Targeted analysis of 116 drugs in hair by UHPLC‐MS/MS and its application to forensic cases

A multi‐target method that can detect a broad range of drugs in human hair, such as hypnotics, anxiolytics, analgesics, benzodiazepines, antihistamines, antidepressants, antipsychotics, and anticonvulsants, was developed based on ultra‐high‐performance liquid chromatography–tandem mass spectrometry (UHPLC‐MS/MS). The drugs were extracted from 10 mg of washed hair by incubation for 18 h in a 25:25:50 (v/v/v ) mixture of methanol/acetonitrile/2 mM ammonium formate (8% acetonitrile, pH 5.3). For 51% of the basic drugs, the lower limits of quantification (LLOQs) were in the range of 0.05–0.5 pg/mg, and the majority (98%) were ≤ 5 pg/mg. Linearity ranged from LLOQs to 100–500 pg/mg for all the basic drugs. For acid and neutral drugs, the LLOQs ranged from 0.4 to 500 pg/mg, and linearity ranged from LLOQs to 80–40 000 pg/mg. According to published reports on concentrations attained in single dose control studies, the present method is sensitive enough to detect single‐dose drug exposure for many of the drugs. The accuracy was within 75–125% for the majority of drugs. Good precision was observed (relative standard deviations [RSD%] < 25%) for most of the compounds, including the prepared quality control (QC) hair samples. The method was applied to forensic cases and concentrations of rarely reported drugs in hair in 25 post‐mortem forensic cases were presented. Hair concentrations of amisulpride, gabapentin, mianserin, mepyramine, orphenadrine, and xylometazoline have not been previously reported. Copyright © 2016 John Wiley & Sons, Ltd.     

First identification of dapagliflozin in human hair: Development of a new liquid chromatography with tandem mass spectrometry method

Sodium-glucose cotransporter 1 inhibitors are a new class of drugs used for the treatment of type II diabetes. Due to their diuretic capabilities and the glycosuria they induce, these molecules cause effective weight loss that could attract the interest of a wider public than diabetics with all the health consequences knowing the adverse effects of these substances. In order to reveal a past exposure to these substances, hair analysis can be very useful especially in the medicolegal context. There are no data in the literature about gliflozin testing in hair. In this study, a method was developed for the analysis of three molecules belonging to the gliflozin family (dapagliflozin, empagliflozin and canagliflozin) using a liquid chromatography system coupled to tandem mass spectrometry. After decontamination with dichloromethane, gliflozins were extracted from hair following incubation in methanol in the presence of dapagliflozin-d5. Validation showed acceptable linearity for all compounds between 10 and 10,000 pg/mg, with limit of detection and limit of quantification at 5 and 10 pg/mg, respectively. Repeatability and reproducibility were below 20% at three concentrations for all analytes. The method was subsequently applied to the hair of two diabetic subjects under dapagliflozin treatment. In one of the two cases, the result was negative, while in the second case, the concentration was 12 pg/mg. Due to the absence of data, it is difficult to explain the absence of dapagliflozin in the hair of the first case. Physico-chemical characteristics of dapagliflozin could explain its bad incorporation in hair, making detection difficult even after daily treatment.     

