Journal of analytical toxicology最新文献

The prevalence of mitragynine (kratom) in forensic toxicology casework has steadily increased over time. Readily available and currently legal, mitragynine is widely used for its stimulant and, depending on concentration, sedative effects. Our laboratory analyzed various fluid and tissue specimens from 51 postmortem cases to investigate the distribution of mitragynine and its active metabolite 7-hydroxymitragynine. Central and peripheral blood concentrations were compared, with an average heart blood to femoral blood ratio being 1.37 for mitragynine and 1.08 for 7-hydroxymitragynine. This ratio >1.0 suggests that mitragynine has some propensity toward postmortem redistribution; however, the difference in concentrations of mitragynine and 7-hydroxymitragynine is not statistically significant. Large average mitragynine to 7-hydroxymitragynine ratios of 30.9 in femoral blood and 32.4 in heart blood were observed compared to average ratios of 14.8 in vitreous humor and 16.9 in urine. In addition, the stability of these two compounds was investigated in both matrix and organic solvent. When stored refrigerated (4°C), mitragynine was stable for up to 30 days and 7-hydroxymitragynine was stable for up to 7 days with an analyte loss of <20%. Following 60 days of refrigerated storage, 7-hydroxymitragynine concentrations dropped over 50% from initial concentrations. Methanolic preparations of mitragynine and 7-hydroxymitragynine were stable following 3 months of storage at -20°C.

In recent years, there has been increasing interest on the use of alternative biological materials in forensic toxicology. Vitreous humor is one of them, which, due to the closed cavity it is contained, has a low degree of contamination and high purity that makes it ideal for use in post-mortem specimens. The aim of this study was to investigate the distribution of tramadol and its active metabolite O-desmethyltramadol in vitreous humor and the usefulness of using this alternative biological matrix in tramadol-related forensic cases. For this purpose, a GC/MS method for the determination of the two analytes in blood and vitreous humor samples, which included solid phase extraction and derivatization using N,O-Bis(trimethylsilyl)trifluoroacetamide with 1% trimethylsilyl chloride, was developed. The method was fully validated according to international guidelines and was applied to blood and vitreous humor samples from 12 forensic cases. Both substances were found to be readily distributed in vitreous humor, since even in cases of very low concentrations of the analytes in blood, their detection was also possible in vitreous humor. In addition, the vitreous humor to blood concentration ratios were calculated for both substances and the mean values were found to be 0.91 for tramadol and 0.94 for O-desmethyltramadol. The results of our study indicate that the information that can be extracted from the analysis of vitreous humor samples is particularly useful during the investigation of tramadol related cases. Nevertheless, the need for further study of this alternative material to establish therapeutic and toxic limits becomes apparent.

Ethyl glucuronide (EtG) and ethyl sulfate (EtS) are mostly analyzed in urine; consequently, most kinetic studies are based on urine samples. In forensic cases, however, it may be necessary to determine these alcohol biomarkers in serum, whole blood or capillary blood. While there are sufficient data on EtG and EtS in serum after alcohol consumption, the amount of data available on whole blood concentrations is small. Therefore, data on corresponding blood-to-serum ratios seem to gain importance. This study provides data on a drinking experiment with 5 drinking episodes, where serum and whole blood samples were taken simultaneously from 11 healthy participants over 10 days. The samples were analyzed for EtG, EtS, and ethanol. EtG and EtS analysis in whole blood and serum were performed by LC -MS/MS; ethanol was determined by GC-FID and an ADH-based method. EtG and EtS reached their maximum concentration 4-7 hours after alcohol consumption. For EtG, a mean blood-to-serum ratio of 0.58 with a range from 0.38 to 0.73 was found; for EtS, the mean ratio was 0.81 with a range from 0.61 to 0.92, indicating a predominant distribution in the serum. For both analytes, high correlation coefficients were obtained when plotting concentrations in serum against concentrations in whole blood. Concerning elimination profiles of the individuals, no time or concentration dependence of EtG or EtS blood-to-serum ratios could be deduced. Neither for EtG nor for EtS a regularity of curve progressions could be observed in our test specimens.

The 2018 Farm Bill legalized hemp and defined it as cannabis plant material having not more than 0.3% ∆9-tetrahydrocannabinol (∆9-THC) by dry weight. This has opened the door for the sale of hemp-derived ∆8-tetrahydrocannabinol (∆8-THC), a psychoactive isomer of ∆9-THC. Hemp has minimal amounts of naturally occurring ∆8-THC; however, the cannabidiol (CBD) found in hemp can be chemically converted into ∆8-THC. Unfortunately, depending on the method of conversion, the amount of ∆8-THC, ∆9-THC and other by-products can vary widely. For many laboratories, the emergence of ∆8-THC products resulted in analytical challenges because of the structural similarity of the isomers resulting in coelution. In response, a novel liquid chromatography-tandem mass spectrometry method was developed to separate the two isomers, with improved limit of detection (LOD) and lower limit of quantification (LLOQ). With this method, clear separation was achieved between ∆9-THC and ∆8-THC, 11-nor-9-carboxy-∆9-tetrahydrocannabinol (∆9-THC-COOH) and 11-nor-9-carboxy-∆8-tetrahydrocannabinol (∆8-THC-COOH) and partial separation of 11-hydroxy-∆9-tetrahydrocannabinol (∆9-THC-OH) and 11-hydroxy-∆8-tetrahydrocannabinol (∆8-THC-OH). While ∆9-THC-OH and ∆8-THC-OH did not achieve baseline separation, sufficient separation was achieved to confidently identify and differentiate the two compounds. LOD and LLOQ were the same for quantitative compounds. Quantitative range of 0.5 ng/mL to 100 ng/mL was achieved for ∆9-THC, ∆8-THC and ∆9-THC-OH and 2.5 ng/mL to 250 ng/mL for ∆9-THC-COOH. Qualitative analysis with LOD of 0.5 ng/mL was achieved for ∆8-THC-OH and 2.5 ng/mL for ∆8-THC-COOH. To achieve the desired LODs and LLOQs, alternate multiple reaction monitoring (MRM) transitions were also explored in addition to those utilized in the laboratory's prior method and other published methods. The method was validated following the American National Standards Institute/Academy Standards Board (ANSI/ASB) Standard 036, Standard Practices for Method Validation in Forensic Toxicology with minor exceptions, and was proven to be reliable and robust.

