Analysis of Cannabinoids and Metabolites in Dried Urine Spots (DUS).

Dried urine spots (DUS) represent a potential alternative sample storage for forensic toxicological analysis. The aim of the current study was to develop and validate a liquid chromatographic tandem mass spectrometric procedure for the detection and quantitative determination of cannabinoids and metabolites in DUS. A two-step extraction was performed on DUS and urine samples. An LC-MS/MS system was operated in multiple reaction monitoring and positive polarization mode. The method was checked for sensitivity, specificity, linearity, accuracy, precision, recovery, matrix effects and carryover. The method was applied to 70 urine samples collected from healthy volunteers and drug addicts undergoing withdrawal treatment. The method was successfully developed for DUS. LODs lower than 2.0 ng/mL were obtained for all the monitored substances. All the validation parameters fulfilled the acceptance criteria either for DUS or urine. Among the real samples, 45 cases provided positive results for at least one compound. A good quali-quantitative agreement was obtained between DUS and urine. A good stability of THC, THCCOOH and THCCOOH-gluc was observed after a 24 h storage, in contrast to previously published results. DUS seems to provide a good alternative storage condition for urine that should be checked for the presence of cannabinoids and metabolites.

Determination of antidepressants and benzodiazepines in paired hair and nail samples.

Hair and nails are keratinized matrices that can be used in Toxicology as matrices for the long-term detection of substances. Whereas hair is an established matrix with decades of use in this field, nails have been less studied, especially including a comparison to hair samples. Specifically in the case of antidepressant and benzodiazepine drugs, very few publications analyzing these drugs in nail samples exist as of yet. For this reason, in the present study a method for the detection of 12 antidepressant and benzodiazepine drugs in hair and nail samples was developed. Samples were decontaminated with 3 washes of dichloromethane, and 25 or 30 mg of hair and nails, respectively, were pulverized. Then, the samples were incubated with 1.5 mL water:ACN (50:50, v/v) with horizontal agitation for 90 min. The supernatant was evaporated and reconstituted in 200 µL of methanol and 2 mL of 2% FA in water, submitted to solid phase extraction (SPE) using Oasis MCX cartridges and analyzed by LC-MS/MS. The method was satisfactorily validated in nail and hair samples for the following parameters: linearity, LOD (0.005-0.02 ng/mg), LOQ (0.01-0.02 ng/mg), selectivity, carryover, accuracy, imprecision, matrix effect, extraction efficiency, process efficiency and autosampler stability. Matched fingernail, toenail and hair samples were obtained from 21 patients under treatment with any of the studied drugs and analyzed with the developed method. The most frequently detected drugs were venlafaxine (n = 11), trazodone (n = 6), zolpidem (n = 5), alprazolam (n = 5) and nordiazepam (n = 5). Concentrations in hair, fingernails and toenails, respectively, were 44.31 ng/mg, 8.05-43.35 ng/mg and 7.02-22.69 ng/mg for venlafaxine; 5.40-19.08 ng/mg, 0.13-1.00 ng/mg and 0.42-1.04 ng/mg for trazodone; 13.86 ng/mg, 5.19 ng/mg and 9.11 ng/mg for fluoxetine; 7.42 ng/mg, 1.85 ng/mg and 0.03-2.81 ng/mg for sertraline; 0.40-1.42 ng/mg, 0.12 ng/mg and 0.16 ng/mg for zolpidem; and 0.02-0.11 ng/mg, 0.07-1.07 ng/mg and 0.05 ng/mg for alprazolam for the patients under active treatment. Hair concentrations were higher than nail concentrations for most drugs in patients under active treatment, with the exception of diazepam (n = 1; 0.12 ng/mg in hair and 0.41 ng/mg in fingernails). Fingernail concentrations were lower than toenail concentrations in patients under active treatment in most compared cases. Comparison of fingernails and toenails of a patient with antifungal treatment did not show an observable effect in concentrations.

Determination and Distribution of Cannabinoids in Nail and Hair Samples.

Hair has been used for decades in toxicology as a biological matrix for long-term detection of substances. Nails are another keratinized matrix that is being studied as an alternative when hair cannot be obtained. Although cannabis is the most prevalent illicit drug in the world, cannabinoid distribution in nails compared with hair has been scarcely studied. In this work, we described two methods for the determination of cannabidiol (CBD), cannabinol (CBN) and Δ9-tetrahydrocannabinol (THC), and main metabolites of THC [11-nor-9-carboxy-THC (THCCOOH), 11-hydroxy-THC (OHTHC) and 8-ß-11-dihydroxyTHC (diOHTHC)] in nail and hair samples. After an alkaline hydrolysis, samples were submitted to solid-phase extraction and analyzed by liquid chromatography with tandem mass spectrometry (LC-MS-MS). The methods were fully validated, with good linearity (r2 > 0.99) in the range of 20 to 100 to 20,000 pg/mg. No endogenous or exogenous interferences were found. Accuracy was from 99.5% to 109.8%, and imprecision was <6.9%. Ion suppression (up to -74.4%) was observed for all the analytes, except for diOHTHC at low concentrations in hair (46.1%). Extraction efficiency ranged from 21.5% to 84.5%. The methods were applied to matched nail and hair specimens from 23 cannabis users to study the incorporation and distribution of the cannabinoids into these matrices. Only CBD, CBN and THC were detected in the samples, with much higher concentrations in fingernails than in toenails and hair. Correlations between analyte concentrations in the different matrices and with reported drug consumption were studied. A preliminary cut-off for THC in toenails was calculated using the cut-off proposed by the Society of Hair Testing in hair for the identification of chronic cannabis use.

Determination of antipsychotic drugs in nails and hair by liquid chromatography tandem mass spectrometry and evaluation of their incorporation into keratinized matrices.

Nail samples are an alternative to hair for long-term monitoring of drug use, although there are a limited number of studies about its applicability. This study presents the development and validation of a LC-MS/MS method for the determination of five antipsychotic drugs (clozapine, haloperidol, levomepromazine, olanzapine and quetiapine) in nail and in hair samples. Samples were washed with dichloromethane, pulverized with a ball mill, and incubated in water:acetonitrile (50:50 v/v) with horizontal agitation for 90 min. Then, samples were purified by solid phase extraction with OASIS MCX cartridges and analysed by LC-MS/MS. The analytical method was fully validated in nails and in hair, including: limits of detection (2.5 pg/mg and 2.5-10 pg/mg, respectively), limits of quantification (LOQ) (10 pg/mg and 10-20 pg/mg, respectively), linearity (LOQ to 10,000 pg/mg), selectivity (no endogenous or exogenous interferences), accuracy (97.4-106.9% and 97.3-108.2%, respectively), imprecision (<7.9% and <8.6%, respectively), extraction efficiency (62.3%-109.8% and 45.1%-83.6%, respectively), matrix effect (-35.6% to 654.4% and -71.0% to -10.8%, respectively) and autosampler stability after 72 h (%loss <12.4%). Moreover, paired fingernail, toenail and hair samples from 13 patients under chronic treatment were analysed, and concentrations in the different matrices were compared. Concentrations in real samples ranged from 11.3-8306 pg/mg in fingernails, from 12.7-1755.4 pg/mg in toenails and from 17.6-24045.4 pg/mg in hair. Hair concentrations were generally higher than nail concentrations; however, a variable pattern was found in fingernails and toenails depending on the case. In addition, concentrations in paired hair and nail samples from a patient under chronic treatment with quetiapine at different doses were studied for 1-year.