Quantitative analysis of methamphetamine in hair of children removed from clandestine laboratories--evidence of passive exposure?

In New Zealand many children have been removed from clandestine laboratories following Police intervention. In the last few years it has become standard procedure that these children have hair samples taken and these samples are submitted to the laboratory for analysis. There are various mechanisms for the incorporation of drugs into hair. The hair follicle has a rich blood supply, so any drug that may be circulating in the blood can be incorporated into the growing hair. Another mechanism is via external contamination, such as spilling a drug on the hair or through exposure to fumes or vapours. Hair samples were analysed for methamphetamine and amphetamine. From the 52 cases analysed 38 (73%) were positive for methamphetamine (>0.1 ng/mg) and amphetamine was detected in 34 of these cases. In no case was amphetamine detected without methamphetamine. The hair washes (prior to extraction) were also analysed (quantified in 30 of the positive cases) and only 3 had a wash to hair ratio of >0.1 (all were <0.5), which may be indicative of a low level of external contamination. This low level of evidence of external contamination suggests that the children are exposed to methamphetamine and are incorporating it into the hair through the blood stream.     

单次摄药毛发分析的研究进展

“迷药犯罪”是指对被害人下药后实施性侵犯、抢劫、诈骗等犯罪行为。单次摄入的镇静催眠类或迷幻类药物在体内迅速代谢,故难以用血液、尿液检测等常规方法来提供摄药证据,而毛发分析的长检测窗特点在解决这类案件时具有重要的价值。单次摄药的毛发分析要求检测方法有很高的灵敏度,需要用二级质谱检测器进行分析,且从头发采样、去污、水解、提取、分析的各环节均应注意防止污染,避免出现假阳性结果。采集的头发必须进行分段分析,除检测对应案发时间的头发段外,还应把其它头发段的检测结果作为对照,据此对分析结果作出严谨、科学的解释。目前已建立了31种镇静催眠药、6种苯丙胺类衍生物、GHB、硫喷妥及其代谢物戊巴比妥以及乙醇代谢物EtG等的检测方法,可以应用于实际案件的毛发分析。     

Detection of phentermine in hair samples from drug suspects

Phentermine (PT) has been widely used as an anti-obesity drug. This drug has to be used with caution due to its close resemblance with amphetamines in its structure and toxicity profile. Recently, PT is in distribution by illegal modes and is found to be available through sources such as the internet, thus their misuse and/or abuse is threatening to be a serious social issue. In the present study, 32 cases of drug suspects were observed for PT abuse, detected using hair samples for drug analysis. PT and other amphetamines, such as methamphetamine (MA), amphetamine (AP), 3,4-methylenedioxyamphetamine (MDMA) and 3,4-methylenedioxyamphetamine (MDA), were extracted using 1% HCl in methanol for 20 h at 38°C. The extracts were derivatized with trifluoroacetic anhydride (TFAA) and analyzed using gas chromatography/mass spectrometry (GC/MS). Among the 32 cases of PT abuse, MA and its main metabolite, AP were identified in seven cases and MDMA and its main metabolite, MDA were detected in two other cases.     

Hair analysis for drug abuse: I. Determination of methamphetamine and amphetamine in hair by stable isotope dilution gas chromatography/mass spectrometry method

Determination of methamphetamine and amphetamine in hair was performed by gas chromatography/mass spectrometry using stable isotope-labeled internal standards, 2-methylamino-1-phenylpropane-2,3,3,3-d4 and 2-amino-1-phenylpropane-2,3,3,3-d4. Extraction of hair with methanol/5M hydrochloric acid (20:1) using ultrasonication was chosen as the standard method. The calibration curves for amphetamines in the hair were linear from 1 to 100 ng/mg (r greater than 0.99). The detection limit was 0.5 ng/mg at the 95% confidence level. The coefficients of variation (CV) (n = 8) of analysis using the spiked hair with methamphetamine were from 0.7 to 6%. The CV (n = 8) of analysis of the methamphetamine abuser's hair was 17.5%. Sectional analysis of monkey and human hair after methamphetamine ingestion suggested a good correlation between the duration of drug use and drug distribution in the hair.     

