敌草快

目的采用固相微萃取结合气相色谱-质谱联用方法快速测定血浆中的敌草快含量。方法采用硼氢化钠/氯化镍还原反应和顶空固相微萃取处理样本,气质联用检测方法,以乙基百草枯为内标,检测血浆中敌草快的含量,对还原反应和萃取的温度、时间等实验条件进行优化,并对方法学进行评价。结果采用m/z 194作为定性定量离子,在0.1~50μg/mL浓度范围内线性关系良好（Y=0.076 9X＋0.274 8,r2=0.997 5）,日内和日间精密度分别小于3.99%、5.64%。回收率为92.59%~101.27%,最低定量限为50ng/mL。敌草快还原产物1,1’-乙基-2,2’-联哌啶存在顺式和反式两种同分异构体,两者比率为1∶5.3,相对标准偏差为13.1%（n=17）。结论本文方法准确、快速、重现性好,可以在血浆等生物样品敌草快含量测定中选用。     

Determination and distribution of clotiapine (Entumine) in human plasma, post-mortem blood and tissue samples from clotiapine-treated patients and from autopsy cases

The antipsychotic drug clotiapine (Entumine^®) has been marketed for more than 35 years, however there is little published data on the therapeutic and toxic concentrations of this drug. To fill this gap, two rapid and sensitive methods were developed for the determination of clotiapine (2-chloro-11-(4-methyl-1-piperazinyl)dibenzo-[b,f][1,4]-thiazepine), in human plasma and post-mortem blood and tissue samples. After simple liquid–liquid extraction at pH 9.5 with n-hexane/dichloromethane (85/15, v/v), clotiapine was quantitated by HPLC-DAD and by GC-NPD. The calibration curve was linear between 10 and 1000 μg/L. The limit of detection (LOD) and the limit of quantification (LOQ) were found to be 2 and 6 μg/L for the GC-NPD method and 5 and 15 μg/L for the HPLC-method, respectively. These methods were applied to 12 plasma samples from patients treated with clotiapine, to seven autopsy cases and to one case of driving under the influence of drugs (DUID). Concentrations ranged for the clotiapine-treated patients between 6 and 155 μg/L (mean 46 μg/L), and for the autopsy cases between 22 and 341 μg/L (mean 123 μg/L).     

HS-SPME-GC/MS联用法测定生物检材中的百草枯

目的建立检测生物检材中百草枯的顶空固相微萃取-气相色谱-质谱联用（HS-SPME-GC/MS）的分析方法。方法尿样中加乙基百草枯作为内标,在氯化镍作催化剂的条件下,用硼氢化钠在碱性条件下进行还原,HS-SPME萃取,提取物经GC/MS分析。全血需先离心,沉淀血细胞提取上清液,再用甲醇沉淀蛋白。最终得到的上清液加内标乙基百草枯,以下操作同尿样。结果尿样和血样中的百草枯的还原产物在1.0μg/mL~100μg/mL范围内线性关系良好,回归方程分别为y=0.0957x-0.0163,r=0.9974（n=6）;y=0.1096x＋0.0871,r=0.9964（n=6）。尿样、血样低、中、高三个质量浓度,RSD值均小于7%。回收率分别为尿样85.49%~100.83%,血样94.72%~99.68%。结论本法操作简便易行、灵敏度高、快速准确。为检测生物检材中的百草枯提供了有效的方法。     

Postmortem tissue concentrations of venlafaxine

Venlafaxine is a phenethylamine antidepressant which inhibits both serotonin and norepinephrine reuptake and is structurally unrelated to the serotonin reuptake inhibitors (SSRIs). Its major metabolite, O-desmethylvenlafaxine (ODV), also inhibits serotonin reuptake. Although metabolized by the cytochrome P-450 (CYP) system, venlafaxine inhibits CYP 2D6 and 3A4 to a far lesser extent than do the SSRIs. Mechanisms of drug action are reviewed and evaluated in the investigation of 12 fatalities occurring over a 6-month-period where venlafaxine was detected.Venlafaxine and ODV were identified by liquid chromatography-mass spectrometry (LC-MS) using atmospheric pressure ionization (API) electrospray in positive mode following an n-butyl chloride extraction. Postmortem tissue concentrations studied in each of 12 postmortem cases for venlafaxine and ODV, were 0.1-36 and <0.05-3.5mg/l (peripheral blood), <0.05-22 and <0.05-9.9mg/kg (liver), <0.05-10 and <0.05-1.5mg/l (vitreous), <0.05-53 and <0.05-6.8mg/l (bile), <0.05-55 and <0.05-21mg/l (urine), respectively, and 0.1-200mg of venlafaxine in the gastric contents. Venlafaxine was typically present with other drugs, including other antidepressants, alcohol, and benzodiazepines. The potential for interaction with each drug is discussed. Over the 6-month-period of this study, there were no deaths ascribed solely to venlafaxine intoxication.     

