Thiol-ene click preparation of a [2+4]-type chiral porous organic cage bonded silica stationary phase and its enantioseparation performance in HPLC.

In this study, a tetraaldehyde ligand was prepared via the Duff formylation of 4,4'-methylenediphenol using trifluoroacetic acid and hexamethylenetetramine, which was further condensed with (1R, 2R)-1,2-cyclohexanediamine to produce a [2 + 4]-type chiral porous organic cage (POC). The obtained POC was covalently bonded on thiol-modified silica via thiol-ene click reaction to fabricate a new chiral stationary phase (CSP). This new CSP was tested for chiral separation in normal-phase (NP) and reversed-phase (RP) modes. 15 racemic samples were enantioseparated in NP mode, and 16 in RP mode. Importantly, this new CSP packed column exhibits favorable enantioseparation complementarity with our previously reported two POC columns ( [3 + 6]-type NC1-R and [4 + 6]-type CC19-R columns). It can effectively resolve several racemates that were unresolved or poorly separated on our earlier reported two POC columns, significantly broadening the scope of separable racemates and improving the versatility of POC-based columns. Under NP-HPLC mode, 9 and 8 of the 15 tested racemates achieved no enantiomeric separation on the NC1-R column and CC19-R column, respectively. Under RP-HPLC mode, 11 and 12 of the 16 tested racemates achieved no enantiomeric separation on the NC1-R column and CC19-R column, respectively. The combined use of these chiral POC columns can separate more chiral compounds than any single column used alone. Furthermore, the newly fabricated POC column is also able to resolve a number of racemates that cannot be fully resolved or are unresolvable on two typical commercial chiral HPLC columns, Chiralcel OD-H [CSP: cellulose tris(3,5-dimethylphenylcarbamate)] and Chiralpak AD-H [CSP: amylose tris(3,5-dimethylphenylcarbamate)] columns. In addition, the influences of column temperature, injection mass, and mobile phase composition on enantioseparation were systematically explored. After hundreds injection runs, the relative standard deviations (RSDs) (n = 5) of retention time and resolution values (Rs) were below 0.83% and below 1.39%, respectively. In addition, the RSDs (n = 3) of retention times and Rs values for column-to-column were below 7.06% and 8.33% respectively. These results demonstrate the good repeatability and stability of the prepared column. The present work confirms that the [2 + 4]-type chiral POC holds great potential as a CSP for HPLC, which not only greatly expands the scope and applicability of POC-based CSPs for chiral separation, but also advances the development of POCs as a new category of HPLC CSPs.