bims-metlip Biomed News
on Methods and protocols in metabolomics and lipidomics
Issue of 2021‒07‒04
forty-three papers selected by
Sofia Costa
Cold Spring Harbor Laboratory

  1. Se Pu. 2020 Nov 08. 38(11): 1294-1301
      Paraquat (PQ) and diquat (DQ) are widely used as non-selective contact herbicides. Several cases involving accidents, suicide, and homicide by PQ or DQ poisoning have been reported. Poising by PQ, which is mainly concentrated in the lungs, causes acute respiratory distress syndrome and leads to multiple organ toxicity. The toxic effects of DQ are similar to those of PQ but relatively less intense. The mortality rates in PQ and DQ poisoning are high. Simultaneous monitoring of the PQ and DQ concentrations in plasma and urine can provide valuable information for early clinical diagnosis and prognosis. High performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) is the main analytical method used to detect PQ and DQ in plasma and urine. As both these compounds are highly polar and water soluble, they cannot be retained effectively on a reversed-phase column with conventional mobile phases. The separation of PQ and DQ by ion-pair chromatography or hydrophilic chromatography has been reported. The use of an ion-pairing reagent helps in improving the retention capabilities of PQ and DQ. However, the sensitivity of MS detection is noticeably decreased because of ion suppression caused by the ion-pairing reagent in the mobile phase; furthermore, ion-pairing reagents may contaminate the MS system. The separation of PQ and DQ by hydrophilic chromatography is easily affected by matrix components in the sample, and their retention times are not stable. Considering PQ and DQ are bicharged cation species in solution, they are more suitable for separation by cation-exchange chromatography. A method based on ion chromatography-triple quadrupole mass spectrometry was established for the determination of PQ and DQ in plasma and urine. The plasma and urine samples were diluted with water, and then purified on a solid-phase extraction column containing a polymer-reversed phase and weak ion-exchange mixed-mode adsorbent (Oasis WCX). PQ and DQ were separated on an IonPac CS 18 analytical column (250 mm×2.0 mm, 6.0 μm) with gradient elution using a methylsulfonic acid solution electrolytically generated from an on-line eluent generation cartridge. An in-line suppressor was used to remove methylsulfonate and other anions from the eluent before the eluent entered the mass spectrometer. Between the suppressor and the ion source in MS, the addition of 3% (v/v) formic acid in acetonitrile as an organic modifier (using an auxiliary pump and a T-piece) aided desolvation in the ion source, resulted in a one-or two-fold improvement of the response, and eliminated the residual effects of the adsorption of PQ and DQ caused by ion source. The analytes were detected by triple quadrupole tandem mass spectrometry using positive electrospray ionization in the multiple reaction monitoring (MRM) mode. PQ-d8 and DQ-d4 were used as internal standards. The calibration curves for PQ and DQ showed good linear relationships in the ranges of 1.0-150 μg/L and 0.5-75 μg/L, respectively, and the correlation coefficients were > 0.999. The average matrix effects of PQ and DQ in plasma were 84.2%-89.3% and 84.7%-91.1%, while the average matrix effects of PQ and DQ in urine were 50.3%-58.4% and 51.9%-59.4%. The average recoveries of PQ and DQ in plasma were 93.5%-117% and 91.7%-112%, respectively, with relative standard deviations (RSDs) of 3.4-16.7% and 2.8%-13.2%, and that in urine were 90.0%-118% and 99.2%-116%, with relative standard deviations of 5.6%-14.9% and 2.4%-17.3% (n=6). The limits of detection of PQ and DQ in plasma and urine were 0.3 μg/L and 0.2 μg/L, respectively, with the corresponding limits of quantification being 1.0 μg/L and 0.5 μg/L. This method is sensitive and accurate, and it can be used to determine PQ and DQ for clinical diagnosis and prognosis in patients.
    Keywords:  diquat; ion chromatography-triple quadrupole mass spectrometry; paraquat; plasma; urine
  2. Se Pu. 2021 Aug;39(8): 845-854
      Human exposure to chemical pollutants in the environment can cause a variety of diseases, including cancer, diabetes, cardiovascular disease, and neurodegenerative diseases (Alzheimer's disease, Parkinson's disease, etc.). Exogenous and environmental pollutant exposure-induced endogenous aldehydes are highly reactive electrophilic compounds that can form covalently modified products with a variety of important biological molecules in the human body, thus inducing toxic effects. Exposome research has become a hotspot since it was first proposed in 2005. Exposure studies can map the complex relationships between biomarkers and disease risk. Therefore, the measurable and characteristic changes of all biomarkers together constitute a key basis for exposome research. Aldehydes are among the main components of chemical exposure. Because of the physical and chemical properties of aldehydes and the existence of multiple matrix interferences in the samples, it is particularly difficult to analyze and characterize them. The analysis and detection methods for aldehydes mainly include sensing analysis, electrochemical methods, fluorescence imaging, chromatography, mass spectrometry (MS), and chromatography-MS. Analytical techniques based on gas chromatography-MS (GC-MS) and liquid chromatography-MS (LC-MS) have emerged as the main methods for chemical exposome research. Chemical derivatization, especially stable isotope labeling derivatization (also known as chemical isotope labeling) combined with LC-MS analytical techniques, can help circumvent the problems encountered in targeted and non-targeted metabolome and exposome analysis. The combination of chemical derivatization with chromatography-MS is one of the most important solutions for the accurate analysis of aldehydes in complex samples. Over the past five years, the development and application of chromatography-MS analytical methods based on chemical derivatization have become key topics in aldehyde analysis. This paper summarizes and reviews the latest progress in GC-MS and LC-MS methods based on chemical derivatization (2015-2020). The review focuses on analytical method development for aldehyde exposure biomarkers in bio-matrices (blood, urine, saliva, biological tissue, etc.). Various derivatization reagents for labeling small-molecule aldehydes, qualitative/quantitative analytical methods and their application value, advantages/disadvantages of different analytical methods for aldehyde exposure biomarkers, and future development trends are also included. The manuscript contents may aid the integrated development of exposome, metabolomics, and lipidomics, as well as research on the environment, ecology, and health. To clarify the complex actions of exogenous and endogenous aldehydes in physiological and pathological events, it is necessary to improve the analysis and characterization techniques and tools for studying the "aldehydome." With the development and application of sophisticated mass spectrometers, advances in high-performance chromatographic separation and bioinformatics, and advent of single-cell analysis and MS imaging, future aldehyde exposome analytical methods will have higher sensitivity and throughput. This in turn would be more useful for screening and identifying unknown aldehyde compounds and discovering new exposome biomarkers.
    Keywords:  aldehydes; biomarkers; chromatography-mass spectrometry; derivatization; isotope labeling; review
  3. Se Pu. 2020 Jun 08. 38(6): 715-721
      A sensitive and accurate method was developed to quantify eight polycyclic aromatic hydrocarbon (PAH) metabolites in human urine by liquid-liquid extraction-high resolution gas chromatography-high resolution dual-focus magnetic mass spectrometry (LLC-HRGC-HRMS). About 2 mL urine was mixed with a deuterium- or 13C-labeled isotopic internal standard, and the conjugated targets were enzymatically digested in the presence of ascorbic acid. The free compounds were extracted with toluene-pentane (1:4, v/v) and condensed to near dryness. Then, the target compounds were redissolved in toluene. After derivatization, they were separated and quantified by HRGC-HRMS. The linear ranges of the 1-hydroxynaphthalene, 1-hydroxyphenanthrene, and 2-hydroxyphenanthrene were 0.14-41.6 μg/L, 0.05-8.33 μg/L, and 0.04-8.33 μg/L, respectively, and those for the other five PAH metabolites were 0.02-8.33 μg/L. The correlation coefficients were greater than 0.99. The limits of detection were in the range of 0.006-0.042 μg/L, and the recoveries were 81.4%-127.0%. The intra-day and inter-day relative standard deviations (RSDs) were less than 6.9% and 10.9%, respectively. This method was utilized for the determination of 330 human urine samples. The results showed that 3-hydroxychrysene and 6-hydroxychrysene were not detected, and the detection rate of the other six PAH metabolites was 100%. This method is sensitive, accurate and stable, and it is suitable for the determination of the eight PAH metabolites in human urine.
    Keywords:  biological monitoring; high resolution gas chromatography-high resolution dual-focus magnetic mass spectrometry (HRGC-HRMS); hydroxyl polycyclic aromatic metabolites; liquid-liquid extraction (LLE); urine
  4. Talanta. 2021 Oct 01. pii: S0039-9140(21)00432-X. [Epub ahead of print]233 122511
      Liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) has become the most versatile analytical tool for profiling small-molecule compounds and increasingly been applied in many fields. Nevertheless, LC-MS based quantification still face some challenges, such as signal drift in LC-MS, which may affect the validity of the obtained data and lead to misinterpretation of biological results. Here, we established a calibration method known as "RIM" to compensate the signal drift of LC-MS. To this end, a mixture of d4-2-dimethylaminoethylamine (d4-DMED)-coded normal fatty acids (C5-C23) was used as calibrants to construct RIM calibration. With the addition of calibrants, not only the MS signal drift, but also the mass accuracy and LC retention time can be calibrated, thereby improving the reliability of quantitative data. The effectiveness of RIM was carefully validated using a human serum extract spiked with 34 standards and then RIM was applied for rat brain untargeted metabolome research. In addition, to expand the functionality and flexibility of RIM for data handling, we generated a MATLAB-based RIM program, which implements the above concepts and allows automatic data process. Taken together, the proposed RIM method has potential application in large-scale quantitative study of complex samples.