Headspace solid-phase microextraction (HS-SPME) and liquid-liquid extraction (LLE): comparison of the performance in classification of ecstasy tablets. Part 2

Headspace solid-phase microextraction (HS-SPME) is assessed as an alternative to liquid-liquid extraction (LLE) currently used for 3,4-methylenedioxymethampethamine (MDMA) profiling. Both methods were compared evaluating their performance in discriminating and classifying samples. For this purpose 62 different seizures were analysed using both extraction techniques followed by gas chromatography-mass spectroscopy (GC-MS). A previously validated method provided data for HS-SPME, whereas LLE data were collected applying a harmonized methodology developed and used in the European project CHAMP. After suitable pre-treatment, similarities between sample pairs were studied using the Pearson correlation. Both methods enable to distinguish between samples coming from the same pre-tabletting batches and samples coming from different pre-tabletting batches. This finding emphasizes the use of HS-SPME as an effective alternative to LLE, with additional advantages such as sample preparation and a solvent-free process.     

Forensic intoxication with clobazam: HPLC/DAD/MSD analysis

Clobazam (Castillium, Urbanil), a benzodiazepine often used as an anxiolytic and in the treatment of epilepsy, is considered a relatively safe drug. The authors present a fatal case with a 49-year-old female, found dead at home. She had been undergoing psychiatric treatment and was a chronic alcoholic. The autopsy findings were unremarkable, except for multivisceral congestion, steatosis and a small piece of a plastic blister pack in the stomach. Bronchopneumonia, bronchitis and bronchiolitis were also diagnosed. Anhigh-performance liquid chromatography (HPLC)/diode array detector (DAD)/mass spectrometry detection (MSD) with electrospray method was developed in order to detect, confirm and quantify clobazam in the post-mortem samples. In the chromatographic separation, a reversed-phase column C18 (2.1 x 150 mm, 3.5 microm) was used with a mobile phase of methanol and water, at a 0.25 ml/min flow rate. Carbonate buffer (pH 10.5) and 20 microl of prazepam (100 microg/ml) as internal standard were added to the samples. A simple and reliable liquid-liquid extraction method for the determination of clobazam in post-mortem samples was described. Calibration curves for clobazam were performed in blood, achieving linearity between 0.01 and 10 microg/ml and a detection limit of 1.0 ng/ml. The clobazam concentration found in post-mortem blood was 3.9 microg/ml, higher than the reported therapeutic concentration (0.1-0.4 microg/ml). The simultaneous acquisition by photodiode array detection and mass spectrometry detection results allowed benzodiazepines to be identified with sufficient certainty. An examination of all the available information suggested that death resulted from respiratory depression due to clobazam toxicity.     

Automated headspace solid-phase microextraction and capillary gas chromatography analysis of ethanol in postmortem specimens

Solid-phase microextraction (SPME) is a relatively new solventless sample preparation technique that allows simultaneous sampling, extraction, pre-concentration, and introduction of analytes from a sample matrix in a single procedure. This methodology has been used for the analysis of several drugs of forensic toxicology interest including volatile compounds. This paper describes a methodology for analysis of ethanol and other volatile compounds using automatic headspace solid-phase microextraction (HS-SPME) and capillary gas chromatography in postmortem specimens. The methodology was initially developed using standard solutions of acetaldehyde, acetone, methanol, and ethanol. Isobutanol was used as internal standard. Postmortem samples of blood, urine, and vitreous humor were obtained during medico-legal autopsies. To date, there are no published paper regarding alcohol analysis in vitreous humor specimens using HS-SPME and limited literature analyzing blood and urine samples. HS-SPME analysis showed that, under optimized conditions, ethanol and isobutanol (internal standard) were well-separated from other volatile compounds such as acetaldehyde, acetone, and methanol considered to be potential interferents in ethanol analysis. The calibration curves for each volatile compound demonstrated good linearity throughout the concentration range from 0.001 to 1.0 g/dl and the detection limit of ethanol in the studied specimens was approximately 0.0001 g/dl.     

