Profiling of illicit fentanyl using UHPLC-MS/MS

Methodology is presented for the profiling of fentanyl in seized drugs using ultra high performance liquid chromatography in combination with tandem mass spectrometry (UHPLC-MS/MS). Target analysis was performed for 40 fentanyl processing impurities, several of which are markers for a specific synthetic route (Siegfried or Janssen). For the separation of these solutes, an Acquity BEH C18 1.7 μm particle column (150 mm × 2.1 mm) with a binary 1% formic acid (pH 2.0)/acetonitrile gradient was used. For MS/MS detection, an atmospheric pressure positive electrospray source was employed with selected reaction monitoring (SRM). The coupling of the high separation power of UHPLC with the highly selective and sensitive detection of MS/MS is amenable to the determination of synthetic route and linking of drug seizures. The technology is also applicable to exhibits containing trace levels of fentanyl in the presence of significantly excess amounts of heroin and/or adulterants.     

Forensic determination of ricin and the alkaloid marker ricinine from castor bean extracts

Liquid chromatography/mass spectrometry (LC/ MS) and matrix assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS methods were developed for the presumptive identification of ricin toxin and the alkaloid marker ricinine from crude plant materials. Ricin is an extremely potent poison, which is of forensic interest due to its appearance in terrorism literature and its potential for use as a homicide agent. Difficulties arise in attempting to analyze ricin because it is a large heterogeneous protein with glycosylation. The general protein identification scheme developed uses LC/MS or MALDI-TOF for size classification followed by the use of the same instrumentation for the analysis of the tryptic digest. Fragments of the digest can be searched in an online database for tentative identification of the unknown protein and then followed by comparison to authentic reference materials. LC fractionation or molecular weight cutoff filtration was used for preparation of the intact toxin before analysis. Extracts from two types of castor beans were prepared using a terrorist handbook procedure and determined to contain 1% ricin. Additionally, a forensic sample suspected to contain ricin was analyzed using the presented identification scheme (data not shown). The identification of the alkaloid ricinine by GC/MS and LC/MS was shown to be a complementary technique for the determination of castor bean extracts.     

Detection of malathion in a victim by gas chromatography/negative ion chemical ionization mass spectrometry

We experienced an autopsy case in which a 53-year-old woman committed suicide by ingesting allegedly a certain agricultural chemical. The blood and stomach contents, after extraction with acetonitrile and chloroform, were subjected to analysis by gas chromatography (GC)/negative ion chemical ionization (CI) mass spectrometry (MS). By total ion monitoring in the negative CI mode, a large peak appeared. The mass spectrum of the peak showed a strong anion at m/z 157, suggesting the presence of an organophosphorus pesticide. By measuring its spectrum in the positive electron impact (EI) mode, it was identified as malathion. The selected ion monitoring in the negative CI mode showed that the malathion peak was not interfered with by any impurities, and its background was very low. The sensitivity in the negative CI mode was about 5-10 times higher than that in the positive EI mode. Our data show that the GC/negative ion CI MS is useful for both screening and sensitive quantitation of organophosphorus pesticides.     

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.     

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。     

GC法和GC/MS法在地区毒品情报分析中的应用

目的探讨气相色谱法(GC)和气相色谱/质谱联用法(GC/MS)在地区毒品情报分析中的应用,并用于实际样品分析。方法采用GC/MS法定性分析毒品样品中的主成分、痕量杂质、掺假剂和稀释剂,并选择GC/FID法和内标标准曲线法定量分析上述各组分,通过定性和定量数据的统计和分析,推断该地区的毒品状况。结果用GC/MS定性分析、GC/FID定量分析,和数据的统计比对,获得了某省某年27起海洛因毒品大案样品主成分、痕量杂质、掺假剂和稀释剂组成及含量的特征,发现该省缴获的大宗海洛因毒品主要以掺假和稀释后的产品出现,且咖啡因和扑热息痛为两种主要的掺假剂和稀释剂。结论GC/MS和GC/FID法在地区毒品情报分析和打击毒品犯罪中可以发挥重要作用。     

Trace Element Analysis of Rough Diamond by LA‐ICP‐MS: A Case of Source Discrimination?*,†

Abstract: Current profiling of rough diamond source is performed using different physical and/or morphological techniques that require strong knowledge and experience in the field. More recently, chemical impurities have been used to discriminate diamond source and with the advance of laser ablation–inductively coupled plasma–mass spectrometry (LA‐ICP‐MS) empirical profiling of rough diamonds is possible to some extent. In this study, we present a LA‐ICP‐MS methodology that we developed for analyzing ultra‐trace element impurities in rough diamond for origin determination (“profiling”). Diamonds from two sources were analyzed by LA‐ICP‐MS and were statistically classified by accepted methods. For the two diamond populations analyzed in this study, binomial logistic regression produced a better overall correct classification than linear discriminant analysis. The results suggest that an anticipated matrix match reference material would improve the robustness of our methodology for forensic applications.     