Prevalence of new psychoactive substances: A retrospective study in hair

New psychoactive substances are conquering the drug scene. Police seize different colourful packages with exceptional names. They are declared as ‘bath salts’, ‘plant food’, or ‘research chemical powders’. Little is known about the actual prevalence of these drugs. Reanalysis of hair samples from routine cases concerning the presence of new psychoactive substances or ‘smart drugs’ should provide insight into changing patterns of designer drugs. All hair samples from 2009 and 2010 that originally tested positive for amphetamines or MDMA (N = 325) were reanalyzed for new or smart drugs such as 4‐fluoroamphetamine, piperazines (BZP, mCPP and TFMPP), cathinones (4‐MMC (mephedrone), methylone, butylone, ethylone, MDPV, methcathinone and cathinone), methylphenidate and ketamine. Hair snippets were extracted using a two‐step extraction procedure. The analytes were analyzed using liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) (electrospray ionization; multiple‐reaction‐monitoring mode – information dependent acquisition – enhanced product ion scan). New psychoactive substances were found in 120 cases (37%). Concerning the piperazine drugs, mCPP was positive in 34 (10.5%) cases and TFMPP in one case. Five mCPP cases were also positive for trazodone, an antidepressant which is metabolized to mCPP. In 11 (3%) cases, 4‐MMC was detected. Concerning the smart drugs, methylphenidate was found in 16 (5%). Ketamine was found in 45 (14%) cases. 4‐Fluoroamphetamine was identified in 12 (4%) cases and methylone in one case.In conclusion, there is a high prevalence of these drugs. Consequently, at least the most common ones (e.g. mCPP, KET, 4‐MMC and 4‐FA) should be included in screening procedures in clinical and forensic toxicology. Copyright © 2012 John Wiley & Sons, Ltd.     

Keratinous matrices for the assessment of drugs of abuse consumption: A correlation study between hair and nails

Keratinous matrices – hair and nails – accumulate substances over time and allow retrospective investigation of past consumption. Analysis of these matrices can provide information complementary to blood and urine analysis or can be used as standalone. So far, research has primarily focused on the detection of substances in hair, while studies in nails are scarce. In this study, we assessed concentrations of drugs of abuse and their metabolites in hair, fingernails, and toenails collected from the same individuals to evaluate differences and correlations between matrices. A total of 26 hair, 24 fingernail, and 18 toenail samples were collected. Samples were analysed by a validated liquid chromatography–tandem mass spectrometry method able to simultaneously detect the following compounds: amphetamine (AMP), methamphetamine, 3,4‐methylenedioxymethamphetamine (MDMA), 3,4‐methylenedioxyethylamphetamine, morphine (MOR), codeine (COD), 6‐monoacetylmorphine (6‐MAM), methadone (MTD), 2‐ethylidene‐1,5‐dimethyl‐3,3‐diphenylpyrrolidine (EDDP), cocaine (COC), benzoylecgonine (BE), and ecgonine methyl ester (EME). Strong positive correlations between hair, fingernails, and toenails were present for COC, BE, EME, AMP and MDMA. MOR, COD, 6‐MAM, MTD and EDDP showed positive trends. Concentrations were generally higher in nails compared to hair. Ratios between parent compounds and their metabolites were assessed for 6‐MAM/MOR, EDDP/MTD, BE/COC and EME/COC. Preliminary cut‐off concentrations for COC, BE, EME and AMP in fingernails and toenails were proposed. In light of these results, nails can be considered as a useful alternative to hair for monitoring of long‐term drug consumption. However, care should be taken regarding the variability in the accumulation of compounds between the matrices.     

Prevalence and concentrations of new designer stimulants,synthetic opioids,benzodiazepines, and hallucinogens in postmortem hair samples: A 13-year retrospective study

Hair samples are frequently analyzed in order to characterize consumption patterns of drugs. However, the interpretation of new psychoactive substance (NPS) findings in hair remains difficult because of lacking data for comparison. In this study, selected postmortem hair samples (n = 1203) from 2008 to 2020 were reanalyzed for synthetic cathinones, piperazines, phenethylamines, hallucinogens, benzodiazepines and opioids to evaluate prevalence data and concentration ranges. Hair samples were extracted using a two-step extraction procedure and analyzed using a validated liquid chromatography–tandem mass spectrometry (LC–MS/MS) method. Overall NPSs were detected in 381 cases (31.6%). Many cases were tested positive for more than one NPS in the same time span. A variety of NPS with a large range of concentrations was observed. For better comparability and interpretation of positive cases in routine work, quantitation data for 13 NPS were calculated as percentiles. The most frequently detected NPS in this study were N-ethylamphetamine, α-pyrrolidinovalerophenone, mephedrone, benzedrone, metamfepramone, and 4-fluoroamphetamine. In conclusion, a high prevalence of these drugs was observed from postmortem hair samples. The results show a growing use of many different NPSs by mainly young drug-using adults. Consequently, NPS screening procedures should be included in forensic toxicology. Our quantitative data may support other toxicologists in their assessment of NPS hair concentrations.     

