Fatal Datura poisoning: identification of atropine and scopolamine by high performance liquid chromatography/photodiode array/mass spectrometry

A forensic method comprising solid phase extraction and HPLC analysis was developed for the detection and confirmation of atropine and scopolamine, the main toxic alkaloids of Datura stramonium and Datura ferox. This method allowed the direct coupling of an electrospray (ZMD) mass selective detector to the HPLC system. Under these conditions, atropine and scopolamine were well separated from other components and detected on the PDA (LOD = 1 microg/ml) and ZMD (LOD(atropine) = 10 pg/ml; LOD(scopolamine) = 100 pg/ml) detectors. Four geographically isolated populations of each of D. stramonium and D. ferox were analysed for seed alkaloids and it was found that the two species were diagnostically different in their atropine-scopolamine ratios. The optimised HPLC method was used to analyse three viscera samples of an adult Caucasian male whose death was ascribed to a fatal heart attack. Atropine and scopolamine were detected in the stomach and its contents, which contained Datura seeds. The chemical profile of the seeds found in the stomach contents was similar to those from four geographically different D. ferox plants.     

Another death due to ingestion of Nicotiana glauca

Deaths attributed to ingestion of Nicotiana glauca are extremely rare. We report here a case where a 43-year-old man was found dead after apparently drinking a water extract of Nicotiana glauca. The primary alkaloid in the plant is anabasine. Toxicological analysis by capillary gas chromatography showed the deceased had a blood anabasine concentration of 2.2 mg/L. Clinically, the features of poisoning are nicotine-like and if death occurs it results from respiratory paralysis. The case further supports the view that, in the human, anabasine is considerably more toxic than nicotine.     

高效液相色谱法测定血浆中雷公藤甲素和雷公藤酮

目的　建立高效液相色谱法定量检测人血浆中雷公藤甲素和雷公藤酮的分析方法。方法　以Oasis HLB固相萃取柱对样品进行提取 ,应用HPLC色谱法二极管阵列检测器测定。结果　该方法的回收率高于 80 % ,线性范围在 10～ 10 0 0ng/ml ,经该方法测得雷公藤甲素的最小检出限为 3 0ng/ml,雷公藤酮的最小检出限为 4 5ng/ml(S/N≥ 3 )。结论　该方法快速灵敏、准确 ,适用于雷公藤中毒的法医学检验。     

LC／MS／MS法测定生物组织中百草枯

目的建立LC／MS／MS检测生物体液中百草枯方法。方法弱阳离子交换固相萃取小柱提取剂，应用LC／MS／MS法对生物样品中百草枯进行定性定量分析。结果经该方法测得百草枯的最小检出限为10ng／ml血（S／N≥3），线性范围为0．02～20μg／ml。结论该方法快速、灵敏、准确，适用于生物检材中百草枯的分析。     

高效液相色谱法测定生物体液中百草枯

目的建立HPLC检测生物体液中百草枯方法。方法弱阳离子交换固相萃取小柱提取,水溶性正相液相色谱分析,结合保留时间和紫外光谱对样品中百草枯进行定性。结果经该方法测得百草枯的最小检出限为10ng/m l血(S/N≥3),线性范围为0.1~10μg/ml。结论该方法快速、灵敏、准确,能有效祛除复杂生物基质中杂质干扰,使待测组分获得满意的峰形,适用于生物检材中百草枯的分析。     

LC／MS／MS法测定全血中马钱子碱和士的宁的含量

目的建立定量分析人全血中马钱子碱和士的宁的高效液相色谱质谱联用法。方法应用Oasis^TM MCX小柱进行固相萃取法提取，XTerra^TM RP18色谱柱分离。结果在该条件下，人血中马钱子碱和士的宁的线性范围为0．01—5．0μg／ml，最小检出限为0．2ng／ml；马钱子碱和士的宁在0．01—5．0μg／ml浓度范围内的回收率均在80％以上。结论高效液相色谱质谱联用法可定量测定血中马钱子碱和士的宁。     

