title: Determination of protein-bound methionine oxidation in the hippocampus of adult and old rats by LC-ESI-ITMS method after microwave-assisted proteolysis.
abstract: Protein-bound methionine (Met) oxidation has been associated with normal aging and a variety of age-related diseases, including Alzheimer's disease and Parkinson's disease. Monitoring the changes of protein-bound methionine content in the brain in response to normal aging and oxidative stress is of great interest and could be used as an indicator of oxidative stress of rats in pathological conditions. We have developed a rapid analytical method for the determination of oxidized products of protein-bound methionine in rat brain. The assay involved rapid acid proteolysis with microwave irradiation and solid-phase extraction of the free amino acids followed by LC-ESI-ITMS analysis. Detection was achieved in positive ionization with an ion trap mass spectrometer operating in multiple-reaction monitoring mode. The calibration curves of the analytes were linear (r(2) > 0.99) in the range between 0.098 and 1.560 μg/mL. Intra- and inter-day relative standard deviation percentages were <9% and <8%, respectively. The assay performance was sufficient to support a rapid analytical tool for monitoring brain protein-bound methionine oxidation levels. The content of protein-bound Met and methionine sulfoxide (MetO) in the hippocampus of adult and old rats with or without H(2)O(2) treatment was determined by employing the new method. The content of protein-bound MetO was significantly increased in old rats after exposure to H(2)O(2). This result indicates increased sensitivity to Met oxidation in the hippocampus of old rats.
authors.affiliation: Department of Pharmacology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
authors.forename: Li-Hong
authors.initials: LH
authors.lastname: Long
authors.forename: Peng-Fei
authors.initials: PF
authors.lastname: Wu
authors.forename: Xin-Lei
authors.initials: XL
authors.lastname: Guan
authors.forename: Jun-Qi
authors.initials: JQ
authors.lastname: Zhang
authors.forename: You
authors.initials: Y
authors.lastname: Jin
authors.forename: Zui
authors.initials: Z
authors.lastname: Zhang
authors.forename: Yue
authors.initials: Y
authors.lastname: Wang
authors.forename: Yi-Yong
authors.initials: YY
authors.lastname: Li
authors.forename: Jian-Guo
authors.initials: JG
authors.lastname: Chen
authors.forename: Fang
authors.initials: F
authors.lastname: Wang
mesh_terms: D000367:Age Factors; D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D006624:Hippocampus / Q000737:chemistry* / Q000378:metabolism; D006868:Hydrolysis; D008297:Male; D008715:Methionine / Q000737:chemistry* / Q000378:metabolism; D008872:Microwaves; D010084:Oxidation-Reduction; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D011506:Proteins; D008715:Methionine