Sudden death caused by 1,1-difluoroethane inhalation

A 20-year-old man was found dead on the floor next to a computer, with a nearly full can of "CRC Duster" dust remover located next to the deceased on the floor, and an empty can of the same product on the computer desk. Toxicologic evaluation using either gas chromatography/mass spectrometry (GC/MS) or gas chromatography/flame ionization detector (GC/FID) method identified the active ingredient 1,1-difluoroethane (Freon 152a) in all tissues analyzed. Tissue distribution studies revealed highest concentration in central blood, lung, and liver. It is believed that the 1,1-difluoroethane inhalation was the cause of death.     

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.     

Drug intelligence based on organic impurities in illicit MA samples

One major objective of the European project "Collaborative Harmonisation of Methods for Profiling of Amphetamine Type Stimulants" (CHAMP), funded by the sixth framework programme of the European Commission, consisted of the harmonisation of a gas chromatography/mass spectrometry (GC/MS) method for the analysis of organic impurities found in illicit methamphetamine (MA) samples in a drug intelligence perspective. Statistical analysis provided a selection of pertinent variables among the 43 organic impurities identified in the chromatograms. As for the 3,4-MethyleneDioxyMethAmphetamine (MDMA) study, correlation coefficients were used as a discrimination tool between populations of linked samples (from the same seizure) and unlinked samples (from different seizures). It was also shown that correlation measurements based on Pearson and cosine functions applied to the data pre-treated by normalisation to the sum of peak responses followed by the square root provided excellent discrimination between the two populations. The organic impurities profiling method was proved to be relevant for the characterization of samples from different seizures and their synthesis route patterns.     

Separation and Detection of Smokeless Powder Additives by Ultra Performance Liquid Chromatography with Tandem Mass Spectrometry (UPLC/MS/MS),

A reversed phase gradient ultra performance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) method has been developed for the analysis of smokeless powders. A total of 20 different components were separated by UPLC and detected by MS/MS in multiple reaction monitoring (MRM) mode. These compounds included diphenylamines, centralites, nitrotoluenes, nitroglycerin, and various phthalates. Simultaneous positive and negative electrospray ionization (ESI) was used along with negative atmospheric pressure chemical ionization (APCI) to detect all compounds in a single analysis. Analysis times were under 8 min with a gradient of 10–73% organic at a flow rate of 0.500 mL/min. With this method, ultraviolet and MRM limits of detection ranging from 0.08 to 2.6 ng and 0.4–64 ng injected were achieved. Commercially available smokeless powders were also extracted with methylene chloride and characterized using the developed UPLC/MS/MS method. The procedure permits the determination of compositional differences between different brands as well as lot‐to‐lot variations.     

Characterization of pentoxyverine metabolites in urine using GC/MS after intoxication with Silomat cough drops

A nearly two and a half year old boy was hospitalized after showing symptoms of disorientation and hallucination. The parents remembered the child playing with a bottle of Silomat cough drops, so that an intoxication was taken into consideration. After liquid/liquid extraction of a urine sample collected in hospital, the underivatized and the acetylated extracts were analyzed by gas chromatography-mass spectrometry (GC/MS) using electron ionization (EI) as well as chemical ionization (CI). In the urine sample high amounts of pentoxyverine (carbetapentane) and several of its metabolites, e.g., different hydrolyzed, desalkylated and ring-hydroxylated products have been identified. The correlation of the results, the observed symptoms, and the access to the Silomat cough drops reveal an intoxication after ingestion of an unknown amount of the antitussive pentoxyverine. Corresponding EI- and CI-GC/MS spectra are presented characterizing the structure of its metabolites.     

