First Report of Pectobacterium polaris Causing Soft Rot on Broccoli in China.

In April 2023, soft rot symptoms were observed in broccoli (Brassica oleracea L. var. italica) commercial fields in Songming County, Yunnan province, China (103°12'E, 25°31'N). The disease incidence in these fields (6 ha in size) was high, exceeding 50%, and it caused significant yield loss. The affected plants displayed characteristic symptoms, with the roots and stems of broccoli becoming soft, yellowish-brown, rotten, and emitting a foul odor. To identify the causal agent, soft rot symptomatic stems were surface sterilized by dipping them in 75% ethanol for 30 seconds, followed by three successive rinses with sterile distilled water. Tissue specimens were then plated onto nutrient agar (NA) plates and incubated at 28°C for 24 hours. (Wang et al. 2022). Three representative bacterial isolates HYC22041801-HYC22041803 from broccoli were selected for further analysis. The colonies on NA plates appeared as white, small, round, and translucent with smooth edges. Physiological and biochemical tests were performed, along with 96 phenotypic screenings using the BIOLOG GENIII microplate system (Biolog, Hayward, CA, USA). Three isolates were negative for D-arabitol, maltose, and sorbitol, but were positive for cellobiose, α-D-glucose, sucrose, glycerol and gentiobiose tests, which are consistent with the reported type strain P. polaris NIBIO1006T (Chen et al. 2021). Total genomic DNA was extracted from three bacterial isolates using the QIAamp DNA Mini Kit (QIAGEN, USA). The 16S rRNA region and nine housekeeping genes (gapA, icdA, mdh, mtlD, pel, pgi, pmrA, proA and rpoS) were amplified with universal primers 27F/1492R (Monciardini et al., 2006) and designed specific primers (Xie et al., 2018), respectively. All amplicons were sequenced and deposited in GenBank with accession numbers ON723841-ON723843 and ON723846-ON723872. The BLASTn analysis of the 16S rRNA amplicons confirmed that the isolates HYC22041801-HYC22041803 belonged to the genus Pectobacterium. Phylogenetic trees based on 16S rRNA gene sequences and multilocus sequence analysis of other nine housekeeping genes of the three isolates were constructed and the results revealed that three isolates clustered with P. polaris type strain NIBIO1006T, which was previously isolated from potato (Dees et al., 2017). To confirm the pathogenicity, nine broccoli seedlings were stab inoculated with a bacterial suspension (108 CFU·ml-1), while sterile distilled liquid LB medium was used as a negative control. The seedlings were kept at 80% relative humidity and 28°C in a growth chamber. Three trials were conducted per isolate (HYC22041801-HYC22041803). After 3 days, the inoculated petioles showed soft rot symptoms similar to those observed initially in the field, while control plants remained asymptomatic. All three isolates were re-isolated successfully from symptomatic tissues to complete Koch's postulates. P. polaris has been previously reported as the causative agent of blackleg in potato in several countries, including Norway, Poland, Russia, and China (Handique et al. 2022; Wang et al. 2022). Additionally, it was reported to cause soft rot in Chinese cabbage in China (Chen et al. 2021). However, this is the first report of P. polaris causing soft rot disease in broccoli in China. This discovery is of great importance for vegetable growers because this bacterium is well established on Cruciferous vegetables in the local area, and effective measures are needed to manage this disease.

[Determination of four amide synthetic cannabinoid isomers by ultra-high performance liquid chromatography-high resolution mass spectrometry].

