γ-羟基丁酸在急性中毒大鼠体液和组织中的检测及分布

目的建立生物检材中γ-羟基丁酸(GHB)的检测方法,研究GHB急性中毒大鼠体内GHB的分布,为GHB中毒的鉴定提供方法和评价依据。方法用GC/MS法检测生物检材中的GHB;以1000mg/kg剂量给大鼠灌胃使其染毒,分别于1h和3h处死,测定体液和组织中GHB的含量。结果测组织中内源性GHB的线性范围是1～20μg/g,R2=0.9974;测组织中外源性GHB的线性范围为100～1500μg/g,R2=0.9958。相对回收率为98%～103%。体内内源性GHB的含量均≤10μg/mL或10μg/g。尿液中GHB含量为最高,其他依次为:胃、血液、肠、肾、肺、脾、心、肝和脑。结论所建方法准确、便捷,适用于GHB中毒的鉴定;尿液是体内检测GHB的最佳检材。     

毛发中GHB的检测及评价

目的探索毛发中外源性GHB的检测及判断的可行性,为涉GHB的鉴定提供方法和依据。方法建立毛发中GHB的GC/MS分析方法,并通过动物实验,考察毛发中内源性GHB的质量分数范围、外源性GHB在毛发中的时间过程以及给药剂量、毛发颜色与毛发中GHB的质量分数关系。结果豚鼠和中国人黑色毛发中内源性GHB质量分数分别为(3.01±1.41)ng/mg(n=28)和(1.02±0.27)ng/mg(n=20);摄GHB后毛发中GHB质量分数明显增加且与给药剂量呈正相关性;GHB在毛干中呈窄带分布;深色毛发中GHB质量分数高于浅色毛发。结论毛发中GHB的检测适用于GHB滥用和中毒的法医毒物学鉴定;根据毛发中的GHB质量分数和毛发分段分析可判断GHB的来源。     

尿液中γ-羟基丁酸及其前体物质的检测和应用

目的建立尿液中γ-羟基丁酸(gamma-hydroxybutyric acid,GHB)及其前体物质1,4-丁二醇(1,4-butanediol,1,4-BD)和γ-丁内酯(gamma-butyrolactone,GBL)的液相色谱-串联质谱法(LC-MS/MS),为相关案件提供依据。方法以GHB-d6、MOR-d3为内标,尿样经甲醇沉淀蛋白后通过液相色谱分离,电喷雾离子源进行离子化,多反应监测模式对各化合物进行检测。结果 GHB及其前体物质1,4-BD、GBL的检出限分别为0.1、0.1和2μg/m L,准确度为87.6%~98.1%,日内及日间精密度均小于15%,基质效应大于80%。结论所建立的分析方法灵敏度高、简便快速、专属性强、可靠性高,可为司法鉴定实践中涉及GHB的案件提供技术支持和基础数据。     

血液、脑脊液中尼可刹米的HPLC法分析

目的建立血液、脑脊液中尼可刹米的HPLC检测方法。方法取空白血液或脑脊液添加不同量的尼可刹米对照液,碱化后用二氯甲烷等有机溶剂提取,用HPLC方法检测。对方法的萃取回收率、线性、精密度等进行考察。结果方法选用二氯甲烷作为萃取溶剂,萃取回收率大于60%。血液中尼可刹米的在0.5~100μg/ml范围内线性良好,R2为0.9993;脑脊液中尼可刹米的线性范围为0.1~100μg/ml,R2为0.9996,最低检出限是5ng(S/N≥3)。检测血液和脑脊液中的尼可刹米方法的精密度均小于6%。结论所建方法准确、简便、灵敏度高,适用于法医毒物分析和临床药物分析。     

