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Related Concept Videos

Mass Spectrometry: Molecular Fragmentation Overview01:20

Mass Spectrometry: Molecular Fragmentation Overview

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The ionization of a molecule into a molecular ion inside the mass spectrometer causes instability in the molecule's structure due to the loss of an electron. This eventually leads to the fragmentation or breaking of some bonds in the molecule. The fragmentation occurs predominantly at specific bonds to yield relatively stable fragments.
One type of fragmentation pattern is the cleavage of a single bond in the molecular ion. The cleavage leads to a radical and a cation. The cleavage can...
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Mass Spectrometry: Carboxylic Acid, Ester, and Amide Fragmentation01:01

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The fragmentation patterns observed for compounds such as carboxylic acids, esters, and amides in the mass spectra include ⍺-cleavage and McLafferty rearrangement. Fragmentation by ⍺-cleavage preferentially occurs at the carbon-carbon bond at the ⍺-position next to the carboxylic group to generate a neutral radical and a cation. Long chain compounds with hydrogen at their γ-carbon undergo McLafferty rearrangement to give a radical cation and a neutral alkene.
For example,...
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Mass Spectrometry: Cycloalkane Fragmentation01:05

Mass Spectrometry: Cycloalkane Fragmentation

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In mass spectrometry, cycloalkanes exhibit distinct fragmentation patterns due to the inherent stability of their molecular ions compared to linear or branched alkanes. The ring structure of cycloalkanes provides additional stability to the molecular ions, often resulting in prominent ion peaks in the mass spectrum.
For example, cyclohexane molecular ions have a mass-to-charge ratio (m/z) of 84, which tends to produce a stronger signal than linear alkanes like hexane. This stability comes from...
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Mass Analyzers: Overview01:13

Mass Analyzers: Overview

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The mass analyzer is a crucial component of the mass spectrometer. In the ionization chamber, the vaporized sample is bombarded with a high-energy electron beam to generate a radical cation and further fragment into neutral molecules, radicals, and cations. A series of negatively charged accelerator plates accelerate the cations into the mass analyzer. The mass analyzer separates ions according to their mass-to-charge (m/z) ratios and then directs them to the detector. The common types of mass...
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Mass Spectrometry: Complex Analysis01:21

Mass Spectrometry: Complex Analysis

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Mass spectrometry is an important technique for the identification of pure compounds. However, it has some limitations for the analysis of complex mixtures, often due to excessive fragmentation making the spectrum too complicated to decipher. Mass spectrometry can be combined with suitable separation methods in sequence, forming hyphenated methods, which are useful in the analysis of complex mixtures.
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...
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Mass Spectrometry: Cycloalkene Fragmentation00:54

Mass Spectrometry: Cycloalkene Fragmentation

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The molecular ions of cycloalkenes undergo fragmentation via a retro-Diels–Alder reaction.
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Collision energy-breakdown curves - An additional tool to characterize MS/MS methods.

Sophie Mörlein1, Carina Schuster1, Michael Paal1

  • 1Institute of Laboratory Medicine, University Hospital, LMU Munich, Germany.

Clinical Mass Spectrometry (Del Mar, Calif.)
|November 25, 2021
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Summary

Collision energy (CE) breakdown curves objectively assess ion yield in tandem mass spectrometry. This method standardizes LC-MS/MS analysis by verifying consistent fragmentation, improving method reliability.

Keywords:
CE, collision energyCLSI, Clinical and Laboratory Standards InstituteCXP, cell exit potentialCollision energy-breakdown curvesDP, declustering potentialISD, internal standardLC-MS/MS, liquid chromatography tandem mass spectrometryLLE, liquid–liquid-extractionMRM, multiple reaction monitoringMethod pre-verificationPP, protein precipitationSIL, stable isotope labeledTandem mass spectrometry (MS/MS)Tes-13C3, testosterone-13C3Tes-d3, testosterone-d3

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Area of Science:

  • Analytical Chemistry
  • Mass Spectrometry

Background:

  • Analyte detection in tandem mass spectrometry relies on collision-induced fragmentation, influenced by collision energy (CE).
  • Empirical optimization of CE, known as "tuning," is common but lacks objectivity.
  • Variations in CE settings cause differential ion yields, impacting quantitative accuracy.

Purpose of the Study:

  • To develop an objective method for evaluating the impact of collision energy settings on analyte ion yield.
  • To establish a systematic approach for optimizing CE in liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods.

Main Methods:

  • Generated collision energy (CE)-breakdown curves by recording numerous mass transitions across a CE range in a single injection.
  • Normalized ion yield against an internal standard with fixed CE and plotted ion yield versus CE.
  • Investigated piperacillin and testosterone in various matrices, exploring sample preparation and internal standard labeling effects.

Main Results:

  • CE-breakdown curves demonstrated characteristic patterns for piperacillin quantifier transitions, varying in maximum ion yield, width, and shape.
  • Divergent curve profiles were noted for piperacillin qualifier transitions.
  • Testosterone analyses showed no significant impact from sample preparation or internal standard labeling on CE selection.

Conclusions:

  • CE-breakdown curves serve as a practical and valuable tool for validating LC-MS/MS methods.
  • This approach ensures consistent fragmentation characteristics between different sample sources and analyte types.
  • Facilitates reliable verification of methods using native and isotope-labeled internal standards.