Isomerization commonly occurs in synthetic cannabinoids (SCs). Owing to the few differences in their structure and properties, it is difficult to simultaneously separate and identify them. Thus, the identification of synthetic cannabinoids is challenging, posing a threat to public security. This study aims to separate and identify four SCs, which are 2-[1-(5-fluoropentyl)-1H-indole-3-formylamino]-3,3-dimethylbutyrate methyl ester (5F-MDMB-PICA), 2-[1-(5-fluoropentyl)-1H-indole-3-formylamino]-3-methylbutyrate ethyl ester (5F-EMB-PICA), N-(1-amino-2,2-dimethyl-1-oxobutyl-2-yl)-1-butyl-1H-indazole-3-formamide (ADB-BINACA), N-(1-carbamoyl-2-methylpropyl)-1-pentyl indazole-3-formamide (AB-PINACA).Supercritical fluid chromatography-mass spectroscopy (SFC-MS) can realize the effective separation of some cannabinoid isomers. However, most laboratories are not equipped with SFC-MS systems. Ultra-high performance liquid chromatography-high resolution mass spectroscopy (UHPLC-HRMS) effectively combines the excellent efficient separation characteristics of liquid chromatography and the powerful qualitative ability of mass spectrometry. It is a commonly used technical method for the detection of amide synthetic cannabinoids and their metabolites in vivo and in vitro because of its advantages of high accuracy and efficiency. Liquid chromatography allows the separation of tested components by exploiting the difference in the partition coefficients between the mobile and stationary phases. When the two phases are in relative motion, the tested components are divided between the two phases, facilitating the separation and analysis of each component. Although the difference in the polarities of the tested amide synthetic cannabinoid isomeric substances is extremely small, liquid chromatography can induce a strong separation effect. The advantages of UHPLC-HRMS include high resolution imparted by mass spectrometry and high sensitivity, allowing its application in the qualitative analysis of various substances. Through UHPLC-HRMS, trace analytes at the nanogram scale as well as pure drugs and their metabolites in biosamples can be detected. This study proposed a method for the determination of two pairs of amide synthetic cannabinoid isomers-5F-EMB-PICA and 5F-MDMB-PICA, ADB-BINACA and AB-PINACA-through UHPLC-HRMS. A Hypersil GOLD C18 column (100 mm×2.1 mm, 1.9 µm) was selected for separation via liquid chromatography, and gradient elution was performed with methanol containing 0.1% formic acid and a 0.1% formic acid aqueous solution containing 10 mmol/L ammonium formate. Full scan/data-dependent secondary mass spectrometry (Full MS/dd-MS2) was conducted in the positive ion mode for detection. The results indicated that the four synthetic cannabinoid isomers could be accurately detected under the abovementioned conditions. The resolution between 5F-EMB-PICA and 5F-MDMB-PICA was 2.06, while that between ADB-BINACA and AB-PINACA was 1.22, indicating the effective separation and detection of both pairs. Furthermore, method validation was conducted to ensure the accuracy of the proposed method. The relationship of the four amide synthetic cannabinoid isomers exhibited excellent linearity. The correlation coefficients (R2) were >0.99. Moreover, the matrix effects of the four SCs in hair samples were between 88.67% and 111.76% and the recoveries were 96.23%-105.11%. The intra-day and inter-day precisions (RSDs) were <10%. The proposed method was used to identify the case materials. AB-PINACA was detected in a hair sample at a content of 0.73 µg/g. 5F-MDMB-PICA was detected in a tobacco sample at a content of 11.3 mg/g. The results indicate that the proposed method can be used for the examination of practical samples conducted by public security organizations. This study provides a reference method for the identification of synthetic cannabinoid isomers.

Analysis on dynamic changes of etizolam and its metabolites and exploration of its development prospect using UPLC-Q-exactive-MS.