Development of microwave-assisted extraction procedure for organic impurity profiling of seized 3,4-methylenedioxymethamphetamine (MDMA)

Abstract: Organic impurity profiling of seized 3,4‐methylenedioxymethamphetamine (MDMA) tablets aims to link tablets to common production sources. Conventionally, organic impurities are extracted from tablets using a liquid–liquid extraction (LLE) procedure prior to analysis by gas chromatography–mass spectrometry (GC‐MS). In this research, the development of an alternative microwave‐assisted extraction/headspace solid‐phase microextraction (MAE/HS‐SPME) procedure is described. The optimal procedure used phosphate buffer (1 M, pH 8), with an HS‐SPME extraction temperature of 70°C for 40 min, using a divinylbenzene/Carboxen?/polydimethylsiloxane (DVB/CAR/PDMS) fiber. Impurities were extracted from seized MDMA exhibits using the MAE/HS‐SPME procedure, as well as HS‐SPME alone, and a conventional LLE procedure. The HS‐SPME procedure was deemed to be the most practical because of the affordability and need for less analyst involvement. Although the LLE was limited in the number of impurities extracted, the procedure is still useful for the extraction of less volatile impurities that are not extracted by HS‐SPME.     

Optimum methamphetamine profiling with sample preparation by solid-phase microextraction

Solid-phase microextraction (SPME) is a relatively new technique in which a small, polymer-coated fiber is employed to extract volatile and semivolatile organic compounds from the sealed headspace above a questioned sample. SPME, coupled with gas chromatography/mass spectrometry (GC/MS), was used to characterize impurities in illicit methamphetamine samples. Trace impurities present in a specimen were tentatively identified using mass-spectral databases and included 1,2-dimethyl-3-phenyl-aziridine (indicating synthesis via a halogenated ephedrine intermediate), ethyl vanillin (a flavoring compound), and caffeine (a stimulant used as cutting agent). The types and numbers of organic compounds sampled by SPME were compared with those collected by various solvent extraction protocols. In addition to unambiguously confirming the presence of methamphetamine, SPME-GC/MS analyses detected approximately 30 more organic analytes than were found by GC/MS following the ethyl acetate extraction method adopted by the United Nations International Drug Control Programme. SPME-GC/MS is a superior method for generating material "fingerprint" profiles in methamphetamine samples. The detection and characterization of increased points of comparison in drug samples provide more detailed chemical signatures for both intelligence and operational information.     

Detection of methamphetamine and amphetamine in a single human hair by gas chromatography/chemical ionization mass spectrometry

A detailed procedure of an extremely sensitive method for quantitation of methamphetamine and amphetamine in human hair by gas chromatography (GC)/chemical ionization (CI) mass spectrometry (MS) is presented. N-methylbenzylamine was used as an internal standard. The samples, after extraction with an organic solvent, were derivatized with trifluoroacetic anhydride before the GC/MS analysis. Quantitation was made with quasi-molecular ions of the derivatives by selected ion monitoring in the CI mode. The detection limit was about 10 pg in an injected volume. The high sensitivity enabled us to measure both stimulants in a single human hair in actual cases.     

Signature Profiling and Classification of Illicit Heroin by GC-MS Analysis of Acidic and Neutral Manufacturing Impurities

Abstract:  The illicit manufacture of heroin results in the formation of trace level acidic and neutral impurities. These impurities are detectable in illicit heroin and provide valuable information about the manufacturing process used. The isolation, derivatization, and semiquantitative analysis of neutral and acidic heroin manufacturing impurities by programmed temperature vaporizing injector-gas chromatography-mass spectrometry (PTV-GC-MS) is described. Trace acidic and neutral heroin impurities were isolated from basic fractions using liquid–liquid extraction. Extracted impurities were treated with N -Methyl- N -trimethylsilyltrifluoroacetamide followed by PTV-GC-MS analyses. Semiquantitative data were obtained using full scan mass spectrometry utilizing unique ions or ion combinations for 36 trace impurities found in crude and/or highly refined heroin samples. Minimum detection limits for acidic and neutral impurities were estimated to be at the 10⁻⁷ level relative to total morphine. Over 500 authentic heroin samples from South America, Mexico, Southwest Asia, and Southeast Asia were analyzed. Classification of illicit heroin based on the presence or absence and relative amounts of acidic and neutral impurities is presented.     