There is a growing interest for quantification of drugs in capillary blood. Phosphatidylethanol (PEth) is a biomarker for alcohol intake measured in whole blood, thus making it a candidate for capillary sampling. Our laboratory has been running a method for PEth quantification in venous blood since 2016 and we aimed to expand this method to also include capillary dried blood spot (DBS) samples. Two 10 µL volumetric absorptive microsampling (VAMS) devices, Capitainer®B Vanadate and Mitra® were included in the method development and validated. Calibrators and quality controls were spiked during the automatic sample extraction without the VAMS devices present, making it possible to extract and analyze both types of VAMS samples in the same set-up. With the Mitra device all pre-established validation criteria were fulfilled in the measuring range 0.03-4.0 µM (21-2812 ng/mL), including method comparison with our venous blood method. Capitainer fulfilled all validation criteria, except for the accuracy of samples with PEth levels ≥ 0.5 µM (≥ 352 ng/mL) (deviation -17.1 to -20.5%). The correlation analysis between Capitainer and the venous blood results showed no constant bias, but an acceptable small proportional mean difference of -7.6%. Overall, the method validation results for both Capitainer and Mitra were considered acceptable. Both devices were found suitable for the analyses of PEth.

Identification of N,N-dimethylpentylone (DMP) in counterfeit "Ecstasy" and "Molly" tablets poses risk to public health due to its adverse effects. Little information is available regarding the pharmacological activity or relevant blood or tissue concentrations of DMP, and even less is known about other structurally related beta-keto methylenedioxyamphetamine analogues on recreational drug markets, such as N-propyl butylone. Here, a novel toxicological assay utilizing liquid chromatography-tandem quadrupole mass spectrometry (LC-QQQ-MS) was developed and validated for the quantitation of DMP and five related synthetic cathinones (eutylone, pentylone, N-ethyl pentylone (NEP), N-propyl butylone, and N-cyclohexyl butylone), with chromatographic resolution from isomeric variants and quantitation performed by standard addition. A forensic series of 125 cases is presented for DMP and related analogs, along with pharmacological activity assessments using monoamine transporter and mouse behavioral assays. The blood concentration range for DMP in postmortem forensic cases was 3.3-4,600 ng/mL (mean: 320±570 ng/mL, median: 150 ng/mL), whereas pentylone, the primary N-desmethyl metabolite of DMP, was identified in 98% of cases with a concentration range 1.3-710 ng/mL (mean±SD: 105±120 ng/mL, median: 71 ng/mL). N-Propyl butylone, a newly identified synthetic cathinone, was quantitated in seven cases (mean±SD: 82±75 ng/mL, median: 50 ng/mL, range: 1.7-200 ng/mL). DMP displayed potent uptake inhibition at the dopamine transporter (IC50=49 nM), with 100-fold weaker potency at the serotonin transporter (IC50=4990 nM). DMP was a locomotor stimulant in mice (ED50=3.5 mg/kg) exhibiting potency relatively similar to eutylone, N-ethyl pentylone, and pentylone. Our results show that DMP is a psychomotor stimulant associated with adverse clinical outcomes leading to death. Forensic laboratories must continue to update testing methods to capture emerging drugs, with specific emphasis on resolution and identification of isomeric species. Following the scheduling of DMP in early-2024, there could be an anticipated market shift towards a new unregulated synthetic stimulant to replace DMP.

Natural toxins present an ongoing risk for human exposure that requires a rapid and accurate diagnosis for proper response. In this study, a qualitative liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method was developed and validated for the detection of a large, diverse selection of natural toxins. Data-dependent acquisition was performed to identify compounds with an in-house mass spectral library of 129 hazardous toxins that originate from plants, animals, and fungi. All 129 compounds were spiked into human urine, extracted, and evaluated for spectral library matching. Of these, 92 toxins met the quality criteria and underwent validation in urine matrix based on American National Standards Institute guidelines. A generalized workflow for method expansion was developed and enables the rapid addition of relevant compounds to the established method. This LC-HRMS method achieves efficient detection of natural toxins in urine, and the created workflow can rapidly increase compound coverage via method expansion.