New developments in SPME, Part 1: The use of vapor-phase deprotonation and on-fiber derivatization with alkylchloroformates in the analysis of preparations containing amphetamines

Due to their high polarity and low vapor pressure, most amine salts of amphetamine-type drugs are not directly amenable to headspace recovery using solid-phase microextraction (SPME). Described in this article is a simple vapor-phase procedure for the conversion of solid drug salt samples into their free bases by the use of triethylamine. This process can be conducted simultaneously with headspace SPME, the outcome being that solid drug salts can be sampled directly for GC-MS without the need for dissolution and chemical processing. Potential applications for this methodology include the noninvasive recovery of drug traces from objects such as banknotes and garments. This new process for recovery of amphetamine-type drugs has been combined with on-fiber derivatization using alkylchloroformates. This extra step was included to improve the chromatographic performance of analytes and allow for the resolution of drug enantiomers.     

GC/MS、GC/NPD法检测毛发中的氯胺酮

目的采用液-液萃取、衍生化和GC/MS、GC/NPD方法,进行毛发中氯胺酮定性定量分析。方法选择4-苯基丁胺为内标,毛发样本用NaOH、HCl及芳基硫酸酯酶/β-葡萄糖醛酸酶等3种方式进行水解,再进行衍生化后,采用GC/MS和GC/NPD方法定性定量分析。对不同水解和衍生化条件以及提取溶剂进行比较优化,并考察方法精密度、稳定性和检出限。结果方法的提取回收率大于95%,精密度和样品稳定性良好,日内和日间标准偏差小于6%;采用GC/NPD和GC/MS直接分析毛发中的氯胺酮,检出限为0.2ng/mg和2.0ng/mg,线性范围为10.0～250.0ng/mg,相关系数均大于0.99;采用酰化衍生化后分析,GC/NPD和GC/MS检出限分别提高至0.1ng/mg和0.2ng/mg。结论该方法回收率高、检测限低,可以用于毛发中氯胺酮的定性定量分析检验。     

Comparison of ethyl glucuronide and fatty acid ethyl ester concentrations in hair of alcoholics, social drinkers and teetotallers

In previous investigations hair analysis for ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEE) proved to be suitable for the detection of excessive alcohol consumption. The aim of this study was to compare EtG and FAEE concentrations in hair of alcoholics, social drinkers and teetotallers. Hair samples from 10 alcoholics in withdrawal treatment, 11 fatalities with documented excessive alcohol consumption, four moderate social drinkers who consumed up to 20 g ethanol per day, and three strict teetotallers were analysed. After external degreasing with n-heptane, extraction with a dimethyl sulfoxide/n-heptane mixture and headspace solid-phase microextraction of the extracts, four fatty acid ethyl esters (FAEEs) (ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate) were analysed by gas chromatography-mass spectrometry (GC-MS) with deuterated internal standards. EtG was determined by GC-MS/NCI after ultrasonication of the samples with H2O, cleanup by SPE with aminopropyl columns and PFP derivatisation. The following concentrations were measured for the four groups: teetotallers EtG < 0.002 ng/mg, FAEE 0.05-0.37 ng/mg, moderate social drinkers EtG < 0.002 ng/mg, FAEE 0.26-0.50 ng/mg, alcoholic patients EtG 0.030-0.415 ng/mg, FAEE 0.65-20.50 ng/mg and the fatalities with alcohol history EtG 0.072-3.380 ng/mg, FAEE 1.30-30.60 ng/mg. The results confirm that by using a cut-off value of the sum of FAEE > 1 ng/mg and/or a positive EtG result in hair, excessive alcohol consumption can be identified using hair analysis. However, no significant correlation between the EtG and FAEE concentrations in the positive cases could be shown. Segmental analysis of some of the specimens did not reveal the same distribution for EtG compared to FAEE in hair, and no chronological accordance compared to the self-reported alcohol consumption could be observed for both parameters. These different results of both methods are discussed in terms of differences between EtG and FAEE in mechanism of formation and incorporation into hair and elimination from hair.     

