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High-Resolution Mass Spectrometry (HRMS)01:15

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Heteronuclear correlation spectroscopy is an analytical technique that investigates the coupling between different types of nuclei, often a proton and an X-nucleus, such as carbon-13 or nitrogen-15. This method is commonly used in nuclear magnetic resonance (NMR) spectroscopy to gain insights into complex chemical compounds' structural and compositional aspects. A typical heteronuclear correlation spectrum displays X-nucleus chemical shifts on one axis and a proton spectrum on the other...
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Double resonance techniques in Nuclear Magnetic Resonance (NMR) spectroscopy involve the simultaneous application of two different frequencies or radiofrequency pulses to manipulate and observe two distinct nuclear spins. One important application of double resonance is spin decoupling, which selectively suppresses coupling with one type of nucleus while observing the NMR signal from another nucleus, simplifying the spectrum and enhancing resolution.
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An HS-MRM Assay for the Quantification of Host-cell Proteins in Protein Biopharmaceuticals by Liquid Chromatography Ion Mobility QTOF Mass Spectrometry
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Streamlined MRM method transfer between instruments assisted with HRMS matching and retention-time prediction.

J J Yang1, Y Han2, C H Mah3

  • 1School of Civil and Environmental Engineering, Nanyang Technological University, 639798, Singapore; Environmental Chemistry and Materials Centre, Nanyang Environment and Water Research Institute, Nanyang Technological University, 637141, Singapore.

Analytica Chimica Acta
|January 29, 2020
PubMed
Summary

This study presents a new platform for transferring multiple reaction monitoring (MRM) methods between mass spectrometry instruments. This approach enhances small molecule analysis without costly authentic standards.

Keywords:
LC-HRMSLC-QqQ-MS/MSMRM method transfersQSRRRetention time prediction

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Area of Science:

  • Analytical Chemistry
  • Mass Spectrometry
  • Environmental Science

Background:

  • Multiple reaction monitoring (MRM) using liquid-chromatography tandem mass spectrometry (LC-QqQ-MS/MS) offers high sensitivity for trace small molecule analysis in complex matrices.
  • The reliance on costly and often unavailable authentic standards limits the widespread adoption of MRM methods.
  • A need exists for practical solutions to enable MRM method transfer and facilitate small molecule quantification without direct standard use.

Purpose of the Study:

  • To develop a practical platform for transferring MRM methods between different LC-QqQ-MS/MS instruments.
  • To enable confident small molecule identification and quantification without the need for authentic standards.
  • To leverage high-resolution mass spectrometry (LC-HRMS) and retention time prediction for method transfer.

Main Methods:

  • Developed a platform integrating LC-HRMS and retention time (RT) prediction for MRM method transfer.
  • Utilized accurate mass measurements from LC-HRMS and MS/MS fragments from literature.
  • Employed retention time matching, peak matching, and a quantitative structure retention relationship (QSRR) model for compound identification.
  • Validated the platform's robustness using spiked environmental chemicals in various complex matrices (sludge water, urine, cell extracts).

Main Results:

  • The platform successfully transfers MRM methods between different LC-QqQ-MS/MS instruments.
  • Integration of LC-HRMS and RT prediction allows for confident small molecule identification without authentic standards.
  • The QSRR model, based on random forest feature selection, achieved a Pearson r² of 0.63 for RT prediction.
  • Robustness was demonstrated across diverse sample types, including environmental, biological, and cellular matrices.

Conclusions:

  • The developed platform provides a practical and cost-effective solution for MRM method transfer.
  • This approach significantly reduces the dependency on expensive and scarce authentic standards for small molecule analysis.
  • The validated platform enhances the reliability and accessibility of trace-level small molecule quantification in complex samples.