As one of the most widely abused designer benzodiazepines in the world, etizolam has been found in many cases in many countries. In this study, UPLC-Q-Exactive-MS was used for the first time to establish a dynamic change model of etizolam and its metabolites in rats. Compared with previous studies, the detection sensitivity and reproducibility of the instrument were higher. In the experiment, we optimized the traditional pharmacokinetic model based on Gauss function. According to the significant difference of etizolam in the plasma elimination phase of rats, a new pharmacokinetic model based on Lorentz function was established to describe the dynamic changes of etizolam more rigorously, which made the error effects lower and the accuracy of the pharmacokinetic parameters was improved. At the same time, the pharmacokinetic parameters of etizolam were compared with four other designer benzodiazepines reported in previous studies in rats, and we found the direct reason for the popularity of etizolam in the NPS market and explored the future development of etizolam for the first time. In addition, 21 metabolites were found through rat experiments to effectively detect etizolam abuse for a long time, of which 4 metabolites had the longest detection window and could be used as long-acting metabolites for experiments, which greatly prolongs the detection window and extends the time range in which etizolam was detected in real cases. This study is the first to conduct a systematic and comprehensive study on the metabolism and pharmacokinetics of etizolam and find out the direct reason for the prevalence of etizolam abuse, and we also discuss the development trend of etizolam in the future market of new psychoactive substances, which is beneficial for forensic experts to assess the trend of drug abuse and can provide reference for relevant drug control and drug treatment.

Predictive value of speckle tracking technique for coronary artery stenosis in patients with coronary heart disease.

OBJECTIVE: To observe the predictive value of speckle tracking technique (STI) for the degree of coronary artery stenosis in patients with coronary heart disease (CHD). METHODS: The clinical data of 120 patients with coronary artery stenosis admitted to Affiliated Hospital of Chengde Medical University from Feb. 2022 to Sep. 2022 was analyzed retrospectively. The other 63 patients who sought for medical help because of chest pain underwent Coronary Arteriography (CAG) examination during the same period but with Gensini score > 0 were selected as the control group. Coronary artery stenosis was divided into three subgroups according to the coronary Gensini score: mild, moderate, and severe stenosis. Routine ultrasound and STI techniques were performed in all patients. In addition, left ventricular global radial peak systolic strain (GRS), left ventricular global longitudinal peak systolic strain (GLS), left ventricular global peak systolic strain (GAS) and left ventricular global circumferential peak systolic strain (GCS) were measured and compared between the two groups and among the three subgroups. RESULTS: There were no marked differences identified in conventional ultrasound parameters between the coronary artery stenosis group and control group, but the absolute values of GLS, GRS, GCS, and GAS were lower in the former group compared to control group; and the severe group had the lowest levels of above indexes, followed by moderate group, then mild group and control group (all P < 0.05). The results showed that the area under the curve (AUC) for GLS, GRS, GCS, and GAS in diagnosing coronary artery stenosis were 0.973, 0.933, 0.947, and 0.901, respectively. The AUCs of GLS, GRS, GCS, and GAS for the diagnosis of moderate/severe coronary artery stenosis were 0.968, 0.908, 0.901, and 0.942, respectively, with GAS and GLS assessed with the largest AUC values and higher sensitivity and specificity than other parameters. CONCLUSION: The global longitudinal strain of left ventricle obtained by STI technique was more sensitive to coronary artery stenosis than that by ultrasound technique, and it had a higher predictive value for coronary artery stenosis.

Metabolic Profile Analysis of Designer Benzodiazepine Etizolam in Zebrafish and Human Liver Microsomes.

As one of the most widely abused designer benzodiazepines worldwide, Etizolam is characterized by its high addiction potential, low production cost, and difficulty in detection. Due to the rapid metabolism of Etizolam in the human body, the probability of detecting the Etizolam parent drug in actual case samples by forensic personnel is low. Therefore, without detecting the parent drug, analysis of Etizolam metabolites can help forensic personnel provide references and suggestions on whether the suspect has taken Etizolam. This study simulates the objective metabolic process of the human body. It establishes a zebrafish in vivo metabolism model and a human liver microsome in vitro metabolism model to analyze the metabolic characteristics of Etizolam. A total of 28 metabolites were detected in the experiment, including 13 produced in zebrafish, 28 produced in zebrafish urine and feces, and 17 produced in human liver microsomes. The UPLC-Q-Exactive-MS technology was used to analyze the structures and related metabolic pathways of Etizolam metabolites in zebrafish and human liver microsomes, and a total of 9 metabolic pathways were identified, including monohydroxylation, dihydroxylation, hydration, desaturation, methylation, oxidative deamination to alcohol, oxidation, reduction acetylation, and glucuronidation. Among them, metabolites involving hydroxylation reactions (including monohydroxylation and dihydroxylation) accounted for 57.1% of the total number of potential metabolites, indicating that hydroxylation may be the major metabolic pathway of Etizolam. Based on the response values of each metabolite, monohydroxylation (M1), desaturation (M19), and hydration (M16) were recommended as potential biomarkers for Etizolam metabolism. The experimental results provide reference and guidance for forensic personnel in identifying Etizolam use in suspects.