Detection and diagnostic interpretation of amphetamines in hair

A review with 22 references on detection and incorporation of amphetamines in hair is presented. This review deals with the detection, incorporation into hair, behavior in the hair shaft, confirmation of past drug use and diagnosis of dependence mainly regarding amphetamine and methamphetamine, along with methoxyphenamine, methylenedioxymethamphetamine, bromomethamphetamine, deprenyl, benzphetamine, fenproporex and mefenorex. First, pretreatment, extraction and analytical methods for amphetamines in hair using immunoassay, HPLC and GC/MS are discussed. This is followed by sections describing the animal experiments, incorporation rates of amphetamines from blood to hair and relationship between drug history and drug distribution in hair. Finally, the diagnosis of amphetamine dependence and confirmation of methamphetamine baby by hair analysis is discussed. The paper concludes with a brief outlook.     

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.     

Performance of immunoassays in screening for opiates,cannabinoids and amphetamines in post-mortem blood

Several immunoassay methods for screening of abused drugs in whole blood were evaluated in post-mortem forensic toxicology. Blood samples known to be positive or negative for opiates, cannabinoids or amphetamines by gas chromatography-mass spectrometry (GC-MS) were analysed by EMIT II Plus and EMIT d.a.u., Syva RapidTest and Triage 8 after acetone precipitation. In these experiments, the EMIT immunoassay method was modified by using the Dade Behring VIVA analyser to detect substances more sensitively. Low concentrations of abused drugs were detected in blood samples. The sensitivities of the modified EMIT method for opiates, cannabinoids and amphetamines were 100, 86 and 98%, respectively, whereas the values were below 86% with the other methods. The specificities of all immunoassay methods for opiates and cannabinoids were 83% or above but 51-85% for amphetamines. Sample rejection occurred in a few cases with the EMIT amphetamine assays. The modified EMIT immunoassay system presented here seems to be useful for screening of drugs of abuse in post-mortem blood samples, especially when urine is not available.     

Hair analysis for drugs of abuse. Hair color and race differentials or systematic differences in drug preferences?

There is currently a debate in the literature on chemical drug analysis concerning the contribution of biophysical attributes associated with specimens and specimen donors to assay outcome. In recent years this debate has focused on hair analysis, but has in the past also been raised in urinalysis interpretation. In this article we examine several aspects of that controversy. First, we present data regarding the effects of hair color on the distribution of positive hair testing results for three drug classes. We compare these results to negative hair samples from comparable donors. This data is derived from head hair from preemployment donors that was classified according to seven visual color categories. We determined the distribution of colors for hair samples devoid of any of three assayed drugs (amphetamines, cocaine, and cannabinoids). Subsequently, this distribution was compared with the distributions for hairs that had tested positive for amphetamines, cocaine or cannabinoids. We examined a total of 2000 randomly selected samples; 500 negative hair samples and 500 positive samples for each of three drugs: cannabinoids, cocaine, and amphetamine. We also evaluated ethnic/racial factors in relation to positive urinalyses for various ethnic/racial groups. We examined approximately 4000 urine specimens from two different groups, each constituting around 2000 specimens. In addition to ethnicity/race and urinalysis outcome, we also examined the relationship between the hair color distributions of urine donors and the corresponding urinalysis results for the three drug classes. We also compared them to drug-negative samples. Our summary impression is that the observed outcome patterns were largely consistent with differences in drug preferences among the various societal groups. There was little evidence of a pattern attributable to hair color bias alone or selective binding of drugs to hair of a particular color. Likewise, there was no discernible pattern associated with race or ethnicity that would lend support to a "race effect" in drug analysis.     

Hair analysis: self-reported use of "speed" and "ecstasy" compared with laboratory findings

Drug use histories were collected from 100 subjects recruited from the "dance scene" in and around Glasgow, Scotland. In addition, each subject donated a hair sample which was analyzed by gas chromatography/mass spectrometry (GC/MS) for amphetamine (AP), methamphetamine (MA), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MD MA) and 3,4-methylenedioxyethylamphetamine (MDEA). The hair samples were analyzed in two 6 cm segments or in full, ranging from 1.5 to 12 cm depending on the length of the hair. Approximately 10 mg of hair was ground to a fine powder before treatment with beta-glucuronidase/aryl sulfatase. A solid-phase extraction procedure was carried out followed by derivatization with pentafluoropropionic anhydride (PFPA). All extracts were analyzed by gas chromatography/mass spectrometry (GC/MS). Of the 139 segments analyzed, 77 (52.5%) were positive for at least one of the five amphetamines. The drug concentrations found in the hair were compared with the self-reported drug histories. A concordance of greater than 50% was found between the self-report data and levels detected in hair. However, no correlation was found between the reported number of "ecstasy" tablets consumed and the drug levels detected in hair. An increase in the average drug levels measured was observed from low to high use (number of "ecstasy" tablets/month). A large number of false negatives and a low number of false positives were observed.     