化学显色法快速筛选饮料及尿液中γ-羟基丁酸和γ-丁内酯

目的建立化学显色法快速筛选饮料及尿液中γ-羟基丁酸(GHB)及其前体γ-丁内酯(GBL)的方法。方法在酸性条件下GHB转化为GBL,GBL和盐酸羟胺在碱性条件下生成γ-羟基丁酰羟胺,γ-羟基丁酰羟胺在酸性条件下和三氯化铁反应,生成紫红色的络合物。结果饮料中GHB最低检出浓度为0.5～2mg/mL,低于常见滥用质量浓度。该方法也可以用于尿液分析,最低检出质量浓度为0.5mg/mL。考察了常见有机溶剂和麻醉镇静药物的干扰。结论该方法简单、安全、快速,为临床和法庭科学实验室快速筛选GHB和GBL提供了便利。     

液相色谱-串联质谱法分析生物检材中的河豚毒素

目的建立血液、尿液以及肝中河豚毒素(tetrodotoxin,TTX)的液相色谱-串联质谱分析方法,并进行方法学验证。方法血液、尿液和肝用1%乙酸甲醇溶液去蛋白后,上清液用固相萃取法净化,LC-MS/MS检测。结果血液、尿液和肝中TTX检出限分别为2ng/mL、2ng/mL和4ng/g。血液和尿液在4～100ng/mL、肝在5～100ng/g的范围内线性关系良好,相关系数r≥0.9973;日内精密度和日间精密度均在12.80%以内;回收率大于47.2%。结论所建方法高效、灵敏、准确,可以为河豚毒素中毒的法医学鉴定、临床诊治以及食品安全的监控提供技术保障。     

LC-MS/MS同时分析血液中五种巴比妥类药物

目的建立血液中巴比妥类药物液相色谱-串联质谱(LC-MS/MS)检测方法。方法血液中的巴比妥类药物用有机溶剂液-液萃取,以乙酰水杨酸为内标,LC-MS/MS测定。结果检测血液中五种巴比妥类药物的最小检出限(LOD)均为10ng/mL,线性范围在0.02￣10μg/mL内良好,方法的回收率70%￣87%。结论所建方法快速、灵敏、准确,可以满足临床毒物和法庭毒物分析的需要。     

液相色谱-串联质谱法测定血液和尿液中秋水仙碱

目的建立检测血液和尿液中秋水仙碱的液相色谱-串联质谱法。方法0.5mL血液或尿液以丁丙诺啡为内标,经pH9.2硼酸盐缓冲溶液碱化后,用乙酸乙酯进行提取,在ZORBAX SB-C18液相柱(150mm×2.1mm×5μm)上以V(甲醇)∶V(20mmol/L乙酸铵和0.1%甲酸缓冲溶液)=80∶20为流动相,流速为0.2mL/min,采用电喷雾正离子模式离子化、多反应监测模式检测秋水仙碱,内标法定量。结果血液、尿液中秋水仙碱与内标丁丙诺啡色谱分离良好,秋水仙碱在0.1~50 ng/mL内均具有良好的线性,相关系数>0.9990,最低检出限为0.05ng/mL,方法回收率为94%~116%,日内与日间精密度(RSD)均小于8.5%。结论所建LC-MS-MS方法灵敏度高、操作简便、快速、准确,适用于血液及尿液等生物检材中痕量秋水仙碱成分的检测。     

LC-MS/MS法测定饮酒者尿液中乙醇和5-羟色胺代谢物

目的建立尿液中5羟(基)-β吲哚乙醇葡萄糖苷酸(5-hydroxytryptophol glucuronide,GTOL)和5羟(基)吲哚-3-乙酸(5-hydroxyindole-3-aceticacid,5-HlAA)的高效液相色谱-串联质谱(LC-MS/MS)检测方法,为疑似饮酒案件的法医学鉴定提供实验依据。方法尿液经乙腈(1%甲酸)沉淀蛋白,经Captiva EMR-Lipid滤柱过滤,以0.1%甲酸水溶液～乙腈(90:10 V/V)等度洗脱,在多反应监测模式下检测GTOL和5-HIAA,并对38例案件尿液样本定性定量检测。结果 GTOL的线性范围是0.025μg/mL～2μg/mL,线性关系良好,R2 0.999,检测限和定量下限分别为0.01μg/mL和0.02μg/mL。5-HIAA的线性范围是0.01μg/mL～50μg/mL,线性关系良好,R2 0.99,检测限和定量下限分别为0.02μg/mL和0.05μg/mL。GTOL和5-HIAA的批内和批间精密度均小于15%,准确度分别在90.52%～111.04%和96.81%～112.30%,稳定性变化率在-11.96%～3.71%。25例乙醇阳性尿液样本GTOL和5-HIAA比值均大于15(pmol/nmol),13例乙醇阴性尿液样本GTOL和5-HIAA比值均小于15(pmol/nmol)。结论本研究建立的尿液中GTOL和5-HIAA定性定量分析方法准确、灵敏,可快速、精确地得到GTOL和5-HIAA比值,为司法鉴定法工作中饮酒判定提供科学依据。     