固相萃取HPLC/MS法测定血中氯硝安定

目的建立测定人血中氯硝安定的HPLC/MS定性、定量方法。方法在血液中添加氯硝安定,采用固相萃取、HPLC法分离、LC/MS法定性、定量。结果标准氯硝安定LC/MS法的工作曲线在0.5~500ng时为Y=0.655×103X+2.15×103,相关系数r=0.9981,最小检出限为0.5ng/m l。结论采用pH8.0的磷酸盐缓冲液进行稀释、C18柱固相萃取氯硝安定回收率较高,优化后固定的LC/MS检测方法简便、快速、准确,适合办案需要。     

人血、尿中富马酸喹硫平的气相色谱分析

目的建立人血、尿中富马酸喹硫平的气相色谱分析方法。方法用乙醚提取血、尿中的富马酸喹硫平，直接对其进行定性、定量分析。以正常人血、尿为空白样本，分别添加标准富马酸喹硫平，确定检材的前处理方法、色谱分析条件、工作曲线、线性范围、方法的精密度、回收率等，并对1例大剂量服用富马酸喹硫平中毒死者的体液浓度进行测定。结果该方法分析血、尿中富马酸喹硫平的线性范围分别为8．0～800．0μg／ml和20．0—800．0μg/ml；最低检测限分别为0．04μg／ml和0．10μg／ml（S／N≥3），日内、日间精密度均小于4％，回收率在97．08％-101．42％之间。结论该分析方法操作便捷、实用、准确度高，适用于富马酸喹硫平的临床血药浓度快速监测和法医毒物鉴定。     

Comparative analysis of gamma-hydroxybutyrate and gamma-hydroxyvalerate using GC/MS and HPLC

This paper describes two analytical techniques used to separate and quantify gamma-hydroxybutyrate (GHB) and gamma-hydroxyvalerate (GHV). The first technique was a N,O-bis(trimethylsilyl)triflouro-acetimide-trimethylchlorosilane derivatization, followed by gas chromatography/mass spectrometry analysis using an HP-5 capillary column at a rate of 1.0 mL/min with a run time of 9.25 min. This technique was found to be sensitive (LOD 1 pg on column) and gave a low average error (5%) in a beverage study. When supplemented by a surrogate spike, the method yielded 97% analyte recovery from beverages. The second technique was high-performance liquid chromatography/UV (HPLC/UV) using a C-18 column with a (20:80% v/v) methanol:dibasic phosphoric buffer (10 mM, pH 3) at a rate of 1.00 mL/min with a run time of 7.5 min. UV detection occurred at 254 nm. This method was found to be less sensitive (LOD 0.05 microg on column) for direct analysis of aqueous samples. To remove interferences seen in the beverage study, a liquid-liquid extraction before HPLC analysis was tested. However, a decreased sensitivity (LOD 100 microg on column) and irreproducible peak profiles resulted.     

头发中5-MeO-DiPT等12种新型色胺类致幻剂的UPLC-MS/MS分析方法研究

目的建立同时分析头发中5-MeO-DiPT等12种新型色胺类致幻剂的超高效液相色谱-串联质谱(UPLC-MS/MS)方法。方法以赛洛西宾-d4和赛洛新-d10为内标,20 mg头发样品加入1 mL提取液(含内标2 ng/mL的0.1%甲酸水溶液)后冷冻研磨,离心取上清液,经Waters Acquity^TM UPLC HSS T3色谱柱分离,采用多反应监测模式同时测定12种新型色胺类致幻剂。结果头发样品中12种色胺类新精神活性物质的检出限为1~10pg/mg,定量限为3~50pg/mg。在相应浓度范围内,12种色胺类新精神活性物质均具有良好的线性关系,相关系数大于0.99。本方法准确度为91.3%~113.5%,日间和日内精密度(RSD)均小于15%,回收率大于80%,无明显基质效应。结论该方法前处理简单、灵敏度高、选择性好,适用于法医毒物分析中新型色胺类致幻剂的鉴定。     

Fatal poisoning from Nicotiana glauca leaves: identification of anabasine by gas-chromatography/mass spectrometry

Death of a worker occurred after ingestion of unknown amounts of Nicotiana glauca G leaves. The leaves were cooked after having been mistakenly considered to be spices of a type which grow in Thailand. After ingestion, two Thai workers collapsed, one with asystolia. Resuscitation efforts were successful only for one of the victims. A GC/MS method was used for the identification of anabasine as the main constituent in the leaves, food extract, blood, and the urine of the deceased. Lacking a standard, it was necessary to interpret the GC/MS spectrum to identify anabasine and establish its presence.     