    Keywords:  Drift correction; LC-MS; Quantitative accuracy; Signal drift
  5. Se Pu. 2020 May 08. 38(5): 529-537
      Flavonoids are polyphenol compounds that are widely distributed in honey; they play an important role in its antioxidant activity and aid in distinguishing various species of honey. An analytical method based on solid phase extraction and high performance liquid chromatography with tandem mass spectrometry (SPE-HPLC-MS/MS) for the simultaneous determination of rutin, myricetin, morin, quercetin, naringenin, hesperetin, luteolin, genistein, kaempferol, isorhamnetin, apigenin, pinocembrin, wogonin, chrysin, galangin, genkwanin, and ferulic acid in honey was developed and validated. The honey samples were diluted with HCl solution (pH 2) and cleaned up on a C18 column by solid phase extraction; the methanol eluent was filtered through a nylon membrane and analyzed by LC-MS/MS on an ODS-3 column (150 mm×4.6 mm, 3 μm). Quantitative detection was performed by LC-MS/MS in the multiple reaction monitoring (MRM) mode under negative electrospray ionization (ESI-) conditions. The external standard method was used for quantitation. The method validation data were obtained. The calibration standard curve solutions were prepared by honey matrix-matching; the linear range was 0-200 μg/kg, with correlation coefficients of the calibration standard curves greater than 0.997. The limits of quantification (LOQs) of this method were 20 μg/kg. The recoveries were in the range of 64.5%-113%, and the relative standard deviations were 1.4%-14.5% at three spiked levels (20, 40, and 100 μg/kg). The developed method is convenient, rapid, and effective, and it is suitable for the determination and confirmation of flavonoids in honey samples.
    Keywords:  flavonoids; high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS); honey; solid phase extraction (SPE)
  6. Se Pu. 2020 May 08. 38(5): 538-546
      A method was established for the rapid determination of 28 prohibited and restricted veterinary drugs of five classes (sulfonamides, quinolones, chloramphenicols, nitrofurans metabolites and triphenylmethane) in fish and shrimp by ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) coupled with simultaneous extraction and purification. After optimizing the sample pretreatment methods and chromatographic conditions, the samples were hydrolyzed with hydrochloric acid. The five kinds of residual drugs were simultaneously extracted with acetonitrile after nitrofuran metabolites derivatized with 2-nitrobenzaldehyde. The extraction solutions were dehydrated with sodium chloride and salted out. The acetonitrile layer was purified with n-hexane and concentrated to near dryness. The residues were dissolved in 1.0 mL methanol. Then, the solutions were purified by dispersive solid phase extraction with C18, N-(n-propyl)ethylenediamine (PSA) and amino bonded silica gel (NH2) sorbents. The residues were separated on a ZORBAX C18 chromatographic column (50 mm×2.1 mm, 1.8 μm). The MS data were acquired in positive and negative multiple reaction monitoring (MRM) modes, and quantitated by the isotope internal standard method. With this approach, the linear relationships of the 28 veterinary drugs were good. The linear correlation coefficients were greater than 0.99. The limits of detection and limits of quantification were in the range of 0.1-0.8 μg/kg and 0.3-2.7 μg/kg, respectively. The average recoveries at three spiked levels (1, 5, and 20 fold of the LOQs) were 70.0%-120% with relative standard deviations of 0.9%-10.0%. The method is simple, rapid, accurate and sensitive, and it is suitable for the rapid determination of various prohibited and restricted drugs in fish and shrimp.
    Keywords:  dispersive solid phase extraction; fish; prohibited and restricted veterinary drugs; shrimp; simultaneous extraction and purification; ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS)
  7. Se Pu. 2020 Dec 08. 38(12): 1456-1464
      A method combining 96-well plate solid-phase extraction with ultra-performance liquid chromatography-tandem mass spectrometry (96-well SPE LC-MS/MS) was developed for the simultaneous determination of eight environmental phenols in urine samples. The samples included seven bisphenol compounds and triclosan. The urine samples were thawed to room temperature, and the target analytes were deconjugated by β -glucuronidase/aryl-sulfatase in ammonium acetate buffer solution at 37℃ overnight. Then, the effects of three kinds of 96-well solid-phase extraction plates and different elution conditions on the purification of the urine samples and the environmental phenol recoveries were compared. The best purification effect was achieved on Oasis HLB 96-well plate (60 mg) solid phase extraction, using 30% (v/v) acetonitrile aqueous solution as the rinse solution. The target analytes were then eluted by methanol solution and evaporated to dryness using a nitrogen blower. After reconstruction with 0.5 mL methanol/water (1:1, v/v) solution, the target compounds were detected by UPLC-MS/MS. To achieve better chromatographic separation, two kinds of analytical columns (C18 and T3) and different types of mobile phases (methanol and acetonitrile as the organic phase) were also compared. The best chromatographic effect was achieved when the treated samples were separated on a C18 column (100 mm×2.1 mm, 1.7 μm) using acetonitrile/water as the mobile phase at a flow rate of 0.3 mL/min. Mass spectra were recorded by negative electrospray ionization under the multiple reaction monitoring (MRM) mode. The sample matrix effect was also evaluated. The absolute matrix effects of bisphenol A, bisphenol F, bisphenol S, bisphenol B, and bisphenol AF were in the range of 3.47% to 15.32%. Since the above mentioned matrix effect was weak, there was no need for compensation measures. On the contrary, tetrachlorobisphenol A, tetrabromobisphenol A, and triclosan showed an absolute matrix effect of 49.58% (moderate), 71.99% (strong), and 86.93% (strong), thus necessitating compensation measures. Therefore, this strategy uses a one-to-one corresponding isotope internal standard method to offset the matrix effect. Six different urine samples were used to evaluate the relative matrix effect. The relative standard deviations (RSDs) of the eight corresponding internal standard peak areas were 3.63%-9.06%, indicating that the relative matrix effect was stable. Under the optimized conditions, linearity ranges were 0.50-50 μg/L for bisphenol A and bisphenol AF; 0.05-50 μg/L for tetrachlorobisphenol A and bisphenol S; 0.01-50 μg/L for bisphenol F and tetrabromobisphenol A; 1.00-50 μg/L for bisphenol B; and 5.00-200 μg/L for triclosan. The correlation coefficients were all greater than 0.9995. At spiked levels of 2.5, 5, and 25 μg/L, the average recovery ratios of the eight target analytes were 81.01%-118.84%, while the intra-day and inter-day precisions were 0.38%-19.41% and 2.54%-17.83%, respectively. The limits of detection (LOD) were 0.002-1.09 μg/L, and the limits of quantitation (LOQ) were 0.007-3.63 μg/L. This method was successfully applied to the determination of the eight environmental phenols in 64 urine samples collected from Beijing area between 2019 and 2020. All the target environmental phenols were detected, except for bisphenol B and bisphenol AF. Bisphenol A and bisphenol S showed the highest detection rates of 100% and 96.9%, respectively. The detection rates of triclosan, tetrabromobisphenol A, tetrachlorobisphenol A, and bisphenol F were 57.8%, 46.9%, 23.4%, and 21.9%, respectively. The medium values of urinary concentration followed the order 1.44 μg/L(triclosan), 0.69 μg/L(bisphenol A), 0.086 μg/L (bisphenol S), 0.0032 μg/L (tetrabromobisphenol A), 0.00050 μg/L (tetrachlorobisphenol A), 0.00 μg/L (bisphenol F, bisphenol B, and bisphenol AF). The aforementioned results imply that the widespread environmental phenolic exposure of Beijing residents is worthy of attention. Compared with traditional solid-phase extraction methods, the method reported in this paper is time-saving, effective, and suitable for the simultaneous analysis of large quantities of samples; moreover, the small sample and organic solvent consumption make this method more environment- and operator-friendly.
    Keywords:  96-well plate; endocrine disrupters; environmental phenols; solid phase extraction (SPE); ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS); urine sample
  8. Bioanalysis. 2021 Jun 30.
      Cortisol is considered a particularly relevant biomarker in the context of stress evaluation. This study aims to review of the available literature on the determination of cortisol in hair using LC-MS/MS. Currently, there is no standardized procedure for the measurement of cortisol concentrations in hair, and different sample preparation, chromatographic separation and mass spectrometric detection conditions were described. Simple methanolic extraction, reversed-phase separation and MRM detection in negative ion mode are the most common employed analytical approaches. Reported assays presented acceptable sensitivity for clinical purposes. The increasing use of mass spectrometry in clinical laboratories may contribute to the establishment of LC-MS/MS as the method of choice for the determination of cortisol concentrations in hair.
    Keywords:  LC-MS/MS; cortisol; hair; liquid chromatography; mass spectrometry
  9. Talanta. 2021 Oct 01. pii: S0039-9140(21)00385-4. [Epub ahead of print]233 122464
      Medium- and long-chain fatty acids (MLFAs) are essential energy sources in cells and possess vital biological functions. Characteristics of MLFAs in biosamples contributes to the understanding of biological process and finding potential biomarkers for relevant diseases. However, there are obstacles of the MLFAs determination due to their poor ionization efficiency in mass spectrometry and structural similarity of the MLFAs. Herein, a derivatization strategy was applied by labeling with d0-N, N-dimethyl-6,7-dihydro-5H-pyrrolo [3,4-d] pyrimidine-2-amine (d0-DHPP) and detecting with ultra-high performance liquid chromatography combined with tandem mass spectrometry (UHPLC-MS/MS) in multiple reaction monitoring (MRM) mode. The parallel isotope labeled internal standards were generated by tagging d6-DHPP to MLFAs. The simple and rapid derivatization procedure and mild reaction conditions greatly reduced the potential of MLFA degradation during the processing procedure. With the methodology, the chromatographic performance was greatly improved, and the mass spectrum response was enhanced up to 1, 600 folds. Finally, the developed derivatization method was applied to serum samples to analyze the alteration of MLFAs induced by 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) exposure in breast cancer nude mice. The semi-quantitative results demonstrated that the BDE-47 exposure significantly influenced the MLFA metabolism in mice.
