Integrating experimental and computational techniques to study chromatographic enantioresolutions of chiral tetrahydroindazole derivatives

With the selection of partially saturated 2H-indazoles as model compounds, we demonstrate the possibility to use Whelk-O1 chiral stationary phases (CSPs) to succeed in efficient small-scale preparative enantioseparations. Runs of three consecutive liquid chromatography injections (about 300 g of racemate repeatedly injected in a 100 microL loop) produced groups of peaks without band contamination (α = 1.2 and RS = 2.57). With this procedure approximately 3.0 mg of each enantiomer, with enantiomeric excess ≥ 97% were obtained. Very profitably, the high volatility of n-hexane used as the sole eluent facilitated the solvent evaporation after the enantiomer recovery. High resolution mass spectrometry analysis confirmed that the chemical identity of the two enantiomers was preserved along the entire process. The ability of Whelk-O1 phases in enantioseparating structurally similar compounds was confirmed with the analysis of other two racemates. Moreover, the relevant chemoselectivity exhibited by the CSP towards the three racemates should allow to simultaneously optimizing the enantioselectivity of different analytes and perform small-scale enantioresolutions of different compounds during the same run. In this study, the integration of experimental off-line electronic circular dichroism analysis with ab initio time-dependent density-functional theory simulations facilitated the assignment of the absolute configuration of the single enantiomers, while a molecular dynamics protocol can be useful to make a priori predictions of the enantioseparation ability of CSP towards selected compounds.