液相色谱-串联质谱法测定血液、尿液中乙基葡萄糖醛酸苷

目的建立血液、尿液中乙基葡萄糖醛酸苷（ethyl glucuronide,EtG）的液相色谱-串联质谱（LC-MS/MS）检测方法。方法血液、尿液用乙腈沉淀蛋白,离心后取上清液用LC-MS/MS检测。结果血液、尿液中EtG的检出限均为0.05μg/mL,线性范围均为0.10～5.00μg/mL（r〉0.999）,检测方法准确度为95%～109%,日间及日内精密度〈12%。对送检案例血液中EtG进行检测,效果良好。结论本方法适用于血液、尿液中EtG的检测。     

Intoxication due to 1,4-butanediol

We report a case of intoxication resulting from the ingestion of a liquid, sold in the illicit market as "liquid ecstasy," which was found to contain 1,4-butanediol, a metabolic precursor of gamma-hydroxybutiric acid (GHB). Identification of the substance in the liquid was performed by gas chromatography-mass spectrometry (GC-MS).The toxicological analysis of blood, urine and gastric content of the victim was performed by immunoassay and gas chromatography with nitrogen-phosphorus detection as screening techniques and by means of GC-MS for confirmation and quantitation of 1,4-butanediol and GHB. The following drug concentrations were found: 82 microg/ml (blood), 401 microg/ml (urine) and 7.4 microg/ml (gastric content) for 1,4-butanediol and 103 microg/ml (blood), 430.0 microg/ml (urine) for GHB. In addition to these, other drugs detected and their blood concentration found in this case were methylenedioxymethylamphetamine (MDMA) 0.23 microg/ml and its metabolite methylenedioxyphenylamphetamine (MDA) 0.10 microg/ml. In the urine, a concentration of 0.10 microg/ml of benzoylecgonine was also found.     

Site-dependent production of gamma-hydroxybutyric acid in the early postmortem period

This study compared endogenous gamma-hydroxybutyric acid (GHB) concentrations in various postmortem fluid samples of 25 autopsy cases. All bodies were stored between 10-20 degrees C until autopsy, and the intervals between death and autopsy were less than 2 days (6-48 h). GHB concentrations were measured by headspace gas chromatography after GHB was converted to gamma-butyrolactone. Endogenous GHB concentrations were significantly higher in femoral venous blood (4.6+/-3.4 microg/ml, n=23) than in cerebrospinal fluid (1.8+/-1.5 microg/ml, n=9), vitreous humor (0.9+/-1.7 microg/ml, n=8), bile (1.0+/-1.1 microg/ml, n=9) and urine (0.6+/-1.2 microg/ml, n=12). GHB concentrations were similar in blood samples taken from different sites. Cut-off limits of 30 and 10 microg/ml are proposed for blood and urine, respectively, to discriminate between exogenous and endogenous GHB in decedents showing no or little putrefaction (postmortem intervals usually 48 h or less). The criterion established for endogenous GHB in postmortem urine may also be applicable to analytical results in cerebrospinal fluid, vitreous humor and bile from deceased persons.     

GHB. Club drug or confusing artifact?