Time‐course measurements of drug concentrations in hair and toenails after single administrations of pharmaceutical products

Hair and nails are often used to prove long‐term intake of drugs in forensic drug testing. The aim of this study was to evaluate the effectiveness of drug testing using hair and nails and the feasibility of determining when drugs were ingested by measuring the time‐courses of drug concentrations in hair and toenails after single administrations of various drugs. Healthy subjects ingested four pharmaceutical products containing eight active ingredients in single doses. Hair and toenails were collected at predetermined intervals, and drug concentrations in hair and nails were measured for 12 months. The administered drugs and their main metabolites were extracted using micropulverized extraction with a stainless steel bullet and were analyzed using liquid chromatography/tandem mass spectrometry. Acidic compounds such as ibuprofen and its metabolites were not detected in both specimens. Acetaminophen, a weakly acidic compound, was detected in nails more frequently than in hair. The maximum concentration of allyl isopropyl acetylurea, a neutral compound, in nails was significantly higher than in hair. Nails are an effective specimen to detect neutral and weakly acidic compounds. For fexofenadine, a zwitterionic compound, and for most basic compounds, the maximum concentrations in hair segments tended to be higher than those in nails. The hair segments showing the maximum concentrations varied between drugs, samples, and subjects. Drug concentrations in hair segments greatly depended on the selection of the hair. Careful interpretation of analytical results is required to predict the time of drug intake. Copyright © 2016 John Wiley & Sons, Ltd.     

Segmental analysis of antidepressant and antipsychotic drugs in the hair of schizophrenic patients

Hair analysis is useful for documenting long‐term exposure to drugs. The potential of hair analysis for therapeutic drug monitoring within the forensic field has been studied, but reference values for some antidepressants and antipsychotics in the hair of individuals undergoing chronic therapy are still lacking. In the present study, a method was developed and validated for the determination of 23 analytes, including antidepressants, antipsychotics, and related metabolites, in human hair by liquid chromatography–tandem mass spectrometry (LC–MS/MS). Hair samples (10 mg) were extracted with a 25:25:50 (v/v/v) mixture of methanol/acetonitrile/2 mM ammonium formate (8% acetonitrile, pH 5.3) utilizing cryogenic grinding. The present method demonstrated sufficient selectivity, robustness, and accuracy. Sixteen analytes in hair were reported in 46 psychiatric patients receiving fixed drug dosages. To the best of our knowledge, the hair concentrations of perphenazine and norolanzapine, as well as the concentrations of amisulpride, aripiprazole and its metabolite dehydroaripiprazole, olanzapine, and sulpiride, in hair from individuals receiving fixed dosages is reported for the first time. A significant relationship between the administered dose and the concentration in the proximal hair segment was found only for clozapine, norclozapine, and chlorpromazine. The results confirmed that the idea of using hair concentrations to monitor a daily dose is inapplicable.     

Development of a sensitive untargeted liquid chromatography–high resolution mass spectrometry screening devoted to hair analysis through a shared MS2 spectra database: A step toward early detection of new psychoactive substances

