Higher paracetamol levels are associated with elevated glucocorticoid concentrations in hair: findings from a large cohort of young adults.

Paracetamol is one of the most commonly used over-the-counter medications. Experimental studies suggest a possible stress-suppressing effect of paracetamol in humans facing experimental stress-inducing paradigms. However, no study has investigated whether paracetamol and steroid hormones covary over longer time frames and under real-life conditions. This study addresses this gap by investigating associations between steroid hormones (cortisol, cortisone, and testosterone) and paracetamol concentrations measured in human hair, indexing a timeframe of approximately three months. The data came from a large community sample of young adults (N = 1002). Hair data were assayed using liquid chromatography-tandem mass spectrometry. Multiple regression models tested associations between paracetamol and  steroid hormones, while adjusting for a wide range of potential confounders, such as sex, stressful live events, psychoactive substance use, hair colour, and body mass index. Almost one in four young adults from the community had detectable paracetamol in their hair (23%). Higher paracetamol hair concentrations were robustly associated with more cortisol (ß = 0.13, ηp = 0.016, p < 0.001) and cortisone (ß = 0.16, ηp = 0.025, p < 0.001) in hair. Paracetamol and testosterone hair concentrations were not associated. Paracetamol use intensity positively correlated with corticosteroid functioning across several months. However, a potential corticosteroid-inducing effect of chronic paracetamol use has yet to be tested in future experimental designs.

Stress Markers, Executive Functioning, and Resilience Among Early Adolescents With Complex Congenital Heart Disease.

Importance: Infants with complex congenital heart disease (cCHD) may experience prolonged and severe stress when undergoing open heart surgery. However, little is known about long-term stress and its role in neurodevelopmental impairments in this population. Objective: To investigate potential differences between early adolescents aged 10 to 15 years with cCHD and healthy controls in physiological stress markers by hair analysis, executive function (EF) performance, and resilience. Design, Setting, and Participants: This single-center, population-based case-control study was conducted at the University Children's Hospital Zurich, Switzerland. Patients with different types of cCHD who underwent cardiopulmonary bypass surgery during the first year of life and who did not have a genetic disorder were included in a prospective cohort study between 2004 and 2012. A total of 178 patients were eligible for assessment at ages 10 to 15 years. A control group of healthy term-born individuals was cross-sectionally recruited. Data assessment was between 2019 and 2021. Statistical analysis was performed from January to April 2023. Exposure: Patients with cCHD who underwent infant open heart surgery. Main Outcomes and Measures: Physiological stress markers were quantified by summing cortisol and cortisone concentrations measured with liquid chromatography with tandem mass spectrometry in a 3-centimeter hair strand. EFs were assessed with a neuropsychological test battery to produce an age-adjusted EF summary score. Resilience was assessed with a standardized self-report questionnaire. Results: The study included 100 patients with cCHD and 104 controls between 10 and 15 years of age (mean [SD] age, 13.3 [1.3] years); 110 (53.9%) were male and 94 (46.1%) were female. When adjusting for age, sex, and parental education, patients had significantly higher sums of hair cortisol and cortisone concentrations (ß, 0.28 [95% CI, 0.12 to 0.43]; P < .001) and lower EF scores (ß, -0.36 [95% CI, -0.49 to -0.23]; P < .001) than controls. There was no group difference in self-reported resilience (ß, -0.04 [95% CI, -0.23 to 0.12]; P = .63). A significant interaction effect between stress markers and EFs was found, indicating a stronger negative association in patients than controls (ß, -0.65 [95% CI, -1.15 to -0.15]; P = .01). The contrast effects were not significant in patients (ß, -0.21 [95% CI, -0.43 to -0.00]; P = .06) and controls (ß, 0.09 [95% CI, -0.11 to 0.30]; P = .38). Conclusions and Relevance: This case-control study provides evidence for altered physiological stress levels in adolescents with cCHD and an association with poorer EF. These results suggest that future studies are needed to better understand the neurobiological mechanisms and timing of alterations in the stress system and its role in neurodevelopment.

Associations between hair-derived cannabinoid levels, self-reported use, and cannabis-related problems.

