LC-QToFMS Presumptive Identification of Synthetic Cannabinoids without Reference Chromatographic Retention/Mass Spectral Information. I. Reversed-Phase Retention Time QSPR Prediction as an Aid to Identification of New/Unknown Compounds

The application of Quantitative Structure-Property Relationship (QSPR) to the prediction of reversed-phase liquid chromatography retention behavior of Synthetic Cannabinoids (SC), and its use in aiding the untargeted identification of unknown SC are described in this paper. 1D, 2D molecular descriptors and fingerprints of 105 SC were calculated with PaDEL-Descriptor, selected with Boruta algorithm in R environment, and used to build-up a multiple linear regression model able to predict retention times, relative to JWH-018 N-pentanoic acid-d5 as internal standard, under the following conditions: Agilent ZORBAX Eclipse Plus C18 (100 mm*2.1 mm I.D., 1.8 mum) column with Phenomenex SecurityGuard Ultra cartridge (C18, 10 mm*2.1 mm I.D., <2mum) kept at 50\ub0C; gradient elution with 5 mM ammonium formate buffer (pH 4 with formic acid) and acetonitrile with 0.01% formic acid, flow rate 0.5 ml/min. The model was validated by repeated k-fold cross validation using 2/3 of the compounds as training set and 1/3 as test set (Q2 0.8593; Root Mean Squared Error, 0.087, ca. 0.56 min; Mean Absolute Error, 0.060) and by predicting rRT of 5 SC left completely out of the modeling study. Application of the model in routine work showed its capacity to discriminate isomers, to identify unexpected SC in combination with mass spectral information, and to reduce the length of the list of candidate isomers to ca. 1/3, thus reducing significantly the time required for predicting high-resolution product ion spectra to be compared to the unknown using a computational MS search/identification approach

LC-QToFMS Presumptive Identification of Synthetic Cannabinoids without Reference Chromatographic Retention/Mass Spectral Information. II. Evaluation of a Computational Approach for Predicting and Identifying Unknown High-Resolution Product Ion Mass Spectra

Despite LC-HR-MS2 enables untargeted acquisition, data processing in toxicological screenings is almost invariably performed in targeted mode. We developed a computational approach based on open source chemometrics software that, starting from a suspected Synthetic Cannabinoid determined formula, searches for isomers in different NPS web databases (NPSWD), predicts retention time (RT) and high-resolution MS2 spectrum, and compares them with the unknown providing a rank-ordered candidates list. R was applied on 105 SC measured data to develop and validate a multiple linear regression QSAR model predicting RT. CFM-ID freeware was used to predict/compare spectra with Jaccard similarity index. Data-dependent acquisition was performed with an Agilent Infinity 1290 LC-6550 iFunnel Q-TOF MS with ZORBAX Eclipse-Plus C18 (100x2.1 mm/1.8 \ub5m) in water/acetonitrile/ammonium formate gradient. Ability of the combined RT/MS2 prediction to identify unknowns was evaluated on SC standards (with leave-one-out from the RT model) and on unexpected SC encountered in real cases. RT prediction reduced the number of isomers retrieved from NPSWD to 1/3 (2792\ub13358\u2192845\ub1983) and differentiated between SC isomers when spectra were not selective (4F-MDMB-BUTINACA, 4F-MDMB-BUTINACA 2'-indazole isomer) or unavailable (4CN-Cumyl-B7AICA, 4CN-Cumyl-BUTINACA). When comparing 30/40 eV measured spectra of 99 SC against RT-selected, CFM-ID predicted spectra of isomers, the right candidate ranked 1st on median and 4th on average; 54% and 88% of times the right match ranked 1st or within the first 5 matches, respectively. To our knowledge, this is the first case of extensive chemometrics application to toxicological screening. In most cases presumptive identification (being based on computation, it requires further information for confirmation) of unexpected SC was achieved without reference measured information. This method is currently the closest possible to true unbiased/untargeted screening. The bottleneck of the method is the processing time required to predict mass spectra (ca. 30-35 s/compound using a 64-bit 2.50-GHz Intel\uae Core\u2122 i5-7200U CPU). However, strategies can be implemented to reduce prediction processing time. Keywords: Untargeted Screening, Synthetic Cannabinoids, High Resolution Mass Spectrometry, MS-MS Spectrum Prediction