    Keywords:  BDE-47; Derivatization; MRM; Medium- and long-chain fatty acid
  10. Se Pu. 2019 Dec 08. 37(12): 1314-1320
      An ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the simultaneous quantitative determination of ten cephalosporins in bee products, including honey, royal jelly, and lyophilized royal jelly powder, has been established. The samples were extracted with an acetonitrile-water (80:20, v/v) solution. After purification by solid phase extraction using an Oasis PRIME HLB cartridge, the extract was blown to dryness under a stream of nitrogen gas and then re-dissolved in 1 mL 0.1% (v/v) formic acid solution and methanol (95:5, v/v). The samples were separated on an Acquity UPLC BEH C18 column, using a mixture of 0.1% (v/v) formic acid solution and methanol as the mobile phase under gradient elution. The analysis was carried out using a positive electrospray ion source in the multiple reaction monitoring mode. The matrix-matched external standard method was applied to quantitative analysis. Good linear relationships were obtained for the ten cephalosporins in certain concentration ranges, and the correlation coefficients were more than 0.999. The limits of detection and limits of quantification for the ten cephalosporins were in the ranges 0.15-1.5 μg/kg and 0.5-5 μg/kg, respectively. The recoveries for the analytes in the bee products were in the range of 75.0%to 89.8%, with relative standard deviations of 1.4% to 4.6%. This method is characterized by a short analysis time and is suitable for the determination of cephalosporins in different bee products by virtue of its simplicity and reliability.
    Keywords:  bee products; cephalosporins; solid phase extraction (SPE); ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)
  11. Anal Chem. 2021 Jul 01.
      Stable-isotope labeling experiments are widely used to investigate the topology and functioning of metabolic networks. Label incorporation into metabolites can be quantified using a broad range of mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy methods, but in general, no single approach can completely cover isotopic space, even for small metabolites. The number of quantifiable isotopic species could be increased and the coverage of isotopic space improved by integrating measurements obtained by different methods; however, this approach has remained largely unexplored because no framework able to deal with partial, heterogeneous isotopic measurements has yet been developed. Here, we present a generic computational framework based on symbolic calculus that can integrate any isotopic data set by connecting measurements to the chemical structure of the molecules. As a test case, we apply this framework to isotopic analyses of amino acids, which are ubiquitous to life, central to many biological questions, and can be analyzed by a broad range of MS and NMR methods. We demonstrate how this integrative framework helps to (i) clarify and improve the coverage of isotopic space, (ii) evaluate the complementarity and redundancy of different techniques, (iii) consolidate isotopic data sets, (iv) design experiments, and (v) guide future analytical developments. This framework, which can be applied to any labeled element, isotopic tracer, metabolite, and analytical platform, has been implemented in IsoSolve (available at and, an open-source software that can be readily integrated into data analysis pipelines.
  12. Metabolites. 2021 Jun 24. pii: 416. [Epub ahead of print]11(7):
      Biochemical methylation reactions mediate the transfer of the methyl group regulating vital biochemical reactions implicated in various diseases as well as the methylation of DNA regulating the replication processes occurring in living organisms. As a finite number of methyl carriers are involved in the methyl transfer, their quantification could aid towards the assessment of an organism's methylation potential. An Hydrophilic Interaction Chromatography-Liquid Chromatography Multiple Reaction Monitoring (HILIC-LC-MRM) mass spectrometry (MS) methodology was developed and validated according to Food & Drug Administration (FDA), European Medicines Agency (EMA), and International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) for the simultaneous determination of nine metabolites i.e., B12, folic acid, 5-methyltetrahydrofolate, S-adenosylmethionine, S-adenosylhomocysteine, betaine, phosphocholine, N,N-dimethylglycine, and deoxythymidine monophosphate in human blood plasma. The sample pretreatment was based on a single step Solid-phase extraction (SPE) methodology using C18 cartridges. The methodology was found to accurately quantitate the analytes under investigation according to the corresponding dynamic range proposed in the literature for each analyte. The applicability of the method was assessed using blood donor samples and its applicability demonstrated by the assessment of their basal levels, which were shown to agree with the established basal levels. The methodology can be used for diagnostic purposes as well as for epigenetic screening.
    Keywords:  Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS); methylation; solid phase extraction; validation
  13. Se Pu. 2020 Jun 08. 38(6): 655-662
      An analytical method was established for the determination of 15 lipid regulators in fish meat by QuEChERS combined with ultra performance liquid chromatography-quadrupole/electrostatic field orbitrap mass spectrometry (UPLC-Q/Orbitrap-MS). The samples were purified by optimized QuEChERS methods. The amounts of the adsorbent materials (primary secondary amine (PSA) (20, 60, 100, 140 and 180 mg), C18 (40, 100, 160, 220 and 280 mg) and sodium acetate (0.2, 0.6, 1.0, 1.4 and 1.8 g)) were optimized by the response surface method to obtain the best purification effect. The target compounds were separated on an XBridge-C18 column (100 mm×2.1 mm, 3.5 μm) using acetonitrile-0.1% (v/v) formic acid aqueous solution (containing 1.5 mmol/L ammonium acetate) as the mobile phases by a gradient elution program. Qualitative and quantitative analysis of the target compounds were performed in the full scan and secondary mass spectrometry scan (dd-MS2) modes with positive and negative ionization. The target compounds showed good linear relationships in their respective ranges, with correlation coefficients (R2) greater than 0.99. The limits of detection (LOD, S/N=3) and limits of quantification (LOQ, S/N=10) were in the range of 0.2-1.0 μg/kg and 0.3-3.1 μg/kg, respectively. The average recoveries were 76.4%-116.0% at LOQ, 2-fold LOQ, and 10-fold LOQ levels. The intra-day relative standard deviations (RSDs) were 1.0%-7.9%, and the inter-day RSDs were 1.7%-18.4%. The method is simple, sensitive and accurate, and it is suitable for the determination and quantification of lipid regulators in fish meat.
    Keywords:  QuEChERS; fish meat; lipid regulators; quadrupole/electrostatic field orbitrap mass spectrometry (Q/Orbitrap-MS); ultra performance liquid chromatography (UPLC)
  14. Se Pu. 2020 Feb 08. 38(2): 218-223
      A method for the determination of chlormequat chloride (CCC) residues in animal derived foods by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was developed. The samples were extracted with acetonitrile containing 1% (v/v) acetic acid and defatted with n-hexane, followed by clean-up on a cationic solid phase extraction column. The analytes were separated on a Venusil MP C18(2) column (150 mm×2.1 mm, 3 μm) under a gradient elution program using acetonitrile and 0.1% (v/v) formic acid aqueous solution as the mobile phases. Then, the analytes were detected by tandem mass spectrometry using a positive electrospray ionization (ESI+) source in the multiple reaction monitoring (MRM) mode. Matrix-matched internal standard calibration curves were used for quantitative analysis. The calibration curves showed good linearity in the range of 0.200-500 μg/L for CCC, with correlation coefficients (r2) no less than 0.9993. The limit of quantification (LOQ) of the method was 0.500 μg/kg. The average recoveries of CCC in pork, beef, mutton, chicken, egg, pig kidney, beef liver, sheep kidney, chicken liver and milk matrices at spiked levels of 0.500-500 μg/kg were 93.4%-101%, and the relative standard deviations were 2.3%-8.0%. The method has less matrix interference, with high sensitivity, accuracy and reliability, and it is suitable for the quantitative detection of CCC residues in animal derived foods.
    Keywords:  animal derived foods; chlormequat chloride (CCC); high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS); solid phase extraction (SPE)
  15. Se Pu. 2020 Feb 08. 38(2): 212-217
      A method was developed for the determination of four protease inhibitors (saquinavir, ritonavir, nelfinavir and indinavir) in chicken using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The samples were extracted by shaking with 30% (v/v) acetonitrile aqueous solution (containing 1% (v/v) trichloroacetic acid), and purified by using mixed-mode cationic-exchanger (MCX) cartridges. The samples were separated on a Luna® C8 column (150 mm×2 mm, 3 μm) using 0.2% (v/v) formic acid aqueous solution (containing 5 mmol/L ammonium acetate) and acetonitrile as the mobile phases with gradient elution. The determination was carried out by using an electrospray ion source in the positive and multiple-reaction monitoring (MRM) modes. The calibration curves showed good linearities in the range of 0.1-20.0 μg/L, and the correlation coefficients (r2) were greater than 0.99. The limits of quantification (LOQs, S/N=10) of the four protease inhibitors varied from 0.20 μg/kg to 0.90 μg/kg. At the spiked levels of 1.0, 2.0, and 10.0 μg/kg, the average recoveries of the four protease inhibitors were ranging from 69.0% to 106.0%. The intra-day and inter-day relative standard deviations (RSDs) were 2.2%-13.8% (n=6) and 3.6%-14.6% (n=3), respectively. The method is simple, efficient, sensitive and accurate, and it can be used to detect residues of saquinavir, ritonavir, nelfinavir and indinavir in chicken.
