A new methodology for simultaneous quantification of total-Aβ, Aβx-38, Aβx-40, and Aβx-42 by column-switching LC/MS/MS.
This article details the development of a novel method that overcomes the drawbacks of sandwich ELISA (sELISA) and allows reliable evaluation of simultaneous quantification of the amyloid (Aβ)-peptides, total-Aβ, Aβx-38, Aβx-40, and Aβx-42, in rat brain by optimized sample purification and column-switching liquid chromatographic-tandem mass spectrometry (LC/MS/MS). This method provides accurate analyses of total-Aβ, Aβx-38, Aβx-40, and Aβx-42 with a linear calibration range between 0.05 and 45 ng/mL. Verification for accuracy and precision of biological samples were determined by a standard addition and recovery test, spiked with synthetic Aβ1-38, Aβ1-40, and Aβ1-42 into the rat brain homogenate. This method showed <20% relative error and relative standard deviation, indicating high reproducibility and reliability. The brain concentrations of total-Aβ, Aβx-38, Aβx-40, and Aβx-42 after oral administration of flurbiprofen in rats were measured by this method. Aβx-42 concentrations (4.57 ± 0.69 ng/g) in rats administered flurbiprofen were lower than those in untreated rats (6.48 ± 0.93 ng/g). This was consistent with several reports demonstrating that NSAIDs reduced the generation of Aβ. We report here a method that allows not only the quantification of specific molecular species of Aβ but also simultaneous quantification of total-Aβ, Aβx-38, Aβx-40, and Aβx-42, thus overcoming the drawbacks of sELISA.
Pharmacokinetics Research Laboratory, Dainippon Sumitomo Pharmaceutical Co., Ltd., Suita-shi, Osaka, Japan. kenichi-watanabe@ds-pharma.co.jp
Ken-ichi
K
Watanabe
Chihiro
C
Ishikawa
Hiroshi
H
Kuwahara
Kimihiko
K
Sato
Setsuko
S
Komuro
Tetsuya
T
Nakagawa
Naruaki
N
Nomura
Shiro
S
Watanabe
Masashi
M
Yabuki
D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000032:analysis*; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002853:Chromatography, Liquid / Q000379:methods; D005480:Flurbiprofen / Q000494:pharmacology; D008297:Male; D008969:Molecular Sequence Data; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
D016229:Amyloid beta-Peptides; D000894:Anti-Inflammatory Agents, Non-Steroidal; D005480:Flurbiprofen