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Updated: Jan 21, 2026

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics
Published on: April 17, 2017
Application of open port sampling interface mass spectrometry (OPSI-MS) to deuterium exchange as an aid for
Andrew D Ray1, Graeme Clemens2, Stephen W Holman2
1Global Product Development, Pharmaceutical Technology & Development, AstraZeneca, Macclesfield Campus, Macclesfield, SK10 2NA, UK.
Rationale:
Deuterium exchange has been demonstrated to provide additional information to accurate mass measurement and collision-induced dissociation on unknown chemical structures. An enhanced method for rapid deuterium exchange could make this technique more routine for structural elucidation. Open port sampling interface mass spectrometry (OPSI-MS) with an aprotic solvent offers a rapid method for performing deuterium incorporation.
Methods:
Samples of standard drug molecules have been analysed by OPSI-MS directly from solids using a make-up flow of acetonitrile + 0.1% trifluoroacetic acid. The resultant spectra were compared with those obtained by OPSI-MS analysis of the samples dissolved in deuterium oxide (D2 O). Solutions of these molecules in acetonitrile/D2 O were analysed using an Atmospheric Solids Analysis Probe (ASAP) at different temperatures to compare the suitability of this technique.
Results:
The number of exchangeable hydrogens was obtained through deuterium exchange using the OPSI source, although there was some incomplete exchange or back-exchange observed. Molecules with one to five exchangeable hydrogens were successfully analysed. ASAP analysis produced more complicated spectra with higher levels of incomplete or back-exchanged ions; this was more pronounced at higher temperatures.
Conclusions:
The use of OPSI provides a method for the rapid determination of the number of exchangeable hydrogens within a molecule. This yields useful information as an aid to the structural elucidation of unknowns. ASAP produces incomplete exchange and cannot be used for incorporation studies.
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