1 Of the possible methods for obtaining enantiomerically pure compounds, chromatographic techniques are most often used for analytical and preparative chiral separation. The mixed library approach involves the synthesis of many library members and subsequent pooling of the members for screening. The disadvantage of this approach is that it is much more difficult to achieve libraries consisting of the extremely large number of members regularly found in mixed libraries.
The resulting chromatograms showed differences in retention times for the library members, indicating that one or more of the library members must interact differently with the two stationary phases. This approach takes advantage of the principle of reciprocity described by Pirkle, Welch, and Lamm. The newer screening method involves the immobilization of the two enantiomers of the racemic analyte of interest separately and the synthesis of a library of potential enantiomerically pure chiral selectors.
An equal amount of the mixture library is allowed to equilibrate with equal amounts of the stationary phases under identical conditions. To assist in recovering the analyte solution from the filter plate after incubation, an Affymax filtration module was used.
DNB-81 Template
Asn Asn
Asn Hyp
Asn Trp
Asn Ser
Asn Thr
Asn Gln
Asn Tyr
Asn Gln 1.150
Asn Tyr- 1.168
The screening of DNB-81 with TBNPO was similar to that of TFAE except 0.8 ml of 0.3mg/ml TBNPO solution in both 1:4 IPA/heptane and 1:4 CHCl3/heptane. The same equilibration times were used as with TFAE: 24 h with 1:4 IPA/heptane and 6 h with 1:4 CHCl3/heptane, and the supernatant was collected by filtration and reconstituted as before. As with TFAE, the TBNPO equilibrations in IPA/heptane (shown in Table 1-4) produced no CSP leads, but when screened in CHCl3/heptane, the library (shown in Table 1-5) had several leads produced.
One of the main members was also a hit in the screening of TFAE: DNB-D-Asn-Thr-Abu-AMPS. This is not surprising since TFAE and TBNPO share some of the same molecular properties that we exploit for potential chiral recognition. This further supports our theory about the influence of solvent choice on the efficiency of CSP due to disruption of hydrogen bonding by protic solvents.
However, in the case of the CHCl3/hexane mobile phase, it would be expected that the potential partition factor of TBNPO using CSP1 would be much higher considering the large difference in the first peak height ratio during screening. Example chromatogram of TBNPO from the f03 equilibrium of library well DNB-81 plotted against the racemic assay.
Asn Trp
Asn Ser
Asn Thr
Asn Tyr
- NA Benzoic acid
Unfortunately, browsing TFAE's 45-member library with a long linker did not yield a significant selector. In the case of TFAE screening of a 45 member library, we had the same problems with reproducibility. Screening the library with silica gel showed no "hits" for TFAE, but the results were reproducible.
Since this procedure appeared to yield more reproducible results, it was used in the screening of the resin-based library. Classically, organic synthesis involved a two-fold strategy: the synthesis step followed by the purification step. Often reactions need to be reoptimized to work on beads due to the heterogeneous nature of the reaction mixture.
In this strategy, an easy isolation process is made possible due to certain properties of the tag. The methylene chloride layers were dried with sodium sulfate and evaporation of the solvent gave a white solid. Included in this list is the terminally terminated version of the tert-butyl protected tyrosine (Figure 3-4).
The chromatographic data indeed correlate well with data in the literature on the binding of various phenolic compounds to beta-cyclodextrin. This experiment demonstrates the potential utility of the 4-t-butylphenyl labeling strategy for solution-phase synthesis processing. The next test to prove the utility of the 4-tert-butylphenyl labeling strategy involves using the label in an actual coupling reaction.
This experiment shows that the tagging motif indeed works in a real synthesis situation, proving the usefulness of the tagging system. These results demonstrate that the carbamate protecting group strategy is a successful way to use the amine functional group of a potential reactant as a site for the facile attachment of the 4-tert-butylbenzyl group. In the case of the described tagging strategy, the most economical way to introduce these ethylene glycol groups is via a linker.
Another thing to note is that the three labels are retained for approximately the same length of time, indicating the stability of the 4-tert-butylphenyl group. While the development of 4-tert-butylphenolic labels was a more straightforward task, there are several considerations that need to be evaluated in the case of lithocholic acid. The alcohol group of the benzyl ester was then protected according to the method shown in Figure 4-3 with a methyl ether group to determine whether the alcohol functional group is an important contributor to the formation of the inclusion complex of lithocholic acid with beta-cyclodextrin. .
The results as shown in Table 4-1 indicate that the alcohol group is not essential for the formation of the inclusion complex and blocking the alcohol functional group actually increases the retention time and thus increases the binding to beta-cyclodextrin.
Compound % Acetonitrile %
