3-BROMOACETYL-7-METHOXYCOUMARIN AS A NEW FLUORESCENT DERIVATIZATION REAGENT FOR CARBOXYLIC-ACIDS IN HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

3-Bromoacetyl-7-methoxycoumarin (I) was synthesized as a highly reactive fluorescent derivatization reagent for use in high-performance liquid chromatographic analysis. Compound I readily reacted with carboxylic acids in acetone at room temperature for 30 min in the presence of potassium bicarbonate and 18-crown-6 (Method A), or an anion-exchange resin (Method B) as catalysts to give the corresponding fluorescent carboxylic acid coumarinacyl esters (II(a-o)). When lauric acid as a test sample was derivatized with I by use of Methods A and B, the detection limits were 0.4 and 0.5 pmol per injection (S/N=3), respectively. These methods were applicable to the measurements of free fatty acids in human plasma (10 - 100 mul). Method B used a weakly basic anion-exchange resin instead of catalysts so in Method A, so it would be useful for the simplification of derivatization.