Simultaneous enantiomeric determination of dansyl-D,L-phenylalanine by fluorescence spectroscopy in the presence of alpha-acid glycoprotein.

Few techniques are amenable to real-time analysis of enantiomers. In this paper, total complexation by alpha-acid glycoprotein (AGP) is shown to discriminate between enantiomers of dansyl-D,L-phenylalanine (DPs) by changing the local environment of the D and L enantiomers (DDP and DLP, respectively) from hydrophilic to hydrophobic. DDP and DLP show the same native fluorescence at lambda ex/lambda em = 200/544 nm in the absence of AGP, but show shifted emissions with a component at lambda ex/lambda em = 220/497 nm in the presence of AGP and in lipophilic solutions. The conditions for an analytical determination have been optimized, and the method has been used to measure the enantiomeric composition of DDP/DLP mixtures with concentration ratios varying over 2 orders of magnitude. The mechanism of chiral recognition for DDP and DLP by AGP is discussed and should be equally applicable to other dansyl-derivative amino acid enantiomers. The association constants for AGP with DDP and with DLP have been determined to be 1.33 x 10(2) L g-1 and 2.29 x 10(2) L g-1, respectively.