RATIONALE: As cannabis potency and cannabis use are increasing in newly legalized markets, it is increasingly important to measure and examine the effects of cannabinoid exposure. OBJECTIVES: The current study aims to assess how hair-derived cannabinoid concentrations - offering insight into three-month cumulative exposure - are associated with common self-report measures of cannabis use and cannabis use-related problems. METHODS: 74 near-daily dependent cannabis users self-reported their quantity of cannabis use, cannabis use-related problems, and estimated cannabis potency. Hair samples were provided to quantify Δ9-THC, CBD, and CBN using LC-MS/MS and THC-consumption was verified by analyzing THC-COOH in hair using GC-MS/MS. RESULTS: Cannabinoids were detectable in 95.95% of the hair samples from individuals who tested positive on a urine screen for cannabis. Δ9-THC concentrations were positively associated with measures of self-reported potency (relative potency, potency category, and perceived 'high'), but Δ9-THC, CBD, CBN concentrations and THC/CBD ratio were not associated with self-reported quantity of use. Self-reported potency, but not hair-derived concentrations, were associated with withdrawal and craving. Self-reported quantity of cannabis use, but not cannabinoid concentrations, were associated with cannabis use-related problems. CONCLUSIONS: The use of hair-derived cannabinoid quantification is supported for detecting cannabis use in near-daily users, but the lack of associations between hair-derived cannabinoid concentrations and self-report measures of use does not support the use of hair analyses alone for quantification of cannabinoid exposure. Further research comparing hair-derived cannabinoid concentrations with other biological matrices (e.g. plasma) and self-report is necessary to further evaluate the validity of hair analyses for this purpose.

Development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method including 25 novel synthetic opioids in hair and subsequent analysis of a Swiss opioid consumer cohort.

Major public health concern is raised by the evidence that common drugs like heroin are now frequently laced or replaced with highly potent novel synthetic opioids (NSOs). The objective of this study was to explore the prevalence and patterns of NSOs in a cohort of Swiss opioid users by hair analysis. Hair analysis is considered an ideal tool for retrospective consumption monitoring. Hair samples from 439 opioid users in Zurich were analyzed. Study inclusion required a previous positive hair test result for heroin metabolites, oxycodone, fentanyl, methadone, or tramadol. The samples were extracted with a two-step extraction procedure, followed by a targeted LC-MS/MS (QTRAP® 6500+) analysis in multiple reaction monitoring mode for a total of 25 NSOs. The method underwent full validation and demonstrated good selectivity and sensitivity with limits of detection (LOD) as low as 0.1 pg/mg. The analyzed sample cohort demonstrated a positivity rate for NSOs of 2.5%, including the following NSOs: butyrylfentanyl, acrylfentanyl, furanylfentanyl, methoxyacetylfentanyl, ocfentanil, U-47700, isobutyrylfentanyl and benzylfentanyl. Furthermore, we were able to identify specific consumption patterns among drug users. The results indicate that hair analysis is a valuable tool for investigating the prevalence of NSOs in drug-using populations, which seems to be low in the case of Swiss opioid users. Nevertheless, the results highlight the need for sensitive analytical detection methods in forensic toxicology to identify and monitor substance distribution in different populations.

Analytical description of adolescent binge drinking patients.

BACKGROUND: Binge drinking is a widespread health compromising behavior among adolescents and young adults, leading to significant health problems, injuries and mortality. However, data on alcohol consumption is often unreliable, as it is mainly based on self-reporting surveys. In this five-year study (2014-2019) at the University Children's Hospital Zurich, we analyzed blood samples from adolescent binge drinking patients to investigate blood alcohol concentrations (BACs), co-ingestion of drugs, assess compliance between self-reported and measured substance use, and test for genetic components of innate alcohol tolerance. Furthermore, hair analysis was performed to retrospectively access drug exposure and to evaluate the potential of hair analysis to assess binge drinking. METHODS: In a prospective, single-center study, patients with alcohol intoxications aged 16 years and younger were included. Blood and hair samples were analyzed by sensitive liquid chromatography - tandem mass spectrometry drug analysis. HTTLPR genotyping was performed with PCR and fragment analysis. RESULTS: Among 72 cases, 72 blood and 13 hair samples were analyzed. BACs ranged from 0.08-3.20‰ (mean 1.63‰, median 1.60‰), while a mean concentration of 3.64 pg/mg hair (median 3.0 pg/mg) of the alcohol marker ethyl glucuronide (EtG) was detected in eleven hair samples, providing no evidence of chronic excessive drinking. In 47% of the cases, co-ingested drugs were qualitatively detected next to ethanol, but only 9% of the detected drugs had blood concentrations classified as pharmacologically active. Cannabis consumption (22%) and stimulant intake (16%) were the most frequently observed drugs. Compliance between patients' statements and measured substances matched well. Although we investigated the genetic contribution to innate alcohol tolerance via the 5-HTTLPR polymorphism, the diverse genetic background of the cohort and small sample size did not allow any conclusions to be drawn. CONCLUSION: Almost half of our binge drinking patients tested positive for other substances, primarily cannabis. We anticipate that our study enhances understanding of consumption behavior of young people and encourage continued efforts to address the harmful effects of binge drinking and co-occurring substance use.