Liquid chromatography/photodiode array detection for determination of strychnine in blood: a fatal case report

An original liquid chromatography method with photodiode-array detection (DAD) is presented for the determination of strychnine in blood. This sensitive method allows the use of only 0.1 ml of sample. The strychnine was isolated from blood using a liquid-liquid extraction procedure and chloroquine as an internal standard. The limits of detection (LOD) and quantification were 0.06 and 0.5 mg/l, respectively. The recovery was 94% and the coefficients of variation (CV) ranged from 5.9 to 10.8%. A fatal case of strychnine poisoning is presented, with a lethal blood concentration of 25 mg/l.     

Determination of phenol and o-cresol by GC/MS in a fatal poisoning case

A fatality due to the ingestion of solution containing phenol and o-cresol is described. The pathological findings were typical of acute substantial poisoning. Blood, urine and stomach content were obtained during post mortem examinations. Phenol and o-cresol were identified using GC/MS. The extractions from autopsy materials were obtained as follows: by gel permeation with cyclohexane/dichloromethane from stomach content, by solid phase extraction (SPE) from urine and by deproteinization with acetonitrile from blood. The phenol and o-cresol concentrations in the samples were found, respectively, as follows: 115.0 and 5.0 microg/g in the stomach contents, 58.3 and 1.9 microg/ml in the blood, 3.3 and 20.5 microg/ml in the urine. Distributions of phenol in fatal poisonings have been reported, but, usually, colorimetry was used as the analytical method and it cannot exclude the interference of other phenolic compounds.     

A rapid method for the extraction of cocaine and benzoylecgonine from body fluids

A method has been developed for the rapid extraction of cocaine and benzoylecgonine from post-mortem blood and urine samples. Solid phase columns containing C8 packing material gave clean, emulsion free extracts from body fluids. The total time for each extraction was approx. 10 min and the system has the capacity for the simultaneous extraction of up to 10 samples. Urine can be extracted directly, blood samples were sonicated before extraction to allow for their easy passage through the columns. The method gave excellent recoveries of cocaine (98-100%) from spiked samples at concentrations of between 50 ng/ml and 10 micrograms/ml. Analysis of the extracts was by high performance liquid chromatography (HPLC).     

血中毒鼠强的固相萃取和GC-NPD法测定

目的 研究人全血中毒鼠强的固相革取(SPE)。方法 用Bond Elute C18固相萃取柱萃取,GC-NPD检测,以甲基对硫磷为色谱内标(CS)。结果 全血中加标0.5μg/ml,毒鼠强回收率为92.5％,变异系数2.3％(CV,n=4)。在0.5～10μg/ml的浓度范围内,线性相关系数为0.9994。检出限(S/N=3)和定量限(S/N=10)分别为6ng/ml和20ng/ml。同一根萃取柱连续使用6次未见性能明显下降(CV=3.6%)。结论 本文方法适用于毒鼠强中毒的全血测定。     

Identification of parent benzodiazepines by gas chromotography/mass spectroscopy (GC/MS) from urinary extracts treated with B-glucuronidase

Urinary glucuronide metabolites of the benzodiazepines were converted back to the parent molecules after treatment with B-glucuronidase. The benzodiazepines were extracted by a one-step liquid/liquid extraction from urine or by a liquid/solid phase extraction. For the limit of detection (LOD), a standard solution of diazepam and oxazepam was serially diluted and analyzed to the point at which a reproducible analytical result was no longer obtained. Using a temperature program and a splitless mode of injection, excellent quantitation was achieved within an 8-min run time. Based upon specimens obtained from patients under a physician's care, we have determined that urinary concentrations of the benzodiazepines > 200 ng/ml are most likely due to abuse rather than to a prescribed ingestion under strict medical surveillance. Therefore, the calibration standard and cutoff concentration for a positive result was set at 200 ng/ml.     

