Simultaneous quantification of opiates, cocaine and cannabinoids in hair

The present paper describes a sensitive method developed in our laboratory for the simultaneous analysis of opiates (morphine, codeine and monoacetylmorphine), cocainics (cocaine and benzoylecgonine) and cannabinoids (Δ⁹-tetrahydrocannabinol and 11-nor-Δ⁹-tetrahydrocannabinol-9-carboxylic acid) in hair samples. After decontaminating the sample with dichloromethane, two consecutive hydrolyses were performed in order to achieve the best conditions for extracting the three kinds of drugs from the protein matrix. First the opiate and cocainic compounds were extracted by means of a soft acidic hydrolysis with 0.1 N HCl at 50 °C overnight and organic solvent extraction at pH 9.2. The cannabinoids need a stronger basic hydrolysis with 11.8 N KOH for 10 min at laboratory temperature. After adding maleic acid, the cannabinoids were extracted with an organic solvent. The derivatization was carried out with heptafluorobutyric anhydride and hexafluoropropanol. Calibration curves were linear between 0.5–100 ng/mg of hair. Recovery and reproducibility were assured. The quantification limits ranged between 0.04–0.26 ng/mg of hair. Seventy hair samples from known drug abusers were cut into 1-cm segments and analyzed by this method. The ranges of measured concentrations (ng/mg) were 0.31–89 for cocaine, 0.1–5.76 for benzoylecgonine, 0.34–45.79 for morphine, 0.45–39.59 for codeine, 0.09–48.18 for monoacetylmorphine, 0.06–7.63 for THC and 0.06–3.87 for THC---COOH. The results of sectional analyses agreed with the self reported drug histories. The usefulness of this method is in assessing earlier drug consumption, and also at the same time obtaining a chronological profile of the consumption of these three types of drugs.     

Radioimmunological determination of cocaine in human hair

A simple procedure for the determination of cocaine in human hair was described. After washing hair samples were crushed in 0.1 m HCl and incubated overnight at 45 degrees C. The acid extracts were neutralized with 1 m NaOH. Phosphate buffer (pH 7.4) was added to the extracts. The cocaine concentrations were measured by radioimmunoassay. Detection in hair was achieved in all hair samples obtained from cocaine users. This method appears to be suitable for the routine determination of cocaine.     

GC／MS同时分析头发中大麻酚类和△^9-四氢大麻酸

目的建立同时分析毛发中THC、CBN、CBD和大麻主要代谢物THC-COOH的方法.方法头发检材去污处理后,加入内标氯灭酸,经NaOH水解和正己烷:乙酸乙酯(9:1)提取,吹干后衍生化,利用GC/MS-SIM方法分析.结果THC、CBN和THC-COOH的最低检出限分别为0.01、0.05和0.01ng/mg.10例阳性头发中均检出THC成分,THC浓度范围为0J 1～8.84ng/mg,有3例未检出THC-COOH,检出者的量亦低于定量下限.结论同时分析头发中的大麻酚类和△9-四氢大麻酚是可行的,头发中THC-COOH浓度明显都低于THC浓度.     

Levels of cocaine and its metabolites in washed hair of demonstrated cocaine users and workplace subjects

In a study of subjects in drug rehabilitation programs, cocaine and cocaine metabolite levels were determined in the hair of 75 subjects who had produced cocaine-positive urine results. The hair was analyzed after being washed with the 3.75 h wash procedure developed by this laboratory. In addition, results of testing 73 non-users are presented, as well as levels of cocaine, benzoylecgonine (BE), cocaethylene, and norcocaine from workplace population samples. The data support a recommendation of reporting as positive a sample with cocaine of 500 pg/mg hair and either a 5% ratio of benzoylecgonine (BE) to cocaine in samples, or the presence of cocaethylene at 50 pg/mg hair, or norcocaine at 50 pg/mg hair for samples < or =2000 pg cocaine/mg hair. For samples with cocaine present at >2000 pg/mg hair, the data indicate that a ratio of 5% BE may be an overly conservative approach. In appropriately washed hair samples, cocaine users can produce hair levels of <5% BE and thus a minimum BE cutoff in lieu of a ratio could be considered.     

Validity testing of commercial urine cocaine metabolite assays: IV. Evaluation of the EMIT d.a.u. cocaine metabolite assay in a quantitative mode for detection of cocaine metabolite

The EMIT d.a.u. cocaine metabolite assay (EMIT dau) was evaluated in a quantitative mode for analysis of clinical specimens obtained after controlled cocaine administration to human subjects. The quantitative results showed high concordance with those of gas chromatography/mass spectrometry (GC/MS) assays of the same specimens for benzoylecgonine, and no false positive or false negative results were obtained. The evaluation also included analysis of standardized solutions containing benzoylecgonine, cocaine, and other cocaine metabolites and isomers. The EMIT dau antibody demonstrated high selectivity for benzoylecgonine. The precision was somewhat less than that reported earlier for other commercial cocaine metabolite immunoassays. Quantitation of initial screening results from EMIT dau testing can serve as a useful guide for confirmation by GC/MS in forensic science urine testing.     

