超高效液相色谱-串联质谱法同时测定尿液中16种抗生素

目的 建立同时分析尿液中16种抗生素的超高效液相色谱-串联质谱(UPLC-MS、MS)方法.方法 以哌拉西林为内标.尿样中的目标化合物经Oasis HLB固相萃取柱富集、纯化,利用ZORBAX SB-C18色谱柱,以0.1%的甲酸溶液-乙腈为流动相经梯度洗脱分离,采用多反应监测(MRM)模式进行分析.结果各化合物的最低...     

Rapid determination of ketamine in urine by liquid chromatography-tandem mass spectrometry for a high throughput laboratory

In recent years, the abuse of ketamine had gained popularity in rave parties in Hong Kong. The Urinalysis Unit of the Government Laboratory of Hong Kong faced a tremendous increase in workload for ketamine analysis. The number of tests performed rose from 10 in 1999 to 15,000 in 2002. As a fully validated immunoassay test for ketamine was not available in the market, most laboratories analyzed ketamine by chromatographic techniques after liquid-liquid extraction. However, these methods reported in the literature are not suitable for high throughput laboratories. Hence, a rapid screening/confirmation method for ketamine by liquid chromatography-tandem mass spectrometry (LC-MS-MS) with a detection limit of 5 ng/ml was developed. After automated solid-phase extraction (SPE), the urine extract was analyzed for ketamine by a 2.5-min chromatographic run, the estimated recovery was 89% and the precision was 11% R.S.D. at 20 ng/ml. With the aid of an in-house developed computer program, the results were presented in spreadsheet format for easy checking. The method has been applied to our laboratory for routine ketamine analysis and a maximum of 200 samples per day can be achieved.     

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。     

Simultaneous determination of eight catechins and four theaflavins in bottled tea by liquid chromatography-tandem mass spectrometry for forensic analysis

Tea, and particularly bottled tea, is widely consumed worldwide and is often encountered at crime scenes in poisoning cases or used in place of urine in drug abuse monitoring. Tea is a rich source of polyphenols, such as catechins and theaflavins, and these compounds are useful for identification of trace quantities of tea samples. However, information on the contents of catechins and theaflavins in bottled tea is limited. In this study, a method was developed for simultaneous analysis of eight catechins and four theaflavins in tea using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The concentrations of these polyphenols were determined in bottled black, oolong, and green teas after a simple pretreatment process by the standard addition method. The developed LC-MS/MS method was rapid and all tested polyphenol compounds were separated within ~14 min. All tea types contained all the catechins, at varying concentrations, but not all the theaflavins were present in all the tea types. This indicates that the theaflavin composition reflects the degree of the fermentation and could be used for discrimination among different types of tea. All the green tea samples contained all eight catechins; however, the concentrations of these compounds varied among the tea samples. Principal component analysis and hierarchical cluster analysis were useful for discrimination of samples. It has been unclear whether the variations of chemical components are useful for forensic discrimination. Our results demonstrate that, in addition to identification of tea varieties, catechins and theaflavins can be used for the discrimination of bottled tea samples.     

Determination of bromadiolone in whole blood by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry

A rapid, sensitive and selective high-performance liquid chromatography tandem mass spectrometric method (HPLC/MS-MS) has been developed and validated for the determination of bromadiolone in whole blood using warfarin as an internal standard (IS). Bromadiolone was extracted from the whole blood samples by liquid-liquid extraction with ethyl acetate. Multiple-reaction monitoring (MRM) was used to detect bromadiolone and IS, using precursor --> product ion combinations at m/z 527 --> 465 and 307 --> 161, respectively. The calibration curve was linear (r2=0.998) in the concentration range of 0.5-100.0 ng/mL with a lower limit of quantification of 0.5 ng/mL in whole blood. Intra- and inter-day relative standard deviations (R.S.D.s) were less than 7.5 and 11.9%, respectively. Recoveries of bromadiolone ranged from 82.1 to 85.2%. This method is found to be determined trace bromadiolone in whole blood and can be used in the diagnosis of the poisoned human beings.     

