Faculty Collaboration Database

Determination of EETs using microbore liquid chromatography with fluorescence detection. Am J Physiol Heart Circ Physiol 2000 Aug;279(2):H857-62

Date

08/03/2000

Pubmed ID

10924087

DOI

10.1152/ajpheart.2000.279.2.H857

Scopus ID

2-s2.0-0033852075 (requires institutional sign-in at Scopus site)   45 Citations

Abstract

Epoxyeicosatrienoic acids (EETs) are cytochrome P-450 metabolites of arachidonic acid involved in the regulation of vascular tone. The method of microbore column high-performance liquid chromatography with fluorescence detection was developed to determine 14,15-EET, 11, 12-EET, and the mixture of 8,9-EET and 5,6-EET. Tridecanoic acid (TA) was used as an internal standard. EETs were reacted with 2-(2, 3-naphthalimino)ethyl trifluoromethanesulfonate (NT) to form highly fluorescent derivatives. A C(18) microbore column and a water-acetonitrile mobile phase were used for separation. Samples were excited at 259 nm, and the fluorescence was detected at 395 nm. The overall recoveries were 88% for EETs and 40% for TA. EETs were detected in concentrations as low as 2 pg (signal-to-noise ratio = 3). The method was used to determine the EET production from endothelial cells (ECs). Bradykinin and methacholine (10(-6) M) stimulated an increase in the production of EETs by ECs two- and fivefold, respectively. This sensitive method may be used for determination of EETs at low concentrations normally detected in complex biological samples.

Author List

Nithipatikom K, Pratt PF, Campbell WB

Author

William B. Campbell PhD Professor in the Pharmacology and Toxicology department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

8,11,14-Eicosatrienoic Acid
Animals
Bradykinin
Cattle
Cells, Cultured
Chromatography, High Pressure Liquid
Coronary Vessels
Endothelium, Vascular
Methacholine Chloride
Microchemistry
Spectrometry, Fluorescence