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Improved peptide mapping using phytic acid as ion-pairing buffer additive in capillary electrophoresis

T Kornfelt1, A Vinther, G N Okafo

  • 1Novo Nordisk A/S, Gentofte, Denmark.

Journal of Chromatography. A
|March 1, 1996
PubMed
Summary

Researchers digested bovine pancreatic trypsin inhibitor (BPTI) and separated fragments using capillary electrophoresis. Phytic acid, an ion-pairing additive, was found to influence peptide mobility based on charge, enabling manipulation of separation order.

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Area of Science:

  • Proteomics and Analytical Chemistry
  • Biochemical Separation Techniques
  • Protein Characterization

Background:

  • Aprotinin and bovine pancreatic trypsin inhibitor (BPTI) are basic polypeptides.
  • Effective separation of peptide fragments is crucial for protein analysis.

Purpose of the Study:

  • To investigate the separation of endoproteinase Lys-C digested BPTI fragments using free solution capillary electrophoresis.
  • To evaluate the impact of phytic acid as an ion-pairing agent on peptide fragment separation.

Main Methods:

  • Digestion of BPTI with endoproteinase Lys-C after unfolding, reduction, and pyridylethylation.
  • Separation of resulting peptide fragments by free solution capillary electrophoresis in a phosphate buffer at neutral pH.
  • Investigation of phytic acid's effect as an ion-pairing buffer additive.

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Main Results:

  • Five peptide fragments were obtained from BPTI digestion.
  • Phytic acid influences the electrophoretic mobility of positively charged peptide fragments through ion-pair formation.
  • Peptide affinity for phytic acid correlates with isoelectric point and charge-to-mass ratios.

Conclusions:

  • Phytic acid effectively modulates peptide fragment mobility in capillary electrophoresis.
  • Adjusting phytic acid concentration allows for manipulation of peptide migration order and separation.
  • This method offers a strategy for optimizing peptide separations in complex mixtures.