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Thiol-ene Monolithic Pepsin Microreactor with a 3D-Printed Interface for Efficient UPLC-MS Peptide Mapping Analyses.

Alexander Jönsson1, Rasmus R Svejdal1, Nanna Bøgelund1

  • 1Department of Pharmacy, Copenhagen University , Universitetsparken 2, Copenhagen E DK-2100, Denmark.

Analytical Chemistry
|March 22, 2017
PubMed
Summary

This study introduces a polymer microfluidic chip for peptide mapping, offering a faster and cheaper alternative to conventional methods. The new immobilized enzyme reactor (IMER) shows comparable digestion efficiency for LC-MS workflows.

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Area of Science:

  • Analytical Chemistry
  • Biochemistry
  • Materials Science

Background:

  • Peptide mapping using liquid chromatography-mass spectrometry (LC-MS) is crucial for protein analysis.
  • Conventional enzymatic digestion methods are time-consuming and costly.
  • There is a need for improved sample handling and reduced preparation time in LC-MS workflows.

Purpose of the Study:

  • To investigate a polymer microfluidic chip-based enzyme reactor as a replacement for conventional enzymatic digestion.
  • To evaluate the digestion efficiency of a thiol-ene based immobilized enzyme reactor (IMER).
  • To assess the direct interfaceability of the microfluidic chip with LC-MS.

Main Methods:

  • Development of a polymer microfluidic chip utilizing off-stoichiometric thiol-ene as bulk material and stationary phase.
  • Immobilization of the proteolytic enzyme pepsin onto the thiol-ene based stationary phase.
  • Comparison of the thiol-ene based IMER with a conventional agarose packed bed pepsin IMER column.
  • Evaluation of digestion efficiency at comparable residence times.
  • Integration with LC-MS using a 3D-printed interface.

Main Results:

  • The microfluidic chip IMER demonstrated comparable digestion efficiency to conventional agarose columns.
  • The thiol-ene based IMER offers potential for reduced cost and preparation time.
  • The chip IMER is directly interfaceable with LC-MS via a 3D-printed interface.

Conclusions:

  • Polymer microfluidic chip-based enzyme reactors are a viable alternative to conventional methods for peptide mapping.
  • This technology can improve sample handling, reduce costs, and decrease preparation time in LC-MS workflows.
  • The developed IMER system shows promise for routine protein analytical research.