METHYLATED N-EPSILON-DANSYL-L-LYSINE AS A FLUOROGENIC REAGENT FOR THECHIRAL SEPARATION OF CARBOXYLIC-ACIDS

Dansyl amino acids having a free amino group and an asymmetric carbon atom were examined as a labeling reagent for chiral compounds containi ng carboxylic moieties to realize enantiomeric separation as well as f luorimetric determination. We tested dansyllysine by reacting it with (+)- and (-)-ibuprofen as a model carboxylic enantiomer. As the intram olecular carboxylic moiety of the dansyl amino acid interfered in the condensation reaction to form an amide bond with the carboxylic acid, the moiety was masked by methylation with trimethylsilyldiazomethane b efore the reaction. These derivatives were reacted with (+)- and (-)-i buprofen and better enantiomeric resolution was achieved with methylat ed dansyllysine on a reversed-phase column. The derivatisation reactio n was facilitated by the use of catalysts that are commonly employed i n peptides synthesis. The reaction was completed within 5 min at room temperature when diethyl phosphorocyanidate was used. Due to the dansy l moiety, methylated dansyl-lysine enables a sensitive determination o f ibuprofen with a fluorescence detector, in addition to the capabilit y of enantiomer resolution. In tests, the detection limits for (+) and (-)-ibuprofen were 4 pmol per injection (S/N=3) at an excitation wave length of 340 nm and an emission wavelength of 523 nm. Linear response s for the determination of (+) and (-)-ibuprofen in human urine were a lso demonstrated (r greater than or equal to 0.998) in the range from 10 to 1000 ng/ml. The precision and accuracy for urine samples spiked with (+)- and (-)-ibuprofen at 10, 100 and 1000 ng/ml were <10.1 and < 14.6% (n=4), respectively. (C) 1998 Elsevier Science B.V. All rights r eserved.