Enantioselective Tail‐to‐Head Terpene Cyclizations by Optically Active Hexameric Resorcin[4]arene Capsule Derivatives

Abstract Molecular capsules enable the conversion of substrates inside a closed cavity, mimicking to some extent enzymatic catalysis. Chirality transfer from the molecular capsule onto the encapsulated substrate has been only studied in a few cases. Here we demonstrate that chirality transfer is possible inside a rather large molecular container of approximately 1400 Å3. Specifically, we present 1) the first examples of optically active hexameric resorcin[4]arene capsules, 2) their ability to enantioselectively catalyze tail‐to‐head terpene cyclizations, and 3) the surprisingly high sensitivity of enantioselectivity on the structural modifications.


General Information
Experimental: All reactions were carried out using standard SCHLENK techniques with Argon (Ar 4.6) as the inert gas. Unless indicated otherwise, glass equipment was dried under a high vacuum (10 −2 mbar) at 500-600 °C using a heat gun. Reactions at low temperatures were performed using cooling baths (−78 °C using dry ice/iPrOH, 0 °C using ice/water). Transfer of liquids with a volume ranging from 1 to 10 μL or from 10 to 100 μL was performed with a microman M1 pipette (Gilson, systematic error: 1.40% − 1.60%) equipped with 10 μL or 100 μL pipette tips, respectively.
Solvents: Anhydrous solvents were purchased from ACROS; solvents for extractions, chromatography, filtrations, and non-anhydrous reactions were purchased from VWR as HPLC grade solvents; NMR solvents were purchased from CAMBRIDGE ISOTOPE LABORATORIES. All solvents were used as received without prior purification.

NMR-Spectroscopy:
DOSY-NMR experiments were performed on a BRUKER Avance III HD four-channel NMR spectrometer operating at 600.13 MHz proton frequency. The instrument was equipped with a cryogenic 5mm four-channel QCI probe (H/C/N/F) with a self-shielded z-gradient. The experiments were performed at 298 K, and the temperature was calibrated using a methanol standard showing accuracy within +/-0.2 K. For the PFGSE (pulsed-field gradient spin echo)

General Information
S4 diffusion experiment, the sample was placed in a 3 mm outer diameter tube, which was then inserted in a standard 5 mm round-bottom tube and securely kept in place as depicted below.
This setup ensured a negligible temperature gradient on the sample even inside a cryogenic probe. The PFGSE experiments were performed using a bipolar gradient pulse sequence. [1] The sigmoidal intensity decrease was fitted with a two-parameter fit (I0 and diffusion coefficient D) with the DOSY routine implemented in topspin 3.6.1 [BRUKER Biospin GmbH]. A typically observed intensity decrease is depicted below.
High-Performance Liquid Chromatography: HPLC was performed on an LC Prominence Liquid Chromatograph system by SHIMADZU equipped with a UV-Vis detector and an ELSD 5  GC Analysis: GC analyses were carried out on a SHIMADZU GC-2010 Plus instrument equipped with an FID detector and an Rtx-5 capillary column (length = 30 m). Hydrogen was used as the carrier gas, and the constant flow mode was used (flow rate = 40 mL/min) with a split ratio of 1:20. The following temperature program was used: 60 °C for 3 min, 15 °C min -1 to 250 °C, and 250 °C for 5 min.
For the determination of enantiopurity via GC, a SHIMADZU GC-2010 Plus instrument equipped with an FID detector and an Rt-bDEXsm capillary column (length = 30 m) was used. Hydrogen was used as the carrier gas, and the constant-flow mode was used (flow rate = 50 mL/min) with a split ratio of 1:20. The following temperature program was used: 60 °C for 1 min, 5 °C min -1 to 70 °C, 70 °C for 10 min, 5 °C min -1 to 220 °C, 220 °C for 10 min.

ESI-HRMS:
High-resolution mass spectra were obtained using the electrospray ionizationtime of flight (ESI-TOF) technique on a BRUKER maXis 4G mass spectrometer.

ECD Spectroscopy of the Obtained Compounds Experimental ECD Spectra
The following ECD spectra of compounds 2a-c and 3a-c (1mM) were measured at 293 K in chloroform (self-assembled capsules), and in chloroform with 5000 eq. of methanol (selfassembly suppressed). All compounds display ECD bands in the range of 250-320 nm. For  Figure S13. Comparison of ECD spectra of (-)-3a-c in chloroform with 5000 eq. of methanol.