Clinical application value of echocardiography in evaluating left ventricular diastolic function in patients with acute pulmonary embolism.

OBJECTIVE: To explore the clinical value of echocardiography in the assessment of left ventricular diastolic function in patients with acute pulmonary embolism (APE). METHODS: APE patients in our hospital from June 2019 to June 2021 were selected as the observation group. They were divided into low-risk group (19 cases), medium-risk group (16 cases) and high-risk group (15 cases). The non-APE people in our hospital during the same period were selected as the control group. All subjects underwent echocardiography to observe the performance of APE patients under echocardiography. The peak velocity ratio S-wave/D-wave (S/D), early diastolic annular velocity/advanced diastolic annular velocity (Ea/Aa), early filling/early diastolic annular velocity (E/Ea), and early filling/early diastolic annular velocity (E/Ea) were compared with Ar and Vp, respectively; receiver operator characteristic (ROC) curve was used to evaluate the value of echocardiography in evaluating left ventricular diastolic function in patients with APE. RESULTS: Echocardiography show different manifestations of APE patients. Compared with the control group, S/D, Ea/Aa, and Vp in the observation group were significantly decreased and E/Ea and Ar in the observation group were significantly increased (p < 0.05). With the increase of risk stratification, S/D, Ea/Aa, and Vp gradually decreased, E/Ea and Ar gradually increased, and the difference was statistically significant (p < 0.05). The sensitivity, specificity, accuracy, positive predictive value, negative predictive value, area under curve (AUC), and 95% CI were 89.52%, 65.57%, 72.86%, 80.82%, 75.38%, 0.730, and 0.629-0.831, respectively. CONCLUSION: Echocardiography can effectively evaluate left ventricular diastolic function in patients with APE, and there are significant differences in left ventricular diastolic function in different risk stratification, which has important reference value for clinical diagnosis and treatment of APE.

Metabolic profiles and screening tactics for MDMB-4en-PINACA in human urine and serum samples.

MDMB-4en-PINACA (Methyl 3,3-dimethyl-2-[1-(pent-4-en-1-yl)-1H-indazole-3-carboxamido] butanoate) is a potent agonist of the CB1 receptor. In 2021, it was one of the most common synthetic cannabinoid receptor agonists (SCRAs) seized by the Beijing Drug Control Agency. MDMB-4en-PINACA can be hard to detect in biological specimens because of ester hydrolysis. In this work, a highly sensitive liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method was developed for the detection of MDMB-4en-PINACA metabolites in urine, serum, and hair samples. Metabolites from authentic samples were compared with those from human liver microsomes (HLMs) in vitro and in zebrafish in vivo. A total of 75 metabolites, including 44 previously unreported metabolites, were identified from urine samples. We found that 11 metabolic pathways were involved in MDMB-4en-PINACA metabolism, including acetylation, a novel metabolic pathway for SCRAs. Our results revealed that ester hydrolysis and hydroxylation were to the major metabolic pathways involved in MDMB-4en-PINACA metabolism. Using serum samples, we detected 9 metabolites along with the parent drug. Only the parent drug was detected using hair samples. The existence of ADB-4en-PINACA makes the currently used biomarkers for MDMB-4enPINACA not very specific for the intake of MDMB-4en-PINACA. Therefore, based on the identified metabolites and their structural features, we propose more sensitive screening tactics for MDMB-4en-PINACA using urine and serum samples.