    Keywords:  chicken; indinavir; nelfinavir; ritonavir; saquinavir; solid phase extraction (SPE); ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)
  16. Se Pu. 2020 Jul 08. 38(7): 817-825
      A rapid method based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed for the simultaneous determination of 37 mycotoxins having various physicochemical properties in grain and animal feed samples. The 37 analytes were extracted from ground samples with acetonitrile-water-formic acid (84:15.9:0.1, v/v/v) by 20 min vibration, and purified using a commercial MLJ-1 pass-through solid-phase extraction (SPE) clean-up cartridge. The analytes were then separated on a reversed-phase BEH RP18 column by a gradient elution program with 0.1 mmol/L ammonium acetate (containing 0.1% (v/v) formic acid) aqueous solution and 0.1% (v/v) formic acid methanol solution as mobile phases. The separated analytes were detected by MS/MS in the multiple reaction monitoring (MRM) mode via ESI+/- ionization. The results showed that the purification was completed in 1 min and that the 37 analytes could be separated on the chromatographic column in 15 min. The 37 mycotoxins showed a linear relationship within their respective linear ranges, and the correlation coefficients of the matrix-matched calibration curves were greater than 0.98. The average recoveries at four spiked levels (limit of quantification (LOQ), LOQ×5, LOQ×10, LOQ×25) for all the targets except fumonisins ranged from 80% to 120%, with the relative standard deviations (RSDs) lower than 20% (n=6). The limits of quantification (LOQs) for all the analytes were between 2 and 40 μg/kg. The proposed method is simple, fast, and accurate, thus being suitable for detecting multiple mycotoxins in grain and animal feed samples.
    Keywords:  feed; grain; impurity adsorption; mycotoxins; solid phase extraction (SPE); ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)
  17. Lab Invest. 2021 Jun 30.
      Stable isotope labeling techniques have been widely applied in the field of metabolomics and proteomics. Before the measured mass spectral data can be used for quantitative analysis, it must be accurately corrected for isotope natural abundance and tracer isotopic impurity. Despite the increasing popularity of dual-isotope tracing strategy such as 13C-15N or 13C-2H, there are no accurate tools for correcting isotope natural abundance for such experiments in a resolution-dependent manner. Here, we present AccuCor2 as an R-based tool to perform the correction for 13C-15N or 13C-2H labeling experiments. Our method uses a newly designed algorithm to construct the correction matrices that link labeling pattern and measured mass fractions, then use non-negative least-squares to solve the labeling patterns. Our results show that the dual-isotope experiments often require a mass resolution that is high enough to resolve 13C and 15N or 13C and 2H. Otherwise, the labeling pattern is not solvable. However, this mass resolution may not be sufficiently high to resolve other non-tracer elements such as oxygen or sulfur from the tracer elements. Therefore, we design AccuCor2 to perform the correction based on the actual mass resolution of the measurements. Using both simulated and experimental data, we show that AccuCor2 performs accurate and resolution-dependent correction for dual-isotope tracer data.
  18. Se Pu. 2020 Jan 08. 38(1): 74-85
      As the main structural components of cellular and sub-cellular membranes, lipids are a major source of energy and play an important role in various biological processes such as cellular signaling. Lipid profiling has attracted increasing attention in recent years, and chromatography-mass spectrometry techniques for lipid profiling occupy a dominant position in this regard. Sample preparation, which aims at enriching trace substances and reducing matrix interferences, is a crucial step in the analysis of lipids for the intricacy of sample matrices. Here, we review recent developments in sample preparation techniques based on chromatography-mass spectrometry and their application to lipid profiling. Various sample preparation techniques are described and summarized. The extraction methods based on liquid phase included liquid-liquid extraction and single organic solvent extraction. The extraction methods based on solid phase involved solid-phase extraction and solid-phase microextraction. The field-assisted extraction methods cover supercritical fluid extraction, pressurized liquid extraction, microwave-assisted extraction, and ultrasound-assisted extraction. Moreover, online coupling sample preparation methods and sample preparation methods for in vivo lipid analysis are presented. Finally, the problems and trends in sample preparation techniques for lipid profiling based on chromatography-mass spectrometry are discussed. It is believed that efficient development of sample preparation techniques would help improve the sensitivity and selectivity of lipid profiling as well as the analysis speed.
    Keywords:  chromatography-mass spectrometry; lipid profiling; sample preparation
  19. Se Pu. 2020 Feb 08. 38(2): 250-254
      A pseudotargeted metabolomics method based on gas chromatography-mass spectrometry (GC-MS) was developed for the analysis of fatty acids in rice. A total of 16 fatty acids were detected in rice. The method was used to identify the difference between fatty acid profiles in rice. The total intensity of the saturated and unsaturated fatty acids was used as the evaluation index. The effects of six extraction methods and four extraction solvents on the extraction efficiencies of the fatty acids were compared. The developed method was used to investigate the fatty acid profiles in five types of rice (Daohuaxiang, Jixing, Jinlangzi, Nongda, and Zhuangyuan). The fatty acid profile in Daohuaxiang rice was different from that in the other four species. The fatty acids in Jinlangzi and Nongda rice were markedly different, while those in Jixing and Jinlangzi rice were similar. The method is simple, stable and accurate, and it can provide basic data for the improvement of rice quality and nutritional value.
    Keywords:  fatty acids; gas chromatography-mass spectrometry (GC-MS); pseudotargeted metabolomics; rice
  20. Int J Mol Sci. 2021 Jun 28. pii: 6972. [Epub ahead of print]22(13):
      Metabolites and enzymes involved in the kynurenine pathway (KP) are highly promising targets for cancer treatment, including gastrointestinal tract diseases. Thus, accurate quantification of these compounds in body fluids becomes increasingly important. The aim of this study was the development and validation of the UHPLC-ESI-MS/MS methods for targeted quantification of biologically important KP substrates (tryptophan and nicotinamide) and metabolites(kynurenines) in samples of serum and peritoneal fluid from gastric cancer patients. The serum samples were simply pretreated with trichloroacetic acid to precipitate proteins. The peritoneal fluid was purified by solid-phase extraction before analysis. Validation was carried out for both matrices independently. Analysis of the samples from gastric cancer patients showed different accumulations of tryptophan and its metabolites in different biofluids of the same patient. The protocols will be used for the evaluation of tryptophan and kynurenines in blood and peritoneal fluid to determine correlation with the clinicopathological status of gastric cancer or the disease's prognosis.
    Keywords:  LC-MS/MS; gastric cancer; kynurenine pathway; peritoneal fluid analysis; serum analysis; targeted metabolomics; tryptophan metabolism
  21. J Am Soc Mass Spectrom. 2021 Jun 30.
      Liquid chromatography-mass spectrometry (LC-MS) is one of the most widely used analytical tools. High analysis volumes and sample complexity often demand more informative LC-MS acquisition schemes to improve efficiency and throughput without compromising data quality, and such a demand has been always hindered by the prerequisite that a minimum of 13-20 MS scans (data points) across an analyte peak are required for accurate quantitation. The current study systematically re-evaluated and compared the impact of different scan numbers on quantitation analysis using both triple quadrupoles mass spectrometry (TQMS) and high-resolution mass spectrometry (HRMS). Contrary to the 13-20 minimal scan prerequisite, the data obtained from a group of eight commercial drugs in the absence and presence of biological matrices suggest that 6 scans per analyte peak are sufficient to achieve highly comparable quantitation results compared to that obtained using 10 and 20 scans, respectively. The fewer minimal scan prerequisite is presumably attributed to an improved LC system and advanced column technology, better MS detector, and more intelligent peak detection and integration algorithms leading to a more symmetric peak shape and smaller peak standard deviation. As a result, more informative acquisition schemes can be broadly set up for higher throughput and more data-rich LC-MS/MS analysis as demonstrated in a hepatocyte clearance assay in which fewer MS scans executed on HRMS led to broader metabolite coverage without compromising data quality in hepatic clearance assessment. The demonstrated acquisition scheme would substantially increase the throughput, robustness, and richness of the nonregulatory analysis, which can be broadly applied in diverse fields including pharmaceutical, environmental, forensic, toxicological, and biotechnological.
  22. Se Pu. 2020 Sep 08. 38(9): 1013-1021
      Capillary electrophoresis-mass spectrometry (CE-MS) has the advantages of higher sensitivity, higher efficiency, and less sample consumption. Moreover, it possesses obvious advantages during the analysis of strongly charged and highly polar samples. CE-MS has been widely applied in life sciences, medicine, and pharmacology. In the past ten years, the main factors affecting its application were system stability, reproducibility, and data accuracy. In order to solve the existing problems of CE-MS, researchers have invested significant effort in technology innovation to further expand CE-MS application. In the fields of medicine and analytical chemistry, substantial research indicates that CE-MS is superior compared to other metabolomic and proteomic approaches. This study aims at reviewing the latest methods and applications developed in the fields of medicine and analytical chemistry since 2015. Furthermore, it also aims at enhancing the technology development-related application value of CE-MS and serving as a reference for future development. Further development of the CE-MS technology is discussed from the aspects of coating-sample interaction, interface types, and data processing methods. Concerning the coating types, neutral coatings had been applied extensively in CE-MS and there should be no limitation to the charge of the analyte. The coating decreased sample adsorption on the inner wall by covering the surface charge, greatly reducing the electroosmotic flow (EOF). A charged capillary coating could modify such an EOF direction. The cationic coating could reduce the hydrophobic interaction between the sample and the capillary column, resulting in higher EOF. If it is applied to the sheathless interface, the resolution could be improved by extending the capillary length. Anionic coatings are predominant among the anionic compounds, shortening the separation time by reducing the interaction between the anionic compounds and the capillary. The coating type should be chosen relative to the analyte characteristics. Concerning the interface technology, all interfaces should be simple, practical, and non-dependent on sheath liquid and background electrolytes. As far as data processing methods are concerned, it is necessary to design and develop a practical method for span space data comparison and processing. The optimized experimental conditions have effectively improved separation efficiency and data comparison analysis. Furthermore, they established a solid foundation for its application development. CE-MS analysis of complex samples in the fields of metabolomics and proteomics (e. g., of tissues, cells, body fluids, etc.) could provide a visualization method for future clinical analysis. It contributes to the development of cancer pathological analysis, drug development, disease surveillance, etc. The characteristic analysis of small molecule metabolites and protein biomarkers directly reflects on enzymatic activity in the biological systems. It could be associated with the development of various diseases/complications. Omics analysis also has an important directive to disease detection and surveillance with obvious advantages in disease diagnosis, staged treatment, drug development, and patient treatment progress. CE-MS is useful in detecting complications and promoting personalized medicine. It provides technical support for future clinical developments. In addition to a comprehensive review of the recent advances of CE-MS research, this paper also indicates the development directions of CE-MS. In order to avoid the problem of omics analysis and obtain the optimized analysis results, future analysis should be improved from the following three aspects:(i) The analysis conditions should be optimized concerning sample preparation methods and separation techniques. (ii) The analytic techniques should be supported to adjust to capillary coating and interface technology. (iii) New ideas should be developed in the fields of clinical research and statistical analysis.
