Figure 7

NMR analysis of bi-functional cello-oligosaccharides. (A) [¹H–¹³C] HSQC spectrum; the sample was in 99.9% D₂O and the spectrum was recorded at 25 °C. Peaks in the proton/carbon signals of the C4ox residue in the non-reducing end are marked by H/C#, where # indicates the carbon number in the residue. Peaks in the proton/carbon signals of the Glc1 (= the former reducing end) spin system are marked by H/C#*. Chemical groups giving rise to signals for the 2-aminooxy group are in bold and underlined. For the sake of simplicity, peaks related to internal monosaccharide residues are not marked (a full assignment of the chemical shifts is provided in Table S1). (B) Correlations between a [¹H–¹³C] HSQC spectrum and a [¹H–¹³C] HMBC spectrum recorded for the bi-functionalized product. The left and the middle insert show correlations (indicated by vertical lines) from H/C-3 and H/C-5 peaks in the HSQC (blue) for the C4-oxidized end to peaks (indicated by a horizontal dotted line) with a common carbon chemical shift of 159.2 ppm in the HMBC (red). The right insert shows a correlation from H/C-2* in the HSQC (blue) for the reducing end to a carbon peak with a carbon chemical shift of 155.4 ppm in the HMBC (red). Both carbon chemical shifts obtained from the HMBC spectrum correspond well to the (expected) presence of an imine (–C=N–) group, which is expected to be formed during the coupling reaction (Fig. 6), thus confirming the structure of the bi-functionalized product. R and R′ represent the non-reducing and the reducing end of the bi-functionalized cello-oligosaccharides, respectively. Further confirmation of the successful coupling can be found in the DOSY spectrum, Figure S3. The spectra were recorded, processed and analyzed using TopSpin 3.2 software (Bruker BioSpin AG), https://www.bruker.com.