Simultaneous Enantiomeric Determination of Dansyl-d,l-Phenylalanine by Fluorescence Spectroscopy in the Presence of α-Acid Glycoprotein

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Abstract
Few techniques are amenable to real-time analysis of enantiomers. In this paper, total complexation by α-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 λex/λem = 200/544 nm in the absence of AGP, but show shifted emissions with a component at λex/λem = 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 × 102 L g-1 and 2.29 × 102 L g-1, respectively.