New method for the quantitative determination of major protein carbonyls, α-aminoadipic and γ-glutamic semialdehydes: Investigation of the formation mechanism and chemical nature in vitro and in vivo

Mitsugu Akagawa, Daisuke Sasaki, Yoshihisa Ishii, Yayoi Kurota, Mari Yamashita, Koji Uchida, Kyozo Suyama

Research output: Contribution to journalArticlepeer-review

45 Citations (Scopus)

Abstract

α-Aminoadipic semialdehyde (AAS) and γ-glutamic semialdehyde (GGS) are identified as the major carbonyl products in oxidized proteins. To elucidate the formation pathway of AAS and GGS in vivo, we developed and validated a new quantification method. AAS and GGS in proteins were derivatized by reductive amination with NaCNBH3 and p-aminobenzoic acid, a fluorescent reagent, followed by acid hydrolysis. It is noteworthy that the fluorescent derivatives were completely stable during acid hydrolysis. The present method permitted the specific, accurate, and sensitive quantification of both semialdehydes by fluorometric high-performance liquid chromatography. Analysis of proteins oxidized by various oxidation systems revealed that AAS and GGS are notably generated by the reaction of proteins with .OH, which is produced by metal-catalyzed oxidation (MCO). Furthermore, exposure of transferrin and human plasma to ascorbic acid and H2O2 significantly promoted the formation of AAS and GGS in vitro, suggesting that both semialdehydes can be generated by MCO in vivo. We also demonstrated their generation through oxidative stress induced by acute iron overload in vivo. In this paper, we describe this analytical technique for simple and precise measurement of AAS and GGS and discuss their formation mechanism in vivo.

Original languageEnglish
Pages (from-to)1059-1065
Number of pages7
JournalChemical Research in Toxicology
Volume19
Issue number8
DOIs
Publication statusPublished - 2006 Aug 1

ASJC Scopus subject areas

  • Toxicology

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