Validity testing of commercial urine cocaine metabolite assays: III. Evaluation of an enzyme-linked immunosorbent assay (ELISA) for detection of cocaine and cocaine metabolite

A validity assessment study was performed on the Genetic Diagnostic Enzyme Immunoassay test kit, a new enzyme-linked immunosorbent assay (GDC ELISA) for detection of cocaine and cocaine metabolite in urine. A set of 290 urine specimens, comprised of clinical cocaine urines collected from 5 male subjects who had received single doses of intravenous cocaine, drug-free urines spiked with cocaine, cocaine metabolites, cocaine isomers, and other drugs of abuse, were assayed by GDC ELISA. The results were compared with results by gas chromatography/mass spectrometry (GC/MS) assay for benzoylecgonine. Concordance was high between the GDC ELISA assay and GC/MS and with results reported earlier for other commercial assays. Detection times and specificity of the GDC ELISA antibody were most similar to those of the Abuscreen radioimmunoassay for cocaine metabolite. Overall, the assay produced no false negative or false positive results and appeared to be a reliable screening test for detection of cocaine and benzoylecgonine in human urine.     

毛发中海洛因及其代谢物的分析综述

对毛发中海洛因及其代谢物—6-单乙酰吗啡和吗啡的提取和检测分析进行了综述。其分析方法大致为6步:收集毛发,清洗毛发,分段,剪(磨)碎毛发,水解,提取净化,检测分析。     

The determination of cocaine in hair: a review

The explosion of literature related to the analysis of hair for cocaine and its products is reviewed. In the commonly accepted applications of hair testing for cocaine, those related to criminal or civil investigations and pharmacotoxicologic studies occupy most of the relevant published work. This review uses detailed, ‘binary’ (yes/no) tables to demonstrate trends in the literature, and allows researchers and caseworkers quick access to the literature most important for answering a variety of questions.     

Disappearance of cocaine from human hair after abstinence

In this work the study of the disappearance of cocaine in hair is reported. The subject of the study is a woman who stopped the consumption of cocaine after a period of drug abuse of over 1 year. Hair samples were collected over a period of 10 months. During this time the absence of cocaine intake was monitored by the toxicological analysis of urine, performed every 2 days. After decontamination with methanol, the hair sample, cut in two segments (0-1.5 and 1.5-3 cm from the hair root) was added with cocaine-D(3) (internal standard), hydrolyzed and extracted with chloroform/isopropanol (9:1). The extract was evaporated to dryness, reconstituted in 25 microl of ethyl acetate and analyzed by GC-MS in SIM mode. The obtained results show that the incorporation of cocaine in hair decreased during the first 3 months after the last consumption and after this period of time no cocaine was found in the hair sections closest to the root.     

Segmental hair analysis for cocaine and heroin abuse determination

Segmental hair analysis was performed to obtain information about the history of drug abuse of subjects in a rehabilitation programme. The analytical data from hair samples were correlated, when possible, with urine analysis and to toxicological anamnesis. Toxicological analysis of hair seems to be a valid tool in this specific field.     

毛发中甲基苯丙胺及代谢产物苯丙胺的分析研究

介绍了毛发中甲基苯丙胺（ MAMP）及代谢产物苯丙胺（ AMP）的 GC/NPD、 GC/MS的定性定量分析方法。毛发用 0.1 mol/L HCl水解, 4－苯基丁胺（ 4－ PBA）为内标,液－液提取,三氟乙酸酐（ TFA）衍生化。毛发用量为 10mg,检出限为 GC/NPD 0.5ng/mg, GC/MS 0.1ng/mg,回收率大于 78％。该方法成功应用于染毒豚鼠毛发中 MAMP及其代谢产物 AMP浓度变化过程的测定。     

Optimized conditions for simultaneous determination of opiates, cocaine and benzoylecgonine in hair samples by GC--MS

The present paper describes a qualitative and quantitative method for the simultaneous detection of opiates, cocaine and benzoylecgonine from human hair samples. Every step of the analytical procedure was studied to find the optimized conditions. Nine different incubation systems were examined. The influence of different pH values of samples on the isolation of analytes from the incubation media by Bond Elut cartridges and the stability of the compounds of interest in the different incubation media and conditions were investigated. The extracting power of different incubation media was studied as well. The phosphate buffer 0.1 N at pH 5 was chosen as the extraction medium in an optimized procedure for simultaneous determination of opiates, cocaine and benzoylecgonine in hair samples. The method developed was validated. Recoveries were 90% for morphine (M), 81% for 6-monoacetylmorphine (6-AM), 90% for codeine (CD), 86% for cocaine (C) and 90% for benzoylecgonine (BE). Relative standard deviation for inter-day precision was better than 12%. The limits of detection resulted as 0.05 ng/mg for M and C, as 0.08 for 6-AM and as 0.2 ng/mg for BE. Forty hair samples collected from drug abusers admitted to centers for detoxification treatment were analyzed obtaining 23 positive results for opiates and/or cocaine. Twelve hair specimens longer than 10 cm were analyzed following a sectional approach. In the six positive cases, it was interesting to find that the 6-AM/M ratio generally decreased for each sample from the proximal segment to the distal segments. Moreover, the 6-AM/M ratio was generally lower than 1 in the intermediate and distal segments.     