Untargeted liquid chromatography–high resolution mass spectrometry (LC–HRMS) techniques have become indispensable tools for systematic toxicological analysis. They allow the research of an almost unlimited number of drugs within a single analytical cycle, but shared mass spectra libraries are still missing to identify newly marketed compounds, along with defined analytical procedures. This article describes the optimization, validation, and application of an untargeted screening method devoted to hair analysis using data‐dependent analysis (DDA) and a shared HRMS database. This method used an ultra‐high performance liquid chromatography coupled to a benchtop Orbitrap. Raw MS data were processed with Compound Discoverer software coupled to the mzCloud? library. Optimizations were performed on blank hair spiked with 19 analytes having different physical and chemical properties. To validate the effectiveness of a shared spectra database, 20 compounds spectra were added and then retrospectively screened. Sensitivity and reliability were evaluated on 317 compounds of interest in toxicology. The method was then applied to 11 hair samples. The matrix effect range by ion suppression/enhancement was 40%–110%. The method allows the detection of 284 among the 317 screened compounds, including 72 new psychoactive substances (NPS). Lower limit of identification (LLOI) and lower limit of detection (LLOD) were 1 to 1000 pg/mg and 1 to 500 pg/mg, respectively. The method was successfully applied to 11 clinical cases and 144 compounds were identified including 24 NPS including AKB48‐5F for the first time in hair. We developed and validated an LC–HRMS untargeted screening of 284 compounds and successfully applied it to 11 real hair samples.     

Distribution pattern of common drugs of abuse,ethyl glucuronide,and benzodiazepines in hair across the scalp

While hair analysis is important and accepted in forensic applications, fundamental knowledge gaps still exist, exacerbated by a lack of knowledge of the incorporation mechanisms of substances into hair. The influence of the hair sampling location on the head on ethyl glucuronide (EtG) and cocaine concentrations was investigated by measuring the complete scalp hair of 14 (2 EtG, 4 cocaine, 8 both EtG and cocaine) study participants in a grid pattern for EtG, drugs of abuse, and benzodiazepines. Head skin perfusion and sweating rates were investigated to rationalize the concentration differences. For EtG, ratios between maximum and minimum concentrations on the scalp ranged from 2.5 to 7.1 (mean 4.4). For cocaine, the ratios ranged from 2.8 to 105 (mean 17.6). EtG concentrations were often highest at the vertex, but the distribution was strongly participant dependent. Cocaine and its metabolites showed the lowest concentrations at the vertex and the highest on the periphery, especially at the forehead. These differences led to hair from some head parts being clearly above conventional cut‐offs and others clearly below. In addition to EtG and cocaine, the distributions of 24 other drugs of abuse and benzodiazepines/z‐substances and metabolites are described. No clear pattern was observed for the head skin perfusion. Sweating rate measurements revealed higher sweating rates on the periphery of the haircut. Therefore, sweat could be a main incorporation route for cocaine. Concentration differences can lead to different interpretations depending on the sampling site. Therefore, the results are highly relevant for routine forensic hair analysis.     

Frequency of new psychoactive substances in hair and urine samples of individuals subject to drug testing in driving license regranting—A toxicological perspective

In recent years, numerous new psychoactive substances (NPS) have emerged on the illicit drug market. The assumed non-detectability of these drugs is often a key motivation for individuals subject to drug testing, such as those in driving license regranting programs. In these programs, NPS are not routinely tested for, and thus, subjects who have to prove abstinence from common drugs of abuse might switch to NPS to avoid positive drug tests. The aim of the study was to determine the frequency of these substances in the hair and urine samples of individuals undergoing drug testing in driving license regranting. A total of 1037 samples (577 hair and 460 urine samples) collected from 949 subjects between February 2017 and December 2018 were retrospectively analyzed for designer drugs and synthetic cannabinoids by liquid chromatography–quadrupole-time-of-flight–mass spectrometry (LC-QTOF-MS). For a more sensitive analysis of synthetic cannabinoids and their metabolites, additional testing was performed by liquid chromatography–tandem mass spectrometry (LC-MS/MS). Overall, 42 hair and two urine samples, which were obtained from 40 subjects, tested positive for NPS resulting in a frequency of 4.2%. While synthetic cannabinoids were detected in all cases, designer drugs were only found in three of these cases. With regard to the 577 hair samples analyzed, 7.3% screened positive, whereas only 0.4% of the 460 tested urine samples contained NPS. The results of this study indicate that synthetic cannabinoid use seems to be popular among this population, and therefore, testing for synthetic cannabinoids should be requested more often preferably using hair analysis.     

