title: Simultaneous determination of selected eicosanoids by reversed-phase HPLC method using fluorescence detection and application to rat and human plasma, and rat heart and kidney samples.
abstract: Eicosanoids are biologically active lipid-derived oxidative metabolites of arachidonic acid. We, herein, present an improved sensitive, selective and robust high performance liquid chromatography (HPLC)-fluorescence assay for simultaneous quantification of eicosanoids in human plasma and rat tissues. Aliquots of 200 μL of plasma or 30 mg of heart or kidney tissues were spiked with 16-hydroxydecanoic acid as internal standard, and extracted with anhydrous acetonitrile using solid phase cartridges. The eluted samples were dried, reconstituted in anhydrous acetonitrile and labeled with 2-(2,3-naphthalimino)ethyl-trifluoromethanesulphonate in the presence of saturated potassium fluoride solution in anhydrous acetonitrile and N,N-diiospropylethylamine as catalyst. The derivatized eicosanoids were extracted with anhydrous acetonitrile using solid phase cartridges. Chromatographic separation was achieved on a C18 reversed phase column using gradient mobile phase of 0.05% of formic acid:acetonitrile:water at 0.8 mL/min flow rate. The analytes were detected at excitation and emission wavelength of 260 and 396 nm, respectively. The assay was linear (r(2)≥ 0.98) in the concentration range of 0.01-2.5 μg/mL. The intra-day and inter-day coefficients variation was less than 19.8%. Using this assay, we were able to quantify arachidonic acid metabolites simultaneously in human and rat biological samples.
authors: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada.
authors: Ali
authors: A
authors: Aghazadeh-Habashi
authors: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada.
authors: Waheed
authors: W
authors: Asghar
authors: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada. Electronic address: fjamali@ualberta.ca.
authors: Fakhreddin
authors: F
authors: Jamali
mesh_terms: D000818:Animals; D001095:Arachidonic Acids / Q000097:blood*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000592:standards; D056148:Chromatography, Reverse-Phase / Q000592:standards; D006801:Humans; D007668:Kidney / Q000737:chemistry*; D057230:Limit of Detection; D016014:Linear Models; D008297:Male; D009206:Myocardium / Q000737:chemistry*; D017207:Rats, Sprague-Dawley; D012015:Reference Standards; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000592:standards
chemical_list: D001095:Arachidonic Acids
keywords: Eicosanoids; Fluorescence detection; High performance liquid chromatography; Human; Rat