Simultaneous determination of post-translational racemization and isomerization of N-terminal amyloid-β in Alzheimer's brain tissues by covalent chiral derivatized ultraperformance liquid chromatography tandem mass spectrometry

Anal Chem. 2014 Jan 7;86(1):797-804. doi: 10.1021/ac403315h. Epub 2013 Dec 9.


Typical markers of protein aging are spontaneous post-translational modifications such as amino acid racemization (AAR) and amino acid isomerization (AAI) during the degradation of peptides. The post-translational AAR and AAI could significantly induce the density and localization of plaque deposition in brain tissues. Alzheimer's disease (AD) is reliably related to the formation and aggregation of amyloid-β peptide (Aβ) plaques in the human brain. No current analytical methods can simultaneously determine AAR and AAI during the degradation of Aβ from AD patients. We now report a covalent chiral derivatized ultraperformance liquid chromatography tandem mass spectrometry (CCD-UPLC-MS/MS) method for the determination of post-translational AAR and AAI of N-terminal Aβ (N-Aβ1-5) in human brain tissues. When subjected to tryptic N-Aβ1-5 from post-translationally modified natural Aβ in focal brain tissues by the CCD procedure, it was monitored at m/z 989.6→637.0/678.9 during electrospray collision-induced dissociation. These N-Aβ1-5 fragments with l-aspartic acid (l-Asp), d-Asp, l-isoAsp, and d-isoAsp could be separated using the UPLC system with a conventional reversed-phase column and mobile phase. The quantification of these peptides was determined using a stable isotope [(15)N]-labeled Aβ1-40 internal standard. The CCD-UPLC-MS/MS assay of potential N-Aβ1-5 allowed for the discovery of the present and ratio levels of these N-Aβ1-5 sequences with l-Asp, d-Asp, l-isoAsp, and d-isoAsp.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • Aged, 80 and over
  • Alzheimer Disease / genetics*
  • Alzheimer Disease / pathology
  • Amyloid beta-Peptides / analysis
  • Amyloid beta-Peptides / genetics*
  • Brain / pathology
  • Brain / physiology*
  • Brain Chemistry / physiology*
  • Chromatography, High Pressure Liquid / methods
  • Female
  • Humans
  • Male
  • Protein Processing, Post-Translational / genetics*
  • Tandem Mass Spectrometry / methods*


  • Amyloid beta-Peptides