Abstract
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; Zn(2)L(3+)] at physiological pH. The monocationic complex [LPA(2-)-Zn(2)L(3+)](+) 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-)-Zn(2)L(3+)](+) against an internal standard [17:0 LPA(2-)-Zn(2)L(3+)](+) 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.
Highlights
Lysophosphatidic acid (LPA) is a lipid mediator that may play an important role in wound healing, embryonic development, and progression of cancer
These results suggest that levels of LPA in the body fluids are of clinical importance as a potential biomarker for ovarian cancer progression [10, 11]
An intense ion peak assigned to [18:1 LPA2Ϫ-68Zn2L3ϩ]ϩ was detected at m/z 1,023.3, but neither the protonated molecule of 18:1 LPA2Ϫ nor the molecular ion of its cation adduct was detected in the mass spectrum, as previously reported by Takeda et al [29]
Summary
2,4,6-Trihydroxy-acetophenone (THAP) and 3,5-dihydroxybenzoic acid (DHB) were purchased from Aldrich Chemical Co. (Milwaukee, WI). 2,4,6-Trihydroxy-acetophenone (THAP) and 3,5-dihydroxybenzoic acid (DHB) were purchased from Aldrich Chemical Co. Phos-tag® (68Zn2L3ϩ) was obtained from the NARD Institute Ltd. Arachidonic acid, oleic acid, and linoleic acid were purchased from Serdary Research Laboratories (London, Ontario, Canada). Palmitic acid and margaric acid were from Nacalai Tesque, Inc. Calf serum was obtained from Gibco BRL and Life Technologies, Inc. Phospholipase A2 from Crotalus adamanteus venom and L-␣-glycerophosphorylcholine were obtained from Sigma Chemical Co. The active fraction of phospholipase D was prepared from cabbage using the method described by Davidson and Long [30]. 1-Alkenyl (alkyl)-2-lyso-phosphatidylcholine (PC) was prepared from PC of bovine heart by mild alkaline hydrolysis [31] The active fraction of phospholipase D was prepared from cabbage using the method described by Davidson and Long [30]. 1-Alkenyl (alkyl)-2-lyso-phosphatidylcholine (PC) was prepared from PC of bovine heart by mild alkaline hydrolysis [31]
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