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Journal of Lipid Research, Vol. 45, 2145-2150, November 2004
Copyright © 2004 by American Society for Biochemistry and Molecular Biology

Methods

Quantitative analysis of lysophosphatidic acid by time-of-flight mass spectrometry using a phosphate-capture molecule

Tamotsu Tanaka^1,*, Hideki Tsutsui^*, Kaoru Hirano^*, Tohru Koike, Akira Tokumura and Kiyoshi Satouchi^*

^* Department of Applied Biological Science, Fukuyama University, Higashimura, Fukuyama, 729-0292, Japan
Department of Functional Molecular Science, Graduate School of Biomedical Sciences, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima, 734-8551, Japan
Faculty of Pharmaceutical Sciences, University of Tokushima, 1-78-1 Shomachi, Tokushima, 770-8505, Japan

¹ To whom correspondence should be addressed. e-mail: tamot{at}fubac.fukuyama-u.ac.jp

Lysophosphatidic acid (LPA) is a lipid mediator that may play an important role in wound healing, embryonic development, and progression of cancer. Here, we report a procedure for the quantification of LPA by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The method is based on a characteristic mass shift with total charge change (from –2 to +1) of the phosphate species due to 1:1 complexation of LPA^2– with a dinuclear zinc (II) complex {1,3-bis[bis(pyridin-2-ylmethyl)amino]propan-2-olato dizinc(II) complex; Zn₂L³⁺} at physiological pH. The monocationic complex [LPA^2–-Zn₂L³⁺]⁺ was detected in the positive mode, in which no other signal of cation adducts of LPA^2– was observed. The detection limit of 18:1 LPA by this method was 0.1 pmol on a sample plate. The intensity ratio of [LPA^2–-Zn₂L³⁺]⁺ against an internal standard [17:0 LPA^2–-Zn₂L³⁺]⁺ increased linearly with their molar ratio. Based on the relative intensities of complex ions, we determined the amounts of LPA homologs in an egg white by this method; the results obtained were in good agreement with those by gas liquid chromatography.

This sensitive and convenient procedure for LPA-specific detection is useful for the quantification of LPA homologs occurring in biological materials.

Abbreviations: DHB, 3,5-dihydroxybenzoic acid; ESI, electrospray ionization; LPA, lysophosphatidic acid; MALDI-TOF MS, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; PC, phosphatidylcholine; THAP, 2,4,6-trihydroxy-acetophenone; Zn₂L³⁺, 1,3-bis[bis(pyridin-2-ylmethyl)amino]propan-2-olato dizinc(II) complex

Supplementary key words egg white • matrix-assisted laser desorption/ionization time-of-flight mass spectrometry • serum

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