Table . Model glycosylation reactions. The stereoselectivity of the reaction is expressed as α:β and based on 1 H-NMR of purified α/β-product mixtures. Bluecolored cells represent α-selectivity, while orange-colored cells represent β-selectivity. The percentage given between brackets represents the yield after purification by column chromatography. Pre-activation based glycosylation conditions: donor 1-4 (1 eq), Tf2O (1.3 eq.), Ph2SO (1.3 eq.), TTBP (2.5 eq.), DCM (0.05 M), -80 to -60 °C, then add nucleophile (2 eq.) at -80 °C and allow to warm to -40 °C. Next, we investigated the stability of the glucosyl aand mannosyl 1,3-dioxanium ions using ion IRIS and computational chemistry (Figure ) as we have previously found the relative stability of the dioxanium ions with respect to the parent oxocarbenium ions to correlate to the stereoselectivity of the acylated glycosyl donors. To aid in spectroscopic elucidation we used 2,3-di-O-methyl-4,6-ethylidene glucose ( ) and mannose ( ) donors. To obtain the cations of the sulfoxide donors, the proton adducts were generated by electrospray ionization (ESI+) and isolated in a Bruker AmaZon Speed ion trap (MS spectra can be found in Supplementary figures 1 and 2). Subsequently, the sulfoxide leaving group was expelled by collision induced dissociation (CID) to generate the glycosyl cations. An IR spectrum of the isolated cations was measured using the freeelectron laser FELIX in the 600-1900 cm -1 range by monitoring the wavelength-dependent IR multiple photon-induced dissociation (IRMPD) yield. The relative stability of the cations was assessed through a density functional theory (DFT) protocol in which the conformational energy landscape (CEL) maps for these cations were generated This method maps the energy of the glycosyl cations as a function of their shape by probing the complete conformational space that these cations can occupy. We plotted the relative stability of the dioxanium ion vs the oxocarbenium ion for the glucosyl (Figure ) and the mannosyl (Figure ) cations. The CEL maps show that both the glucosyl and mannosyl 1,3-dioxanium ions are significantly more stable than their oxocarbenium ion counterparts, with a larger difference being found for the mannosyl ions (+9.7 kcal mol -1 vs +5.7 kcal mol -1 ). The identified lowest energy conformers of the dioxanium and oxocarbenium cations were in accordance with local minima identified through an independent protocol, and were subsequently used to compute reference IR-spectra to aid in interpretation of the experimentally obtained spectra. Figure /b show the computed spectra (filled) and the experimental spectra (black line) to confirm the formation of the dioxanium ions, as indicated by a characteristic dioxanium O-[C-O] + stretch (∼1550 cm -1 ) and the absence of the oxocarbenium [C1=O5] + (∼1565 cm -1 ) and benzoyl C=O (∼1775 cm -1 ) stretches. Overall, these results show that in the gas phase, in the absence of counterions, both the 3-acyl glucosyl and mannosyl donors form bridged dioxanium ions. The intrinsic stability of the dioxanium ions versus the corresponding oxocarbenium ions does not provide an adequate explanation for the observed stereoselectivity reported in Table , and the absence of observeable 3-O-benzoyl-2-O-benzyl-4,6-Obenzylidene glucose dioxanium ions in C CEST-NMR. We therefore probed the pathways that lead to the formation of the dioxanium ions from the parent anomeric triflates using DFT computations the mannoside (+9.6) than the glucoside (+8.3), as a result of torsion between the mannosyl benzoate ester and the axial C-2 group. The conformation of the ring then changes via TSII to give a B2,5 conformation (RCII). The order of these two events was also changed (chair to boat transition before benzoate rotation), but this provided higher energy pathways (see Supplementary Information figure ). Finally, the benzoate ester displaces the anomeric triflate (TSIII, Figure and) to form the triflate anion stabilized dioxanium ion (PC). This step requires significantly more energy for the glucoside (+19.9) than for the mannoside (+14.0 kcal mol -1 ). Notably, for the mannoside the height of this transition state (TSIII-man) is similar to the TSII indicating that expulsion of the triflate is rather favorable when the mannoside adopts the correct geometry. Repositioning of the triflate towards the carbonyl carbon of the benzoate group then forms the final triflate stabilized dioxanium ion (P), in which the alpha face of the dioxanium ion is available for nucleophilic attack. Similar to the PC and the bare dioxanium ion (Supplementary table ), the Pman (9.9) is much more stable than the P-glu (13.0).