Associations of psychoactive substances and steroid hormones in hair: Findings relevant to stress research from a large cohort of young adults.

OBJECTIVE: Epidemiological studies increasingly use hair samples to assess people's cumulative exposure to steroid hormones, but how the use of different psychoactive substances may affect steroid hormone levels in hair is, so far, largely unknown. The current study addresses this gap by establishing the substance exposure correlates of cortisol, cortisone, and testosterone in hair, while also accounting for a number of relevant covariates. METHOD: Data came from a large urban community-sample of young adults with a high prevalence of substance use (N = 1002, mean age=20.6 years, 50.2% female), who provided 3 cm of hair samples. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) quantified cortisol, cortisone, and testosterone, as well as delta-9-tetrahydrocannabinol (THC), 3,4-methylenedioxymethamphetamine (MDMA, "Ecstasy"), cocaine, several opioids, and their respective metabolites. Multiple linear regression models with covariates were used to predict steroid hormone levels from substance exposure in a four-step approach: In the full sample, low and high substance hair concentrations (median split) were first tested against no use for each substance individually (step 1) and for all substances together (step 2). Then, within the participants with any substance in hair only, the continuous hair concentration of each substance in pg/mg (step 3) and finally of all substances together, were regressed (step 4). RESULTS: Low, high, and continuous levels of THC in hair were robustly associated with higher levels of cortisol (sig. in step 1 low THC: ß = 0.29, p = .021; high THC: ß = 0.42, p = .001; step 2: low THC: ß = 0.27, p = 0.036, and high THC: ß = 0.40, p = .004, and step 4: ß = 0.12, p = .041). Participants with high MDMA levels had higher levels of cortisone without adjusting for other substances (step 1: ß = 0.34, p = .026), but this effect was not significant in the other models. While high THC levels were associated with lower levels of testosterone in step 2 (ß = -0.35, p = .018), MDMA concentration was positively related to testosterone concentration with and without adjusting for other substances (step 3: ß = 0.24, p = .041; step 4: ß = 0.17, 95%, p = .015) in male participants. CONCLUSION: The use of psychoactive substances, especially of cannabis and ecstasy, should be considered in studies investigating steroid hormones in hair.

Advances in testing for sample manipulation in clinical and forensic toxicology-part B: hair samples.

As a continuation of part A, focusing on advances in testing for sample manipulation of urine samples in clinical and forensic toxicology, part B of the review article relates to hair, another commonly used matrix for abstinence control testing. Similar to urine manipulation, relevant strategies to manipulate a hair test are lowering drug concentrations in hair to undercut the limits of detection/cut-offs, for instance, by forced washout effects or adulteration. However, distinguishing between usual, common cosmetic hair treatment and deliberate manipulation to circumvent a positive drug test is often impossible. Nevertheless, the identification of cosmetic hair treatment is very relevant in the context of hair testing and interpretation of hair analysis results. Newly evaluated techniques or elucidation of specific biomarkers to unravel adulteration or cosmetic treatment often focused on specific structures of the hair matrix with promising strategies recently proposed for daily routine work. Identification of other approaches, e.g., forced hair-washing procedures, still remains a challenge in clinical and forensic toxicology.

How Do Road Traffic Noise and Residential Greenness Correlate with Noise Annoyance and Long-Term Stress? Protocol and Pilot Study for a Large Field Survey with a Cross-Sectional Design.

