title: Characterization of lysophosphatidic acid subspecies produced by autotaxin using a modified HPLC ESI-MS/MS method.
abstract: Lysophosphatidic acid (LPA) is a bioactive lipid with a plethora of biological functions including roles in cell survival, proliferation, and migration. Although high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC ESI-MS/MS) technology has been used to measure the levels of LPA in human blood, serum and plasma, current methods cannot readily detect the minute levels of LPA from cell culture. In this study, a modified HPLC ESI-MS/MS method with enhanced sensitivity was developed, which allows accurate measurements of LPA levels with a limit of quantitation at approximately 10 femtomoles. The method was validated by quantitation of LPA levels in the media of previously characterized cell lines ectopically expressing autotaxin. Specifically, autotaxin overexpression induced an increase in the 16:0, 18:2, 18:1, 18:0, and 20:4 subspecies of LPA, but not the 22:6 LPA subspecies. Lastly, this HPLC ESI-MS/MS method was cross-validatedviabiological assays previously utilized to assay LPA levels. Hence, this HPLC ESI-MS/MS method will allow researchers to measurein vitroLPA levels and also distinguish between specific LPA subspecies for the delineation of individual biological mechanisms.
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614 ; Research and Development, Hunter Holmes McGuire Veterans Administration Medical Center, Richmond, Virginia, 23249.
authors.forename: Dayanjan S
authors.initials: DS
authors.lastname: Wijesinghe
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614.
authors.forename: Eric K
authors.initials: EK
authors.lastname: Mayton
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Mietla
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614.
authors.forename: Abir
authors.initials: A
authors.lastname: Mukherjee
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614.
authors.forename: Jinhua
authors.initials: J
authors.lastname: Wu
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614 ; The Massey Cancer Center, Virginia Commonwealth University, Richmond, Virginia, 23298.
authors.forename: Xianjun
authors.initials: X
authors.lastname: Fang
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, 1101 East Marshall Street, P.O. Box 980614, Richmond, Virginia, 23298-0614 ; Research and Development, Hunter Holmes McGuire Veterans Administration Medical Center, Richmond, Virginia, 23249 ; The Massey Cancer Center, Virginia Commonwealth University, Richmond, Virginia, 23298.
authors.forename: Charles E
authors.initials: CE
authors.lastname: Chalfant