Highly sensitive determination of free fatty acids in human serum by high-performance liquid chromatography with fluorescence detection.

A highly sensitive and rapid high-performance liquid chromatographic method for the determination of free fatty acids in human serum is described. The fatty acids are converted into the corresponding fluorescent derivatives by the reaction with 3-bromomethyl-6,7-dimethoxy-1-methyl-2(1H)-quinoxalinone in the presence of potassium carbonate and 18-crown-6 in acetonitrile. The derivatives are separated simultaneously within 44 min on a reversed-phase column (YMC-Pack C8) with a gradient elution of aqueous methanol and detected fluorimetrically. The detection limits are 0.5-2 fmol in a 10-microliters injection volume. This sensitivity permits precise determination of free fatty acids including lauric, myristoleic and linolenic acids, which occur in serum at very low concentrations, in 5 microliters of sera from healthy subjects and patients with diabetes.