Critical comparison of mass analyzers for forensic hair analysis by ambient ionization mass spectrometry.

RATIONALE: Recently, several direct and/or ambient mass spectrometry (MS) approaches have been suggested for drugs of abuse imaging in hair. The use of mass spectrometers with insufficient selectivity could result in false-positive measurements due to isobaric interferences. Different mass analyzers have been evaluated regarding their selectivity and sensitivity for the detection of Δ9-tetrahydrocannabinol (THC) from intact hair samples using direct analysis in real time (DART) ionization. METHODS: Four different mass analyzers, namely (1) an orbitrap, (2) a quadrupole orbitrap, (3) a triple quadrupole, and (4) a quadrupole time-of-flight (QTOF), were evaluated. Selectivity and sensitivity were assessed by analyzing secondary THC standard dilutions on stainless steel mesh screens and blank hair samples, and by the analysis of authentic cannabis user hair samples. Additionally, separation of isobaric ions by use of travelling wave ion mobility (TWIM) was investigated. RESULTS: The use of a triple quadrupole instrument resulted in the highest sensitivity; however, transitions used for multiple reaction monitoring were only found to be specific when using high mass resolution product ion measurements. A mass resolution of at least 30,000 FWHM at m/z 315 was necessary to avoid overlap of THC with isobaric ions originating from the hair matrix. Even though selectivity was enhanced by use of TWIM, the QTOF instrument in resolution mode could not indisputably differentiate THC from endogenous isobaric ions in drug user hair samples. CONCLUSIONS: Only the high resolution of the (quadrupole) orbitrap instruments and the QTOF instrument in high-resolution mode distinguished THC in hair samples from endogenous isobaric interferences. As expected, enhanced selectivity compromises sensitivity and THC was only detectable in hair from heavy users. Copyright © 2016 John Wiley & Sons, Ltd.

(Un)targeted Scanning of Locks of Hair for Drugs of Abuse by Direct Analysis in Real Time-High-Resolution Mass Spectrometry.

Forensic hair evidence can be used to obtain retrospective timelines of drug use by analysis of hair segments. However, this is a laborious and time-consuming process, and mass spectrometric (MS) imaging techniques, which show great potential for single-hair targeted analysis, are less useful due to differences in hair growth rate between individual hairs. As an alternative, a fast untargeted analysis method was developed that uses direct analysis in real time-high-resolution mass spectrometry (DART-HRMS) to longitudinally scan intact locks of hair without extensive sample preparation or segmentation. The hair scan method was validated for cocaine against an accredited liquid chromatography/tandem mass spectrometry (LC/MS/MS) method. The detection limit for cocaine in hair was found to comply with the cutoff value of 0.5 ng/mg recommended by the Society of Hair Testing; that is, the DART hair scan method is amenable to forensic cases. Under DART conditions, no significant thermal degradation of cocaine occurred. The standard DART spot size of 5.1 ± 1.1 mm could be improved to 3.3 ± 1.0 mm, corresponding to approximately 10 days of hair growth, by using a high spatial resolution exit cone. By use of data-dependent product ion scans, multiple drugs of abuse could be detected in a single drug user hair scan with confirmation of identity by both exact mass and MS/HRMS fragmentation patterns. Furthermore, full-scan high-resolution data were retrospectively interrogated versus a list of more than 100 compounds and revealed additional hits and temporal profiles in good correlation with reported drug use.

Evidence based decontamination protocols for the removal of external Δ9-tetrahydrocannabinol (THC) from contaminated hair.