Urban areas are continuously growing, and densification is a frequent strategy to limit urban expansion. This generally entails a loss of green spaces (GSs) and an increase in noise pollution, which has negative effects on health. Within the research project RESTORE (Restorative potential of green spaces in noise-polluted environments), an extended cross-sectional field study in the city of Zurich, Switzerland, is conducted. The aim is to assess the relationship between noise annoyance and stress (self-perceived and physiological) as well as their association with road traffic noise and GSs. A representative stratified sample of participants from more than 5000 inhabitants will be contacted to complete an online survey. In addition to the self-reported stress identified by the questionnaire, hair cortisol and cortisone probes from a subsample of participants will be obtained to determine physiological stress. Participants are selected according to their dwelling location using a spatial analysis to determine exposure to different road traffic noise levels and access to GSs. Further, characteristics of individuals as well as acoustical and non-acoustical attributes of GSs are accounted for. This paper presents the study protocol and reports the first results of a pilot study to test the feasibility of the protocol.

Associations of different hormonal contraceptive methods with hair concentrations of cortisol, cortisone, and testosterone in young women.

Hair concentrations of cortisol, cortisone, and testosterone are non-invasive measures of cumulative steroid hormone levels. Use of contraceptives co-varies with levels of cortisol and cortisone in women's hair. It is unclear, however, how different contraceptive methods (i.e., that differ in their steroid hormone composition) affect corticosteroid and testosterone hair levels. The current study examines associations of contraceptives with hair steroid hormone concentrations in females from the community (N = 464, M = 20.6 years old, age range = 19-22). Self-reported contraceptives were first categorized as combined estrogen-progestin or progestin-only, and then analyzed individually in follow-up analyses. Multiple regressions adjusting for body mass index (BMI) and hair characteristics revealed that levels of hair cortisol, cortisone, and testosterone were significantly lower in women who used combined estrogen-progestin methods than in women who did not use hormonal contraception (ßcortisol(log) = -0.29; ßcortisone(log) = -0.28; ßtestosterone(log) = -0.36), showing moderate to large effect sizes (d = 0.64, d = 0.71, and d = 0.81, respectively). Concentrations of hair cortisol were lower in women who used progestin-only contraceptives (ß = -0.49) compared to no contraceptive use, with a large effect size (d = 1.67). Follow-up analyses revealed that the association of the three steroid hormones with estrogen-progestin methods was strongest for the combined oral "micro-pill." Future studies of hair steroid hormones should take into account the specific type of contraceptive used, as this may affect study results.

Single sample preparation for the simultaneous extraction of drugs, pharmaceuticals, cannabinoids and endogenous steroids in hair.

Recently, we published a multi-analyte method for the simultaneous analysis of 116 drugs and pharmaceuticals including different substance groups like opioids, stimulants, benzodiazepines, z-drugs, antidepressants and neuroleptics based on a single sample workup followed by a single analytical measurement with LC-MS/MS. However, in some cases, additional analysis of further substance groups, such as cannabinoids and endogenous steroids, is required, which are analyzed in our laboratory using separate sample preparation and separate analytical methods. The goal of this study was to use the knowledge from the different sample preparations and combine them into a single sample preparation and extraction workflow for the simultaneous extraction of drugs, pharmaceuticals, cannabinoids, and endogenous steroids to be analyzed with the appropriate analytical methods. A partial validation of selected parameters such as selectivity, linearity, limit of quantification (LOQ), accuracy, precision and robustness for the different analytical methods was carried out and revalidated. In addition, comparative measurements of quality controls and authentic pools were performed and statistically evaluated using the unpaired t-test or the non-parametric Mann-Whitney test. The results using the newly established sample preparation and extraction were in good agreement with the original data. In conclusion, the newly established sample preparation is suitable for the combined extraction of drugs, pharmaceuticals, cannabinoids and endogenous steroids, and gives reliable results for quantification of various substances.

Endocannabinoid and steroid analysis in infant and adult nails by LC-MS/MS.

