De P, Salvat J, et al. 2024, Available at SSRN 4999887.
This work demonstrates the possibility of using enzyme promiscuity as a tool to obtain multiple enantiomerically pure ketoses in a predictive manner. In addition, analytical methods to characterize the enantiomeric identity of the product ketoses are proposed. Quantitative mass spectrometry (GC/MS) and 13C-NMR spectroscopy were used to analyze the enantiomeric ketose products. Among them, 13C-labeled xylitol (meso, 2R 3R 4S) is the only representative pentitol studied, which is oxidized to 4-13C-L-xylulose via C2 oxidation catalyzed by galactitol dehydrogenase (G2DH).
2-13C-D-xylitol for ketose product analysis of xylitol
The peak observed at δ 72.0 ppm is identified as C2 of the substrate 2-13C-D-xylitol (23a). When 2-13C-D-xylitol (23a) is reacted with G2DH, two additional peaks appear at δ 75.9 and δ 74.9 ppm, alongside the peak at δ 72.0 ppm (Fig A-D). This clearly suggests that 2-13C-D-xylulose (24) is not the resulting product. Additionally, the methoxime-TMS GC/MS analysis confirmed the presence of xylulose (Fig A). The newly identified peaks, which do not fall within the anomeric region, were attributed to C4 of the 4-13C-L-xylulose (25) anomers (25a, 25b).