Analysis of fatty acid ethyl esters in hair as possible markers of chronically elevated alcohol consumption by headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS)

Fatty acid ethyl esters (FAEE) are products of the nonoxidative ethanol metabolism, which are known to be detectable in blood only about 24h after the last alcohol intake. After deposition in hair they should be suitable long-term markers of chronically elevated alcohol consumption. Therefore, a method for the analysis of ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate from hair was developed based on the extraction of the hair sample by a dimethylsulphoxide (DMSO)/n-hexane mixture, separation and evaporation of the n-hexane phase and application of headspace solid-phase microextraction (HS-SPME) in combination with gas chromatography-mass spectrometry (GC-MS) to the extract. For use as internal standards, the corresponding D(5)-ethyl esters were prepared. The HS-SPME/GC-MS measurements were automatically performed using a multi-purpose sampler. The detection limits of the FAEE were between 0.01 and 0.04ng/mg and the reproducibility was between 3.5 and 16%. By application of the method to hair samples of 21 fatalities with known heavy alcohol abuse 0.045-2.4ng/mg ethyl myristate, 0.35-13.5ng/mg ethyl palmitate, 0.25-7.7ng/mg ethyl oleate and 0.05-3.85ng/mg ethyl stearate were measured. For social drinkers (30-60g ethanol per week), the concentrations were about one order of magnitude smaller. For 10 teetotalers negative results or traces of ethyl palmitate were found. It was shown by supplementary investigations in single cases that FAEE are also present in sebum, that there is no strong difference in their concentrations between pubic, chest and scalp hair, and that they are detectable in hair segments after a 2 months period of abstinence. From the results follows that the measurement of FAEE concentrations in hair is a useful way for a retrospective detection of alcohol abuse.     

Use of solid-phase microextraction (SPME) for the determination of methadone and EDDP in human hair by GC-MS

Solid-phase microextraction (SPME) is a new extraction technique with many advantages: small sample volume, simplicity, quickness and solvent-free. It is mainly applied to environmental analysis, but is also useful for the extraction of drugs from biological samples. In this paper the use of SPME is proposed for the determination of methadone and its main metabolite EDDP in hair by GC-MS. The hair samples were washed, cut into 1-mm segments, and incubated with Pronase E for 12 h. A 100-micron polydimethylsiloxane (PDMS) film fibre was submerged for 30 min in a diluted solution of the hydrolysis liquid (1:4 with borax buffer) containing methadone-d3 and EDDP-d3 as internal standards. Once the microextraction was concluded the fibre was directly inserted into the CG injection port. Linearity was found for methadone and EDDP in the range studied, 1.0-50 ng/mg hair, with correlation coefficients higher than 0.99. Interassay relative standard deviation (R.S.D) was determined to be less than 13.30% for methadone and less than 8.94% for EDDP, at 3.0 and 30.0 ng/mg. Analytical recoveries were close to 100% for both compounds on spiked samples. The method was applied to the analysis of real hair samples from eight patients of a methadone maintenance programme. The concentration of methadone in hair ranged from 2.45 to 78.10 ng/mg, and for EDDP from 0.98 to 7.76 ng/mg of hair.     

Determination of opiates and cocaine in hair using automated enzyme immunoassay screening methodologies followed by gas chromatographic-mass spectrometric (GC-MS) confirmation

The objective of this study was to develop a two-step strategy for analysis of opiates and cocaine in hair samples involving an immunological screening procedure followed by confirmation of results using gas chromatography-mass spectrometry (GC-MS). A semi-quantitative automated competitive enzyme-linked immunosorbent assay (ELISA) methodology using Oral Fluid Micro-Plate Enzyme Immunoassays (Orasure Technologies, Inc.) was developed and validated. Applicability was proven by analysis of authentic head hair samples from drug users (n=103) and from opiate associated fatalities (n=21). The optimum cutoff values for the ELISA tests were 0.1 ng cocaine-equivalents/mg hair and 0.05 ng morphine-equivalents/mg hair using a 50 mg hair sample. Both ELISA tests had a sensitivity of 100%, the specificity was 66% for cocaine-equivalents and 42% for morphine-equivalents. The intraassay precision was 11% for the cocaine and 3% for the opiates ELISA, while interassay precision was 12% for the cocaine and 4% for the opiates ELISA test. The actual analyte concentrations in the hair samples were determined using GC-MS and were between 0.04 and 5.20 ng/mg for heroin (HER), between 0.04 and 30.01 ng/mg for 6-monoacetylmorphine (MAM), between 0.03 and 11.87 ng/mg for morphine (MOR), between 0.02 and 1.84 ng/mg for codeine (COD), between 0.02 and 2.48 ng/mg for acetylcodeine (AC), between 0.01 and 21.37 ng/mg for cocaine (COC), between 0.03 and 10.51 ng/mg for benzoylecgonine (BE) and between 0.05 and 1.26 ng/mg for cocaethylene (CE). The automated ELISA tests were proven to be valid screening procedures for the detection of cocaine and opiates in hair as confirmed by GC-MS. Screening methods provide rapid and inexpensive automated pre-test procedures to detect drugs in hair or other matrices. For forensic purposes screening therefore represents an ideal complement to routinely applied GC-MS procedures.     