GHB can be produced either as a pre- or postmortem artifact. The authors describe two cases in which GHB was detected and discuss the problem of determining the role of GHB in each case. In both cases, NaF-preserved blood and urine were analyzed using gas chromatography. The first decedent, a known methamphetamine abuser, had GHB concentrations similar to those observed with subanesthetic doses (femoral blood, 159 microg/ml; urine, 1100 microg/ml). Myocardial fibrosis, in the pattern associated with stimulant abuse, was also evident. The second decedent had a normal heart but higher concentrations of GHB (femoral blood, 1.4 mg/ml; right heart, 1.1 mg/ml; urine, 6.0 mg/ml). Blood cocaine and MDMA levels were 420 and 730 ng/ml, respectively. Both decedents had been drinking and were in a postabsorptive state, with blood to vitreous ratios of less than 0.90. If NaF is not used as a preservative, GHB is produced as an artifact. Therefore, the mere demonstration of GHB does not prove causality or even necessarily that GHB was ingested. Blood and urine GHB concentrations in case 1 can be produced by a therapeutic dose of 100 mg, and myocardial fibrosis may have had more to do with the cause of death than GHB. The history in case 2 is consistent with the substantial GHB ingestion, but other drugs, including ethanol, were also detected. Ethanol interferes with GHB metabolism, preventing GHB breakdown, raising blood concentrations, and making respiratory arrest more likely. Combined investigational, autopsy, and toxicology data suggest that GHB was the cause of death in case 2 but not case 1. Given the recent discovery that postmortem GHB production occurs even in stored antemortem blood samples (provided they were preserved with citrate) and the earlier observations that de novo GHB production in urine does not occur, it is unwise to draw any inferences about causality unless (1) blood and urine are both analyzed and found to be elevated; (2) blood is collected in NaF-containing tubes; and (3) a detailed case history is obtained.     

GC法检测血液和尿液中甲基苯丙胺和咖啡因

目的建立同时测定血、尿中甲基苯丙胺和咖啡因含量的方法。方法应用GC／NPD技术,以4-苯基丁胺为内标,直接碱化,用氯仿提取,三氟乙酸酐衍生化,8CB熔融石英毛细管柱（30m×0．25mm×0．25μm）分析。结果生物样品中甲基苯丙胺与咖啡因在0．012—7．5μg/mL浓度范围内线性关系良好,检测限（S／N=3）依次为1．2ng／mL,0．6ng／mL（血）;1．6ng／mL,0．8ng／mL（尿）。苯丙胺在0．017—10．0μg／mL浓度范围内线性关系良好,检测限为1．6mg／mL（血）,3．2ng／mL（尿）。所有样本回收率均大于85％。结论本方法准确、灵敏,适用于血、尿中甲基苯丙胺及其代谢物苯丙胺的三氟乙酸酐衍生化物和咖啡因的同时检测,为判定滥用毒品种类、追查毒品来源以及研究生物体内甲基苯丙胺和咖啡因的交互影响提供了检测手段。     

Quantitative Analysis of Gamma‐Hydroxybutyrate at Endogenous Concentrations in Hair using Liquid Chromatography Tandem Mass Spectrometry

Abstract: A method capable of quantifying endogenous concentrations of gamma‐hydroxybutyrate (GHB) in human head hair was developed and validated using liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS). Hair was digested under alkaline conditions, and GHB was isolated using liquid–liquid extraction. LC/MS/MS was performed using atmospheric pressure chemical ionization in the negative mode, multiple reaction monitoring, and deuterated internal standard (GHB‐D₆). Linearity was observed between 0.1 and 100 ng/mg GHB (R² = 1.000). The limits of detection and quantitation in human hair were 0.2 and 0.4 ng/mg, respectively. Accuracy at 2 ng/mg and 10 ng/mg was determined to be 97% and 94%, and intra‐assay CVs at these concentrations were 5.2% and 7.4% (n = 4). Beta‐hydroxybutyrate (BHB), alpha‐hydroxybutyrate, gamma‐butyrolactone, and 1,4‐butanediol did not produce an interference, and there was negligible ion suppression or enhancement from the matrix.     