A mixed-drug intoxication involving venlafaxine and verapamil

This case report describes the suicide of a 52-year-old woman whose cause of death was attributed to a mixed-drug intoxication involving venlafaxine and verapamil. Venlafaxine is prescribed for the treatment of depression and should be used with caution in patients with cardiovascular disease. Verapamil is a calcium channel blocker primarily used for treatment of cardiovascular disorders. The following drug concentrations were determined in postmortem fluids: verapamil--3.5 mg/L (femoral blood), 9.4 mg/L (subclavian blood), and 1.0 mg/L (vitreous fluid); norverapamil--1.0 mg/L (femoral blood), 2.1 mg/L (subclavian blood), and 0.20 mg/L (vitreous fluid); verapamil and norverapamil could not be detected in bile or urine due to the high levels of erythromycin present; venlafaxine--6.2 mg/L (femoral blood), 8.6 mg/L (subclavian blood), 5.3 mg/L (vitreous fluid), 54.0 mg/L (bile), and 72.3 mg/L (urine); and O-desmethylvenlafaxine--5.4 mg/L (femoral blood), 8.3 mg/L (subclavian blood), positive (vitreous fluid), 29.2 mg/L (bile), and 9.5 mg/L (urine). The cause of death was determined to be a mixed-drug intoxication resulting from an overdose of verapamil and venlafaxine. The manner of death was determined to be suicide.     

生物检材中阿维菌素的HPLC—MS／MS分析

目的建立生物检材包括血、肝组织、胃组织中阿维菌素（Avermectins）的HPLC—MS／MS分析方法。方法采用Oasis HLB固相萃取柱进行提取,以XTerra^TM RP18柱（2．1mm×100mm,3．5μm）色谱柱分离,以甲醇-0．1％冰醋酸水溶液（75：25）为流动相,流速为0．2mL／min。结果线性范围10ng／mL～3μg／mL,最小检出限为0．1ng／mL。结论本方法准确、快速,可用于生物检材血中阿维菌素的定性定量分析,肝及胃组织中阿维菌素的定性分析。     

植物酶在解剖组织中有机磷、氨基甲酸酯农药检测中的应用

本文报导了有机磷和氨基甲酸酯检测的一个新的酶抑制法.该法基于有机磷和氨基甲酸酯化合物对植物酶水解底物2,6-二氯乙酰靛酚的抑制作用.本法简单,快速,适于农药中毒的现场检验.其最小检出量分别为0.2～1μg/μ1和1～2μg/ml.     

固相萃取──紫外导数光谱法测定脏器中敌鼠钠含量

肝、肾、肺、脑等脏器中敌鼠钠含量的题示测定法，使用国产GDX403树脂为革取材料，溶剂消耗少，操作简便，回收率约70％，检测下限小于1μg．g-1，适合做为法医毒物分析常规方法。     

HPLC／MS检测生物样品中的丁丙诺啡

目的建立了生物样品中丁丙诺啡的高效液相色谱-电喷雾串联质谱检测方法。方法样品经固相萃取提取净化、反相液相色谱分离后进行质谱检测，根据保留时间及特征离子进行定性分析，以母离子m／z468进行定量分析。结果在10-500ng／ml（ng／g）范围内峰面积与质量浓度的线性关系良好（r^2〉0．993）。在50、100、500ng／ml（ng／g）3个添加水平，尿、血、肝中丁丙诺啡的平均回收率为74％～94％，日内测定结果的相对标准偏差小于8％，日间测定结果的相对标准偏差小于10％。结论该方法简单、灵敏，特异性强，适用于生物样品中丁丙诺啡的分析检验。     

生物检材中吗啡类生物碱的LC-MS／MS分析

目的针对滥用药物分析鉴定实践中亟待解决的问题,开展LC-MS/MS分析生物检材中吗啡类生物碱的应用研究。方法满足不同的鉴定需要,分别建立血液、尿液、唾液和头发等生物检材的样品前处理方法,确定同时分析海洛因、单乙酰吗啡、吗啡、可待因、乙酰可待因、二氢可待因酮和氢吗啡酮等吗啡类生物碱的LC-MS/MS方法。将方法应用于实际案例。结果所建立的方法对吗啡类生物碱分离良好。尿液稀释法、尿液提取法和头发中吗啡的最低检测限(LOD)分别为10ng/mL、0.01ng/mL和0.01ng/mg。结论所建立的方法简便、快速、特异性强、灵敏度高。目标物中加入二氢可待因酮和氢吗啡酮扩大了方法的实用范围。     

QuEChERS-LC/MS快速检测兽用麻药“舒泰”

