bims-metlip Biomed News
on Methods and protocols in metabolomics and lipidomics
Issue of 2020‒12‒27
five papers selected by
Sofia Costa
Cold Spring Harbor Laboratory

  1. J Pharm Biomed Anal. 2020 Dec 08. pii: S0731-7085(20)31709-X. [Epub ahead of print] 113823
      A highly sensitive and accurate electrospray liquid chromatography tandem-mass spectrometry (ESI-LC-MS/MS) method for determination of testosterone in human serum and saliva was developed and validated. Accurate quantification of testosterone in human matrices is essential in diagnosis and management of androgen status in men, women and children, and in forensic investigations of suspected abuse of anabolic androgenic steroids. Chromatography was performed on an HSS-T3 C18 column with a total run-time of 5.5 min. The tandem mass spectrometry was operated in positive electrospray ionization mode with multiple reaction monitoring. Serum and saliva samples of 200 μL, were prepared by solid-phase extraction using a 96-well plate following precipitation with 200 μL methanol. 13C labeled testosterone was used as internal standard for quantification. The standard curve was linear within the range of 4-1000 pg/mL and the limit of quantification of both serum and salivary testosterone was 4 pg/mL. Accuracy were 99-101 % and 93-95 % with between-run imprecision in serum and saliva, respectively, and inter- and intra-assay coefficients of variation were less than 9.2 %. The method proved to be applicable for determination of testosterone over a wide range of concentrations in serum and saliva samples from clinical patients with various androgen disorders, healthy male and female adults as well as from forensic cases.
    Keywords:  LC–MS/MS; Saliva; Serum; Testosterone
  2. J Chromatogr A. 2020 Dec 05. pii: S0021-9673(20)31059-1. [Epub ahead of print]1636 461785
      Metabolomics systematically studies the changes of metabolites in biological systems in the temporal or spatial dimensions. It is a challenging task for comprehensive analysis of metabolomics because of diverse physicochemical properties and wide concentration distribution of metabolites. Used as enrichment sorbents, chemoselective probes, chromatographic stationary phases, MS ionization matrix, nanomaterials play excellent roles in improving the selectivity, separation performance, detection sensitivity and identification efficiency of metabolites when mass spectrometry is employed as the detection technique. This review summarized the recent development of nanoparticle-assisted metabolites analysis in terms of assisting the pretreatment of biological samples, improving the separation performance and enhancing the MALDI-MS detection.
    Keywords:  MALDI imaging; Metabolomics; chemoselective probe; enrichment; nanoparticles; stationary phase
  3. Analyst. 2020 Dec 21.
      Microfluidic chips can perform a broad range of automated fluid manipulation operations for chemical analysis including on-line reactions. Derivatization reactions carried out on-chip reduce manual sample preparation and improve experimental throughput. In this work we develop a chip for on-line benzoyl chloride derivatization coupled to microdialysis, an in vivo sampling technique. Benzoyl chloride derivatization is useful for the analysis of small molecule neurochemicals in complex biological matrices using HPLC-MS/MS. The addition of one or more benzoyl groups to small, polar compounds containing amines, phenols, thiols, and certain alcohols improves reversed phase chromatographic retention, electrospray ionization efficiency, and analyte stability. The current derivatization protocol requires a three-step manual sample preparation, which ultimately limits the utility of this method for rapid sample collection and large sample sets. A glass microfluidic chip was developed for derivatizing microdialysis fractions on-line as they exit the probe for collection and off-line analysis with HPLC-MS/MS. Calibration curves for 21 neurochemicals prepared using the on-chip method showed linearity (R2 > 0.99), limits of detection (0.1-500 nM), and peak area RSDs (4-14%) comparable to manual derivatization. Method temporal resolution was investigated both in vitro and in vivo showing rapid rise times for all analytes, which was limited by fraction length (3 min) rather than the device. The platform was applied to basal measurements in the striatum of awake rats where 19 of 21 neurochemicals were above the limit of detection. For a typical 2 h study, a minimum of 120 pipetting steps are eliminated per animal. Such a device provides a useful tool for the analysis of small molecules in biological matrices which may extend beyond microdialysis to other sampling techniques.
  4. J Sep Sci. 2020 Dec 21.
      Creatinine and purines are gout related metabolites commonly quantified by liquid chromatography coupled with ultraviolet and mass spectrometry. However, the high cost of liquid chromatography coupled with mass spectrometry hindered its extensive use in ordinary hospitals and clinical laboratories. Using the traditional liquid chromatography method, the full separation of these metabolites in complex biological samples is still not achieved. In this study, an improved ultra-high-performance liquid chromatography with ultraviolet spectroscopy method was reported for quantitative determination of five gout related metabolites (i.e. creatinine, uric acid, hypoxanthine, xanthine, and inosine) in human serum within 10 min. A UPLC system equipped with a hydrophilic C18 column was used to improve separation, shorten analysis time, and increase analysis throughput. The performance of the method was validated by evaluating linearity (squared correlation coefficient > 0.9991), recovery (92.8-100.0%, with relative standard deviation < 4.7%), accuracy (relative errors < 14.6%), precision (0.2-4.1% for intra-day and 2.1-7.3% for inter-day) and stability (-14.1-8.3% in auto-sampler for 12 h and -13.3-2.2% for freeze-thaw cycles). This method was successfully applied to quantify gout-related metabolites in serum samples of healthy controls and gout patients, which was expected to be used in the clinical investigation of gout at different stages. This article is protected by copyright. All rights reserved.
    Keywords:  creatinine and purines; gout; human serum; quantification; ultra-performance liquid chromatography