Fatal intoxication with 1,4-butanediol: Case report and comprehensive review of the literature

A fatal case of 1,4-butanediol (1,4-BD) oral ingestion is reported here, in which a 51-year-old man was found dead in his bed. According to the police report, the deceased was a known drug user. A glass bottle labeled (and later confirmed to be) “Butandiol 1,4” (1,4-BD) was found in the kitchen. Furthermore, the deceased's friend stated that he consumed 1,4-BD on a regular basis. The autopsy and histological examination of postmortem parenchymatous organ specimens did not revealed a clear cause of death. Chemical-toxicological investigations revealed gammahydroxybutyrat (GHB) in body fluids and tissues in the following quantities: femoral blood 390 mg/L, heart blood 420 mg/L, cerebrospinal fluid 420 mg/L, vitreous humor 640 mg/L, urine 1600 mg/L, and head hair 26.7 ng/mg. In addition, 1,4-BD was qualitatively detected in the head hair, urine, stomach contents, and the bottle. No other substances, including alcohol, were detected at pharmacologically relevant concentrations. 1,4-BD is known as precursor substance that is converted in vivo into GHB. In the synoptic assessment of toxicological findings, the police investigations and having excluded other causes of death, a lethal GHB-intoxication following ingestion of 1,4-BD, can be assumed in this case. Fatal intoxications with 1,4-BD have seldom been reported due to a very rapid conversion to GHB and, among other things, non-specific symptoms after ingestion. This case report aims to give an overview to the published of fatal 1,4-BD-intoxications and to discuss the problems associated with detection of 1,4-BD in (postmortem) specimens.     

尿样中苯骈二氮杂Zhu类药物筛选分析研究

目的建立尿样中苯骈二氮杂类药物两种筛选分析方法.方法用GC/ECD、GC/MS直接测定苯骈二氮杂类药物原体和GC/ECD、GC/MS测定1,4-苯骈二氮杂类药物的酸水解产物苯甲酮同系物.结果GC/ECD直接测定苯骈二氮杂类药物方法,大部分药物的回收率为60%～90%,线性范围为20～200ng/ml尿,线性相关系数大于0.99,最低检出限达0.5ng/ml～10ng/ml.结论所建两种方法各有其特点又可相互补充,已成功地应用于司法鉴定实践.     

血液和尿液中草甘膦的净化富集及检测

目的建立血液、尿液中草甘膦的净化富集及定性定量方法。方法取适量血、尿样品,用80%甲醇水溶液稀释,经Agilent Accu Bond NH2（弱阴离子交换柱）净化富集,用3m L 5%氨水溶液洗脱后进行ESI/LC/MRM对草甘膦的定性及阴离子色谱法（IC）的定量检验分析。结果血液、尿液中草甘膦的IC法检验回收率均大于90%,方法的线性范围均在1-100μg/m L,血检测限为0.1μg/m L,尿液为0.08μg/m L。血液、尿液中草甘膦的LC/MRM的方法检测限均在5ng/m L左右。结论建立的血、尿中草甘膦的前处理方法和LC/MRM定性分析及IC定量分析方法,可满足司法检验要求。     

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。     

Gamma hydroxybutyric acid (GHB) concentrations in humans and factors affecting endogenous production

The endogenous nature of the drug of abuse gamma hydroxybutyric acid (GHB) has caused various interpretative problems for toxicologists. In order to obtain data for the presence of endogenous GHB in humans and to investigate any factors that may affect this, a volunteer study was undertaken. The GHB concentrations in 119 urine specimens from GHB-free subjects and 25 urine specimens submitted for toxicological analysis showed maximal urinary GHB concentrations of 3mg/l. Analysis of 15 plasma specimens submitted for toxicological analysis detected no measurable GHB (less than 2.5mg/l). Studies in a male and female volunteer in which different dietary food groups were ingested at weekly intervals, showed significant creatinine-independent intra-individual fluctuation with overall urine GHB concentrations between 0 and 2.55, and 0 and 2.74mg/l, respectively. Urinary concentrations did not appear to be affected by the particular dietary groups studied.The concentrations measured by gas chromatography with flame ionisation detection (GC-FID) and gas chromatography with mass spectrometry (GC-MS) lend further support to the proposed urinary and plasma interpretative cut-offs of 10 and 4mg/l, respectively, where below this it is not possible to determine whether any GHB detected is endogenous or exogenous in nature.