Determination of barbiturates in hair samples by using a validated UHPLC-HRMS method: application in investigation of drug-facilitated sexual assault.

In recent years, benzodiazepines and benzodiazepine-like drugs are the most common substances associated with drug-facilitated sexual assaults (DFSA); however, barbiturates are also detected occasionally. Segmental hair analysis provides useful information on the historic pattern of drug use, enabling differentiation between single exposure in DFSA cases and chronic use. However, sensitive and specific methods for barbiturate analysis in hair samples are needed. Herein, we present an ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS) method for qualitative and quantitative determination of seven barbiturates in hair samples. Firstly, a hair strand was decontaminated and then freeze-milled in liquid nitrogen. Next, 50 mg of powdered hair was extracted with methanol in an ultrasonic bath for 10 min in the presence of 10 ng phenobarbital-d5. The supernatant was dried under nitrogen gas, and the pellet was dissolved in 100 µL mobile phase. Afterwards, 10 µL of the suspension was injected into the UHPLC-HRMS system. The present method involved two UHPLC conditions for determination of barbiturates (I) and identification of the structural isomers amobarbital and pentobarbital (II). This method showed satisfactory linearity in a range of 0.02-20.00 ng/mg for UHPLC conditions I and II, both with a high determination coefficient (0.9991-0.9999). The selectivity, intra- and interday precision, accuracy and matrix effect of the method were acceptable. Next, the validated method was applied to investigate an authentic DFSA case. Hair samples (black, approximate 25 cm long) were collected 3 months after the assault, and the proximal segments (0-5 cm from the root; each segment was 1 cm long) were analyzed. Amobarbital was detected at a concentration of < LOQ (limit of quantification) and 0.09 ng/mg in the second and third 1 cm hair segment but not in the other segments. Thus, our method was successful in determining barbiturate concentration in human hair after a single-dose exposure, showing its potential for application in the investigation of DFSA cases.

Analysis of Five Mushroom Toxins in Blood by UPLC-HRMS. / 血液中5ç§è˜‘è‡æ¯’ç´ çš„UPLC-HRMS分析.

OBJECTIVES: To develop a method for the simultaneous and rapid detection of five mushroom toxins (α-amanitin, phallacidin, muscimol, muscarine and psilocin) in blood by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS). METHODS: The blood samples were precipitated with acetonitrile-water solution(Vacetonitril∶Vwater=3∶1) and PAX powder, then separated on ACQUITY Premier C18 column, eluted gradient. Five kinds of mushroom toxins were monitored by FullMS-ddMS2/positive ion scanning mode, and qualitative and quantitative analysis was conducted according to the accurate mass numbers of primary and secondary fragment ions. RESULTS: All the five mushroom toxins had good linearity in their linear range, with a determination coefficient (R2)≥0.99. The detection limit was 0.2-20 ng/mL. The ration limit was 0.5-50 ng/mL. The recoveries of low, medium and high additive levels were 89.6%-101.4%, the relative standard deviation was 1.7%-6.7%, the accuracy was 90.4%-101.3%, the intra-day precision was 0.6%-9.0%, the daytime precision was 1.7%-6.3%, and the matrix effect was 42.2%-129.8%. CONCLUSIONS: The method is simple, rapid, high recovery rate, and could be used for rapid and accurate qualitative screening and quantitative analysis of various mushroom toxins in biological samples at the same time, so as to provide basis for the identification of mushroom poisoning events.

A high-throughput metabolomics approach for the comprehensive differentiation of four Pulsatilla Adans herbs combined with a nontargeted bidirectional screen for rapid identification of triterpenoid saponins.

Pulsatilla Adans (PSA) herbs (Ranunculaceae) have been widely used in traditional medicine in China and other countries. However, the authentication and quality control of PSA herbs have always been a challenging task due to their similar morphological characteristics and the diversity of the multiple components that exist in the complicated matrix. Herein, a novel integrated strategy combining UHPLC/Q-Orbitrap-MS techniques with chemometrics analysis is proposed for the discrimination of PSA materials. We developed a comprehensive method integrating a nontargeted bidirectionally screened (NTBDS) MS data set and a targeted extraction peak area analysis for the characterization of triterpenoid saponins of PSA from different species. After that, partial least-squares discriminant analysis (PLS-DA) was performed on the obtained MS data set and the parameter variable importance for the projection (VIP) value and P value were employed to screen the valuable MS features to discriminate PSA from different species. In addition, the receiver operating characteristic (ROC) curve is used to verify the reliability of MS features. Finally, heatmap visualization was employed to clarify the distribution of the identified triterpenoid saponins, and four medicinal species of PSA were successfully differentiated. Additionally, 34 constituents were reported in PSAs for the first time, 81 triterpenoid saponins were identified as differential components, and 12 chemical ingredients were characterized as potential chemical markers to differentiate the four officinal PSA herbs. This is the first time that the differences in different PSA herbs have been observed systematically at the chemical level. The results suggested that using the identified characteristic components as chemical markers to identify different PSA herbs was effective and viable. This method provides promising perspectives in the analysis and identification of the ingredients of Chinese herbal medicines, and the identification of similar herbs from the same species.