Sample preparation method for the combined extraction of ethyl glucuronide and drugs of abuse in hair

Often in hair analysis, a small hair sample is available while the analysis of a multitude of structurally diverse substances with different concentration ranges is demanded. The analysis of the different substances often requires different sample preparation methods, increasing the amount of required hair sample. When segmental hair analysis is necessary, the amount of hair sample needed is further increased. Therefore, the required sample amount for a full analysis can quickly exceed what is available. To combat this problem, a method for the combined hair sample preparation using a single extraction procedure for analysis of ethyl glucuronide with liquid chromatography‐multistage fragmentation mass spectrometry/multiple reaction monitoring (LC–MS³/MRM) and common drugs of abuse with LC–MRM was developed. The combined sample preparation is achieved by separating ethyl glucuronide from the drugs of abuse into separate extracts by fractionation in the solid‐phase extraction step during sample clean‐up. A full validation for all substances for the parameters selectivity, linearity, limit of detection, limit of quantification, accuracy, precision, matrix effects, and recovery was successfully completed. The following drugs of abuse were included in the method: Amphetamine; methamphetamine; 3,4‐methylenedioxy‐N‐methylamphetamine (MDMA); 3,4‐methylenedioxyamphetamine (MDA); 3,4‐methylenedioxy‐N‐ethylamphetamine (MDE); morphine; 6‐monoacetylmorphine; codeine; acetylcodeine; cocaine; benzoylecgonine; norcocaine; cocaethylene; methadone; 2‐ethylidene‐1,5‐dimethyl‐3,3‐diphenylpyrrolidine (EDDP) and methylphenidate. In conclusion, as only 1 sample preparation is needed with 1 aliquot of hair, the presented sample preparation allows an optimal analysis of both ethyl glucuronide and of the drugs of abuse, even when the sample amount is a limiting factor.     

Screening for basic drugs in hair of drug addicts by liquid chromatography/time-of-flight mass spectrometry

Hair analysis in forensic and clinical toxicology has been strongly focused on drugs of abuse, and comprehensive, drug class-independent screening methods based on mass spectrometric detection have not been applied to date. In this study, a qualitative drug screening method by liquid chromatography coupled to time-of-flight mass spectrometry, earlier developed and evaluated for forensic toxicological urine analysis, was adapted for screening of basic drugs in hair. The method included alkaline hydrolysis, purification with mixed-mode solid phase extraction, and analysis by liquid chromatography coupled to time-of-flight mass spectrometry with automated data analysis and reporting. Identification was based on accurate mass, isotopic pattern fit, and retention time, if available. Analysis of 32 hair samples from deceased drug addicts revealed 35 different drugs. The drug classes identified included antidepressants, antipsychotics, antiepileptics, amphetamines, opioids, beta-blockers, a benzodiazepine, a hypnotic, a local anesthetic, an antiemetic, and an antipyretic analgesic. The findings were in good agreement with the findings in blood and urine by other methods. Moreover, information about previous drug use not evident in the analysis of other matrices was obtained in the majority (72%) of the cases. Tramadol was an especially predominant finding, suggesting tramadol abuse as an opioid substitute. One apparent false-positive finding was identified. The mean and median mass accuracies of positive findings were 2.3 and 1.8 ppm, corresponding to 0.5 and 0.4 mDa, respectively. Cutoff values for tramadol and methamphetamine in hair were 100 and 200 pg/mg, respectively. The method proved to be a simple and straightforward tool for comprehensive screening of basic drugs in hair.     