Simple and simultaneous analysis of fenfluramine, amphetamine and methamphetamine in whole blood by gas chromatography-mass spectrometry after headspace-solid phase microextraction and derivatization

A simple and sensitive method for the simultaneous analysis of fenfluramine, amphetamine and methamphetamine in whole blood was developed using a headspace-solid phase microextraction (SPME) and derivatization. A 0.5 g whole blood sample, 5 microl d(5)-methamphetamine (50 micrig/ml) as an internal standard, and 0.5 ml sodium hydroxide (1 M) were placed into a 12 ml vial, and sealed rapidly with a silicone septum and an aluminum cap. Immediately after the vial was heated to 70 degrees C in an aluminium block heater, the needle of the SPME device was inserted through the septum of the vial, and the extraction fiber was exposed in the headspace for 15 min. First, heptafluorobutyric anhydride was injected into the injection port of the GC-MS, and the compounds extracted by the fiber were then desorbed and derivatized simultaneously by exposing the fiber in the injection port. The calibration curves, using an internal standard method, demonstrated good linearity throughout the concentration range from 0.01 to 1.0 microg/g. The detection limits of this method were 5.0 ng/g for fenfluramine and methamphetamine, and 10 ng/g for amphetamine. No interferences were found, and the time for analysis was about 30 min for one sample. This method was applied to a suicide case in which the victim ingested fenfluramine. Fenfluramine was detected in the blood sample collected from the victim at the concentration of 7.7 microg/g.     

Solid-phase microextraction for the determination of cocaine and cocaethylene in human hair by gas chromatography-mass spectrometry

A method for the simultaneous determination of cocaine (COC) and cocaethylene (CE) in human hair was developed, using solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) as analytical technique to identify and quantify the drugs. Selected ion monitoring (SIM) mode was used to obtain higher sensitivity. The deuterated-labeled analogues were used as internal standards. The detector response was linear for the drugs studied over the range 0.4-15 ng/mg, with correlation coefficients higher than 0.995. The coefficients of variation oscillated between 0.65% and 14.18% and the accuracy was in the range from 0.73% to 11.20%. The limits of quantitation and detection were found to be acceptable. Finally, this method was applied to 15 hair samples from cocaine users, obtaining positive results in all cases. The mean concentrations were 5.39 ng/mg (range: 0.43-8.98 ng/mg) for cocaine and 1.11 ng/mg (range: 0.42-2.23 ng/mg) for cocaethylene.     

Determination of cathinone,cathine and norephedrine in hair of Yemenite khat chewers

A sensitive and reproducible method for the quantitative determination of cathinone (CTN), norpseudoephedrine (NPE, cathine) and norephedrine (NE) from hair was developed. The compounds were extracted for 4 hours with phosphate buffer pH 2.0, followed by a standard solid phase extraction procedure on a mixed phase column, derivatization with heptafluorobutyric anhydride (HFBA) and GC-MS separation and quantification using D(3)-ephedrine (D(3)-E) and alpha-aminoacetophenone (AAP) as the internal standards. The diastereomers NPE and NE were satisfactorily separated. In the validation, the limits of detection and of quantification were determined at 0.03-0.08 ng/mg and 0.10-0.24 ng/mg, respectively and the interday standard deviation was between 10 and 15%. The method was applied to hair samples of 24 Yemenite khat chewers. All three compounds were detected in 23 of these cases. The concentrations ranged from 0.57 to 23.9 ng/mg for NPE, 0.19-25.0 ng/mg for NE and 0.11-22.7ng/mg for CTN. A highly significant correlation was found between the self-reported data about the khat consumption habits of the volunteers (4-56h chewing per week) and the concentrations of norephedrine and norpseudoephedrine in hair.     

