The role of LiOTf in the formation of 1 is worthy for further investigation. Similar to other template-induced macrocyclization reactions, we envision Li ion pre-organizes an acyclic furylmethane tetramer (7) to furnish a more efficient ring-closing reaction to form 1. The simulated structure of 7•Li exhibits a tetradentate structure with all four furyl groups simultaneous To verify this experimentally, we synthesized and isolated 7 and subjected it to NMR titration experiment between 7 and LiOTf in CD3CN (same condition as formation of 1) (Scheme 2a). However, the derived 1:1 binding constant (Ka) for LiOTf is 0.21 M -1 (Figure ), which is too weak to provide any template effect that improves the chemical yield of 1. On the contrary, the electrostatic potential (ESP) map , the calculated binding energy and distortion energy (Table ) reveal a highly favorable binding in 7•Li complex, due to the high positive charge density in Li ion (Scheme 2b), and is thus consistent with our experimental result (Table , entry 5 vs. 13/14). The contrasting data from titration and calculated binding energy suggest that Li-template effect is a kinetic phenomenon. Thermodynamically, 7•Li is unfavored. However, once it forms, it rapidly condenses with 6a to give 1. When 7 was applied to macrocycle formation condition, 1 was obtained in 38.4% with LiOTf, but only 20.0% without it, highlighting the critical role of an appropriate Li template (Figure ). With sufficient amount of 1 in hand, we turned our attention to its post-synthetic transformation to pillar [4]naphthalene (2a). When (trimethylsilyl)phenyl trifluoromethanesulfonate (8a) was added to a reaction mixture of 1 and CsF (5 equiv.) in CH3CN/CH2Cl2 (8:5) at 40 °C for 16 h, cycloaddition product, pillar [4](1,4-epoxynapthalene) (4a), was smoothly obtained as a white solid in 92% yield (Figure ). 1 H NMR spectrum of crude product 4a showed a mixture of species (Figure ), suggesting that 4a exists in multiple conformations. In principle, four approach trajectories of benzyne to 1 are possible that would lead to epoxyl group pointing either inside or outside with respect to the resulting macrocycle as well as pointing up or down (see section S5e in ESI). After taking consideration of both symmetry and steric, this results in a potentially 18 different conformations of 4a. Our speculation was then confirmed by reverse phase analytical HPLC experiments. In the LC chromatogram, multiple broad peaks were observed with different response factors (Figure ). We next attempted to isolate different fractions by column chromatography. Interestingly, although most fractions we isolated contain more than one species, MALDI-TOF spectra of these fractions as well as crude 4a show the exclusive formation of 4a, which unambiguously confirmed the formation of different conformations of 4a under reaction condition (Figure ). Fortunately, we isolated one fraction that contains a single product, confirmed by HPLC chromatogram of the isolated species (Figure ). Based on NMR spectroscopic analysis (Figure ), we assigned this isolated isomer 4a with three epoxy groups pointing up and one pointing down as well as all epoxy groups pointing inside the ring (BBBD conformation, see section S5e in ESI for detailed assignments). In order to evaluate the energetics of different conformers of 4a, a DFT-conformational scan was first performed to determine all possible conformers of 4a, and each conformer was then subjected to single point calculation under M06-2X /def2-TZVP level theory together with Grimme's D3 dispersion correction for more accurate energy. The result is summarized in Table . The lowest energy conformer 4a is with an alternate exo-endo-exo-endo sequence (BDBD conformation), with energy >5 kcal/mol lower than the next stable conformer (BBBD).