Development of a new GC–MS/MS method for the determination of metformin in human hair

Diabetes mellitus is one of the most important public health challenges. Metformin (1,1‐dimethylbiguanide) represents the “gold standard” for the treatment of diabetes mellitus type 2. Despite its important role in reducing mortality and morbidity in the diabetic population, metformin is associated with an increased risk of stroke. To document exposure to a drug, hair is considered to be the specimen of choice to complement blood and urine, since it provides historical detail of a subject's chronic exposure to drug(s). Measuring hair concentration of metformin can be important for forensic toxicologists investigating criminal poisoning or Munchausen's syndrome by proxy. In clinical toxicology, drug monitoring using hair to document metformin observance has not yet been described. To document the interest of hair analysis for metformin, the authors have developed and validated a method using a gas‐chromatography tandem mass spectrometry system and applied it to authentic hair obtained from 9 diabetic patients under daily treatment. The validation procedure demonstrated a LOD an LOQ of 1 and 100 pg/mg, respectively and acceptable linearity, repeatability and reproducibility. The hair of the 9 patients tested positive in the low ng/mg range with concentrations ranging from 0.3 to 3.8 ng/mg. It seems obvious, in comparison with other drugs, that metformin is badly incorporated into hair, as the daily dosage varied from 1 to 3 g. Although limited in the number of subjects, the study allowed to postulate a possible correlation between daily dose and concentration in dark hair, while for light hair no correlation was found.     

Detection of prohibited substances in equine hair by ultra‐high performance liquid chromatography–triple quadrupole mass spectrometry – application to doping control samples

The detection of drugs in human hair samples has been performed by laboratories around the world for many years and the matrix is popular in disciplines, such as workplace drug testing. To date, however, hair has not become a routinely utilised matrix in sports drug detection. The analysis of hair samples offers several potential advantages to doping control laboratories, not least of which are the greatly extended detection window and the ease of sample collection and storage. This article describes the development, validation, and utilisation of a sensitive ultra‐high performance liquid chromatography – triple quadrupole mass spectrometry (UHPLC – MS/MS) method for the detection of 50 compounds. This provides significantly improved coverage for those analytes which would be of particular interest if detected in hair, such as anabolic steroid esters and selective androgen receptor modulators (SARMs). Qualitative validation of the method resulted in estimated limits of detection as low as 0.1 pg/mg for the majority of compounds, with all being detected at 2 pg/mg or below. The suitability of the method for the detection of prohibited substances in incurred material was demonstrated by the successful detection of several compounds, such as stanozolol, boldenone undecylenate, clenbuterol, and GW‐501516, in genuine equine hair samples. Estimated concentrations of the detected substances ranged from 0.27 to 8.6 pg/mg. The method has been shown to be fit‐for‐purpose for routine screening of equine hair samples by the analysis of over 400 genuine hair samples.     

Psychotropic drugs in mothers' milk: a comprehensive review of assay methods, pharmacokinetics and of safety of breast-feeding

Many mentally ill women want to breast-feed their babies but, if they are taking psychotropic drugs, there is very little systematic data upon which to base decisions about whether or not it is safe to do so. We therefore attempt to provide a comprehensive and critical summary of existing case reports and of studies of breast-feeding in relation to commonly used psychotropic drugs. The literature review focuses on the following drugs: antidepressants: tricyclics and serotonin selective reuptake inhibitors (SSRIs); antipsychotic drugs: chlorpromazine, perphenazine, haloperidol and clozapine; mood stabilizers: lithium and carbamazepine; and benzodiazepines. The research literature consists mainly of single case reports and there have been very few attempts at controlled, longitudinal investigations. Findings are often difficult to compare because of differences in methods or because of lack of key information. Most data are available about the tricyclic antidepressants but even here we have found that the reports cover only a grand total of 66 mother-infant pairs. Dilemmas about whether or not to contraindicate breast-feeding arise most commonly in relation to postnatal depression. The findings to date suggest that provided that infants are healthy at the outset it is likely that the benefits of breast-feeding will outweigh potential hazards if their mothers are taking established tricyclic drugs at recommended dose levels. Much less is known about risks associated with SSRI antidepressants or about antipsychotic drugs such as phenothiazines and butyrophenones or mood stabilizers such as carbamazepine, all of which enter breast-milk. Safeguards are suggested for future single case studies, which, as they accumulate, will provide a platform for mounting controlled prospective studies properly to test for any acute toxic effects and for possible long-term adverse effects of such drugs on infants' development. Appendix 1 is a review of assay methods. Appendix 2 examines pharmacokinetic factors in newborn preterm and sick infants with special reference to contraindications to breast-feeding. Appendix 3 is a review of methods for assessing infant health and development.     