    Keywords:  capillary electrophoresis-mass spectrometry (CE-MS); metabolomics; proteomics; review; technical improvement
  23. Se Pu. 2020 Nov 08. 38(11): 1348-1354
      Police officers currently use the colloidal gold rapid testing method to detect heroin in the urine of drug abusers, but the results are often rendered erroneous due to the presence of antitussive drugs, which contain opioids. The traditional manual liquid-liquid extraction method for urine testing has low efficiency and poor sensitivity, and hence, it fails to meet the requirements of the public security department to crack down on drug abusers. Therefore, to avoid punishment, most rapid-test-positive people make false claims about intaking cough suppressants. It is imperative to establish a highly efficient automatic method for the simultaneous determination of multiple opioids in urine, to rule out the use of heroin. A method based on solid-phase extraction and derivatization coupled with gas chromatography-mass spectrometry (GC-MS) has been developed for the simultaneous detection of morphine, O6-acetylmorphine, codeine, and acetyl codeine in urine. Since these four opioids exists as cations in acidic aqueous solution, the urine samples collected from dead bodies or drug addicts were adjusted to pH 6 by using phosphate buffer, enriched, and purified by MCX-SPE columns. Then, morphine, O6-acetylmorphine, and codeine were derivatized by N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA) for GC-MS testing. The effects of sample loading and elution flow rate, percentage of formic acid in the wash solvent (methanol), percentage of ammonia in the eluent (methanol), volume of the wash solvent, and drying time of the cartridge on the extraction efficiency were investigated in detail. The best results were obtained under the following conditions:sample loading and elution flow rate, 1.0 mL/min; volume fraction of formic acid in the wash solvent, 3%; volume fraction of ammonia in the eluent solvent, 5%; volume of 3% (v/v) formic acid in methanol (eluent), 1 mL; and drying time of the cartridge, 1 min. The GC-MS results showed good linearity in the range of 0.02-0.8 μg/mL with correlation coefficients (r2) ≥ 0.998. The limits of detection (LODs) and limits of quantification (LOQs) were 0.0016-0.0039 μg/mL and 0.0054-0.0128 μg/mL, respectively. The recoveries of the target analytes were between 93.0% and 110.3% at spiked levels of 0.02, 0.1, and 0.2 μg/mL. As opposed to similar reported methods, our method showed high sensitivity and recovery; furthermore, the matrix interference was eliminated, and the chromatographic peaks of the analytes were completely separated from the impurity peaks at the level of 0.2 μg/mL. The automatic solid-phase extraction equipment is convenient to operate and allows one to process samples in batches. The conditions for solid-phase extraction can be precisely controlled, and the detection accuracy is greatly improved. In addition, a large number of sample tests can be performed by a few experimenters. Hence, this method facilitates simple and rapid forensic toxicology testing and drug abuse monitoring on a large scale.
    Keywords:  derivatization; gas chromatograph-mass spectrometry (GC-MS); morphine; opioid; solid-phase extraction (SPE); urine
  24. Se Pu. 2020 Dec 08. 38(12): 1396-1401
      Streptomycin (STR) and dihydrostreptomycin (DSTR) are two of the most common aminoglycoside antibiotics used in veterinary medicine. STR is produced by some streptomyces griseus strains, and DSTR is a derivative of STR. In recent years, STR has been widely used in grapes to induce denuclearization. However, high levels of STR may have adverse effects like serious ototoxicity and nephrotoxicity. Therefore, to ensure the quality of grapes and the health of consumers, the regulation of STR and DSTR levels in grapes is required. An analytical method was developed for the identification and quantification of STR and DSTR in grapes by liquid chromatography-tandem mass spectrometry (LC-MS/MS). STR and DSTR are highly polar compounds due to the presence of various amino and hydroxyl groups in their structure. The determination of STR and DSTR poses a considerable analytical challenge, both during sample preparation and instrument analysis. In this study, the main factors governing the response, recovery, and sensitivity of these compounds, such as the type of chromatographic column, the type and proportion of the mobile phase and extraction solvent, the dosage of sodium 1-hexane sulfonate solution, and elution solvent and its volume, were investigated during sample pretreatment and instrument analysis. The STR and DSTR residues in the grape sample were extracted by ultrasonication with a phosphoric acid solution (pH 2), and cleanup and enrichment was performed using an Oasis HLB solid phase column. The analysis was performed using a UPLC Waters HSS T3 column (100 mm×2.1 mm, 1.8 μm) at the column temperature of 35℃. The injection volume was 2 μL. The mobile phase consisted of 0.1% formic acid aqueous solution and methanol with a volume ratio of 60:40. ESI-MS/MS was operated in multiple reaction monitoring (MRM) mode. External standard calibration curves were used for quantification. Based on the optimized method, both analytes displayed good linearity between 2 and 400 μg/L. The correlation coefficients were 0.9991-0.9997. Recoveries in spiked blank grape samples (5, 10, 20, and 40 μg/kg) ranged from 76.8% to 91.9%, with the relative standard deviations (RSDs) less than 10.2%, in compliance with the current legislation. The limits of detection and the limits of quantification of both analytes were 1 μg/L and 5 μg/kg, respectively. To assess the feasibility and potential of the proposed approach for routine analyses of STR and DSTR in other kinds of grape samples, the developed method was applied to the analysis of these compounds in red grapes, xinyu grapes, and xiahei grapes. The recoveries of STR and DSTR in the three kinds of blank grape samples were 77.2%-83.9% and 70.8%-78.9%, respectively, and the RSDs ranged from 3.0% to 15.6%. The results showed that the optimized methods can yield satisfactory recoveries for the analytes in grapes. In this method, the combination of Waters HSS T3 column to overcome the difficulties of the retention and separation of these highly polar compounds in the reverse phase, avoids the use of an ion-pair additive in the mobile phase to increase their retention, which is known to cause severe contamination of the column and serious ion suppression with electrospray ionization detection. In addition, the ideal enrichment and purification effect can be achieved by adding a sodium 1-hexane sulfonate solution to the superstratum extract with the use of only Oasis HLB for sample treatment. The method described herein has the advantages of easy operation, accuracy, and selectivity, making it feasible for the identification and quantification of STR and DSTR residues in grapes.
    Keywords:  dihydrostreptomycin; grape; liquid chromatography-tandem mass spectrometry (LC-MS/MS); solid phase extraction (SPE); streptomycin
  25. Se Pu. 2020 Feb 08. 38(2): 206-211
      A rat model with isoproterenol (ISO)-induced myocardial ischemia was obtained. Liquid chromatography-mass spectrometry (LC-MS)was conducted on the serum and heart tissue metabolites of normal and model rats. Principal component analysis (PCA) and differentiation analysis of supervised partial least-squares method (PLS-DA) were applied to the metabolomics data for multidimensional statistical analysis, and the resulting biomarkers were screened. Compared with the normal group, 18 different endogenous metabolites in plasma and tissues were detected, including the products of arginine and proline metabolism; glycine, serine, and threonine metabolism; glutamine and glutamate metabolism; and taurine and hypotaurine metabolism. These metabolites can be used as important biomarkers for myocardial ischemia. The results of this study can help reveal the pathogenesis of myocardial ischemia and aid clinical diagnosis.
    Keywords:  biomarkers; liquid chromatography-mass spectrometry (LC-MS); metabolomics; myocardial ischemia
  26. Se Pu. 2020 Feb 08. 38(2): 224-231
      An ultra performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-MS/MS) method was developed for the determination of seven heterocyclic aromatic amines (HCAs) in oil and fried food samples. The samples were extracted with acetonitrile containing 1% (v/v) ammonia water, defatted by n-hexane saturated with acetonitrile, and cleaned up with PCX solid phase extraction column. The extracts were separated on a Waters ACQUITY UPLC BEH C18 reversed-phase column (50 mm×2.1 mm, 1.7 μm) with gradient elution by using acetonitrile-10 mmol/L ammonium formate aqueous solution as the mobile phase. Quantitative detection was performed by using a positive-ion electrospray ionization source in the multiple reaction monitoring (MRM) mode, employing an internal isotope standard. The method showed good linear relationships in the range of 0.5-100 μg/L for the seven HCAs with correlation coefficients (R2)>0.999. The average recoveries ranged from 64.31% to 113.8% with the relative standard deviations of 0.18%-9.26% at the three spiked levels in oil and fried food samples. The limits of detection (LODs) and limits of quantification (LOQs) were 0.01-0.14 ng/g and 0.09-0.38 ng/g, respectively. The method is sensitive, accurate, and suitable for the determination and confirmation of the seven HCAs in oil and fried food samples.