LC-MS/MS-MRM筛选血液中22种有毒生物碱成分

目的建立液相色谱-串联质谱法对血液中的22种常见有毒生物碱成分进行筛选分析。方法血液以丁丙诺菲为内标经液液提取后,用液相色谱-串联质谱仪以电喷雾电离(ESI )、多反应监测(MRM)方式进行分析。结果以化合物的保留时间、两对母离子/子离子对定性,最低检出限为0.1～20ng/mL。结论该方法选择性佳、灵敏度高,适用于法医毒物分析和临床毒物分析中有毒生物碱的分析。     

液相色谱/质谱法检测血液中的艾司佐匹克隆

目的建立手性分离-液相色谱/质谱法,用于分离检测血液中艾司佐匹克隆。方法血液经HLB小柱固相萃取后,采用手性分离柱,用电喷雾正离子模式离子化、多反应监测模式检测艾司佐匹克隆,外标法定量。结果采用本文方法可有效分离佐匹克隆的左旋、右旋异构体,血中艾司佐匹克隆在1～100ng/mL范围内线性关系良好,相关系数为0.996 4,最低检出限为0.8ng/mL,回收率为76%～88%,日内与日间精密度均小于8%。结论本文所建方法简便、快速、分离度好,适用于血液中艾司佐匹克隆的检测。     

液相色谱-串联质谱法检测血液中的美西律

目的建立测定血液中美西律(mexiletine)的液相色谱-串联质谱联用法(LC-MS/MS)。方法采用简便的乙腈蛋白沉淀法对血液进行预处理,应用Allure PFP Propyl液相柱分离,用电喷雾正离子模式离子化,多反应监测模式对美西律进行分析。结果美西律与内标纳洛酮分离良好,在0.02～10.00μg/mL内线性关系良好,相关系数为0.9999,回归方程为y=0.0283x-0.0151,日内与日间精密度的RSD均小于15%,最低检测限为0.01μg/mL。结论建立的LC-MS/MS方法简单、灵敏、可靠,可同时适用于美西律临床药物监测和法医毒物分析的需要。     

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。     

生物检材中苯丙胺类兴奋剂和氯胺酮的LC-MS/MS分析

目的建立生物检材中苯丙胺类兴奋剂和氯胺酮的液相色谱-串联质谱(LC-MS/MS)分析方法。方法生物检材包括血液、尿液和毛发,采用稀释法和液液提取的前处理方法,应用两个不同的液相柱,优化LC-MS/MS分析方法,并考察了血液和尿液基质的离子抑制作用。结果同时分析苯丙胺和MDA,液相1在3m in内完成,液相2可用于确认分析或复杂基质分离。尿液稀释法检材用量少,前处理简便快速。毛发中苯丙胺类兴奋剂和氯胺酮的最低检测限(LOD)为0.005~0.05ng/mg。对送检案例检材产妇头发和胎毛进行苯丙胺类兴奋剂和氯胺酮的分析。结论本方法可用于生物检材中苯丙胺类兴奋剂和氯胺酮的同时分析,血、尿等生物检材的离子抑制作用是影响本方法灵敏度的主要原因。     

超高效液相色谱-MS/MS法测定血中11种苯丙胺类物质

目的应用超高效液相色谱-质谱法对全血中11种苯丙胺类毒品进行定量测定。方法全血样品经1%(v/v)甲酸-乙腈提取,采用Ostra磷脂过滤板净化处理,使用ACQUITY UPLC BEH Phenyl(100mm×2.1mm,1.7μm)色谱柱,以0.3%(v/v)甲酸溶液-乙腈为流动相进行梯度洗脱。在多反应监测模式下测定全血样本中苯丙胺、甲基苯丙胺、二亚甲基双氧苯丙胺、替苯丙胺、3,4-亚甲基二氧基乙基苯丙胺、N-甲基-1-(3,4-亚甲二氧基苯基)-2-丁胺、副甲氧基甲基苯丙胺、麻黄碱、甲基麻黄碱、卡西酮、甲卡西酮,并进行方法学考察。结果 11种苯丙胺类物质的检出限(S/N≥3)为0.01~0.4ng/m L,在0.5~50μg/L范围内线性关系良好(r0.999);回收率在75.8%~103.4%之间,相对标准偏差在1.6%~13.0%之间。结论本文建立的超高效液相色谱-质谱法快速、简便、灵敏,适用于中毒案件检验及吸毒人员排查。     

