Anabolic androgenic steroids （AASs） are a class of synthetic steroid hormones that mimic androgens, and they rank as the most widely abused doping agents worldwide. High resolution mass spectrometry （HRMS） has unique advantages in the detection of AASs due to its high resolution, high sensitivity, high selectivity and data traceability. HRMS can not only be used for the qualitative and quantitative analysis of AASs and their metabolites in different biological samples, effectively improving the ability to analyze complex samples and increasing the reliability of analytical results, but can also infer AASs metabolites and reveal metabolic pathways by combining in vitro and in vivo metabolic models. This paper reviews the research progress of HRMS in AASs analysis methods, in vitro and in vivo metabolism of AASs, and also explores its application prospects in the field of forensic science.

Mass spectrometry imaging （MSI） is an emerging high-tech “photography” method combining mass spectrometry and image visualization technologies, which can simultaneously monitor the spatial distribution of multiple molecules in biological samples. In recent years, the characteristics of this technology have been used to visually study the distribution of drugs in hair, among which matrix-assisted laser desorption ionization-mass spectrometry imaging（MALDI-MSI） technology has been most widely applied in hair analysis. Based on the principle of MALDI-MSI technology, this paper discusses the key technical elements of MALDI-MSI technology applied to hair analysis and reviews the research achievements of MSI in hair analysis. It concludes that improving the ionization efficiency, resolution, sensitivity and stability of MALDI-MSI technology remains the future exploration direction, and simplifying data analysis and establishing databases will help promote MALDI-MSI as a routine analytical technique for hair analysis.

Objective To establish a method to identify an unknown substance based on the combined use of gas chromatography-quadrupole time-of-flight mass spectrometry （GC-QTOF-MS）, ultra-high performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry （UPLC-Q/Orbitrap HRMS）and nuclear magnetic resonance （NMR） techniques. Methods The unknown substance was dissolved in methanol and was detected by GC-QTOF-MS and UPLC-Q/Orbitrap HRMS, and was dissolved in methanol-d4 to be detected by NMR. Results The main characteristics ion peaks of components with retention time of 9.67 min in GC-QTOF-MS measured were 84.080 8, 110.999 7, 128.107 0 （base peak）, 138.994 7, etc. The protonated molecular ion peak m/z in UPLC-Q/Orbitrap HRMS was 268.109 3. It was inferred that the unknown substance was an analog of the synthetic cathinone substance 2-methyl-1-[4-（methylthio）phenyl]-2-morpholinopropan-1-one （MTMP） by comparing the mass spectrum information and molecular structure of MTMP. NMR analysis confirmed it as a novel N-morpholine substituted synthetic cathinone substance 1-（4-chlorophenyl）-2-methyl-2-morpholinopropan-1-one （CMMP）. Conclusion The method established in this study can be used for structural confirmation of CMMP.

Objective To explore the characteristic fragment ions, ion abundance ratios and mass spectrometry fragmentation rules of etomidate and its structural analogues by using gas chromatography-quadrupole/orbitrap mass spectrometry （GC-Q/Orbitrap MS） and liquid chromatography-linear ion trap quadrupole-orbitrap mass spectrometry （LC-LTQ-Orbitrap MS） techniques, providing important data support for the identification and prediction of etomidate structural analogues. Methods GC-Q/Orbitrap MS and LC-LTQ-Orbitrap MS were used to analyze metomidate, etomidate, isopropoxate and propoxate, to obtain their GC high resolution mass spectra and LC high resolution mass spectra. Results Under the electron impact （EI） ion source mode, etomidate, metomidate and the other two analogues all produced their molecular ions and seven identical fragment ions （m/z 77.038 6, 79.054 2, 95.024 0, 105.069 9, 143.073 0, 172.099 5 and 199.086 6）, among which isopropoxate and propoxate also produced characteristic fragment ions m/z 216.089 3. In the collision cell of the electrospray ionization （ESI） source mode, etomidate, metomidate and the other two analogues all produced three identical fragment ions （m/z 95.024 0, 105.069 9 and 113.034 6）. Meanwhile, each substance produced one characteristic fragment ion （metomidate: m/z 127.050 2; etomidate: m/z 141.065 9; isopropoxate and propoxate: m/z 155.081 5）. Conclusion Common fragment ions exist among etomidate and its structural analogues, and characteristic ion fragments are generated based on the different carbon numbers of their side chains. The structure of side chains can affect the abundance ratio of each fragment ion, providing a basis for the structural identification and prediction of such compounds.

Objective To establish a rapid screening method for 60 types of natural toxins in whole blood by ultra-high performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry （UPLC-Q/Orbitrap HRMS). Methods The chromatographic and mass spectrometric information of 60 standard samples of natural toxins were analyzed and recorded, and a screening database was built. Whole blood was pretreated by protein precipitation method combined with ultrasonic-assisted dispersion, and then a Hypersil GOLD^TM C₁₈ column was used with 5 mmol/L ammonium formate aqueous solution （containing 0.1% formic acid） and acetonitrile as mobile phase for gradient elution. In the positive ion mode, the data were collected in full scan/data-dependent secondary scan （Full MS/dd-MS²） mode. Based on the established screening library, the rapid screening of 60 types of natural toxins in whole blood was realized by TraceFinder software. Results A UPLC-Q/Orbitrap HRMS method was developed for the screening of 60 types of natural toxins in whole blood. Except for the limit of detection （LOD） of oxymatrine （20 ng/mL） and strophanthidin （40 ng/mL）, the LOD for the other 58 natural toxins was in the range of 0.05-5 ng/mL. Conclusion This method has a simple and efficient pretreatment process and can achieve rapid screening of 60 types of natural toxins in whole blood.