    Keywords:  fried foods; heterocyclic aromatic amines (HCAs); oils; solid phase extraction (SPE); triple quadrupole mass spectrometry (MS/MS); ultra performance liquid chromatography (UPLC)
  27. Plant Biotechnol (Tokyo). 2021 Mar 25. 38(1): 167-171
      Depository of low-molecular-weight compounds or metabolites detected in various organisms in a non-targeted manner is indispensable for metabolomics research. Due to the diverse chemical compounds, various mass spectrometry (MS) setups with state-of-the-art technologies have been used. Over the past two decades, we have analyzed various biological samples by using gas chromatography-mass spectrometry, liquid chromatography-mass spectrometry, or capillary electrophoresis-mass spectrometry, and archived the datasets in the depository MassBase ( As the format of MS datasets depends on the MS setup used, we converted each raw binary dataset of the mass chromatogram to text file format, and thereafter, information of the chromatograph peak was extracted in the text file from the converted file. In total, the depository comprises 46,493 datasets, of which 38,750 belong to the plant species and 7,743 are authentic or mixed chemicals as well as other sources (microorganisms, animals, and foods), as on August 1, 2020. All files in the depository can be downloaded in bulk from the website. Mass chromatograms of 90 plant species obtained by LC-Fourier transform ion cyclotron resonance MS or Orbitrap MS, which detect the ionized molecules with high accuracy allowing speculation of chemical compositions, were converted to text files by the software PowerGet, and the chemical annotation of each peak was added. The processed datasets were deposited in the annotation database KomicMarket2 ( The archives provide fundamental resources for comparative metabolomics and functional genomics, which may result in deeper understanding of living organisms.
    Keywords:  database; mass spectrometry; metabolome
  28. Se Pu. 2020 Jan 08. 38(1): 66-73
      Lipids play an important physiological role in living system. They are the major components of cellular membranes, in addition to participating in energy storage as well as signal recognition and transmission. In recent years, an increasing number of studies have shown that disturbances in lipid metabolism are closely related to the development of some diseases. Lipid analysis is of great significance for understanding the mechanism and the process of diseases. Because of the matrix interference and low concentration of analytes, existing techniques require sample preparation steps to ensure good analytical performance. In the present study, sample preparation methods in lipidomics used in various fields are reviewed, including lipid extraction methods, such as liquid-liquid extraction and solid phase extraction, as well as the chemical derivatization techniques for different lipid categories. In addition, the development of the sample preparation in lipidomics is also prospected.
    Keywords:  chemical derivatization; lipid extraction; lipidomics; review; sample preparation
  29. Rapid Commun Mass Spectrom. 2021 Jun 28. e9154
      RATIONALE: Accurate and reliable measurements are mandatory in the field of environmental monitoring. Matrix effects are often depicted as the Achilles heel of LC-MS analysis since they may be prejudicial for analytical performances such as detection capability and accuracy if not documented nor compensated. Here a methodological work for the evaluation and compensation of matrix effects is described.METHODS: Natural and synthetic representative water samples were used for the evaluation of matrix effects with the post-extraction addition technique. Samples were analysed by an ultra-performance liquid chromatography separation coupled to tandem mass spectrometry (LC-MS/MS) and electrospray ionization (ESI). Isotopic dilution was investigated as a way to allow compensation of signal alteration and therefore satisfactory quantification. When this approach was not possible a methodological work has been done to choose the more appropriate internal standard.
    RESULTS: The matrix effects were dependent on both matrix composition and nature of analyte. They ranged from a total signal suppression to a signal enhancement of +27% but were independent of compound concentration. The correction of matrix effects by internal standards was satisfactory, particularly for compounds benefiting from isotope dilution leading to acceptable quantification performances.
    CONCLUSION: Even if no exhaustive nor agreed criteria exist for the final interpretation of matrix effects, this study highlights the interest of isotope dilution to reduce their inherent prejudicial effects in quantification and the need to conduct this type of study in representative matrices. Moreover, a methodological approach is proposed to choose the more appropriate available internal standard when isotope dilution is not possible.
  30. Se Pu. 2020 Dec 08. 38(12): 1388-1395
      The presence of 3-chloro-1,2-propanediol fatty acid esters (3-MCPDE) in food and processed materials has recently become a topic of concern because of the toxicity of their metabolites. 3-MCPDE structurally similar to glyceride, which makes it difficult to separate or extract them from oils and fritters. A method based on ultra performance convergence chromatography-tandem mass spectrometry (UPC2-MS/MS) was established for the determination of 15 3-MCPDE in vegetable oils and fritters. Amino-packed columns were used to purify the samples. The analytical conditions were optimized, and the matrix effect was investigated. The sample was treated by column chromatography to remove glyceride and free fatty acids, which induce strong matrix effects. The amino-packed column was eluted with hexane and hexane-ethyl acetate (6:4, v/v). Every 1 mL of the eluent was analyzed using a UPC2 and ACQUITY QDa detector. Elution curves were drawn based on the testing data and used to determine the collection volume. The collection volumes for 3-chloro-1,2-propanediol diesters and monoesters according to the elution curves were 7-14 mL and 3-9 mL. The collected eluent was mixed and dried under nitrogen flow at a temperature of 60℃. A hexane-isopropanol (98:2, v/v, 1 mL) mixture was used to dissolve the residue. The resulting solution was separated on a Viridis HSS C18 SB column (150 mm×2.1 mm, 1.8 μm) under gradient elution. Supercritical carbon dioxide and methanol (containing 40% acetonitrile and 0.1% formic acid) were used as the mobile phases, and the flow rate was 1 mL/min. The separated compounds were analyzed by tandem MS with an electrospray ionization (ESI) source in positive and multiple reaction monitoring modes. Water (containing 97% isopropanol and 0.2% ammonia water) was used as the auxiliary pump mobile phase, and the flow rate was 0.2 mL/min. The method showed good linear relationships in the range of 0.5-100 μg/L (r2 ≥ 0.9973). The limits of detection (LODs) and limits of quantification (LOQs) were 0.01-0.68 μg/L (S/N=3) and 0.04-1.74 μg/L (S/N=10), respectively. The average recoveries (n=9) at the three spiked levels were in the range of 81.6%-98.5%. The relative standard deviations were in the range of 1.8%-6.4%. The matrix effects in the case of the oils and fritters were weak. The developed method was used to detect 44 oil samples and eight fritter samples. Meanwhile, some suspect 3-MCPDE compounds outside the scope of the investigation were analyzed based on their primary and secondary mass spectra. The detection rates of 3-MCPDE in oils and fritters were 84.1% and 87.5%, and their amounts were in the range of 0.024-4.481 mg/kg and 0.018-1.144 mg/kg, respectively. The detection rates of 3-MCPDE in rapeseed oil were higher compared to those for other kinds of oil. The method is specific, fast, simple, accurate, reliable, and environmentally friendly, in addition to being more sensitive than other methods and showing better matrix compatibility for oils. This method has been successfully used to determine the types and amounts of 3-MCPDE in vegetable oils and fritters. The research findings provided accurate data to assess the exposure risk of 3-MCPDE. The results of our experiment also provided valuable information for elucidating the formation mechanism of 3-MCPDE. The proposed method can be used to analyze waste edible oil based on large amounts of analysis data. However, this method has some limitations. The resolution ratio of the mass spectrometer used in this method is too low for the qualitative analysis of unknown compounds. The qualitative results for the suspect 3-MCPDE compounds are not particularly accurate, and a large variety of monomer standards are required for the quantitative determination of 3-MCPDE. The 3-MCPDE standards are expensive, and there is limited choice of these standards; moreover, they are difficult to synthesize. The poor ionization yield of 3-chloro-1,2-propanediol monoesters under the ESI conditions resulted in high LODs. Hence, it is necessary to develop a method for increasing the ionization of monoesters, for example, via derivatization.
    Keywords:  3-chloro-1,2-propanediol fatty acid esters (3-MCPDE); fritters; ultra performance convergence chromatography-tandem mass spectrometry (UPC2-MS/MS); vegetable oil
  31. Se Pu. 2020 Jun 08. 38(6): 672-678
      A method based on dispersive solid-phase extraction-high performance liquid chromatography-tandem mass spectrometry (DSPE-HPLC-MS/MS) was developed for the simultaneous determination of five analytes scutellarein, 4'-hydroxywogonin, norwogonin, baicalein, and wogonin in healthy product tablets. The samples were extracted with 10 mL acetone and purified on 75 mg C18. The types and dosages of the extracted solvents and adsorbents were optimized. The results showed that these five analytes exhibited good linear relationships in their respective linear ranges. The correlation coefficients (r) were more than 0.99. The detection limits and quantitative limits were 0.5-40 μg/kg and 2.0-120 μg/kg, respectively. Recovery tests were carried out using three kinds of health product tablet matrices. The concentrations were one, five, and ten times the quantitative limit. The average recovery of these five targets was 83.1% to 106.5%, with relative standard deviations (RSDs) ranging from 0.97% to 4.52%. This method is easy to operate, and it shows high sensitivity and good reproducibility. Thus, it is suitable for the simultaneous determination of the target compounds in health tablets.
    Keywords:  baicalein; dispersive solid-phase extraction (DSPE); health product; high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS)
  32. Se Pu. 2019 Dec 08. 37(12): 1261-1267
      Sialic acids are a family of 9-carbon carboxylated monosaccharides. They are widely found in vertebrates, as well as some invertebrates, fungi and bacteria. Sialic acids in the organisms exist in free form or as important components at the terminal positions of glycoconjugates. They are involved in a variety of physiological activities, and are closely related to disease, such as inflammation and cancer. Analytical methods based on chromatography and/or mass spectrometry are the most widely used for characterizing sialic acids from biological samples. In order to improve the detection sensitivity and/or efficiency of chromatographic separation, it is necessary to derivatize the sialic acids prior to analysis. A variety of derivatization methods have been developed in the past few decades. The present review focuses on the derivatization methods for the analysis of sialic acids by chromatography and/or mass spectrometry at the monosaccharide, free sialic acid, N/O-glycan and glycosphingolipid levels. The applications and development trends in this field are also prospected.