Quantitative determination of amphetamines, cocaine, and opiates in human hair by gas chromatography/mass spectrometry

Hair of young subjects (N = 36) suspected for drug abuse was analysed for morphine, codeine, heroin, 6-acetylmorphine, cocaine, methadone, amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and 3,4-methylenedioxyethylamphetamine (MDEA). The analysis of morphine, codeine, heroin, 6-acetylmorphine, cocaine, and methadone in hair included incubation in methanol, solid-phase extraction, derivatisation by the mixture of propionic acid anhydride and pyridine, and gas chromatography/mass spectrometry (GC/MS). For amphetamine, methamphetamine, MDA, MDMA, and MDEA analysis, hair samples were incubated in 1M sodium hydroxide, extracted with ethyl acetate, derivatised with heptafluorobutyric acid anhydride (HFBA), and assayed by GC/MS. The methods were reproducible (R.S.D. = 5.0-16.1%), accurate (85.1-100.6%), and sensitive (LoD = 0.05-0.30ng/mg). The applied methods confirmed consumption of heroin in 18 subjects based on positive 6-acetylmorphine. Among these 18 heroin consumers, methadone was found in four, MDMA in two, and cocaine in two subjects. Cocaine only was present in two, methadone only in two, methamphetamine only in two, and MDMA only in seven of the 36 subjects. In two out of nine coloured and bleached hair samples, no drug was found. Despite the small number of subjects, this study has been able to indicate the trend in drug abuse among young people in Croatia.     

Evaluation of cocaine, amphetamines and cannabis use in university students through hair analysis: preliminary results

The evaluation of drug abuse in a defined population was performed through toxicological hair analysis. Hair samples from university students ranging from 18 to 25 years of age were anonymously collected and screened for cocaine, amphetamines and cannabinoids by radioimmunoassay (RIA). Positive results (cut-off values adopted were 2 ng/mg for cocaine and amphetamines and 0.5 ng/mg for cannabinoids) were confirmed by GC/MS. Preliminary results showed 19% of positive results for cocaine on 200 samples analysed. No confirmed positive results were obtained for amphetamine analysis. RIA technique demonstrated its unsuitability for cannabinoids preliminary screening on hair, giving a high percent of false positive results.     

Detection of cocaine in blood stains]

Cocaine is rapidly degraded in blood samples, and its degradation was found to be highly dynamic in nature. The analysis of blood spots dried on filter paper may provide a method to minimize the break-down of cocaine and to largely preserve the analytical profile of the parent drug and its hydrolysis products at the time of sampling. The short term stability of cocaine in 100 microL blood spots prepared from unpreserved and preserved (sodium fluoride, 0.25%) blood samples was compared to the stability of the particular whole blood specimens stored in tubes at ambient temperature and at -20 degrees C. Due to dehydration, both the chemical and the enzymatic hydrolysis of cocaine and its products could be stopped in dried blood spots. More than 75% of the initial cocaine concentration could be detected in the blood spots, and the analytical profile was ensured for 17 days. Provided its practical suitability, the spot technology should offer a simple approach to detect actual impairment of motorists taken in police custody in the view of section 24a of the German traffic act as well as in cocaine associated criminal cases.     

Correlation of the incidence of cocaine and cocaethylene in hair and postmortem biologic samples

Hair samples are useful as a matrix for drug testing because drugs can be detected in hair for longer periods than in blood or urine. The authors report a prospective comparison of the detection of cocaine and cocaethylene in routine postmortem biologic specimens to the detection of cocaine and cocaethylene in hair. The authors collected hair samples from various areas of the head in 53 autopsy cases, prepared them, and analyzed them by gas chromatography/mass spectrometry (GC/MS) for cocaine and cocaethylene. The authors compared the results of hair analysis with the results of toxicologic analysis performed on routine postmortem samples by enzyme multiplied immunoassay technique and GC/MS. Cocaine was found in either biologic fluids or in hair in 16 of 53 samples tested. Nine samples were positive for cocaine in both biologic fluids and hair. Five samples contained cocaine only in biologic fluids, and two contained cocaine only in hair. Cocaethylene was present in two cases. Drug screening of hair provides additional information in some autopsy cases, but the authors have not made hair analysis a routine practice. It may prove useful to save hair samples in all cases for later analysis if warranted by additional history or autopsy findings.