Case report: Etizolam and its major metabolites in two unnatural death cases

A simultaneous analytical method for etizolam and its main metabolites (alpha-hydroxyetizolam and 8-hydroxyetizolam) in whole blood was developed using solid-phase extraction, TMS derivatization and ion trap gas chromatography tandem mass spectrometry (GC-MS/MS). Separation of etizolam, TMS derivatives of alpha-hydroxyetizolam and 8-hydroxyetizolam and fludiazepam as internal standard was performed within about 17 min. The inter-day precision evaluated at the concentration of 50 ng/mL etizolam, alpha-hydroxyetizolam and 8-hydroxyetizolam was evaluated 8.6, 6.4 and 8.0% respectively. Linearity occurred over the range in 5-50 ng/mL. This method is satisfactory for clinical and forensic purposes. This method was applied to two unnatural death cases suspected to involve etizolam. Etizolam and its two metabolites were detected in these cases.     

Methadone and EDDP in hair from human subjects following a maintenance program: results of a pilot study

A specific method has been developed for the quantitative determination of methadone (MTD) and its major metabolite, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), in hair.An amount of 50mg hair samples were incubated in 0.01M HCl overnight at 60 degrees C and deuterated internal standards of MTD and EDDP were added before extraction. Hydrolyzed solutions were extracted by automated solid-phase extraction procedure and analyzed on a gas chromatography (GC) coupled to a ion trap mass spectrometer (MS). Positive chemical ionization was used with acetonitrile as liquid reagent. The different validation parameters, linearity, repeatability, recovery and detection limits are presented. A relative standard deviation (R.S.D.) of 12 and 11% was obtained for the repeatability of MTD and EDDP, respectively. The limits of quantification (LOQ) was 0.05ng/mg for MTD and 0.2ng/mg for EDDP.A number of 26 hair samples from human subjects following a long-term MTD therapy were analyzed by this method. Blood samples of these subjects were analyzed with a routine method using a liquid-liquid extraction and GC/nitrogen phosphorus detector (NPD). MTD was quantified in blood and hair samples and EDDP found in 50% of the hair sample.A comparison was made between the concentrations found in blood or in hair and the dose administrated. This study could demonstrate that there is no relation between the administrated dose and MTD or EDDP concentrations in hair.     

A solid-phase microextraction method for the detection of ignitable liquids in fire debris

A solid-phase microextraction (SPME) procedure involving direct contact between the SPME fibers and the solid matrix and subsequent gas chromatography/mass spectrometric analysis for the detection of accelerants in fire debris is described. The extraction performances of six fibers (100 mum polydimethylsiloxane, 65 mum polydimethylsiloxane-divinylbenzene, 85 mum polyacrylate, 85 mum carboxen-polydimethylsiloxane, 70 mum Carbowax-divinylbenzene, and 50/30 mum divinylbenzene-Carboxen-polydimethylsiloxane) were investigated by directly immersing the fibers into gasoline, kerosene, and diesel fuel. For simulated fire debris, in the direct contact extraction method, the SPME fiber was kept in contact with the fire debris matrix during extraction by penetrating plastic bags wrapping the sample. This method gave comparable results to the headspace SPME method in the extraction of gasoline and kerosene, and gave an improved recovery of low-volatile components in the extraction of diesel fuel from fire debris. The results demonstrate that this procedure is suitable as a simple and rapid screening method for detecting ignitable liquids in fire debris packed in plastic bags.     

Detection of thiopental and pentobarbital in head and pubic hair in a case of drug-facilitated sexual assault

The quali-quantitative determination of two barbiturates, thiopental and its metabolite pentobarbital, in head and pubic hair samples of a woman who had been sexually assaulted during hospitalisation, is reported. Hair was analysed by means of solid-phase microextraction (SPME) and gas chromatography-multiple mass spectrometry (GC-MS-MS), in chemical ionisation conditions. Thiopental and pentobarbital were found in three proximal head hair segments (sample 1A: 0.30 and 0.40 ng/mg; sample 1B: 0.20 and 0.20 ng/mg; sample 3: 0.15 and 0.20 ng/mg) and pubic hair sample. Two distal head hair segments were negative for both barbiturates. Despite the lack of collection and toxicological analysis of blood or urine samples within the hospital setting, analytical findings from hair revealed the use of the anaesthetic agent thiopental to sedate the victim quickly and deeply and commit sexual assault.     