Proficiency test for the analysis of hair for drugs of abuse,organized by the Society of Hair Testing

Eighteen laboratories interested in the analysis of human hair for drugs of abuse participated in a proficiency test (PT) organized by the Society of Hair Testing (SoHT) in 2001.Samples sent to the participants included one drug-free hair sample and two samples from drug users, sent in the form of short segments previously checked for homogeneity by three reference laboratories. Participants were requested to analyze the samples following the standard procedure used routinely in their laboratories.The compounds present in the samples included opiates, cocaine and metabolite, cannabinoids and amphetamines. All the laboratories analyzed opiates, cocaine and benzoylecgonine (BE); only 10 analyzed amphetamines, and 9 cannabinoids. Various methods were used to extract drugs from the hair-enzyme treatment, acidic, basic and methanol extractions. All the laboratories employed GC-MS, with the exception of two which used GC-MS/MS and LC-MS/MS, respectively. Six laboratories performed initial screening tests by RIA, ELISA or EMIT.Results show that the laboratories performed well qualitatively, since they successfully identified all the analytes that they tested, with the exception of eight false results. However, the scatter of quantitative results was high.     

Patterns in drug use in the United Kingdom as revealed through analysis of hair in a large population sample

This paper presents an overview of the most common sectioning patterns utilised in the analysis of hair for drug use; report on the major user groups (sectors) that currently make use of hair analysis in the United Kingdom (UK); present the results for the different drug groups analysed in samples of hair samples analysed at TrichoTech between 2001 and 2005.A total of 186,084 tests on 34,626 hair samples were performed for the commonly requested drug groups. There were 145,799 enzyme-linked Immunosorbent positive screening tests (ELISA), which were subsequently confirmed by gas chromatography equipped with mass spectrometry detection (GC–MS). The two major sectors were the Medico-Legal sector (65%) and Workplace (20%). Police (Forensics), Clinical Monitoring, Schools, Research and Insurance accounted together for the remaining 15% of the samples. Combinations of several sections patterns were requested covering periods from the most recent month up to 24 months. The most common sectioning pattern was one single section measuring 3 cm, to cover the most recent 3 months (44%), which in some cases was complemented by a further 3 cm to cover together 6 months (13%). The second most common sectioning pattern was the analysis of three sections of 1 cm each to cover the most recent 3 months (28%), when a more detailed evaluation of drug use pattern was relevant. Samples collected from other areas of the body such as axilla, pubic, chest, beard and leg, constituted 6% of the samples. The analysis of monthly sections plays an important role in the evaluation and interpretation of drug use, particularly in certain Medico-Legal cases. The sectors with the highest rates of positive results were Police (Forensics) (78%), Medico-Legal (62%) and Clinical (54%). The common drugs in each group were cannabinol (27%), cocaine (25%), morphine (17%), amphetamine (13%) and diazepam (15%). The positive rate for the Workplace sector was 10%. The most common drugs detected in the Workplace samples in each group were: THC (4%), codeine (2%), cocaine (2%), MDMA (0.5%) and diazepam (0.1%). The concentration levels of drugs found in samples from the workplace were lower than in the other sectors (95% of cases). The exceptions were for dihydrocodeine and MDMA, where levels were 170 and 143% higher, respectively. However, the maximum levels detected in the Workplace samples were lower. The Medico-Legal sector is the most prevalent sector using hair analysis in the UK but the rate of Workplace sector use of hair testing is increasing. One in 10 workplace hair tests detected the presence of at least one drug, which is twice the rate of detection using urine, which is a 1 in 20 urine sample. This means that the chances of identifying people on drugs in the workplace by testing hair samples are twice as likely than urine samples.     

Use of headspace solid-phase microextraction (HS-SPME) in hair analysis for organic compounds