Analysis of biofluids for flunitrazepam and metabolites by electrospray liquid chromatography/mass spectrometry

A rapid and sensitive liquid chromatography/electrospray ionization mass spectrometry (LC/MS) procedure has been developed for the analysis of biofluids containing flunitrazepam and its metabolites. Specimens were spiked with deuterated analogs of the analytes. Urine specimens were enzymatically hydrolyzed and blood specimens were untreated. Extractions were carried out using CleanScreen DAU SPE cartridges. The drugs were separated on a C18 column using a methanol-water-ammonium hydroxide (60:40: 0.03 v/v) mobile phase. After determination of base peaks using full scan mass spectrometry, the mass spectrometry method was optimized to operate in selected-ion monitoring (SIM) mode for the base peak of each analyte. Positive findings were confirmed by LC/MS/MS using the same mobile phase and column. This analytical procedure allows for the detection of low levels of flunitrazepam and metabolites in biofluids. It is useful for ascertaining the role of flunitrazepam in cases of drug-facilitated sexual assault.     

Validated method for the simultaneous determination of Δ-THC and Δ-THC-COOH in oral fluid, urine and whole blood using solid-phase extraction and liquid chromatography–mass spectrometry with electrospray ionization

A fully validated, sensitive and specific method for the extraction and quantification of Δ⁹-tetrahydrocannabinol (THC) and 11-nor-9-carboxy-Δ⁹-THC (THC-COOH) and for the detection of 11-hydroxy-Δ⁹-THC (11-OH THC) in oral fluid, urine and whole blood is presented. Solid-phase extraction and liquid chromatography–mass spectrometry (LC–MS) technique were used, with electrospray ionization. Three ions were monitored for THC and THC-COOH and two for 11-OH THC. The compounds were quantified by selected ion recording of m/z 315.31, 329.18 and 343.16 for THC, 11-OH THC and THC-COOH, respectively, and m/z 318.27 and 346.26 for the deuterated internal standards, THC-d₃ and THC-COOH-d₃, respectively. The method proved to be precise for THC and THC-COOH both in terms of intra-day and inter-day analysis, with intra-day coefficients of variation (CV) less than 6.3, 6.6 and 6.5% for THC in saliva, urine and blood, respectively, and 6.8 and 7.7% for THC-COOH in urine and blood, respectively. Day-to-day CVs were less than 3.5, 4.9 and 11.3% for THC in saliva, urine and blood, respectively, and 6.2 and 6.4% for THC-COOH in urine and blood, respectively. Limits of detection (LOD) were 2 ng/mL for THC in oral fluid and 0.5 ng/mL for THC and THC-COOH and 20 ng/mL for 11-OH THC, in urine and blood. Calibration curves showed a linear relationship for THC and THC-COOH in all samples (r² > 0.999) within the range investigated.The procedure presented here has high specificity, selectivity and sensitivity. It can be regarded as an alternative method to GC–MS for the confirmation of positive immunoassay test results, and can be used as a suitable analytical tool for the quantification of THC and THC-COOH in oral fluid, urine and/or blood samples.     

Simultaneous determination of trace benzodiazepines from drinks by using direct electrospray probe/mass spectrometry (DEP/MS).

This study presents a novel means of rapidly determining benzodiazepines in various drinks. Electrospray mass spectrometry with a direct probe design is used as the analytical instrument. Samples are treated only by a simple liquid-liquid extraction prior to direct electrospray probe/mass spectrometry (DEP/MS) analysis. The proposed method provides a relatively easy and efficient means of identifying the drugs in a very short time interval. On average, sample analysis is completed in less than five minutes including sample pretreatment. It is especially useful in forensic science since the technology can provide a rapid identification for unknown samples. Thus, the method proposed herein is highly appropriate for rapidly screening a large number of samples.     