Calculated ECD Spectra
Experimental ECD spectra of compounds (−)-3a-c have been compared with TD-DFT calculations performed for M-(−)-3d, a truncated version of compounds (−)-3a-c with methyl residues instead of longer alkyl chains. As common practice, one should consider different conformers of given molecule if a fluxional nature at the experimental temperature is expected and, eventually, consider a Boltzmann averaging of the results to reproduce the experimental observations. For this compound, molecular mechanics (GHEMICAL force field) optimizations calculated interactively using Avogadro provided evidence of the existence of two relatively stable conformers in vacuum, a crown and a chair conformation (see Fig. S14). These two conformers have been subsequently optimized at the B3LYP level with Grimme's D4 corrections, [5] a def2-TZVP basis set, using implicit chloroform within the Conductor-like Polarizable Continuum Model (CPCM) [6] (see Appendix for XYZ input files). All calculations were conducted using the ORCA 5.0 code. [7] The crown conformer resulted to be 35 kJ/mol more stable than the chair one and their Boltzmann factor at room temperature is chair:crown=7 x 10 -7 . Thus, we can conclude that the only relevant conformation for calculating the spectrum is the crown one.
Using the B3LYP optimized structure we calculated the ECD spectrum using TD-DFT formalism and the ωB97X functional, the def2-TZVP basis set and CPCM implicit chloroform. The vertical excitations have been subsequently post-processed using the SpecDis 1.71 program [8] using a Gaussian broadening factor of 0.16 eV a scaling of the relative intensities of 0.08. The whole spectrum has been red-shifted of 40 nm to better match the experimental one, as common practice in the comparison of ECD calculated spectra. [9] The comparison of the experimental and calculated spectra is reported in Fig. S15. The HCl stock solution was kept in the fridge, and the titration was repeated immediately before each use.

Determination of the water content of the resorcin[4]arenes:
The water content of each capsule was determined via 1 H-NMR measurements using tetraethylsilane (TES) as the internal standard. The corresponding resorcin [4]arene (0.01 mmol, 6 eq., 20 mM) was dissolved in 450 μl of CDCl3, and TES was added (50 L, 33.4 mM, 634 g, 1.67 mol, 1.0 eq.). The water content of CDCl3 was determined in a separate experiment with TES as the internal standard.
The water content of CDCl3 was deducted from the water content of the capsule experiment, to arrive at the water content of the resorcin [4]arene.

S29
followed by the addition of n-decane (1.34 μL, 6.89 μmol, 1.5 eq.). Additional chloroform was added to bring the total volume to 137.5 μL. An aliquot (2 μL) of the reaction mixture was diluted with 0.3 mL of n-hexane (containing 0.08% DMSO). The diluted sample was put in the freezer for at least 15 min, centrifuged to remove the corresponding resorcin [4]arene, and subjected to GC analysis (initial sample). HCl stock solution in chloroform (0.03 eq.) was added to the reaction mixture, and the mixture was briefly shaken. The sealed GC vial was kept at 30 °C (±1 °C) using a thermostated heating block made from aluminum. Further samples were taken at the indicated times and analyzed by gas chromatography. Conversions and yields were calculated by employing the equations described in our previous work [10] and plotted against time (see profiles for the nerol cyclization reactions inside supramolecular capsules based on resorcin [4]arenes 2a-c and 3a-c below; only chiral products are depicted). Enantiomeric excesses were determined with chiral GC measurements (see Table 2 of the manuscript).

Observation of the enantiomeric excess of limonene and α-terpineol over time for the nerol cyclization reaction with (-)-3b
No significant changes were observed. In previous work, we demonstrated that both compounds will convert further to achiral products. [10] Therefore, some small changes in ee over time are expected.

Determination of the Products Configuration
The configuration of the formed cyclization products, limonene and α-terpineol, was determined by chiral GC measurements. As references, the commercially available racemic mixture and the enantiopure sample were analyzed and compared with chromatograms of the reaction mixture. Chiral GC traces of commercial samples of limonene and α-terpineol and the cyclization reactions of NOH inside the enantioenriched capsules based on resorcin [4]arene 3b are shown in Fig. S25 and S26.

Cyclization Reactions in Chloroform with Different Water Content
To investigate the influence of the water content on the reaction, the cyclization of nerol was performed under three different conditions (6.3, 8.0, and 15.4 eq. of water/capsule).
Resorcin [4]arene (−)-3b containing 6 eq. of water per capsule in CDCl3 of varying water content (determined by NMR with TES as internal standard, see chapter 4.1) was used.
 Dry CDCl3 was prepared by drying the commercial CDCl3 with activated molecular sieves.
 Water saturated CDCl3 was prepared according to the following procedure: to 5 mL of CDCl3 0.1 mL of Milli-Q water was added. The mixture was sonicated for 10 seconds and then left standing until complete phase separation. A clear chloroform phase was then decanted to a clean vial.
 Reactions in both dry and water saturated CDCl3 were compared to the reaction in commercially available CDCl3, which was used for all other cyclization reactions described in this work. The total water content for the three different conditions is reported in Table S2.

Cyclization Reactions at Low Temperature
Reactions at 4 °C were performed in a fridge according to the General Procedure described in Section 4.1.