Amphetamines in Workplace Testing Hair Samples over the Years 2011-2020.

Amphetamines (AMPs) in hair were investigated with thousands of workplace testing head and body hair samples collected and analyzed over 10 years and tabulated by year. All samples were washed by a published extensive method prior to confirmation by liquid chromatography-mass spectrometry-mass spectrometry. Presented are concentrations of parent methamphetamine (METH), 3,4-methylenedioxymethamphetamine (MDMA) and methylenedioxyamphetamine as metabolite and AMP as metabolite and without the presence of parent drug. Some differences in METH concentrations from year to year were significant, and some ratios of metabolite to parent drug for both METH and MDMA also varied significantly. While rates of METH use may not have changed significantly, some aspects of the drugs ingested as demonstrated by hair analysis varied over the 10-year period.

Cannabinoids Tetrahydrocannabinol, Cannabinol, Cannabidiol, Tetrahydrocannabivarin and 11-nor-9-carboxy-∆9-THC in Hair.

The cannabinoids tetrahydrocannabinol (THC), tetrahydrocannabivarin (THCV), cannabidiol (CBD), cannabinol (CBN) and (-)-11-nor-9-carboxy-∆9-tetrahydrocannabinol (THC-COOH) were determined in 4,773 hair samples. Confirmation of THC-COOH was by GC-MS-MS (gas chromatography--mass spectrometry-mass spectrometry). Confirmation of THC, THCV, CBN and CBD was by LC-MS-MS (liquid chromatoraphy--mass spectrometry-mass spectrometry) on an AB Sciex QTRAP 6500+ LC-MS-MS. The purpose of this work was not to utilize any analyte other than THC-COOH as indicative of ingestion, but to assess the absence or presence, and relative concentrations, of the other cannabinoid analytes in hair of marijuana users vs. primarily CBD users. In this regard, 10% of samples contained significantly higher concentrations of CBD relative to THC than the other 90%. A concentration of CBD that is five times greater than that of THC was proposed as good evidence of primarily CBD ingestion. THC concentrations in the samples ranged from below the limit of detection (5 pg/mg) to 47,808 pg/mg hair, varying widely in the relationship between parent THC and the metabolite THC-COOH. CBN was present in most samples, but concentrations relative to THC decreased with increasing THC concentrations. Only 26% of the samples contained THCV detectable by the method. When present, THCV concentrations averaged 1.77% of THC. A limitation of this study is the lack of subject histories to determine the types and amounts of products used and the mode of ingestion. Also, not all THC from external contamination may be removed. Nonetheless, the data provide a useful guide as to what cannabinoids may be found in hair, at what concentrations and under conditions of marijuana use vs. likely primarily CBD use.

Hydroxycocaines as Metabolic Indicators of Cocaine Ingestion.

Hydroxycocaines in hair were investigated with many hundreds of head and body hair samples. All samples were washed by a published extensive aqueous method prior to confirmation by LC-MS/MS. Concentrations, percent of cocaine, and ratios of para- and meta-hydroxycocaines to ortho-hydroxycocaine are presented. Hydroxycocaines as percent of cocaine did not appear to be affected by cocaine concentrations, but were shown to increase with cocaethylene concentrations. Stability of hydroxycocaines over a year of ambient storage was demonstrated. Ortho-hydroxycocaine was shown to be formed by exposure of cocaine-positive hair to peroxide, while para- and meta-hydroxycocaines were not. Presence of para- or meta-hydroxycocaine at > 0.05% of cocaine is proposed as indicating ingestion of cocaine. This indicator prevents black hair from being more likely interpreted as positive for ingestion than lighter colored hair.

A Retrospective Analysis of Selected Opioids in Hair of Workplace Drug Testing Subjects.