The determination of lysergide (LSD) in urine by high-performance liquid chromatography-isotope dilution mass spectrometry (IDMS)

The use of isotope dilution mass spectrometry (IDMS) has been investigated for the forensic confirmation of lysergic acid diethylamide (LSD) in urine by LC-MS. The advantages of using a deuterated analog of LSD as an internal standard over methysergide are discussed. This study includes a comparison of the electrospray mass spectra of LSD, LSD-d3 and methysergide, and discusses the choice of suitable ions for use in selected ion monitoring (SIM) mode. An IDMS method is presented for the LC-MS confirmation of LSD in urine, with a limit of quantification (LOQ) of 0.5 ng/mL, reflecting the forensic requirement at this laboratory. Under some circumstances the LOQ can be improved to 0.1 ng/mL. This method is linear in the range tested (up to 10 ng/mL LSD in urine) and has been validated in terms of accuracy and precision.     

Validated method for the simultaneous determination of Delta9-THC and Delta9-THC-COOH in oral fluid, urine and whole blood using solid-phase extraction and liquid chromatography-mass spectrometry with electrospray ionization

A fully validated, sensitive and specific method for the extraction and quantification of Delta(9)-tetrahydrocannabinol (THC) and 11-nor-9-carboxy-Delta(9)-THC (THC-COOH) and for the detection of 11-hydroxy-Delta(9)-THC (11-OH THC) in oral fluid, urine and whole blood is presented. Solid-phase extraction and liquid chromatography-mass spectrometry (LC-MS) technique were used, with electrospray ionization. Three ions were monitored for THC and THC-COOH and two for 11-OH THC. The compounds were quantified by selected ion recording of m/z 315.31, 329.18 and 343.16 for THC, 11-OH THC and THC-COOH, respectively, and m/z 318.27 and 346.26 for the deuterated internal standards, THC-d(3) and THC-COOH-d(3), respectively. The method proved to be precise for THC and THC-COOH both in terms of intra-day and inter-day analysis, with intra-day coefficients of variation (CV) less than 6.3, 6.6 and 6.5% for THC in saliva, urine and blood, respectively, and 6.8 and 7.7% for THC-COOH in urine and blood, respectively. Day-to-day CVs were less than 3.5, 4.9 and 11.3% for THC in saliva, urine and blood, respectively, and 6.2 and 6.4% for THC-COOH in urine and blood, respectively. Limits of detection (LOD) were 2 ng/mL for THC in oral fluid and 0.5 ng/mL for THC and THC-COOH and 20 ng/mL for 11-OH THC, in urine and blood. Calibration curves showed a linear relationship for THC and THC-COOH in all samples (r(2)>0.999) within the range investigated. The procedure presented here has high specificity, selectivity and sensitivity. It can be regarded as an alternative method to GC-MS for the confirmation of positive immunoassay test results, and can be used as a suitable analytical tool for the quantification of THC and THC-COOH in oral fluid, urine and/or blood samples.     

Selective and sensitive detection of chromium(VI) in waters using electrospray ionization mass spectrometry

From 2000 through 2011, there were 14 criminal cases of violations of the Clean Water Act involving the discharge of chromium, a toxic heavy metal, into drinking and surface water sources. As chromium(VI), a potential carcinogen present in the environment, represents a significant safety concern, it is currently the subject of an EPA health risk assessment. Therefore, sensitive and selective detection of this species is highly desired. This study reports the analysis of chromium(VI) in water samples by electrospray ionization mass spectrometry (ESI-MS) following its reduction and complexation with ammonium pyrrolidinedithiocarbamate (APDC). The reduction and subsequent complexation produce a characteristic [Cr(III)O]–PDC complex which can be detected as a protonated ion of m/z 507 in the positive ion mode. The detection is selective to chromium(VI) under acidic pH, even in the presence of chromium(III) and other metal ions, providing high specificity. Different water samples were examined, including deionized, tap, and river waters, and sensitive detection was achieved. In the case of deionized water, quantification over the concentration range of 3.7 to 148 ppb gave an excellent correlation coefficient of 0.9904 using the enhanced MS mode scan. Using the single-reaction monitoring (SRM) mode (monitoring the characteristic fragmentation of m/z 507 to m/z 360), the limit of detection (LOD) was found to be 0.25 ppb. The LOD of chromium(VI) for both tap and river water samples was determined to be 2.0 ppb. A preconcentration strategy using simple vacuum evaporation of the aqueous sample was shown to further improve the ESI signal by 15 fold. This method, with high sensitivity and selectivity, should provide a timely solution for the real-world analysis of toxic chromium(VI).     

