Determination of Fatty Acids in Saliva of Smokers and Nonsmokers by HPLC with Fluorescence Detection Using a Hydrazine-Based Difluoro-boraindacene Reagent

1,3,5,7-Tetramethyl-8-daminozide-difluoroboradiaza- s-indacence (TMBODIPY-H), a pre-column fluorescent derivatization reagent, was applied to the analysis of fatty acid by high-performance liquid chromatography with fluorescence detection. Using this reagent, 12 fatty acids from propionic acid (C3) to stearic acid (C18) can be derivatized at room temperature in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide within 2 h. The baseline separation of these derivatives can be achieved within 46 min by gradient elution. The detection limits of these fatty acids are in the range of 0.1-1 nM and the linear ranges of most of them are 0.5-100 nM. This was the first application of hydrazine-based difluoro-boraindacene reagent for the analysis of fatty acids, and the proposed method has been successfully used for the determination of saliva samples of smokers and nonsmokers with recoveries of 88-110 %.