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Current State of Oligonucleotide Characterization Using Liquid Chromatography-Mass Spectrometry: Insight into

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Analytical techniques for oligonucleotide therapeutics are crucial for drug development. This review covers chromatographic methods, mass spectrometry challenges, and impurity analysis for regulatory approval.

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Area of Science:

  • Biochemistry
  • Analytical Chemistry
  • Pharmaceutical Science

Background:

  • Growing interest in oligonucleotide therapeutics necessitates robust analytical methods.
  • Accurate analysis and quantitation of these biomolecules are essential for drug development and regulatory approval.

Purpose of the Study:

  • To review current chromatographic techniques for oligonucleotide analysis.
  • To discuss mass spectrometry (MS) applications and challenges for oligonucleotide therapeutics.
  • To outline strategies for characterizing impurities and degradation products.

Main Methods:

  • Anion exchange chromatography (AEX)
  • Hydrophilic interaction liquid chromatography (HILIC)
  • Ion pair chromatography (IPC)
  • Mass spectrometry (MS)

Main Results:

  • Key advantages and challenges of AEX, HILIC, and IPC are discussed.
  • Colloid formation in specific mobile phases presents an analytical challenge.
  • MS sensitivity issues, including salt adduction and charge state envelopes, are reviewed.
  • Oligonucleotide ionization mechanisms (CRM, IEM, CEM) are analyzed.

Conclusions:

  • Characterization of impurities and degradation products is vital for regulatory approval.
  • Optimal analytical strategies are proposed for determining oligonucleotide-related substances.
  • Future improvements in the field are suggested to enhance oligonucleotide therapeutic analysis.