A possible new oxidation marker for hair adulteration: Detection of PTeCA (1H‐pyrrole‐2,3,4,5‐tetracarboxylic acid) in bleached hair

Hair analysis has become a valuable tool in forensic toxicology to assess drug or alcohol abstinence. Yet, hair adulteration by cosmetic products presents a major challenge for forensic hair analysis. Oxidative treatments, e.g. bleaching, may lead to analyte loss and thereby to false negative results. Currently, the eumelanin degradation product 1H‐pyrrole‐2,3,5‐tricarboxylic acid (PTCA) serves as a marker for oxidative hair treatment, but requires the definition of cut‐off values. To investigate further eumelanin degradation products as markers for oxidative hair treatment, hair samples with and without in vitro bleaching (hydrogen peroxide (H₂O₂) concentrations 1.9% up to 12%; incubation times 15 min, 30 min, 60 min) were analyzed by liquid chromatography coupled to high‐resolution time of flight mass spectrometry (HPLC‐HRMS). The distribution of eumelanin degradation products along the hair shaft was investigated for routine applicability after segmentation of cosmetically untreated hair samples and authentically treated hair samples. The signals of the eumelanin degradation products PTCA, 1H‐pyrrole‐2,3,4‐tricarboxylic acid (isoPTCA), and 1H‐pyrrole‐2,3,4,5‐tetracarboxylic acid (PTeCA) were found to be significantly elevated after in vitro bleaching already with low H₂O₂ concentrations and after short incubation times. In contrast to PTCA and isoPTCA, PTeCA was not detectable in cosmetically untreated segments up to 12 cm from hair root and was only formed through the oxidation process. The results of the study show that the detection of PTeCA within the proximal 3 to 6 cm segment can be applied to reliably detect hair adulteration attempts through hair bleaching.     

Segmental hair analysis for differentiation of tilidine intake from external contamination using LC‐ESI‐MS/MS and MALDI‐MS/MS imaging

Segmental hair analysis has been used for monitoring changes of consumption habit of drugs. Contamination from the environment or sweat might cause interpretative problems. For this reason, hair analysis results were compared in hair samples taken 24 h and 30 days after a single tilidine dose. The 24‐h hair samples already showed high concentrations of tilidine and nortilidine. Analysis of wash water from sample preparation confirmed external contamination by sweat as reason. The 30‐day hair samples were still positive for tilidine in all segments. Negative wash‐water analysis proved incorporation from sweat into the hair matrix. Interpretation of a forensic case was requested where two children had been administered tilidine by their nanny and tilidine/nortilidine had been detected in all hair segments, possibly indicating multiple applications. Taking into consideration the results of the present study and of MALDI‐MS imaging, a single application as cause for analytical results could no longer be excluded. Interpretation of consumption behaviour of tilidine based on segmental hair analysis has to be done with caution, even after typical wash procedures during sample preparation. External sweat contamination followed by incorporation into the hair matrix can mimic chronic intake. For assessment of external contamination, hair samples should not only be collected several weeks but also one to a few days after intake. MALDI‐MS imaging of single hair can be a complementary tool for interpretation. Limitations for interpretation of segmental hair analysis shown here might also be applicable to drugs with comparable physicochemical and pharmacokinetic properties. Copyright © 2014 John Wiley & Sons, Ltd.     

