Synthesis of toluene-4-sulfonic acid 2-(2-thiophen-2-yl-phenanthro[9,10-]imidazol-1-yl)-ethyl ester and its application for sensitive determination of free fatty acids in ginkgo nut and ginkgo leaf by high performance liquid chromatography with fluorescence detection.

A novel fluorescent labeling reagent toluene-4-sulfonic acid 2-(2-thiophen-2-yl-phenanthro[9,10-]imidazol-1-yl)-ethyl ester has been designed and synthesized. It was used to label twenty-six fatty acids (C5-C30) successfully in the presence of KCO catalyst in ,-dimethylformamide solvent. The reaction conditions were optimized by employing a three-factor, three-level Box-Behnken design. Derivatives were sufficiently stable to be efficiently analyzed by high-performance liquid chromatography with fluorescence detection. All fatty acid derivatives were separated on a hypersil BDS-C8 column in conjunction with a gradient elution with a good baseline resolution. Good linear correlations were observed for all fatty acids with correlation coefficients > 0.993. The established method exhibited high sensitivity and excellent repeatability. The limit of detection (at a signal-to-noise ratio of 3 : 1) was 8.8-45.5 fmol. The method was used to quantify free fatty acids in ginkgo nut and ginkgo leaf samples with satisfactory results.