Page 89 - Chiral Separation Techniques

3.6 Library of Cyclic Oligopeptides as Additives to Background Electrolyte … 65
Table 3-1. Values of enantiomeric resolution of DNP-amino acids in a running electrolyte containing
the three fractions 1, 2, and 3 of the cyclo(Arg-Lys-X-Pro-X-β Ala) sublibrary separated by preparative
HPLC.
Resolution
Analyte
c(Arg-Lys-X-Pro-X-β Ala) Fraction 1 Fraction 2 Fraction 3

DNP-D,L-Glu 2.05 1.09 4.05 1.69
DNP-D,L-Ala 0.80 0 5.54 2.45
DNP-D,L-Leu 0 0 3.53 0.89

This led to the conclusion that these amino acids were essential for the resolution
capability and only 6 new libraries of 18 compounds had to be synthesized with
these amino acid residues to define the position 3. Surprisingly, the separation abil-
ities of all six libraries were very similar. Therefore, tyrosine was chosen for contin-
uing deconvolution, since it is convenient as its aromatic ring can easily be detected
by UV spectrometry. The last step, defining position 5, required the synthesis and
testing of 6 individual hexapeptides.

30

25

20
Resolution 15

10

5

0 RKYPYßA
DNP-Glu DNP-Ala DNP-n-Val DNP-h-Ser DNP-Gln DNP-Leu DNP-Asp RKXXXßA
RKYPXßA
RKXPXßA
DNP-Pro

Fig. 3-3. Comparison of the values of enantiomeric resolution of different DNP-D,L-amino acids at dif-
ferent deconvolution stages of a cyclic hexapeptide sublibrary. Resolution values in a cyclo(Arg-Lys-X-
X-X-β-Ala) sublibrary, in the first line, are compared to those obtained in sublibraries with a progres-
sively increasing number of defined positions. All the sublibraries were 30 mM in the running buffer while
the completely defined cyclo(Arg-Lys-Tyr-P-Tyr-β–Ala) peptide is used at 10 mM concentration. Condi-
tions: cyclopeptide sublibrary in 20 mM sodium phosphate buffer, pH 7.0; capillary, 50 µm i.d., 65 cm
total length, 57 cm to the window; V = –20 kV, I = 40; electrokinetic injection, –10 kV, 3 s; detection at
340 nm. (Reprinted with permission from ref. [75]. Copyright 1998, American Chemical Society.)