Surveillance of drug abuse in Hong Kong by hair analysis using LC‐MS/MS

The aim of this study is to reveal the habits of drug abusers in hair samples from drug rehabilitation units in Hong Kong. With the application of liquid chromatography–tandem mass spectrometry (LC–MS/MS) technology, a total of 1771 hair samples were analyzed during the period of hair testing service (January 2012 to March 2016) provided to 14 drug rehabilitation units including non‐governmental organizations (NGOs), rehabilitation centers, and medical clinics. Hair samples were analyzed for abused drugs and their metabolites simultaneously, including ketamine, norketamine, cocaine, benzoylecgonine, cocaethylene, norcocaine, codeine, MDMA, MDA, MDEA, amphetamine, methamphetamine, morphine, 6‐acetylmorphine, phencyclidine, and methadone. The results showed that ketamine (77.2%), cocaine (21.3%), and methamphetamine (16.5%) were the frequently detected drugs among those drug abusers, which is consistent with the reported data. In addition, the usage of multiple drugs was also observed in the hair samples. About 29% of drug‐positive samples were detected with multiple drug use. Our studies prove that our locally developed hair drug‐testing method and service can be a valid tool to monitor the use of abused drugs, and which could facilitate rehabilitation program management.     

Assessment of Pregabalin Use by Hair Testing

Background: The detection and analysis of drugs in hair has progressively emerged as a consequence of the enhanced sensitivity of analytical techniques used in forensic toxicology; a greater advantage in using this matrix respect to classical ones (i.e., urine and blood) is an easier and noninvasive sample collection, even when the careful supervision of law-enforcement officers is required to avoid the risk that the sample may be adulterated or replaced. Moreover, according to the length of the hair, the history of drug exposure can be retrospectively monitored from few weeks up to months or years since sample collection. Objective: Given the potential negative effects of pregabalin, an antiepileptic and analgesic drug with a high risk of misuse and abuse, the laboratory was asked to test for the drug in hair. Method: A new ultrahigh-performance liquid chromatography coupled to tandem mass spectrometry was developed. The method involves incubation of 25 mg of cut hair in acetonitrile for 2 h in an ultrasonic bath and separation on an Acquity HSS C18 column (150 × 2.1 mm × 1.8 µm) maintained at 50°C in a thermostatically controlled oven. A gradient elution was performed. Results: The method was fully validated according to international standards. The limit of quantitation of the test was 10 pg/mg. Five authentic cases of pregabalin in hair segments were tested using the method and the results were in the range 17–1487 pg/mg. Conclusion: This new method was found suitable to monitor both patients under pregabalin therapy and dependent subjects.     

Single hair analysis: Validation of a screening method for over 150 analytes and application on documented single-dose cases

Hair is the matrix of choice in forensic toxicology when retrospective analysis is needed. Nonetheless, due to misalignment, different growth stages and segmentation lengths of 0.5–1 cm, resolution of time is limited. By segmental analysis of single hairs, most of these factors can be compensated and resolution of time is enhanced. A method for manually segmenting single hairs in 2-mm sections and screening for 156 analytes by liquid chromatography coupled to tandem mass spectrometry has been developed and validated. The method was applied to 15 single-dose cases concerning different pharmaceuticals by analyzing 10 hairs each, sampled 1 and 2 months after ingestion in most cases. The validation showed a lower limit of quantification of ≤1.25 pg/segment for ~90% of analytes and good accuracy. Many substances could be detected in the presented cases, whereas detection of benzodiazepines and low dosed opioids remains challenging. In positive cases, characteristic peak-shaped concentration profiles across the hairs were obtained. The segment with most coinciding peak maxima can be allocated to the time of ingestion. A method for the determination of individual hair growth rate was applied and revealed a gap between expected and actual position of peak maxima. Additionally, different localization of simultaneously administered substances was observed. These findings were tried to be explained by different routes of incorporation and may contribute to current knowledge. The presented method may directly be applied to similar questions in hair analysis, and the findings are considered important for interpreting further results in single hair analysis.