Clinical Application of Echocardiography in Evaluating Left Ventricular Diastolic Function in Patients with Acute Pulmonary Embolism.

Objective: To explore the clinical application of echocardiography in assessing left ventricular diastolic function in patients with acute pulmonary embolism (APE). Methods: About 90 patients with APE treated in our hospital from March 2016 to March 2019 were enrolled as the study group. The enrolled patients were assigned into three groups in accordance with the risk stratification of APE, with 30 patients in each group, including high-risk group, medium-risk group, and low-risk group. Approximately 90 healthy persons who underwent physical examination in the same period were enrolled as the control group. The two groups were used to analyze the clinical value of echocardiography in evaluating left ventricular diastolic function in patients with APE and to explore the correlation between left ventricular diastolic function and different risk stratification of APE. Results: First of all, we compared the side values of echocardiography. Compared to the control group, right ventricular end-diastolic diameter/left ventricular end-diastolic diameter (RVED/LVED) and pulmonary artery systolic pressure (PASP) in the study group were higher, while left ventricular end-diastolic volume (LVEDV), left ventricular end-systolic volume (LVESV), left ventricular volume per stroke (LVSV), and left ventricular ejection fraction (LVEF) in the study group were lower (P < 0.05). Secondly, we compared the echocardiographic side values of patients with different APE risk stratifications: (1) RVED/LVED, PASP: low-risk group < medium-risk group < high-risk group; (2) LVEDV, LVESV, LVSV, LVEF: low-risk group > medium-risk group > high-risk group (P < 0.05). The routine echocardiographic findings of APE patients: M-type left ventricular wave group of pulmonary embolism (PE) indicated right ventricular enlargement and interventricular septum shifted to the left ventricle; short-axis section of PE artery indicated thrombosis at the opening of the right pulmonary artery (indicated by arrow); long-axis section of the left ventricle of PE indicated right ventricular enlargement (RV 3.5 cm, LV 3.9 cm); four-chamber view of PE indicated enlargement of the right atrium and the right ventricle, and interventricular septum shifted like left ventricle. In terms of mitral annulus displacement, early diastolic ESD and ELD in the study group were lower compared to those in the normal control group, while late diastolic ASD and ALD in the study group were higher compared to those in the normal control group (P < 0.05). The results of Pearson correlation analysis indicated that there was a positive correlation between RVED/LVED, PASP, and APE risk stratification, while APE risk stratification was negatively correlated with LVEDV, LVESV, LVSV, and LVEF (P < 0.05). Conclusion: Echocardiography attaches importance to the evaluation of left ventricular diastolic function in APE. The side value of echocardiography can sensitively reflect the changes of left ventricular diastolic function in APE. The predictive value in the diagnosis of APE is better than other traditional parameters of left ventricular diastolic function, especially in patients with different APE risk stratifications. There is a remarkable correlation between the changes of left ventricular diastolic function and different APE risk stratifications. Echocardiographic measurement can be employed to assess left ventricular diastolic function in APE, which can be developed in clinical treatment and prognosis evaluation to provide a new evaluation method.

A retrospective analysis of data from illicit drug abuse cases in Beijing between 2018 and 2020.

The overall number of drug abuse cases has been on the rise around the world, causing it a disaster in many countries. In this retrospective study, we analyzed the characteristics of 11,903 drug abuse cases in Beijing, the capital of China, in the perspective of age, gender, nationality, region, season, type of specimens and various drugs, respectively. The case information was collected by the national-level forensic toxicology laboratory, which belongs to the China University of Political Science and Law. It was shown that the overall number of drug abuse cases had increased sharply from 2018 to 2019 and had a decrease of 25% in 2020. The incidence of drug abuse cases involving men was much higher than that of the cases involving women. The adolescents and young adults accounted for the largest share of drug abusers in our study. Haidian, as a national center of scientific and technological innovation with global influence, had the largest percentage of toxic substance-related cases, and the change was more pronounced than other districts during the 3-year-period. Chaoyang was second to Haidian in the number toxic substance-related cases. As a key window of economic, Chaoyang had the highest number of foreigner drug cases. June to August had the highest number of drug abuse cases, while amphetamines, opioids, cannabis and cocaine were the most common toxic substances involved in the cases we detected. To our knowledge, this is the first study to provide an overall analysis of drug abuse cases in Beijing, China. The study indicated the presence of a wide number of drug abuse cases in the capital city and confirmed the threat for the public safety and health. Such forensic information can assist the government to devise intelligence-based preventive and repressive measures and policies.