Objective To establish and optimize an in vitro incubation system with human liver microsomes and investigate the in vitro metabolites and possible metabolic pathways of mirtazapine. Methods Three major metabolites of mirtazapine were selected to optimize the incubation conditions of liver microsomes. The metabolites of mirtazapine were analyzed by liquid chromatography-high resolution mass spectrometry （LC-HRMS） to identify the in vitro metabolites and metabolic pathways of mirtazapine. Results Ten metabolites, including nine phase Ⅰ metabolites and one phase Ⅱ metabolite, were identified in the in vitro liver microsome incubation. Among them, five new metabolites and one new metabolic pathway were discovered. The pathways involved in phase Ⅰ metabolic included methylation, hydroxylation, oxidation, reduction, etc., while the phase Ⅱ biotransformation was mainly glucuronidation. Conclusion The metabolites discovered in this study are consistent with the main metabolites of mirtazapine reported in literature, which are N-desmethylmetazapine, 8-hydroxy mirtazapine and mirtazapine-N-oxide. The results can provide basis for the confirmation of mirtazapine cases and provide reference for the study of other substances.

Objective To analyze the independent influencing factors of intimate partner homicide （IPH） cases, construct an IPH prediction model, and provide a basis for criminal profiling. Methods A total of 476 convicted homicide cases in Guangdong Province from January 1, 2014, to December 31, 2020, were collected as modeling dataset. They were divided into the IPH group （n=180） and the non-intimate partner homicide （N-IPH） group （n=296） based on whether the offender and victim were intimate partners. Logistic regression was used to build the model, the model was evaluated through the receiver operating characteristic （ROC） curve analysis and a nomogram was drawn. Internal validation was conducted using ten-fold cross-validation method. A total of 126 court judgments from outside Guangdong Province from January 1, 2011, to December 31, 2020, were randomly collected for external validation. Results Through multi-factor Logistic regression analysis, 7 variables were ultimately selected for inclusion in the model. The Hosmer-Lemeshow goodness of fit test result of the model was χ²=13.158, P=0.068. The ROC area under the curve （AUC） of the model was 0.939 （95% CI: 0.919-0.959）, the cut-off value was 0.292, the sensitivity was 0.900, and the specificity was 0.865. The calibration curve was close to the ideal curve. The ten-fold cross-validation showed the accuracy of 0.863 and a Kappa value of 0.708. The external validation results showed an AUC of 0.922 （95% CI: 0.872-0.971）, a cut-off value of 0.292, a sensitivity of 0.890, and a specificity of 0.886. The calibration curve tended to the ideal curve. Conclusion The IPH prediction model based on forensic field indicators has good predictive ability, reliable accuracy and stability, and can provide a scientific method for criminal profiling.

Objective To investigate the value of wrist MRI in bone age estimation for male adolescents in Shanghai, Zhejiang and Jiangsu. Methods A total of 124 Han male adolescents aged 6.0 to 18.0 years from Shanghai, Zhejiang and Jiangsu were selected as subjects. Their weight and height were measured, and T1WI and T2WI sequences of the wrist were scanned. The distal ends of the radius and ulna, and the first to five metacarpal epiphyses and corresponding metaphyses were selected as observational indexes after MRI images of the wrist were obtained. The development of each index was classified （0-2 grades） by a deputy senior imaging expert, then the maximum width of each index was measured by another deputy senior expert. Height, weight, classification and maximum width of indexes were used as input variables, and age was used as the target variable. Support vector machine, random forest, current reality tree, and linear regression models were established to estimate the bone age, and the model with the highest accuracy was selected. Results The height, weight, classification of wrist bone epiphysis development, maximum width of each bone metaphysis and epiphysis were all correlated with age （P<0.05）. The accuracies of the support vector machine were the highest when the differences between bone age and actual chronological age were within 1.0 and 1.5 years （88.7% and 96.0%, respectively）. Conclusion It is feasible to estimate bone age by using MRI images. Quantifying the maximum width of the epiphysis and corresponding metaphysis of bone and combining it with MRI image classification can effectively reduce the estimation error.

Nitazenes are a class of synthetic opioids that have recently emerged in the illegal drug market. They are characterized by high potency and a narrow therapeutic window. Compared with traditional opioids, these substances are more likely to cause severe respiratory depression. In recent years, the abuse of nitazenes has occurred frequently, causing hundreds of overdose deaths worldwide, and posing a serious threat to individual health and public safety. As more nitazenes continue to emerge, research on the identification of these substances has received increasing attention. This paper reviews the structural characteristics, in vivo and in vitro pharmacological characterizations and analysis methods of common nitazenes, aiming to provide a basis and reference for the study of these substances in forensic toxicology.