Abstract: A simple and sensitive method for the determination of long-chain free fatty acids (FFAs) using 1-［2-(p-toluenesulfonate)ethyl］-2-phenylimidazole ［4,5-f］ 9,10-phenanthrene (TSEPIP) as pre-column derivatization reagent by reversed-phase high performance liquid chromatography with fluorescence detection and mass spectrometric identification was developed. Eleven long-chain (C20-C30) FFA derivatives were separated on Eclipse XDB-C8 column with a good baseline resolution. Studies on derivatization conditions indicate that FFA reacts rapidly and smoothly with TSEPIP in the presence of K2CO3 catalyst at 90 ℃ in N, N-dimethylformamide solvent to give the corresponding sensitively fluorescent derivatives with maximal yields close to 100% with a 5-fold molar reagent excess. The identification of 11 FFA derivatives was carried out by online post-column mass spectrometry with atmospheric pressure chemical ionization source in positive-ion mode. The contents of 11 FFAs in soil and three bryophytes (Homomallium connexum (Card.) Broth., Actinothuidium hookeri, Neckera pennata) were determined. The results indicate that the bryophyte plants enrich an abundance of FFAs from soil. The excitation and emission wavelengths of fluorescence detection were set at 260 nm and 380 nm, respectively. Linear correlation coefficients for the most FFA derivatives were higher than 0.9996, and the detection limits (at signal-to-noise of 3∶1) were 26.19-76.67 fmol. The method is sensitive and reproducible for the determination of long-chain FFAs in real samples.

Key words: 2.College of Chemistry Science, Qufu Normal University, Qufu 273165, China, 3.Graduate School of the Chinese Academy of Sciences, Beijing, 1.Northwest Plateau Institute of Biology, the Chinese Academy of Sciences, Xining 810001, China