    Keywords:  chromatography; derivatization; mass spectrometry (MS); review; sialic acids
  33. Toxins (Basel). 2021 Jun 11. pii: 418. [Epub ahead of print]13(6):
      In the course of assessing the human exposure to mycotoxins, biomarker-based approaches have proven to be important tools. Low concentration levels, complex matrix compositions, structurally diverse analytes, and the large size of sample cohorts are the main challenges of analytical procedures. For that reason, an online solid phase extraction-ultra high-performance liquid chromatography-tandem mass spectrometry (online SPE-UHPLC-MS/MS) method was developed, allowing for the sensitive, robust, and rapid analysis of 11 relevant mycotoxins and mycotoxin metabolites in human urine. The included spectrum of analytes comprises aflatoxin M1 (AFM1), altenuene (ALT), alternariol monomethyl ether (AME), alternariol (AOH), citrinin (CIT) and its metabolite dihydrocitrinone (DH-CIT), fumonisin B1 (FB1), ochratoxin A (OTA), and zearalenone (ZEN) as well as α- and β-zearalenol (α- and β-ZEL). Reliable quantitation was achieved by means of stable isotope dilution, except for ALT, AME and AOH using matrix calibrations. The evaluation of method performance displayed low limits of detection in the range of pg/mL urine, satisfactory apparent recovery rates as well as high accuracy and precision during intra- and interday repeatability. Within the analysis of Zimbabwean urine samples (n = 50), the applicability of the newly developed method was shown. In addition to FB1 being quantifiable in all analyzed samples, six other mycotoxin biomarkers were detected. Compared to the occurrence rates obtained after analyzing the same sample set using an established dilute and shoot (DaS) approach, a considerably higher number of positive samples was observed when applying the online SPE method. Owing to the increased sensitivity, less need of sample handling, and low time effort, the herein presented online SPE approach provides a valuable contribution to human biomonitoring of mycotoxin exposure.
    Keywords:  HPLC-MS/MS; biomonitoring; metabolite; mycotoxin; online solid phase extraction; urine
  34. Se Pu. 2020 Nov 08. 38(11): 1302-1307
      Dacarbazine (DTIC) is a first-line chemotherapy drug that is widely used in clinical practice for malignant melanoma. DTIC is metabolized by the liver in vivo. Some drugs are excreted in urine in the form of a prototype. Hence, DTIC in urine can be monitored to evaluate its utilization and conversion rate in the human body, and then to determine its therapeutic effect. Urine is the only body fluid that can be obtained in large quantities without damage, and it plays an important role in the analysis of body functions. However, the composition of urine is complex and there is large matrix interference, because of which trace analysis or trace component analysis is difficult. At present, the main analytical methods for DTIC are high performance liquid chromatography (HPLC) with/without mass spectrometry (MS). HPLC and HPLC-MS have the advantages of good separation effect, good selectivity, high detection sensitivity, automatic operation, and wide application range. Unfortunately, DTIC is a strongly polar and weakly basic compound; thus, it is difficult to achieve good separation and obtain good peak shapes by conventional reversed-phase chromatography. To overcome these defects, it is necessary to develop a novel method for the analysis of DTIC. In this study, mice were subjected to 12 h of fasting; then, blueberry anthocyanin was administered by gavage, and DTIC was administered by intraperitoneal injection. Then, morning urine was collected in a metabolic cage. Urine collection was continued every 4 days for a total of 5 times. Within 2 h, the collected urine was centrifuged (3000 g, 4℃) for 10 min to remove solids. The supernatant was stored in a refrigerator at-80℃. Before analysis, the urine samples were removed from the refrigerator and thawed naturally at room temperature. Then, the samples were treated by the acetone-sediment method, freeze-dried, dissolved in the mobile phase, and subjected to HPLC analysis with isocratic elution. The separation was performed on a Shimadzu-GL ODS column (250 mm×4.6 mm, 5 μm). The mobile phase was methanol/acetonitrile (1:1, v/v)-0.01 mol/L NaH2PO4 (pH 6.5; 20:80, v/v) at a flow rate of 1 mL/min. The detection wavelength, column temperature, and running time were 280 nm, 30℃, and 15 min, respectively. Under the optimized conditions, the retention time of DTIC was 5.3 min, and a good peak shape was obtained. The linearity ranged from 0.25 to 1000 μg/mL (r2=0.999). The limits of detection and quantification were calculated to be 0.12 μg/mL and 0.25 μg/mL based on signal-to-noise ratios of 3 and 10, respectively. At three spiked levels (50.0, 375, and 500 μg/mL), the average recoveries were 98.9%, 102%, and 99.1% with relative standard deviations (RSDs) of 3.2%, 1.3%, and 1.2% (n=5), respectively. The RSDs of the interday and intraday measurements were lower than 3.8% and 4.4%, respectively. The proposed method allowed for the accurate determination of DTIC in urine using a mixed organic solvent/phosphate buffer solution as the mobile phase, with equivalent elution for 15 min. This method was successfully applied to monitor the change in DTIC concentration in the urine of C57BL/6 mice in various stages of melanoma. The results demonstrate that the method is simple, reliable, and easy to apply.
    Keywords:  blueberry anthocyanin; dacarbazine; high performance liquid chromatography (HPLC); melanoma; nutritional adjuvant; urine
  35. Front Chem. 2021 ;9 707382
      The retention time provides critical information for glycan annotation and quantification from the Liquid Chromatography Mass Spectrometry (LC-MS) data. However, the variation of the precise retention time of glycans is highly dependent on the experimental conditions such as the specific separating columns, MS instruments and/or the buffer used. This variation hampers the exploitation of retention time for the glycan annotation from LC-MS data, especially when inter-laboratory data are compared. To incorporate the retention time of glycan across experiments, Glucose Unit Index (GUI) can be computed using the dextrin ladder as internal standard. The retention time of glycans are then calibrated with respect to glucose units derived from dextrin ladders. Despite the successful application of the GUI approach, the manual calibration process is quite tedious and often error prone. In this work, we present a standalone software tool GlycanGUI, with a graphic user interface to automatically carry out the GUI-based glycan annotation/quantification and subsequent data analysis. When tested on experimental data, GlycanGUI reported accurate GUI values compared with manual calibration, and thus is ready to be used for automated glycan annotation and quantification using GUI.
    Keywords:  GUI; annotation; glycan; mass spectrometry; quantification
  36. Se Pu. 2021 Aug;39(8): 878-888
      The complexity of the soil matrix, as well as the wide spectrum and trace levels of antibiotic residues in soil, make highly sensitive instrumental methods, efficient purification and enrichment methods, and simultaneous determination of multiple antibiotics key and challenging aspects in the analysis of antibiotics in soil. In this study, a solid phase extraction-ultra-high performance liquid chromatography-tandem mass spectrometry (SPE-UHPLC-MS/MS) method was developed for the simultaneous determination of 30 antibiotics (grouped into seven categories: sulfonamides, fluoroquinolones, tetracyclines, macrolides, β-lactams, amphenicols, and lincosamides) in soil samples. In the UHPLC-MS/MS experiment, florfenicol and chloramphenicol were analyzed in the negative ionization multiple-reaction monitoring (MRM) mode, and the other 28 target analytes were analyzed in the positive MRM mode. Sensitive MS conditions were realized by optimizing the instrumental parameters such as collision energy and declustering potential. The effects of the injection solvent (proportion of methanol to water) and mobile phase (types and compositions of the solvents) on the shape and intensity of the chromatographic peaks were studied. The optimized UHPLC conditions were as follows: injection solvent, 10%(v/v) methanol aqueous solution; chromatographic column, BEH-C18 column; mobile phase, methanol and water both containing 0.1%(v/v) formic acid; flow rate, 0.4 mL/min; sample injection volume, 5.0 μL. The effects of the extraction solution (the types and compositions of solvents) and clean-up processes (pH of the loading solution, as well as the types and compositions of the rinse solution and elution solvent) on the method performance were investigated. The acetonitrile/Na2EDTA-McIlvaine buffer showed better extraction efficiency for fluoroquinolones than did the methanol/Na2EDTA-McIlvaine buffer. Improved recoveries of sulfonamides, macrolides, tetracyclines, and β-lactams were observed when the pH of the loading solution was set to 8.0. The recoveries of sulfadiazine and amoxicillin increased with a decrease in the proportion of methanol to water for the rinse solution. Compared to individual methanol or acetonitrile, the methanol-acetonitrile (1∶1, v/v) mixture showed better elution efficiency for the target analytes. The optimized pretreatment conditions were determined as follows: the soil sample was spiked with mixed internal standards, and then extracted with 10 mL of acetonitrile/Na2EDTA-McIlvaine buffer (1∶1, v/v) by shaking for 30 min and ultrasonication for 15 min. The extraction was repeated three times. The sample extract was adjusted to pH 8.0 and loaded onto an Oasis HLB cartridge for purification. The cartridge was rinsed with 10 mL of water to remove impurities and eluted with 10 mL of methanol-acetonitrile (1∶1, v/v). Quantitative analysis was conducted using the isotope internal standard method. The method limits of detection and quantification were in the range of 0.013-1.21 and 0.043-4.04 μg/kg, respectively. The correlation coefficients of the calibration curve were 0.992-1.00, suggesting good linearity of the method. At three spiked levels (20, 100, and 200 μg/kg), the average recoveries of most target antibiotics were 44.8% to 164%, and the relative standard deviations were 0.700% to 14.8%. The method was successfully applied to the analysis of the 30 antibiotics in six soil samples. Seventeen antibiotics were detected in the soil samples, and the total contents of the antibiotics in each sample ranged from 73.4 to 184 μg/kg. Twelve antibiotics with a detection frequency of 100% included roxithromycin, clarithromycin, ciprofloxacin, norfloxacin, enrofloxacin, ofloxacin, fleroxacin, lomefloxacin, oxytetracycline, doxycycline, tetracycline, and penicillin G. Ciprofloxacin and norfloxacin were the predominant antibiotics in the soils, with contents in the range of 13.7-32.1 and 15.6-43.6 μg/kg, respectively. The developed method is simple, rapid, and solvent-saving, and it shows promise for use in the simultaneous determination of trace levels of the 30 antibiotics in soil.
