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Determination of free fatty acids in blood, tagged with 4‐(2‐carbazoylpyrrolidin‐1‐yl)‐7‐(N, N‐dimethylaminosulfonyl)‐2, 1, 3‐benzoxadiazole, by high‐performance liquid chromatography with fluorescence detection
Author(s) -
Toyo'oka Toshimasa,
Takahashi Maki,
Suzuki Asako,
Ishii Yasuko
Publication year - 1995
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.1130090403
Subject(s) - chemistry , chromatography , palmitic acid , high performance liquid chromatography , fluorescence , fatty acid , detection limit , palmitoleic acid , fluorescence spectroscopy , organic chemistry , physics , quantum mechanics
Abstract The free fatty acids in blood were determined by high‐performance liquid chromatography (HPLC) after pre‐column tagging with 4‐(2‐carbazoylpyrrolidin‐1‐yl)‐7‐( N,N ‐dimethylaminosulphonyl)‐2, 1, 3‐benzoxadiazole (DBD‐ProCZ). The tagging conditions were optimized with palmitic acid (C 16:0 ) and linoleic acid (C 18:2 ) as representative free fatty acids, saturated and unsaturated, respectively. Under the mild reaction conditions of room temperature for 90 min in dimethylformamide (DMF) containing 1‐ethyl‐3‐(3‐dimethylaminopropyl)‐carbodiimide hydrochloride (EDC, 0.2 M )/pyridine (2%), all the fatty acids tested were tagged with the DBD‐ProCZ to produce highly fluorescent derivatives which emit light at 550 nm (excitation at 450 nm). The fluorescence wavelenghts were essentially the same for all fatty acids, whereas the intensities were different for individual fatty acids. The derivatives obtained from ten free fatty acids were completely separated by reversedphase chromatography with two isocratic elution conditions. The on‐column detection limit (signal‐to‐noise ratio of 3) with proposed HPLC separation and fluorescence detection is in the range of 19 (palmitic acid) −176 fmol (palmitoleic acid). The free fatty acids in rat serum and human plasma were successfully determined using the present method.