International Journal of Tryptophan Research

6-Chlorotryptophan possesses unique bioactivity and can be used as a precursor for several bioactive compounds in medicinal chemistry. It was enantioselectively synthesized by condensing 6-chloroindole with racemic N-acetylserine, followed by enzymatic hydrolysis with l-aminoacylase (EC 3.5.1.14). The optical purity was examined by conducting high-performance liquid chromatography with a Cinchona alkaloid-based zwitterionic chiral stationary phase (CSP) [CHIRALPAK® ZWIX(+)], which bears a chiral trans-2-aminocyclohexanesulfonic acid moiety tagged at C-9 of the Cinchona alkaloid. The zwitterionic CSP enabled efficient enantiomeric separations of monosubstituted tryptophan derivatives 1-methyltryptophan, 5-methyltryptophan, 6-methyltryptophan, 5-methoxytryptophan, and 6-chlorotryptophan with a methanol/H2O (98/2) mobile phase containing formic acid (FA) and diethylamine (DEA) additives. The mobile phase contains 25–75 mM FA and 20–50 mM DEA, enabling good separation of the enantiomers of each tryptophan derivative (α > 1.25). Thus, the optical purity of the synthesized 6-chloro- L-tryptophan was easily determined (greater than 99.0%) using HPLC with the zwitterionic CSP.