Headspace solid phase microextraction (HS-SPME) has advantages of high purity of the extract, avoidance of organic solvents and simple technical manipulation and can be used in combination with gas chromatography-mass spectrometry (GC-MS) in the hair analysis of a number of drugs. HS-SPME coupled with the hydrolysis of the hair matrix by 4% sodium hydroxide in the presence of excess sodium sulphate and of a suitable internal standard proved to be a convenient one-step method for the measurement of many lipophilic basic drugs such as nicotine, amphetamine derivatives, local anaesthetics, phencyclidine, ketamine, methadone, diphenhydramine, tramadol, tricyclic antidepressants and phenothiazines. Detection limits were between 0.05 and 1.0 ng/mg. From spiked 10-mg hair samples absolute recoveries between 0.04 and 5.7% were found. These recoveries decreased considerably if larger sample amounts were used, perhaps due to increased drug solubility in the aqueous phase or to elevated viscosity in the presence of dissolved hair proteins. Because of the phenolic hydroxyl group a change of pH after alkaline hair digestion (by adding excess orthophosphoric acid) was necessary for the detection of delta 9-tetrahydrocannabinol (delta 9-THC), cannabinol (CBN) and cannabidiol (CBD) by HS-SPME. Nevertheless, the detection limits were such that only CBN could be detected in hair of a consumer. Clomethiazole, a compound hydrolysed in alkali, was measured by HS-SPME after extraction with aqueous buffer. The detection limit was 0.5 ng/mg. Cocaine could not be detected by HS-SPME. The application of HS-SPME to hair samples from several forensic and clinical cases is described.     

Evaluation of the IDS One-Step ELISA kits for the detection of illicit drugs in hair

This work presents the validation of a new immunological assay, the One-Step enzyme-linked immunosorbent assay (ELISA) tests from International Diagnostic Systems Corp. for the screening of drugs of abuse (cannabis, amphetamines, opiates, and cocaine) in human hair, with subsequent GC-MS confirmation. After decontamination and segmentation into small pieces, 50 mg of hair sample were incubated in 1 ml of methanol during 16 h at 40 degrees C. A 100 microL aliquot was collected and evaporated to dryness in presence of 100 microL of methanol/hydrochloric acid (99:1, v/v) to avoid amphetamines loss. The dried extract was dissolved in 100 microL of the "sample and standard diluent" solution included in the kit. This solution was submitted to analysis according to the recommended instructions of the manufacturer. During the validation phase, GC-MS confirmations were conducted according to our fully validated and published methods for opiates, cocaine, cannabis, and amphetamines determinations in hair. In a last development step, these procedures were slightly modified to directly confirm ELISA results by GC-MS using the methanolic extract. Ninety-three specimens were simultaneously screened by the ELISA tests (103 for tetrahydrocannabinol (THC)) and confirmed by GC-MS. Twenty were found positive for cannabis (THC: 0.10-6.50 ng/mg), 21 for cocaine (0.50-55.20 ng/mg), 24 for opiates (6-acetylmorphine (6-AM): 0.20-11.60 ng/mg, MOR: 0.20-8.90 ng/mg, codeine (COD): 0.20-5.90 ng/mg), and 13 for amphetamines (AP: 0.20 and 0.27 ng/mg, methamphetamine (MAP): 0.30 and 1.10 ng/mg, methylenedioxymethamphetamine (MDMA): 0.22-17.80 ng/mg). No false negative results were observed according to the Society of Hair Testing's (SoHT) cutoffs (0.5 ng/mg for cocaine, 0.2 ng/mg for opiates and amphetamines, and 0.1 ng/mg for THC). The One-Step ELISA kits appear suitable due to their sensitivity and specificity for drug of abuse screening in hair. This technology should find interest in workplace drug testing or driving license regranting, especially when many samples have to be tested with a high rate of negative samples, as ELISA is an easy and high-throughput method.     

Hair testing and self-report of cocaine use

Hair analysis is a useful tool in both clinical and forensic fields: it allows information about drugs of abuse (DOA) consumption to be obtained. However, in spite of analytical results, sometimes patients continue to deny using drugs or, on the contrary, insist on describing themselves as severe drug addicts; indeed there are often considerable difficulties in getting truthful statements about the real amount of drugs used. In this study we have tried to compare cocaine concentration in hair samples with self-reported drug intake. We enrolled 113 subjects (61 Africans, 52 Caucasians) who had been recently sent to jail. They were asked to tell about their use of illicit drugs during the last three months and then submitted to hair analysis. Hair segments (3 cm) were analyzed by GC-MS for amphetamines, cocaine and opiates. Useful data was obtained from 82 subjects, separated into two main groups on account of ethnic origin (African or Caucasian) and divided further into daily, weekly and monthly users. The results showed qualitative results and self-reported consumption to be in good agreement, although the correlation between frequency of consumption and concentration in hair revealed sometimes higher concentrations in contrast with the admission of low consumption. There was a definite separation between occasional and daily use (especially in Caucasian people), while concentrations found where weekly use was reported were more variable. Concentrations of cocaine measured in Africans' hair were much higher than in Caucasians'. Even if this study is exclusively based on self-report, it provides some interesting information in order to differentiate the frequency of consumption, and especially underlines the great importance of ethnic bias on hair analysis.     