高效液相色谱法测定肝组织中4种蟾蜍二烯内酯类化合物

目的建立高效液相色谱法检测肝组织中蟾蜍二烯内酯的定量方法。方法 以OasisHLB固相萃取柱对样品进行提取,应用HPLC色谱法分离、二极管阵列检测器测定。结果该方法的回收率高于70％,线形范围在12ng/g-1200ng/g,经该方法测得华蟾毒它灵、蟾毒灵的最小检出量为O.4ng,华蟾毒精、脂蟾毒配基的最小检出量为0.5ng(S/N≥3)。结论该方法快速、灵敏、准确,可用于蟾酥中毒的法医学检验。     

LC-MS/MS analysis of pholedrine in a fatal intoxication case

Pholedrine (4'-hydroxymethamphetamine) is a cardiovascular agent exerting hypertensive and adrenergic effects. High doses may cause a drop in the peripheral circulation blood flow and increase blood pressure, heart rate and body temperature up to a state of central respiratory paralysis. A 15-year-old girl who suffered from heavy agitation and hallucinations was admitted to the intensive care unit in a comatose state. The clinical findings included a maximum heart rate of 170 bpm and a body temperature of 43.8 degrees C. Resuscitation measures were in vain and abandoned after approximately 2h. A toxicological emergency analysis using GC/MS revealed a considerable amount of pholedrine in blood and urine. A method for determining pholedrine in human body fluids utilizing high-performance liquid chromatography (HPLC)/tandem mass spectrometry (LC-MS/MS) with a turbo ion-spray source was developed, using D11-methamphetamine and D5-methylenedioxymethamphetamine as internal standards. Samples were prepared by SPE extraction using SPEC-C18AR/MP3((R)) columns, which yielded the best extraction recovery (67%). Chromatographic separation was achieved at pH 5 on an RP-18 stationary phase applying gradient elution from 50 to 70% of B (methanol/acetonitrile 3/1 (v/v), 0.02% acetic acid) in A (5mM ammonium acetate/acetonitrile 95/5 (v/v), 0.02% acetic acid). Supra-pure acetic acid was added to the post-column effluent with a flow rate of 0.2 microl/min to optimize ionization. Detection was carried out in the positive ionization, multiple reaction monitoring (MRM) mode. The chromatograms showed no interference from other substances. The limit of detection (LOD, S/N=3) of pholedrine was 0.8 ng/ml and its lower limit of quantification (LLOQ, S/N=10) 3ng/ml. The calibration curve was linear (r=0.999) in the range 1-100 ng/ml. Samples with higher concentrations were diluted to suit the working range. The intra-day R.S.D. between 5 and 80 ng/ml were 3.8-8.7% and the inter-day R.S.D. between 5 and 100 ng/ml were 6.7-10.7%. The pholedrine concentrations in blood and urine collected when the girl was still alive were 16.1 microg/ml (R.S.D. 10.5%) and 1120 microg/ml (R.S.D. 8%), respectively. In post-mortem samples, they were 23.0 microg/ml (R.S.D. 5.1%) in heart blood and 27.3 microg/g (R.S.D. 6.6%) in the liver.     

固相萃取-GC/MS法检测生物检材中的百草枯

目的采用固相萃取-气相色谱/质谱分析方法检测血液、尿液和脏器组织中的百草枯。方法人血液、尿液和猪肺组织样品经三氯乙酸去除蛋白后,取上清用十二烷基三甲基溴化铵和十二烷基硫酸钠处理过的C18小柱提取,提取物用硼氢化钠在碱性条件下还原,产物用气相色谱/质谱法分析,外标法定量。结果生物检材中百草枯回收率为78%～87%,最低检出限为0.1μg/mL,在0.5～1mg/mL范围内线性关系良好,可对实际案例检材进行定量检测。结论本文固相萃取-气相色谱/质谱分析方法能满足中毒生物检材检验及临床毒物检验需要。     

高效液相色谱法测定人血液、尿液中的2,4-D丁酯

目的建立检测血液、尿液中2,4-D丁酯的高效液相色谱分析方法。方法采用正己烷为样品萃取溶剂,色谱柱为Zorbax SB-Aq柱,流动相为V（甲醇）∶V（水）=60∶40。结果 2,4-D丁酯在血液和尿液中的线性范围分别为0.10～10.00μg/mL（r≥0.999 8）和0.08～8.00μg/mL（r≥0.999 5）,检测限分别为0.002 0μg/mL和0.001 8μg/mL,准确度为94.5%～104.5%,日内、日间精密度≤4.5%。结论本研究建立的血液、尿液中2,4-D丁酯的提取和HPLC检测方法,可应用于2,4-D丁酯中毒的快速检验和中毒死亡的法医学鉴定。