Polymer-based lab-on-a-molecule for the selective and sensitive recognition of fluoride and cyanide.

A polymer chemical sensor based on lab-on-a-molecule was synthesised by amine-aldehyde polymerisation for selective detection of fluorine ions (F-) and cyanide (CN-). The polymer chemical sensor shows significant absorption and fluorescence changes upon the addition of F- and CN-, which can be observed by the naked eye and optical reactions. The polymer has a higher fluorescence enhancement effect than its monomer and the distance of wavelength of each anion increased which could be applied to better distinguish the two anions.

Metabolic Profiles of 5F-MDMB-PICA in Human Urine, Serum and Hair Samples Using LC-Q Exactive HF-MS.

In 2020, 5F-MDMB-PICA (5F-MDMB-2201) was one of the most common synthetic cannabinoids (SCs) identified in drugs seized by the Beijing Drug Control Agency, and it was categorized as Schedule II by the United Nations Office on Drugs and Crime in March 2020. It is difficult to detect 5F-MDMB-PICA in biological matrices due to its fast metabolic rate in vivo. In this work, 5F-MDMB-PICA metabolic profiles were investigated by liquid chromatography--quadrupole exactive high field orbitrap mass spectrometer (LC-Q Exactive HF MS), with accurate mass measurements in human urine, serum and pubic hair. To obtain intact metabolites, solid-phase extraction for urine and serum and direct ultrasonic extraction for pubic hair were applied to clean the samples without enzymatic hydrolysis. The differences in 5F-MDMB-PICA metabolism in the three different matrices were compared for the first time to determine the best detection biomarkers for monitoring 5F-MDMB-PICA misuse. Urine samples were determined to be the preferred biological material for identifying 5F-MDMB-PICA abuse. Forty-seven intact metabolites were detected in human urine, the ester hydrolyzed with glucuronidated metabolite in urine samples can be used as the primary biomarker to identify drug misuse. Fifteen metabolites were found in serum samples. Ester hydrolysis was considered to be the major metabolic pathway, and a large number of metabolites were involved with it. Zero metabolites apart from the parent drug were detected in pubic hair samples. Twenty-eight new metabolites and their metabolic pathways were characterized and tentatively identified by LC-QE-HF-MS, and a new potential biomarker (M5 ester hydrolysis + propionic acid) was also identified.

Brief Report: Predictors of Relapse for Patients With Dextromethorphan Dependence.

BACKGROUND AND OBJECTIVES: The recreational use of dextromethorphan (DXM) triggers dependence. The pattern and clinical predictors of relapse are unclear. METHODS: Here, we retrospectively analyzed the clinical information of 28 patients with DXM dependence (20 males and 8 females). RESULTS: The mean age at admission was 25.89 years (standard deviation [SD] = 7.84), and the average duration of DXM abuse was 24.96 months (SD = 17.40). The relapse rates at 1 month, 3 months, 6 months, and 1 year were 7.14%, 25%, 71.43%, and 89.29%, respectively. Depression and anxiety status at baseline were positive predictors of relapse (r = -.539, P = .006, R2 = 0.290; r = -.449, P = .024, R2 = 0.202, respectively). DISCUSSION AND CONCLUSIONS: Our results suggest that patients with DXM dependence are at high risk of relapse and that measuring affective disturbance is important for predicting the outcomes of their treatment. SCIENTIFIC SIGNIFICANCE: This study first identified a high relapse rate among patients with DXM dependence and found that the baseline depression and anxiety status was correlated with the time length of relapse. These findings not only warn us of the vulnerability of DXM-dependent relapse but also reveal the importance of measuring affective disturbance in relapse prevention. (Am J Addict 2020;00:00-00).