External contamination can cause false positive results in forensic hair testing for drugs of abuse and is therefore a major concern when hair evidence is used in court. Current literature about decontamination strategies is mainly focused on external cocaine contamination and no consensus on the best decontamination procedure for hair samples containing cannabinoids has been reached so far. In this study, different protocols with solvents, both organic as well as aqueous, were tested on blank and drug user hair for their performance on removing external cannabis contamination originating from either smoke or indirect contact with cannabis plant material. Smoke contamination was mimicked by exposing hair samples to smoke from a cannabis cigarette and indirect contact contamination by handling hair with cannabis contaminated gloves or hands. Δ9-tetrahydrocannabinol (THC) levels in the hair samples and wash solvents were determined using liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. Aqueous surfactant solutions removed more THC contamination compared to water, but much less than organic solvents. Methanol, dichloromethane and chloroform were most efficient in removing THC contamination. Due to its lower environmental impact, methanol was chosen as the preferred decontamination solvent. After testing of different sequential wash steps on externally contaminated blank hair, three protocols performed equally well, removing all normal level and more than 99% of unrealistically high levels of external cannabis contamination. Thorough testing on cannabis users' hair, both as such and after deliberate contamination, showed that using these protocols all contamination could be washed from the hair while no incorporated THC was removed from truly positive samples. The present study provides detailed scientific evidence in support of the recommendations of the Society of Hair Testing: a protocol using a single methanol wash followed by a single aqueous SDS solution wash, followed by a Milli-Q water rinsing step, is suggested as the preferred decontamination protocol to remove external cannabis contamination from hair while preserving the incorporated compounds.

Ultratrace LC-MS/MS analysis of segmented calf hair for retrospective assessment of time of clenbuterol administration in Agriforensics.

In agriforensics, time of administration is often debated when illegal drug residues, such as clenbuterol, are found in frequently traded cattle. In this proof-of-concept work, the feasibility of obtaining retrospective timeline information from segmented calf tail hair analyses has been studied. First, an ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) hair analysis method was adapted to accommodate smaller sample sizes and in-house validated. Then, longitudinal 1 cm segments of calf tail hair were analyzed to obtain clenbuterol concentration profiles. The profiles found were in good agreement with calculated, theoretical positions of the clenbuterol residues along the hair. Following assessment of the average growth rate of calf tail hair, time of clenbuterol administration could be retrospectively determined from segmented hair analysis data. The data from the initial animal treatment study (n = 2) suggest that time of treatment can be retrospectively estimated with an error of 3-17 days.

Rapid analysis of Δ-9-tetrahydrocannabinol in hair using direct analysis in real time ambient ionization orbitrap mass spectrometry.

RATIONALE: Forensic hair analysis methods are laborious, time-consuming and provide only a rough retrospective estimate of the time of drug intake. Recently, hair imaging methods using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) were reported, but these methods require the application of MALDI matrix and are performed under vacuum. Direct analysis of entire locks of hair without any sample pretreatment and with improved spatial resolution would thus address a need. METHODS: Hair samples were attached to stainless steel mesh screens and scanned in the X-direction using direct analysis in real time (DART) ambient ionization orbitrap MS. The DART gas temperature and the accuracy of the probed hair zone were optimized using Δ-9-tetrahydrocannabinol (THC) as a model compound. Since external contamination is a major issue in forensic hair analysis, sub-samples were measured before and after dichloromethane decontamination. RESULTS: The relative intensity of the THC signal in spiked blank hair versus that of quinine as the internal standard showed good reproducibility (26% RSD) and linearity of the method (R(2) = 0.991). With the DART hair scan THC could be detected in hair samples from different chronic cannabis users. The presence of THC was confirmed by quantitative liquid chromatography/tandem mass spectrometry. Zones with different THC content could be clearly distinguished, indicating that the method might be used for retrospective timeline assessments. Detection of THC in decontaminated drug user hair showed that the DART hair scan not only probes THC on the surface of hair, but penetrates deeply enough to measure incorporated THC. CONCLUSIONS: A new approach in forensic hair analysis has been developed by probing complete locks of hair using DART-MS. Longitudinal scanning enables detection of incorporated compounds and can be used as pre-screening for THC without sample preparation. The method could also be adjusted for the analysis of other drugs of abuse.