Topic on High Resolution Mass Spectrometry Technology YAN Hui
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 GOLDTM 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.
Topic on High Resolution Mass Spectrometry Technology