    Keywords:  antibiotics; soil; solid phase extraction (SPE); ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS)
  37. Metabolites. 2021 Jun 08. pii: 365. [Epub ahead of print]11(6):
      Lipidomics aim to quantify lipid species in all kinds of samples, including tissues. To subject a fixed amount of sample to various workflows, tissue homogenates were frequently prepared at defined concentrations in water or by addition of organic solvents. Here, we investigated this first step of tissue lipidomics by quantitative flow injection analysis coupled to Fourier-Transform mass spectrometry (FTMS). The influence of sample concentration, solvent composition, and homogenization procedure on the recovery of lipids was studied in murine liver. Liver homogenates were prepared either by grinding tissue in liquid nitrogen or by bead-based homogenization. Ground samples were dissolved at different concentrations in water, methanol, and water/methanol = 1/1 (v/v). Here, lipid recovery depends on solvent composition and sample concentration. The recovery of nonpolar lipid classes, including triglycerides and cholesteryl ester, was decreased in methanolic homogenates. In contrast, due to superior dispersion of precipitates, bead-based homogenization resulted in efficient lipid recovery independent of the solvent composition. However, lipid distribution within samples, i.e., lipid content of supernatant and pellet following centrifugation, was altered substantially by solvent composition. In conclusion, accurate lipid quantification of tissue homogenates requires evaluation of solvent composition, sample concentration, as well as the homogenization method to guarantee efficient lipid recovery. Due to a potential loss of lipids, removal of precipitates by centrifugation prior to lipid extraction should be avoided.
    Keywords:  extraction; lipidomics; lipids; mass spectrometry; preanalytics; quantification; recovery; solvent; tissue homogenization
  38. Se Pu. 2020 May 08. 38(5): 491-501
      Dispersive liquid-liquid microextraction (DLLME) is a mini-extraction technology with merits of low cost, environment friendliness, and high extraction efficiency; moreover, this method is easy to operate and requires only a short operation time. In this paper, several extraction models, including normal dispersive liquid-liquid microextraction, ionic liquid dispersive liquid-liquid microextraction, and ultrasound-assisted dispersive liquid-liquid microextraction, are reviewed according to the type of extraction solvents and dispersive methods. This paper also summarizes the application of DLLME to the analysis of biological samples in the last five years.
    Keywords:  biological samples; dispersive liquid-liquid microextraction (DLLME); dispersive methods; extraction solvents; review
  39. Se Pu. 2020 Aug 08. 38(8): 906-913
      A method based on liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) was developed for the simultaneous screening and determination of fentanyl and its 26 analogs in liquid and solid powder drugs. The established method involves successive extraction by 5 mL 75% (v/v) acetonitrile aqueous solution and 5 mL acetonitrile, followed by clean-up using the hydrophilic lipophilic balance (HLB) solid-phase extraction method. Detection was achieved by electrospray ionization (ESI) in the positive mode, TOF-MS, and information-dependent acquisition (IDA)-MS/MS acquisition; the external standard method was adopted for quantification. Two databases of accurate mass and fragment ions were created. The standard matrix-matched calibration curves of the 27 target compounds were linear in the range of 5.00-100 μg/L, with the correlation coefficients (r2)>0. 99. The limits of quantification for the 27 target compounds were 10.0 μg/kg. The recoveries for all the target compounds in vitamin C tablet, headache powder, cough syrup, and transdermal patch samples were in the ranges of 82.9%-106%, 84.8%-106%, 86.9%-109%, and 83.1%-106%, respectively, with relative standard deviations ranging from 0.38% to 8.71% (n=6). The results demonstrated that the developed method is rapid and sensitive for the simultaneous monitoring and determination of fentanyl and 26 its analogs in liquid and solid powder drugs.
    Keywords:  drugs; fentanyl and its analogues; high resolution mass spectrometry (HRMS); liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF-MS); screening
  40. Molecules. 2021 Jun 16. pii: 3669. [Epub ahead of print]26(12):
      Saccharides are the most common carbon source for Streptococcus thermophilus, which is a widely used bacterium in the production of fermented dairy products. The performance of the strain is influenced by the consumption of different saccharides during fermentation. Therefore, a precise measurement of the concentrations of saccharides in the fermentation media is essential. An 18-min long method with limits of quantitation in the range of 0.159-0.704 mg/L and with 13C labelled internal standards employing hydrophilic interaction chromatography coupled to mass spectrometric detection-(HILIC-LC-MS) allowed for simultaneous quantification of five saccharides: fructose, glucose, galactose, sucrose, and lactose in the fermentation samples. The method included a four-step sample preparation protocol, which could be easily applied to high-throughput analysis. The developed method was validated and applied to the fermentation samples produced by Streptococcus thermophilus.
    Keywords:  HILIC-LC-MS; Streptococcus thermophilus; fermentation; saccharides
  41. J Chromatogr A. 2021 Jun 15. pii: S0021-9673(21)00469-6. [Epub ahead of print]1652 462345
      Cannabis sativa is commonly used worldwide and is frequently detected by forensic laboratories working with biological specimens from potentially impaired drivers or pilots. To address the problem of limited published methods for cannabinoids quantification in postmortem specimens, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to quantify Δ9-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), 11-nor-9-carboxy-THC (THCCOOH), 8β,11-dihydroxy-THC (8β-diOH-THC), 8β-hydroxy-THC (8β-OH-THC), THC-glucuronide (THC-g), THCCOOH-glucuronide (THCCOOH-g), cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG), Δ9-tetrahydrocannabivarin (THCV), and 11-nor-9-carboxy-THCV (THCVCOOH). Solid phase extraction concentrated analytes prior to analysis on a biphenyl column coupled to a mass spectrometer in electrospray positive ionization mode using multiple reaction monitoring. Linearity ranged from 0.25-50 ng/mL (THC-g), 0.5-100 ng/mL (CBN), 0.5-250 ng/mL (THC, 11-OH-THC, THCCOOH, CBD, and CBG), 1-100 ng/mL (8β-diOH-THC, THCVCOOH, 8β-OH-THC, and THCV) and 1-250 ng/mL (THCCOOH-g). Within-run imprecision was <11.2% CV, between-run imprecision <18.1% CV, and bias was less than ±15.1% of target concentration in blood for all cannabinoids at three concentrations. No carryover or interferences were observed. All cannabinoids were stable in blood at room temperature for 24 h, refrigerated (4°C) for 96 h, and following three freeze/thaw cycles. Matrix effects greater than 25% were observed for most analytes in tissues. The proof of concept for method applicability involved measurement of cannabinoids in a pilot fatally injured in an aviation crash. This new analytical method is robust and sensitive, enabling collection of additional cannabinoid postmortem distribution data to improve interpretation of postmortem cannabinoid results.
    Keywords:  Cannabinoids; Distribution; LC-MS/MS; Postmortem
  42. Se Pu. 2020 Jul 08. 38(7): 782-790
      A method was developed for the simultaneous determination of 16 mycotoxins in drug and food homologous products by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) combined with accelerated solvent extraction (ASE) and QuEChERS. The target mycotoxins in drug and food homologous products were extracted by ASE. After concentration, the extracts were purified by QuEChERS. Then, the target compounds were analyzed by UPLC-MS/MS in both positive and negative electrospray ionization and MRM modes. Aflatoxin B1 and fumonisin B1 were quantified by the internal standard method, and the remaining mycotoxins were quantified by the matrix-matched external standard method. The proposed method showed a good linear relationship, with correlation coefficients greater than 0.99. The limits of detection (LODs) and limits of quantification (LOQs) of the 16 mycotoxins ranged from 0.008 μg/kg to 0.3 μg/kg and from 0.03 μg/kg to 1.0 μg/kg, respectively. The blank samples were spiked at three levels, and the recoveries ranged from 70.8% to 118%, with the RSDs being 2.5% to 10.2%. The developed method was successfully applied to mycotoxin analysis in 30 scutellaria, puerarin and sea buckthorn samples bought from local markets. Different levels of mycotoxins were detected in some of the products. The proposed method is simple, rapid and sensitive, and it can be applied to the simultaneous determination of multi-mycotoxins in drug and food homologous products.
    Keywords:  QuEChERS; accelerated solvent extraction (ASE); drug and food homologous products; mycotoxins; ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)
  43. Metabolites. 2021 Jun 07. pii: 364. [Epub ahead of print]11(6):
      Gut microbiota and their metabolic products are increasingly being recognized as important modulators of human health. The fecal metabolome provides a functional readout of the interactions between human metabolism and the gut microbiota in health and disease. Due to the high complexity of the fecal matrix, sample preparation often introduces technical variation, which must be minimized to accurately detect and quantify gut bacterial metabolites. Here, we tested six different representative extraction methods (single-phase and liquid-liquid extractions) and compared differences due to fecal amount, extraction solvent type and solvent pH. Our results indicate that a minimum fecal (wet) amount of 0.50 g is needed to accurately represent the complex texture of feces. The MTBE method (MTBE/methanol/water, 3.6/2.8/3.5, v/v/v) outperformed the other extraction methods, reflected by the highest extraction efficiency for 11 different classes of compounds, the highest number of extracted features (97% of the total identified features in different extracts), repeatability (CV < 35%) and extraction recovery (≥70%). Importantly, optimization of the solvent volume of each step to the initial dried fecal material (µL/mg feces) offers a major step towards standardization, which enables confident assessment of the contributions of gut bacterial metabolites to human health.
    Keywords:  LCMS (liquid chromatography mass spectrometry); fecal metabolites; gut microbiota; metabolomics; sample preparation