A novel approach for oxidation analysis of therapeutic proteins

Iva Turyan*, Nikhil Khatwani, Zoran Sosic, Shiranthi Jayawickreme, Daniel Mandler

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

4 Scopus citations


Measuring and monitoring of protein oxidation modifications is important for biopharmaceutical process development and stability assessment during long-term storage. Currently available methods for biomolecules oxidation analysis use time-consuming peptide mapping analysis. Therefore, it is desirable to develop high-throughput methods for advanced process control of protein oxidation. Here, we present a novel approach by which oxidative protein modifications are monitored by an indirect potentiometric method. The method is based on adding an electron mediator, which enhances electron transfer (ET) between all redox species and the electrode surface. Specifically, the procedure involves measuring the sharp change in the open circuit potential (OCP) for the mediator system (redox couple) as a result of its interaction with the oxidized protein species in the solution. Application of Pt and Ag/AgCl microelectrodes allowed for a high-sensitivity protein oxidation analysis. We found that the Ru(NH3)62+/3+ redox couple is suitable for measuring the total oxidation of a wide range of therapeutic proteins between 1.1 and 13.6%. Accuracy determined by comparing with the known percentage oxidation of the reference standard showed that percentage oxidation determined for each sample was within ±20% of the expected percentage oxidation determined by mass spectrometry.

Original languageAmerican English
Pages (from-to)108-113
Number of pages6
JournalAnalytical Biochemistry
StatePublished - 1 Feb 2016

Bibliographical note

Publisher Copyright:
© 2015 Elsevier Inc. All rights reserved.


  • Indirect potentiometry
  • LC-MS
  • Oxidation analysis
  • Therapeutic proteins
  • Water-soluble raw materials composed of tween


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