Opioids, both naturally occurring and semisynthetic, are effective pain management medications, but also possess the potential for abuse. Analyses of over 37,000 head and body hair samples containing codeine, morphine, hydrocodone, hydromorphone, oxycodone or oxymorphone provide a view of use habits of workplace-testing subjects that cannot be obtained from fluid matrices results. Testing was performed using FDA cleared immunoassays using either 2 ng morphine or oxycodone per 10 mg hair as calibrators. Non-negative screening samples were washed with an extended aqueous wash procedure followed by LC-MS-MS confirmation at a cutoff concentration of 2 ng opioid per 10 mg hair. The LC-MS-MS method measured codeine, morphine, 6-acetylmorphine, hydrocodone, hydromorphone, oxycodone and oxymorphone with an administratively established LOQ of 0.50 ng opioid per 10 mg hair. The linear range was 0.50-100 ng morphine per 10 mg hair, and 0.50-150 ng opioid per 10 mg hair for all other measured analytes. For all analytes, within run precision was ≤5.4%, and between-run precision was ≤6.4%. Analysis of samples containing metabolites found that, among codeine positive samples, 97% contained less than 10% morphine metabolite and 88% less than 20% hydrocodone metabolite, among hydrocodone positive samples, 97% contained less than 10% hydromorphone metabolite and 95% of oxycodone positive samples contained less than 10% oxymorphone metabolite. Our analysis of opioid-positive samples may provide guidelines for interpretation of hair opioid levels typically observed in workplace testing.

Nail Analysis for Drugs: A Role in Workplace Testing?

Analysis of nail clippings may be a useful back-up for hair analysis when hair is unavailable. One aspect of using nails or hair is the ability to analyze whether drug present is from ingestion or from contamination. A common method of three 15-s rinses in methanol failed to remove drug from nails that had been soaked in either 5 or 50 µg/mL cocaine, methamphetamine or morphine for 1 h. While methanol rinsing did not remove contaminating drug, washing the nails soaked with 5 and 50 µg/mL of these drugs with an extended wash, a method developed for hair analysis and consisting of a 15-min isopropanol wash, and three 30-min and two 60-min phosphate buffer-0.1% albumin washes, when applied to nails did remove most of the contaminating drug. The drug left in the nails after extended washing could be interpreted as contamination by applying a wash criterion that is routinely applied in hair analysis. Successful decontamination of the soaked contaminated nail model was followed by applying this extended wash method to presumptive positive nail samples identified in workplace testing. While the extended buffer wash and wash criterion distinguish contamination from ingestion with hair, we failed to demonstrate that the method effectively differentiates contamination from ingestion with nails.

Carboxy-THC in Washed Hair: Still the Reliable Indicator of Marijuana Ingestion.

The presence of the metabolite 11-nor-9-carboxy-delta-9-tetrahydrocannabinol (C-THC) in hair is generally accepted as the definitive proof of delta-9-tetrahydrocannabinol (THC) ingestion. During hair analysis, the removal of any potential C-THC external contamination that could result from marijuana smoke or close personal contact via a wash procedure is critical. Here, we performed a series of experiments to demonstrate that C-THC is the reliable indicator of marijuana ingestion when paired with the correct washing procedure to remove potential external contamination.

Synthesis/biological evaluation of hydroxamic acids and their prodrugs as inhibitors for Botulinum neurotoxin A light chain.

Botulinum neurotoxin A (BoNT/A) is the most potent toxin known. Unfortunately, it is also a potential bioweapon in terrorism, which is without an approved therapeutic treatment once cellular intoxication takes place. Previously, we reported how hydroxamic acid prodrug carbamates increased cellular uptake, which translated to successful inhibition of this neurotoxin. Building upon this research, we detail BoNT/A protease molecular modeling studies accompanied by the construction of small library of hydroxamic acids based on 2,4-dichlorocinnamic hydroxamic acid scaffold and their carbamate prodrug derivatization along with the evaluation of these molecules in both enzymatic and cellular models.

Dynamic vaccine blocks relapse to compulsive intake of heroin.