High-sensitivity tunneling magneto-resistive micro-gyroscope with immunity to external magnetic interference.

Micro-electro-mechanical system (MEMS) gyroscopes have numerous potential applications including guidance, robotics, tactical-grade navigation, and automotive applications fields. The methods with ability of the weak Coriolis force detection are critical for MEMS gyroscopes. In this paper, we presented a design of MEMS gyroscope based on the tunneling magneto-resistance effect with higher detection sensitivity. Of all these designed parameters, the structural, magnetic field, and magneto-resistance sensitivity values reach to 21.6 nm/°/s, 0.0023 Oe/nm, and 29.5 mV/Oe, thus, with total sensitivity of 1.47 mV/°/s. Multi-bridge circuit method is employed to suppress external magnetic interference and avoid the integration error of the TMR devices effectively. The proposed tunneling magneto-resistive micro-gyroscope shows a possibility to make an inertial grade MEMS gyroscope in the future.

Characterization and tentative identification of new flunitrazepam metabolites in authentic human urine specimens using liquid chromatography-Q exactive-HF hybrid quadrupole-Orbitrap-mass spectrometry (LC-QE-HF-MS).

Flunitrazepam (FNZ) is a potent hypnotic, sedative, and amnestic drug used to treat severe insomnia. In our recent study, FNZ metabolic profiles were investigated carefully. Six authentic human urine samples were purified using solid phase extraction (SPE) without enzymatic hydrolysis, and urine extracts were then analyzed by liquid chromatography-Q exactive-HF hybrid quadrupole-Orbitrap-mass spectrometry (LC-QE-HF-MS), using the full scan positive ion mode and targeted MS/MS (ddms2) technique to make accurate mass measurements. There were 25 metabolites, including 13 phase I and 12 phase II metabolites, which were detected and tentatively identified by LC-QE-HF-MS. In addition, nine previously unreported phase II glucuronide conjugates and four phase I metabolites are reported here for the first time. Eight metabolic pathways, including N-reduction and O-reduction, N-glucuronidation, O-glucuronidation, mono-hydroxylation and di-hydroxylation, demethylation, acetylation, and combinations, were implicated in this work, and 2-O-reduction together with dihydroxylation were two novel metabolic pathways for FNZ that were identified tentatively. Although 7-amino FNZ is widely considered to be the primary metabolite, a previously unreported metabolites (M12) can also serve as a potential biomarker for FNZ misuse.

Analysis of AMB-FUBINACA Biotransformation Pathways in Human Liver Microsome and Zebrafish Systems by Liquid Chromatography-High Resolution Mass Spectrometry.

In this study, the metabolic profiles of a new illicit drug AMB-FUBINACA were investigated using both human liver microsome and zebrafish models. Liquid chromatography Q Extractive HF Hybrid Quadrupole-Orbitrap mass spectrometry (LC-QE-HF-MS) was employed to analyze the metabolic sites and pathways. AMB-FUBINACA was added to the in vitro liver microsome incubation model to simulate the metabolic processes in human body. The results showed that a total of 17 metabolites were generated in the human liver microsome model; the main metabolic pathways of the phase I metabolism included ester hydrolysis, methylation, ester hydrolysis combined with decarboxylation, hydroxylation, ester hydrolysis combined with indazole ring hydroxylation, etc. while glucuronidation served as the main metabolic pathway of the phase II metabolism. The zebrafish system produced a similar result with 16 of the same 17 metabolites identified. The phase I metabolites M3.1 (ester hydrolysis), M1.2 (alkyl chain hydrolysis) and the phase II metabolite M3.2 (M3.1 glucuronide) were recommended to be the potential poisoning markers.