分子印迹固相萃取法在血中苯丙胺类毒品分析中的应用

目的建立分子印迹固相萃取（MISPE）、GC/MS分析方法,用于血液中苯丙胺类毒品检测。方法 10mmol/L醋酸铵缓冲液（pH8.0）4倍稀释空白添加血液,1mL甲醇,1mL10mmol/L醋酸铵缓冲液（pH8.0）活化苯丙胺类分子印迹固相萃取柱;2×1mL去离子水、1mL60%的乙腈去离子水、1mL1%醋酸乙腈洗涤杂质;2×1mL1%甲酸/甲醇洗脱,洗脱液挥干定容,经GC/NPD、GC/MS分析检测。结果各种苯丙胺类毒品回收率均在90%以上,在20～5 000ng/mL浓度范围内线性关系良好,r2为0.995 7～0.998 9,LOQ在16～30ng/mL之间,LOD在8～15ng/mL之间。结论本方法回收率高,净化效果显著,稳定性好,杂质干扰少,可用于血液中低浓度苯丙胺类毒品的分析检测。     

High-performance liquid chromatography-ultraviolet-visible spectroscopy-electrospray ionization mass spectrometry method for acrylic and polyester forensic fiber dye analysis

A critical point of comparison between a fiber collected from a crime scene and a fiber from a known source is the color. Fiber dye analysis using thin-layer chromatography or ultraviolet (UV)-visible (Vis) microspectrophotometry provides useful, although limited, data for comparison. High-performance liquid chromatography-electrospray ionization mass spectrometry (LC/MS) overcomes these limitations by integrating chromatography, ultraviolet-visible spectroscopy, and mass spectrometry into a single instrument. In order to evaluate the applicability of the LC/MS to forensic fiber dye analysis, a multi-stage chromatographic method using acidified water and acidified acetonitrile was developed that separated and identified a mixture of 15 basic and 13 disperse dye standards. The LC/MS also detected and analyzed dyes extracted from individual 0.5 cm acrylic and polyester fibers, demonstrating its applicability to this type of analysis. With regard to the analysis of disperse dyes in polyester fibers, the replacement of pyridine with acetonitrile in the extraction system allowed direct injection of the extracts into the LC/MS. The advantage of the LC/MS over other instrumental methods of textile dye analysis is demonstrated by the analysis and differentiation of three black acrylic fibers: two fibers had similar UV-Vis spectra but were differentiated with chromatography and two had similar UV-Vis spectra and chromatograms but were differentiated using the mass spectrometer.     

Effective injection in pulsed splitless mode for impurity profiling of methamphetamine crystal by GC or GC/MS

Gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS) are commonly used for the impurity profiling of illegal drugs. For the impurity profiling of methamphetamine, it is very important to obtain information about impurities related to the manufacturing route and the precursor chemicals [B. Remberg, A.H. Stead, Drug characterization/impurity profiling, with special focus on methamphetamine: recent work of the United Nations International Drug Control Programme, Bull. Narcotics LI (1999) 97-117 ]. There are many artifact impurities arising from the preparation of samples and conditions of GC. Moreover, some impurities pose a barrier to the statistical processing of methamphetamine profiling. We investigated capillary GC analysis using pulsed splitless (PS/L) injection to minimize the thermal decomposition of impurities at the injection port and improve the transfer of samples into the column. We confirmed that the optimal conditions of PS/L-mode are 230 degrees C (injection temperature), 50 psi (pulsed pressure) and 1.1 min (pulsed time) for the methamphetamine profiling. Based on the impurity profiles of 48 methamphetamine crystals in PS/L-mode, we can achieve very easy handling and obtained, good results.     

Analysis of drugs of abuse in hair by automated solid-phase extraction, GC/EI/MS and GC ion trap/CI/MS

In our laboratory, analysis of human hair for the detection of drugs of abuse was first performed in 1995. Initially, requests for hair analysis were few, and it is only since 1997 that these analyses have become routine. As demand grew, we developed an automatic solid-phase extraction method; the use of a robot ASPEC allowed us to drop certain fastidious manipulations, and to treat a large number of samples at a time. This method is described, along with analysis by gas-chromatography-mass spectrometry (GC/MS) in selected ion monitoring mode (SIM), for the following drugs: codeine, 6-monoacetylmorphine (6-MAM), morphine, cocaine, methadone, ecstasy (MDMA) and Eve (MDE). This requires prior derivatization with propionic anhydride. The different validation parameters, linearity, repeatability, recovery and detection limits are described, as well as the application of this method to some real cases. Analysis of these cases is also performed by an ion trap GC/MS in chemical ionization mode (GC/IT/CI/MS) in order to demonstrate the usefulness of this technique as a complement to routine analysis. Analysis by GC/IT/CI/MS indeed avoids the risk of false-positive results by the identification of metabolites.