title: Quantification of intracellular and extracellular prostanoids stimulated by A23187 by liquid chromatography/electrospray ionization tandem mass spectrometry.
abstract: Prostanoids are bioactive substances that contribute to various biological and pathological processes. To evaluate both extracellular and intracellular levels of prostanoids at the same time, we developed methods for quantification of extracellular and intracellular levels of prostanoids, including prostaglandin E(2) (PGE(2)), PGD(2), PGF(2α), 6-keto PGF(1α), and TXB(2), in cultured cells using liquid chromatography/tandem mass spectrometry (LC/MS/MS), and we validated the LC/MS/MS methods. A solid-phase extraction cartridge was used for extraction of prostanoids. The prostanoids were separated by a C(18) column with an isocratic flow of acetonitrile/water/acetic acid (40:60:0.1, v/v/v). Calibration curves of extracellular measurement for the prostanoids were linear in the range from 0.1 to 100 ng/mL (r(2)>0.999), and those of intracellular measurement were linear in the range from 0.05 to 50 ng (r(2)>0.999). Validation assessment showed that both methods of extracellular and intracellular measurements were highly reliable with good accuracy and precision. We also applied the methods to human airway epithelial Calu-3 cells and human lung adenocarcinoma epithelial A549 cells.
authors.affiliation: Laboratory of Clinical Pharmaceutics & Therapeutics, Division of Pharmasciences, Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-ku, Sapporo, Japan.
authors.forename: Ayako
authors.initials: A
authors.lastname: Furugen
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Yamaguchi
authors.forename: Nobuaki
authors.initials: N
authors.lastname: Tanaka
authors.forename: Hajime
authors.initials: H
authors.lastname: Ito
authors.forename: Kazuaki
authors.initials: K
authors.lastname: Miyamori
authors.forename: Asuka
authors.initials: A
authors.lastname: Fujikawa
authors.forename: Natsuko
authors.initials: N
authors.lastname: Takahashi
authors.forename: Jiro
authors.initials: J
authors.lastname: Ogura
authors.forename: Masaki
authors.initials: M
authors.lastname: Kobayashi
authors.forename: Takehiro
authors.initials: T
authors.lastname: Yamada
authors.forename: Nariyasu
authors.initials: N
authors.lastname: Mano
authors.forename: Ken
authors.initials: K
authors.lastname: Iseki
mesh_terms: D000001:Calcimycin / Q000494:pharmacology; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D045604:Extracellular Fluid / Q000737:chemistry* / Q000187:drug effects; D006801:Humans; D007424:Intracellular Fluid / Q000737:chemistry* / Q000187:drug effects; D016018:Least-Squares Analysis; D011453:Prostaglandins / Q000032:analysis*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013929:Thromboxane B2 / Q000032:analysis
chemical_list: D011453:Prostaglandins; D000001:Calcimycin; D013929:Thromboxane B2