A common method to quantify chronic stress is the analysis of stress markers in keratinized matrices such as hair or nail. In this study, we aimed to validate a sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the combined quantification of steroid hormones and endocannabinoids (eCBs) in the keratinized matrix nail. Furthermore, we aimed to investigate the suitability of the nail matrix for the detection of these stress markers in a pilot study. An LC-MS/MS method was used for the simultaneous identification and quantification of four eCBs (2-arachidonoylglycerol (2-AG), anandamide (AEA), oleoylethanolamide (OEA), palmitoylethanolamide (PEA)) and five steroid hormones (cortisol, cortisone, androstenedione, progesterone, testosterone) in human nails using a surrogate analyte method for each analyte. The method was validated in terms of selectivity, response factor, linearity, limit of quantification (LOQ), precision, accuracy, matrix effect, recovery, robustness, and autosampler stability. Nail samples were extracted for 1 h with methanol following a clean-up with a fully automated supported liquid extraction (SLE). The influence of nail weight on the quantification was investigated by using 0.5-20 mg of nail sample. As a proof of concept, nail samples (N = 57) were analyzed from a cohort representing newborns (1 month old), children (between 1 and 10 years), and adults (up to 43 years). It could be shown that the established workflow using a 1 hour extraction and clean-up by SLE was very robust and resulted in a short sample preparation time. The LC-MS/MS method was successfully validated. Matrix effects with ion enhancement occurred mainly for 2-AG. Sample weights below 5 mg showed variations in quantification for some analytes. Certain analytes such as PEA and progesterone could be accurately quantified at a sample weight lower than 5 mg. This is the first study where steroids and eCBs could be simultaneously detected and quantified in infant and adult nails. These results show that nails may serve as an alternative keratinized matrix (compared to hair) for the retrospective monitoring of cumulative eCB and steroid hormone levels. The combined assessment of eCBs and steroids from nails could provide a new approach to gain new insights into stress exposure in newborns and adults.

Alterations of Stress-Related Glucocorticoids and Endocannabinoids in Hair of Chronic Cocaine Users.

BACKGROUND: Previous research in animals and humans has demonstrated a potential role of stress regulatory systems, such as the hypothalamic-pituitary-adrenal (HPA) axis and the endocannabinoid (eCB) system, in the development of substance use disorders. We thus investigated alterations of HPA and eCB markers in individuals with chronic cocaine use disorder by using an advanced hair analysis technique. METHODS: We compared hair concentrations of glucocorticoids (cortisone, cortisol) and the eCBs 2-arachidonylglycerol, anandamide (AEA), oleoylethanolamide (OEA), and palmitoylethanolamide (PEA) between 48 recreational cocaine users (RCU), 25 dependent cocaine users (DCU), and 67 stimulant-naïve controls. Self-reported substance use and hair concentrations of substances were also assessed. RESULTS: Significantly higher concentrations of hair cortisone were found in RCU and DCU compared with controls. Hair concentrations of OEA and PEA were significantly lower in DCU compared with RCU and controls. Additionally, within cocaine users, elevated cocaine hair concentration was a significant predictor for increased glucocorticoid and decreased OEA hair levels. Moreover, higher 3,4-methyl​enedioxymethamphetamine hair concentration was correlated with elevated cortisone and AEA, OEA, and PEA levels in hair within cocaine users, whereas more self-reported cannabis use was associated with lower eCBs levels in hair across the total sample. CONCLUSION: Our findings support the hypothesis that the HPA axis and eCB system might be important regulators for substance use disorders. The mechanistic understanding of changes in glucocorticoid and eCB levels in future research might be a promising pharmacological target to reduce stress-induced craving and relapse specifically in cocaine use disorder.

A novel bedtime pulsatile-release caffeine formula ameliorates sleep inertia symptoms immediately upon awakening.

Sleep inertia is a disabling state of grogginess and impaired vigilance immediately upon awakening. The adenosine receptor antagonist, caffeine, is widely used to reduce sleep inertia symptoms, yet the initial, most severe impairments are hardly alleviated by post-awakening caffeine intake. To ameliorate this disabling state more potently, we developed an innovative, delayed, pulsatile-release caffeine formulation targeting an efficacious dose briefly before planned awakening. We comprehensively tested this formulation in two separate studies. First, we established the in vivo caffeine release profile in 10 young men. Subsequently, we investigated in placebo-controlled, double-blind, cross-over fashion the formulation's ability to improve sleep inertia in 22 sleep-restricted volunteers. Following oral administration of 160 mg caffeine at 22:30, we kept volunteers awake until 03:00, to increase sleep inertia symptoms upon scheduled awakening at 07:00. Immediately upon awakening, we quantified subjective state, psychomotor vigilance, cognitive performance, and followed the evolution of the cortisol awakening response. We also recorded standard polysomnography during nocturnal sleep and a 1-h nap opportunity at 08:00. Compared to placebo, the engineered caffeine formula accelerated the reaction time on the psychomotor vigilance task, increased positive and reduced negative affect scores, improved sleep inertia ratings, prolonged the cortisol awakening response, and delayed nap sleep latency one hour after scheduled awakening. Based on these findings, we conclude that this novel, pulsatile-release caffeine formulation facilitates the sleep-to-wake transition in sleep-restricted healthy adults. We propose that individuals suffering from disabling sleep inertia may benefit from this innovative approach.Trials registration: NCT04975360.