Higher Creatinine Concentrations in Ethyl Glucuronide-Positive Urine Specimens Collected from Subjects in a Controlled Alcohol Abstinence Program: Is Serum Creatinine a Good Marker of Renal Function in Drinkers?

Creatinine is higher in ethyl glucuronide-positive urines of drivers under controlled alcohol abstinence program, accounting for a late antidiuretic effect of alcohol. The consequent temporary decrease in serum creatinine may account for its negative trend at the increase in alcohol intake observed in epidemiological studies.AbstractAims The aim of this study was to examine urine creatinine concentrations in drivers submitted to controlled alcohol abstinence programs.Methods Urine samples (n = 32,210) were screened for ethyl glucuronide (EtG) by immunoassay during a 2-year period. Non-negatives underwent EtG and ethyl sulfate (EtS) confirmation by coupled-column Liquid Chromatography-Tandem Mass Spectrometry. Urine samples were tested for dilution by the analysis of creatinine content with <0.2 g/l indicating a dilute specimen.Results The mean urine creatinine was significantly higher in EtG positives compared to negatives (1.47 0.98 vs. 1.17 +/- 0.79 g/l). The difference between positives and negatives was consistent within genders and age groups (<45; 45). The higher urinary creatinine in EtG positives is explained by a late antidiuretic effect of alcohol.Conclusion Attempts to dilute urine specimens by drinking water or other liquids before voiding are less effective for EtG/EtS compared with illicit drugs excreted in urine. If the temporary decrease in serum creatinine as a consequence of the late antidiuretic effect of alcohol is confirmed by controlled studies, serum creatinine as an indicator of kidney function should be reconsidered in drinkers

Synthetic cannabinoids in hair—Prevalence of use in abstinence control programs for driver's license regranting in Germany

Although the use, structural variety, and prevalence of synthetic cannabinoids (SCs) have steadily increased on the drug market, they are rarely analyzed in abstinence control programs for driver's license regranting. The aim of this study was to determine the SC prevalence in these programs by analyzing hair samples collected between March 2020 and March 2021 from various regions in Germany, mainly Bavaria (40%). Specimens were analyzed quantitatively for drugs of abuse and qualitatively for 107 SCs. Hair samples were screened by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and to search for unknown SC analogs, positive samples were additionally screened by liquid chromatography-high resolution time of flight mass spectrometry (LC-qTOF/MS). The analysis of 5097 hair samples resulted in 181 SC detections (3.6%), showing a wide range of 44 SCs, with up to 13 different compounds found in a single sample. The most prevalent compounds were 5F-MDMB-PICA and MDMB-4en-PINACA; furthermore, 10 new substances not initially covered by LC-MS/MS analysis were detected by LC-qTOF/MS. The SC positivity rate was comparable to cocaine (5.4%) and amphetamine (2.6%). Only in 35 cases (0.7%), SC analysis was requested by the clients, highlighting the insufficient coverage of SC consumption in the studied collective. In summary, hair sample analysis proved to be a valuable tool to monitor the use of SCs. In order to keep pace with newly emerging SC analogs, an up-to-date analytical method is essential. Prospectively, SCs should be more routinely screened in hair analysis for abstinence control to avoid cannabis substitution by SCs.Synthetic cannabinoids (SCs) have steadily increased on the drug market but are rarely analyzed in abstinence control programs for driver's license regranting. Between March 2020 and March 2021, hair samples from this collective were retrospectively screened for SCs by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, and those testing positive were additionally screened by liquid chromatography-high resolution time of flight mass spectrometry (LC-qTOF/MS).imag