液相电喷雾离子质谱技术在法医DNA中的应用

生物质谱技术是近年来发展较为迅速的生物分析技术之一,在对大分子生物质量测定中,其以快速、准确、灵敏等优点受到研究者的青睐。其中,液相电喷雾离子电离质谱(LC-ESI-MS)在法医DNA分析中对STR以及SNP遗传标记的检测具有独特的应用价值和广阔的应用前景。本文对ESI-MS的现状和在法医DNA分析中的应用等方面进行了综述,分析了LC-ESI-MS技术在法医学DNA分析中的优缺点,以促进其在国内法医DNA分析中的应用与发展。     

Analysis of Δ-tetrahydrocannabinol in oral fluid samples using solid-phase extraction and high-performance liquid chromatography–electrospray ionization mass spectrometry

An analytical method using solid-phase extraction (SPE) and high-performance liquid chromatography–mass spectrometry (LC–MS) has been developed and validated for the confirmation of Δ⁹-tetrahydrocannabinol (THC) in oral fluid samples. Oral fluid was extracted using Bond Elut LRC-Certify solid-phase extraction columns (10 cm³, 300 mg) and elution performed with n-hexane/ethyl acetate. Quantitation made use of the selected ion-recording mode (SIR) using the most abundant characteristic ion [THC + H⁺], m/z 315.31 and the fragment ion, m/z 193.13 for confirmation, and m/z 318.00 for the protonated internal standard, [d₃-THC + H⁺]. The method proved to be precise for THC, in terms of both intra-day and inter-day analyses, with coefficients of variation less than 10%, and the calculated extraction efficiencies for THC ranged from 76 to 83%. Calibration standards spiked with THC between 2 and 100 ng/mL showed a linear relationship (r² = 0.999). The method presented was applied to the oral fluid samples taken from the volunteers during the largest music event in Portugal, named Rock in Rio-Lisboa. Oral fluid was collected from 40 persons by expectoration and with Salivette^®. In 55% of the samples obtained by expectorating, THC was detected with concentration ranges from 1033 to 6552 ng/mL and in 45% of cases THC was detected at concentrations between 51 and 937 ng/mL. However, using Salivette^® collection, 26 of the 40 cases had an undetectable THC.     

Quantitative analysis of multiple illicit drugs in preserved oral fluid by solid-phase extraction and liquid chromatography-tandem mass spectrometry

We present a validated method for the simultaneous analysis of basic drugs which comprises a sample clean-up step, using mixed-mode solid-phase extraction (SPE), followed by LC-MS/MS analysis. Deuterated analogues for all of the analytes of interest were used for quantitation. The applied HPLC gradient ensured the elution of all the drugs examined within 14 min and produced chromatographic peaks of acceptable symmetry. Selectivity of the method was achieved by a combination of retention time, and two precursor-product ion transitions for the non-deuterated analogues. Oral fluid was collected with the Intercept, a FDA approved sampling device that is used on a large scale in the US for workplace drug testing. However, this collection system contains some ingredients (stabilizers and preservatives) that can cause substantial interferences, e.g. ion suppression or enhancement during LC-MS/MS analysis, in the absence of suitable sample pre-treatment. The use of the SPE was demonstrated to be highly effective and led to significant decreases in the interferences. Extraction was found to be both reproducible and efficient with recoveries >76% for all of the analytes. Furthermore, the processed samples were demonstrated to be stable for 48 h, except for cocaine and benzoylecgonine, where a slight negative trend was observed, but did not compromise the quantitation. In all cases the method was linear over the range investigated (2-200 microg/L) with an excellent intra-assay and inter-assay precision (coefficients of variation <10% in most cases) for QC samples spiked at a concentration of 4, 12 and 100 microg/L. Limits of quantitation were estimated to be at 2 microg/L with limits of detection ranging from 0.2 to 0.5 microg/L, which meets the requirements of SAMHSA for oral fluid testing in the workplace. The method was subsequently applied to the analysis of Intercept samples collected at the roadside by the police, and to determine MDMA and MDA levels in oral fluid samples from a controlled study.