Evaluation of the catalytic decomposition of H2O2 through use of organo-metallic complexes--a potential link to the luminol presumptive blood test

Hair testing for drugs of abuse is performed in Lombardy by eleven analytical laboratories accredited for forensic purposes, the most frequent purposes being driving license regranting and workplace drug testing. Individuals undergoing hair testing for these purposes can choose the laboratory in which the analyses have to be carried out. The aim of our study was to perform an interlaboratory exercise in order to verify the level of standardization of hair testing for drugs of abuse in these accredited laboratories; nine out of the eleven laboratories participated in this exercise. Sixteen hair strands coming from different subjects were longitudinally divided in 3-4 aliquots and distributed to participating laboratories, which were requested to apply their routine methods. All the participants analyzed opiates (morphine and 6-acetylmorphine) and cocainics (cocaine and benzoylecgonine) while only six analyzed methadone and amphetamines (amphetamine, methamphetamine, MDMA, MDA and MDEA) and five Δ(9)-tetrahydrocannabinol (THC). The majority of the participants (seven labs) performed acidic hydrolysis to extract the drugs from the hair and analysis by GC-MS, while two labs used LC-MS/MS. Eight laboratories performed initial screening tests by Enzyme Multiplied Immunoassay Technique (EMIT), Enzyme-linked Immunosorbent Assay (ELISA) or Cloned Enzyme Donor Immunoassay (CEDIA). Results demonstrated a good qualitative performance for all the participants, since no false positive results were reported by any of them. Quantitative data were quite scattered, but less in samples with low concentrations of analytes than in those with higher concentrations. Results from this first regional interlaboratory exercise show that, on the one hand, individuals undergoing hair testing would have obtained the same qualitative results in any of the nine laboratories. On the other hand, the scatter in quantitative results could cause some inequalities if any interpretation of the data is required.     

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。     

Amphetamine derivative related deaths

Amphetamine its methylendioxy (methylendioxyamphetamine methylenedioxymethylamphetamine, methylenedioxyethylamphetamine) and methoxy derivatives (p-methoxyamphetamine and p-methoxymethylamphetamine) are widely abused in Spanish society. We present here the results of a systematic study of all cases of deaths brought to the attention of the Madrid department of the Instituto Nacional de Toxicologia from 1993 to 1995 in which some of these drugs have been found in the cadaveric blood. The cases were divided into three categories: amphetamine and derivatives, amphetamines and alcohol, amphetamines and other drugs. Data on age, sex, clinical symptoms, morphological findings, circumstances of death, when known, and concentration of amphetamine derivatives, alcohol and other drugs in blood are given for each group. The information provided here may prove to be useful for the forensic interpretation of deaths which are directly or indirectly related to abuse of amphetamine derivatives.     

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。     

Analysis of drugs of abuse in hair: evaluation of the immunochemical method VMA-T vs. LC-MS/MS or GC-MS

Hair analysis is an elaborate and time-consuming multi-step process. The immunometric test VMA-T from Comedical has been evaluated as screening assay for hair analysis. From routine work, authentic samples were selected that were positive for opiates, cocaine, MDMA-type drugs, amphetamines, methadone or THC. These hair samples were investigated by LC-MS or GC-MS and the VMA-T procedure, respectively. Using the cut-off values recommended by the Society of Hair Testing, the VMA-T method discriminates with good sensitivity between negative and positive hair samples and is an expedient screening method for drugs in keratinized matrices such as hair. In order to save time and resources, the residue of the VMA-T extraction solution can be reused for confirmation analysis by LC-MS except for cocaine.     

Detection of drugs in human hair using Abbott ADx, with confirmation by gas chromatography/mass spectrometry (GC/MS).

The authors suggest use of the fluorescence polarization immunoassay (FPIA) technique in evaluation of chronic drug abuse using human hair. Hair was decontaminated in 5 mL of ethanol for 15 min at 37 degrees C and then incubated in 3 mL of 1M sodium hydroxide (NaOH) for 1 h at 100 degrees C. Afterwards, the aliquots were neutralized and analyzed using Abbott ADx for a negative or positive response for the following drugs: benzodiazepines, barbiturates, antidepressants, opiates, cocaine, amphetamine, and cannabis. All the positive samples were confirmed by gas chromatography/mass spectrometry (GC/MS). Only one false positive was detected (caused by interference of a phenothiazine with the antidepressants kit), clearly demonstrating the capability of ADx for toxicological screening of human hair.