Highly sensitive analysis of methamphetamine and amphetamine in human whole blood using headspace solid-phase microextraction and gas chromatography-mass spectrometry

A simple and highly sensitive method for analysis of derivatized methamphetamine (MA) and amphetamine (AM) in whole blood was developed using headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry electron impact ionization selected ion monitoring (GC-MS-EI-SIM). A whole blood sample, deuterated-MA (d(5)-MA), as an internal standard (IS), tri-n-propylamine and pentafluorobenzyl bromide were placed in a vial. The vial was heated and stirred at 90 degrees C for 30min. Then the extraction fiber of the SPME was exposed at 90 degrees C for 30min in the headspace of the vial while being stirred. The derivatives adsorbed on the fiber were desorbed by exposing the fiber in the injection port of a GC-MS. The calibration curves showed linearity in the range of 0.5-1000ng/g for both MA and AM. The time for analysis was about 80min per sample. In addition, this proposed method was applied to two autopsy cases where MA ingestion was suspected. In one case, MA and AM concentrations in the mixed left and right heart blood were 165 and 36.9ng/g, respectively. In the other case, MA and AM concentrations were 1.79 and 0.119 microg/g in the left heart blood, and 1.27 and 0.074 microg/g in the right heart blood, respectively.     

Aqueous phase hexylchloroformate derivatization and solid phase microextraction: determination of benzoylecgonine in urine by gas chromatography-quadrupole ion trap mass spectrometry.

A derivatization/solid phase microextraction (SPME) method for the determination of benzoylecgonine in urine was developed. The derivatization is conducted directly in 1 mL of urine while sonicating for 3 min with 12 microL of hexyl chloroformate and 70 microL of a mixture containing acetonitrile:water:hexanol:2-dimethylaminopyridine (5:2:2:1 v/v), yielding benzoylecgonine hexyl ester (BHE) as the product. After the 3 min period, an aliquot of 250 microL is transferred to a vial for SPME. After the desired extraction time the 100 microns polydimethylsiloxane SPME fiber was transferred to the GC-MS for separation and analysis with a quadrupole ion trap mass spectrometer. The hexyl chloroformate derivatization and SPME procedures were optimized for compatibility and sensitivity. The method was found linear for 0.10 to 20.0 micrograms/mL (r2 = 0.999) of benzoylecgonine in urine using benzoylecgonine-d3 as an internal standard (1.5 micrograms/mL). Intra-day precisions were 8.8 and 6.8% RSD for 0.30 microgram/mL and 17 micrograms/mL benzoylecgonine standards in urine (n = 6), respectively. Inter-day precision (n = 3) were < or = 3.3% RSD, indicating good reproducibility. A detection limit of 0.03 microgram/mL (S/N = 3) was achieved, thus making the SPME method a simplified alternative to SPE for GC-MS confirmation after EMIT tests for benzoylecgonine which have a cutoff of 0.30 microgram/mL. Quantitative results by SPME and SPE of two clinical urine specimens known positive for cocaine by EMIT were in excellent agreement. Benzoylecgonine was detected by the derivatization/SPME method in 22 out of 22 other urine specimens known positive for cocaine.     

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.     

HS-SPME-GC/MS法检测尿液及毛发中苯丙胺类毒品

目的采用顶空固相微萃取（HS-SPME）、GC/MS分析方法,对生物样品中苯丙胺（AM）、甲基苯丙胺（MAM）、3,4-亚甲二氧基苯丙胺（MDA）和3,4-亚甲二氧基甲基苯丙胺（MDMA）4种苯丙胺类毒品进行定性定量分析。方法在碱性和饱和盐处理状态下,采用100μm聚二甲基硅氧烷（PDMS）萃取纤维,于顶空瓶中进行生物样品AM、MAM、MDA、MDMA 4种毒品萃取,以2-甲基苯乙胺为内标,经气-质联用选择离子检测（GC/MS/SIM）模式进行定性定量分析。对HS-SPME条件优化,对方法的精密度、准确度和检出限进行测定。结果 AM、MAM、MDA、MDMA 4种毒品尿液中的最低检出限为5ng/mL,毛发中的最低检出限为0.5ng/mg。尿液中线性关系范围为0.05μg/mL~5μg/mL,r〉0.991,回收率为82%~108%,RSD为2.6%~6.1%（n=5）;毛发中线性关系范围为5ng/mg~500ng/mg,r〉0.992,回收率为80%~113%,RSD（%）为1.4%~6.8%（n=5）。结论 HS-SPME-GC/MS各项定量参数符合分析要求。该方法简单、灵活、经济、快速、无溶剂,适用于生物检材中该类毒品的分析。     

