Chiral analysis of β-alanyl-D,L-tyrosine and its derivatives and estimation of binding constants of their complexes with 2-hydroxypropyl-β-cyclodextrin by capillary electrophoresis

Chiral CE methods were developed for the elucidation of L- or D-configuration of tyrosine residue in antimicrobial dipeptide beta-alanyl-tyrosine (beta-Ala-Tyr) isolated from the hemolymph of larvae of fleshfly Neobellieria bullata and for the evaluation of enantiopurity of its synthetic isomers (beta-Ala-D-Tyr and beta-Ala-L-Tyr), and enantiomers of their amidated and acetylated derivatives, beta-Ala-D,L-Tyr-NH2 and N-Ac-beta-Ala-D,L-Tyr, respectively. Baseline separations were achieved for all three pairs of enantiomers: (i) for beta-Ala-D,L-Tyr in acidic background electrolyte composed of 32/50 mM tris(hydroxymethyl)aminomethane/H3PO4, pH 2.5, and 20 mg/mL 2-hydroxypropyl-beta-cyclodextrin as chiral selector; (ii) for beta-Ala-D,L-Tyr-NH, enantiomers in acidic background electrolyte consisting of 48/50 mM tris(hydroxymethyl)aminomethane/H3PO4, pH 3.5, and 30 mg/mL 2-hydroxypropyl-beta-cyclodextrin; and (iii) for enantiomers of N-Ac-beta-Ala-D,L-Tyr in alkaline background electrolyte composed of 50/49 mM Na2B4O7/NaOH, pH 10.5, and 60 mg/mL 2-hydroxypropyl-beta-cyclodextrin. From CE analyses of mixed samples of isolated beta-Ala-Tyr and synthetic standards beta-Ala-L-Tyr and beta-Ala-DTyr, it turned out that isolated beta-Ala-Tyr was pure L-enantiomer. In addition, the average apparent binding constants, K-b, and average actual ionic mobilities of the complexes of beta-Ala-D,L-Tyr and its above derivatives with 2-hydroxypropyl-Acyclodextrin were determined. These complexes were weak, with K-b values ranging from 11.2 to 79.1 L/mol. Their cationic mobilities were equal to (5.6-9.2) x 10(-9) m(2)/V/s, and anionic mobilities to (-1.3-1.6) x 10(-9) m(2)/V/s.

Automated summary

Chiral CE methods were used to separate and identify the configuration of the tyrosine residue in β-Ala-Tyr, and to evaluate the enantiopurity of its synthetic isomers and derivatives. The results showed that the isolated β-Ala-Tyr was pure L-enantiomer.