目的本文对兽药"舒泰"中有效成分进行了结构确证,并建立了生物检材中替来他明和唑拉西泮的快速检验方法。方法在血液和尿液的生物检材中,通过加标实验,经QuEChERS萃取后,进行LC/MS对替来他明和唑拉西泮的定性定量检测分析。结果在血液和尿液的生物样品的加标实验中,替来他明的RSD%在0.5%~3.5%,唑拉西泮的RSD在0.5%~1.1%;替来他明的回收率在75.8%~100.3%,唑拉西泮的回收率在68.8%~76.6%,其中血液中替来他明的方法检出限为0.16ng/mL,尿液中为0.20ng/mL,唑拉西泮在血液中的方法检出限0.17ng/mL,尿液中为0.22ng/mL。结论建立的QuEChERS萃取方法,操作流程简便,方法重现性好,只需100μL取样量,更适合于痕量生物检材中替来他明和唑拉西泮的检验分析。     

生物检材中甲基苯丙胺及苯丙胺的分析研究进展

对近几年国内外22篇有关生物检材中甲基苯丙胺及苯丙胺测定的文献进行了综述。介绍了血、尿、毛发等生物检材的收集与预处理方法,比较了生物检材中甲基苯丙胺及苯丙胺的液-液萃取(LLE)、固相萃取(SPE)、固相微萃取(SPME)和顶空固相微萃取(HS-SPME)等提取方法,以及内标的选取、不同的衍生化方法和包括免疫、GC/MS、GC/NPD、GC/ECD、GC/FID、HPLC、HPCE在内的各种检测方法。最后,对分析结果的评定进行了讨论。     

Use of headspace solid-phase microextraction (HS-SPME) in hair analysis for organic compounds

Headspace solid phase microextraction (HS-SPME) has advantages of high purity of the extract, avoidance of organic solvents and simple technical manipulation and can be used in combination with gas chromatography-mass spectrometry (GC-MS) in the hair analysis of a number of drugs. HS-SPME coupled with the hydrolysis of the hair matrix by 4% sodium hydroxide in the presence of excess sodium sulphate and of a suitable internal standard proved to be a convenient one-step method for the measurement of many lipophilic basic drugs such as nicotine, amphetamine derivatives, local anaesthetics, phencyclidine, ketamine, methadone, diphenhydramine, tramadol, tricyclic antidepressants and phenothiazines. Detection limits were between 0.05 and 1.0 ng/mg. From spiked 10-mg hair samples absolute recoveries between 0.04 and 5.7% were found. These recoveries decreased considerably if larger sample amounts were used, perhaps due to increased drug solubility in the aqueous phase or to elevated viscosity in the presence of dissolved hair proteins. Because of the phenolic hydroxyl group a change of pH after alkaline hair digestion (by adding excess orthophosphoric acid) was necessary for the detection of delta 9-tetrahydrocannabinol (delta 9-THC), cannabinol (CBN) and cannabidiol (CBD) by HS-SPME. Nevertheless, the detection limits were such that only CBN could be detected in hair of a consumer. Clomethiazole, a compound hydrolysed in alkali, was measured by HS-SPME after extraction with aqueous buffer. The detection limit was 0.5 ng/mg. Cocaine could not be detected by HS-SPME. The application of HS-SPME to hair samples from several forensic and clinical cases is described.     

Quantitative determination of strychnine in blood and urine by gas chromatography with mass-selective detector

A method for the quantitative determination of strychnine in biological fluids by gas chromatography--mass spectrometry is proposed. The preparation of samples for the analysis included extraction of strychnine from blood and urine with the use of AccuBond(II) EVIDEX cartridges for solid-phase extraction and SPEC MP3 disks respectively. The efficiency of extraction was estimated at 0.05 mg/l for blood and 0.02 mg/l for urine. The detection limit was 0.10 mg/l in blood and 0.05 mg/l in urine.     

Colchicine Extract Suicidal Lethal Poisoning Confirmation Using High‐Resolution Accurate Mass Spectrometry: A Case Study

A case of suspected acute and lethal intoxication caused by colchicine has been reported. The woman was hospitalized after her suspicion of suicidal poisoning by a rare autumn crocus (Colchicum autumnale). Suspected colchicine poisoning was confirmed using a novel UHPLC method with a modern reversed‐phase stationary phase with a sub 2‐micron superficial porous particle size combined with a QTOF mass spectrometer. Sample preparation procedure included the addition of propiverine as internal standard, protein precipitation using methanol and solid phase extraction. High‐resolution MS only and targeted MS/MS modes are reported for the qualitative analysis and screening of other potential drugs of abuse in blood samples. All Ion MS mode was used for quantitative determination of colchicine afterward. The concentration of colchicine in the blood sample was approximately 41 ng/mL, and more than 200 μg/mL of the plant extract used for the suicide.