Systemic detectability of dexamethasone and prednisolone after eye drop application in horses.

BACKGROUND: Equine sport agencies list steroids as prohibited substances for competing horses. OBJECTIVES: The objective of this study was to investigate if the controlled substances dexamethasone and prednisolone are detectable in equine serum and urine samples during and after treatment with eye drops and if this can generate a positive doping test. STUDY DESIGN: Prospective cohort study. METHODS: The study cohort included 11 horses. One eye of the horses was treated with either dexamethasone (Maxitrol® 0.1%, n = 5 eyes) or prednisolone (Pred forte® 1%, n = 6 eyes) eye drops 3 times daily for 14 days. Dexamethasone and prednisolone concentrations were determined in serum and urine at day 0 (negative control), 1, 7, 14, 15, 17 and 21 using liquid chromatography-tandem mass spectrometry. Blood samples were collected within 2 hours post application. Urine samples were collected during spontaneous urination. RESULTS: All serum samples (range: 0.7-43 ng/mL, mean 2.1 ng/mL) and urine samples (range 1.2-5 ng/mL, mean 0.8 ng/mL) showed measurable amounts of dexamethasone during the course of treatment. Concentrations in both serum and urine samples were below limit of detection (LOD) 24 hours after the last dexamethasone treatment (day 15). All serum samples (range 1.1-32.5 ng/mL, mean 6.4 ng/mL) and urine samples (range 3.7-19 ng/mL, mean 4.6 ng/mL) were positive for prednisolone during treatment. Urine samples were below LOD on day 15; serum samples on day 21. CONCLUSIONS: Dexamethasone and prednisolone eye drops can induce detectable drug levels in serum and urine samples of horses after a 14-day treatment plan. This can lead to a positive doping result. All samples tested negative (below LOD of the analytical method) for dexamethasone one day and for prednisolone one week after treatment cessation.

Simultaneous quantification of steroid hormones and endocannabinoids (ECs) in human hair using an automated supported liquid extraction (SLE) and LC-MS/MS - Insights into EC baseline values and correlation to steroid concentrations.

Endogenous steroid hormones and endocannabinoids (ECs) are important regulators in the stress response of the human body. For the measurement of chronic stress, hair analysis has been established as method of choice for long-term and retrospective determination of endogenous stress markers. A sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of five steroid hormones (cortisone, cortisol, androstenedione, testosterone, progesterone) and four endocannabinoids (anandamide, palmitoylethanolamide, 2-arachidonylglycerol, oleoylethanolamide) in hair was developed and validated. The hair samples were extracted with methanol and cleaned up with a fully automated supported liquid extraction (SLE) before analysis. Special attention was paid to the difficulties accompanying the quantification of endogenous analytes in hair. Five different strategies for endogenous compound quantification in hair (surrogate analyte, standard addition, background correction, stripped matrix and solvent calibration) were tested and compared. As a result, the approach of the surrogate analyte was used for the quantification of steroid hormones whereas background correction was used for endocannabinoids. The measurement of 58 samples from healthy young adults allowed insights into endocannabinoid ranges in hair and the correlation to steroid hormones. No significant differences in steroid and EC concentration levels of male and female in hair were found, except for testosterone (p < 0.001) and androstenedione (p < 0.0001). Cortisol to cortisone and testosterone to androstenedione concentrations were significantly and positively correlated. There were significant intercorrelations between endocannabinoids.

Quantification of Classic, Prescription and Synthetic Opioids in Hair by LC-MS-MS.

The current use and misuse of synthetic and prescription opioids in the USA has reached epidemic status. According to the US Department of Health and Human Services, every day more than 130 people in the USA die after overdosing on opioids, and 2.1 million had an opioid use disorder in 2018. Hair is becoming an alternative matrix of increasing interest in forensic toxicology to investigate drug use and abuse patterns due to its long window of detection. The focus of this project was to develop and validate a method that simultaneously detects and quantifies 27 classic, prescription and synthetic opioids in hair by liquid chromatography-tandem mass spectrometry (LC-MS-MS). Hair samples were decontaminated and pulverized in a bead mill. Twenty-five milligrams of hair powder were incubated in a buffer overnight. Mixed mode cation exchange solid phase extraction was carried out before undergoing reversed-phase chromatographic separation, successfully resolving isobaric opioids. We used two multiple reaction monitoring transitions in positive mode to identify each analyte. The linearity range was 1-500 pg/mg for fentanyl and synthetic opioids and 10-500 pg/mg for prescription and classic opioids. Imprecision was <17.5% and bias ranged from -13.6 to 12.0%. Majority of compounds showed extraction efficiency >50%, and ion suppression from -89.2 to -26.6% (CV < 19%, n = 10). This method was applied to 64 authentic cases, identifying 13 compounds from our panel. A sensitive and specific method was developed for the identification and quantification of 27 classic, prescription and synthetic opioids in hair by LC-MS-MS.