Heroin addiction, a chronic relapsing disorder characterized by excessive drug taking and seeking, requires constant psychotherapeutic and pharmacotherapeutic interventions to minimize the potential for further abuse. Vaccine strategies against many drugs of abuse are being developed that generate antibodies that bind drug in the bloodstream, preventing entry into the brain and nullifying psychoactivity. However, this strategy is complicated by heroin's rapid metabolism to 6-acetylmorphine and morphine. We recently developed a "dynamic" vaccine that creates antibodies against heroin and its psychoactive metabolites by presenting multihaptenic structures to the immune system that match heroin's metabolism. The current study presents evidence of effective and continuous sequestration of brain-permeable constituents of heroin in the bloodstream following vaccination. The result is efficient blockade of heroin activity in treated rats, preventing various features of drugs of abuse: heroin reward, drug-induced reinstatement of drug seeking, and reescalation of compulsive heroin self-administration following abstinence in dependent rats. The dynamic vaccine shows the capability to significantly devalue the reinforcing and motivating properties of heroin, even in subjects with a history of dependence. In addition, targeting a less brain-permeable downstream metabolite, morphine, is insufficient to prevent heroin-induced activity in these models, suggesting that heroin and 6-acetylmorphine are critical players in heroin's psychoactivity. Because the heroin vaccine does not target opioid receptors or common opioid pharmacotherapeutics, it can be used in conjunction with available treatment options. Thus, our vaccine represents a promising adjunct therapy for heroin addiction, providing continuous heroin antagonism, requiring minimal medical monitoring and patient compliance.

The use of small molecule probes to study spatially separated stimulus-induced signaling pathways.

Simultaneous activation of signaling pathways requires dynamic assembly of higher-order protein complexes at the cytoplasmic domains of membrane-associated receptors in a stimulus-specific manner. Here, using the paradigm of cellular activation through cytokine and innate immune receptors, we demonstrate the proof-of-principle application of small molecule probes for the dissection of receptor-proximal signaling processes, such as activation of the transcription factor NF-κB and the protein kinase p38.

A vaccine strategy that induces protective immunity against heroin.

Heroin addiction is a wide-reaching problem with a spectrum of damaging social consequences. A vaccine capable of blocking heroin's effects could provide a long-lasting and sustainable adjunct to heroin addiction therapy. Heroin, however, presents a particularly challenging immunotherapeutic target, as it is metabolized to multiple psychoactive molecules. To reconcile this dilemma, we examined the idea of a singular vaccine with the potential to display multiple drug-like antigens; thus two haptens were synthesized, one heroin-like and another morphine-like in chemical structure. A key feature in this approach is that immunopresentation with the heroin-like hapten is thought to be immunochemically dynamic such that multiple haptens are simultaneously presented to the immune system. We demonstrate the significance of this approach through the extremely rapid generation of robust polyclonal antibody titers with remarkable specificity. Importantly, both the antinociceptive effects of heroin and acquisition of heroin self-administration were blocked in rats vaccinated using the heroin-like hapten.

A Diels-Alder Reaction Conducted Within the Parameters of Aqueous Organocatalysis: Still Just Smoke and Mirrors.

Conducting reactions using water as solvent is a highly prized goal for the organic chemist. Based upon recent literature and our continuing interest in the field of aqueous organocatalysis, we tested the scope of an enamine based Diels-Alder reaction using (±)-nornicotine, proline and a proline derivative as aqueous organocatalysts. Unfortunately, none of the examined catalysts under aqueous conditions proved useful, leaving the aqueous Diels-Alder reaction as an elusive goal.

Chirality holds the key for potent inhibition of the botulinum neurotoxin serotype a protease.

Botulinum neurotoxin serotype A (BoNT/A) is the most toxic protein known to man and also a bioterrorism agent. As defined by our previous research targeting the etiological agent responsible for BoNT/A intoxication, a protease, we now report on the asymmetric synthesis of four new BoNT/A inhibitors; the most potent of this series is roughly 2-fold more active than the best small molecule inhibitor currently known.