Preferential extraction of hydrocarbons from fire debris samples by solid phase microextraction

Headspace analysis by extraction/GC-MS is a common method of detecting volatile hydrocarbon accelerants in fire debris samples. Solid-phase microextraction was tested to determine if there is selective extraction of chemically distinct compounds. It was found that both the polydimethylsiloxane (PDMS) and Carboxen/PDMS solid phase microextraction fibers show preferential extraction of aliphatic or aromatic compounds from the headspace depending on fiber type and temperature. The Carboxen/PDMS fiber type showed particular (although not exclusive) selectivity for extraction of aromatic hydrocarbons. Other experimental considerations of SPME are noted.     

Improved gas chromatography-negative ion chemical ionization tandem mass spectrometric method for determination of 11-nor-Δ9-tetrahydrocannabinol-9-carboxylic acid in hair using mechanical pulverization and bead-assisted liquid-liquid extraction

A gas chromatography-negative ion chemical ionization tandem mass spectrometric (GC-NCI-MS/MS) method was developed and validated for the determination of 11-nor-Δ(9)-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in human hair. After decontamination, hair samples were weighed (25mg), mechanically pulverized with a bead mill, and incubated in 0.7 mL of 1.0M sodium hydroxide at 95 °C for 30 min. Bead-assisted liquid-liquid extraction was performed with n-hexane:ethyl acetate (9:1, v/v), a method developed in our laboratory. The extract was evaporated to dryness, derivatized with pentafluoropropanol and pentafluoropropionic anhydride, and analyzed by GC-MS/MS in the negative ion chemical ionization mode using methane as the reagent gas. The linear ranges were 0.05-10.0 pg/mg for THC-COOH with the coefficient of determination (r(2) = 0.9976). The intra-day and inter-day precisions were within 1.7 and 13.8%, respectively. The intra-day and inter-day accuracies were -4.8 to 10.0% and -3.9 to 3.8%, respectively. The limit of detection and quantification were 0.015 and 0.05 pg/mg, respectively. The recoveries were in the range of 79.4-87.1%. The results indicate that the proposed method is simple, rapid, accurate, and precise for determination of THC-COOH in hair. The method identified THC-COOH in hair specimens from suspected marijuana abusers.     

A study into the rate of incorporation of eight benzodiazepines into rat hair

The incorporation of eight benzodiazepines (chlordiazepoxide, diazepam, estazolam, flunitrazepam, flurazepam, medazepam, oxazepam and triazolam) into rat hair was investigated by HPLC and GC-MS. Each of the benzodiazepines was injected daily into three Dark Agouti (DA) rats for 10 days at 10mg/kg. The back hair of the rats was removed by shaving prior to the first injection and again on the 28th day after the initial administration.To investigate optimum extraction conditions, 10mg aliquots of rat hair incorporated with diazepam, flurazepam or medazepam were extracted by seven different methods (Proteinase K, methanol-ammonia, methanol-trifluoroacetic acid, Soerensens buffer, 1M NaOH, beta-glucuronidase/arylsulfatase, Biopurase). The method found to yield the highest recoveries, for all three drugs, was the acidic methanol extraction. Using this extraction procedure, the incorporation rates (ICR: the ratio of the hair concentration to the plasma AUC) of eight benzodiazepines into rat hair were investigated. The ICRs ranged from 0.002 (flunitrazepam) to 0.049 (flurazepam).The major metabolites of flurazepam were investigated in rat hair. The mean hair concentrations of desalkylflurazepam and 2-hydroxyethylflurazepam were 3.31 and 0.05 ng/mg, respectively, which are 24 and 0.36% of the parent compound in hair.