Towards Best Practice in Hair Metabolomic Studies: Systematic Investigation on the Impact of Hair Length and Color.

Untargeted metabolomic studies are used for large-scale analysis of endogenous compounds. Due to exceptional long detection windows of incorporated substances in hair, analysis of hair samples for retrospective monitoring of metabolome changes has recently been introduced. However, information on the general behavior of metabolites in hair samples is scarce, hampering correct data interpretation so far. The presented study aimed to investigate endogenous metabolites depending on hair color and along the hair strand and to propose recommendations for best practice in hair metabolomic studies. A metabolite selection was analyzed using untargeted data acquisition in genuine hair samples from different hair colors and after segmentation in 3 cm segments. Significant differences in metabolites among hair colors and segments were found. In conclusion, consideration of hair color and hair segments is necessary for hair metabolomic studies and, subsequently, recommendations for best practice in hair metabolomic studies were proposed.

Cheating on forensic hair testing? Detection of potential biomarkers for cosmetically altered hair samples using untargeted hair metabolomics.

Hair analysis has become an integral part in forensic toxicological laboratories for e.g. assessment of drug or alcohol abstinence. However, hair samples can be manipulated by cosmetic treatments, altering drug concentrations which eventually leads to false negative hair test results. In particular oxidative bleaching of hair samples under alkaline conditions significantly affects incorporated drug concentrations. To date, current techniques to detect cosmetic hair adulterations bear limitations such as the implementation of cut-off values or the requirement of specialized instrumentations. As a new approach, untargeted hair metabolomics analysis was applied to detect altered, endogenous biomolecules that could be used as biomarkers for oxidative cosmetic hair treatments. For this, genuine hair samples were treated in vitro with 9% hydrogen peroxide (H2O2) for 30 minutes. Untreated and treated hair samples were analyzed using liquid-chromatography high-resolution time-of-flight mass spectrometry. In total, 69 metabolites could be identified as significantly altered after hair bleaching. The majority of metabolites decreased after bleaching, yet totally degraded metabolites were most promising as suitable biomarkers. The formation of biomarker ratios of metabolites decreasing and increasing in concentrations improved the discrimination of untreated and treated hair samples. With the results of this study, the high variety of identified biomarkers now offers the possibility to include single biomarkers or biomarker selections into routine screening methods for improved data interpretation of hair test results.

Hair cortisol measurement in older adults: Influence of demographic and physiological factors and correlation with perceived stress.

AIMS: This study aimed to investigate correlation between hair cortisol levels and perceived stress scale (PSS) in addition to a range of demographic and physiological factors in a sample of older adults. EXPERIMENTAL: Hair cortisol concentrations were established from 42 older adults aged between 60 and 80 years old. Age, sex, hair colour, smoking status, employment status, daytime sleeping, medication, waist to hip ratio (WHR) and PSS scores were assessed through a questionnaire. Hair cortisol concentration was assessed through liquid chromatography coupled to tandem mass-spectrometry (LC-MS/MS). RESULTS: Amongst the older adult group there was no statistically significant correlation between hair cortisol concentration and age, employment status, daytime sleep duration, WHR or PSS. When compared to previous data assessing hair cortisol in toddlers (7 months to 3 years old), adolescents (12-17 years old) and adults (18-60 years old) it is observed that there is a trend for higher hair cortisol in older adults (60-80 years old). Hair cortisol concentrations were significantly higher in males (n = 20) than in females (n = 22) and in participants with dark brown hair (n = 8). No relationship was investigated between hair cortisol concentration and smoking status or medication intake. CONCLUSIONS: The results confirm that hair samples are a useful alternative to the current mediums that are used to analyse biological cortisol. The results also validate the use of LC-MS/MS as an effective analytical method for the quantitation of hair cortisol concentrations.