Reaction Steps
The product from the previous step (89.6 mg; 0.546 mmol) was dissolved in Tris.HCl buffer (50 mM, pH 7.5; 2 mL) containing mercaptoethanol (2 mM) and MnCl2 (2 mM). An aliquot of the L-fucose isomerase crude extract (preparation described infra), (1 mL, 820 units) was added and the mixture was stirred overnight at room temperature. Methanol (2 vol) was added and the mixture was filtered through Celite. The solvent was distilled under reduced pressure and the residue chromatographed on Dowex 50W-X8 resin (Barium form, 400 mesh, 2.5×9.5 cm) with water/ethanol (1:1). Compound 8c (75.2 mg, 84%) was obtained as a yellowish oil: (89 mg, 0.50 mmol, 25%). [α]D=−47.6 (c=0.5, MeOH, 30 min), [Lit. [α]Deg=−26.2° (c=1, pyridine), Lit [α]Deg=−33° (c=1.2, water)]. 1H-NMR (400 MHZ, D2O) δ (ppm): 5.03 (H1α, d, JH1α,H2α=3.7 Hz), 4.39 (H1β, d, JH1β,H 2β=8 Hz), 3.73 (H5α, m), 3.69 (H4α, H4β, d, J=3 Hz), 3.66 (H3α, d, JH2α,H3α=3.5 Hz), 3.62 (H2α, d, JH2α,H3α=3.5 Hz), 3.45 (H3β, dd, JH2β,H3β=10 Hz, JH3β,H4β=3-5 Hz), 3.35 (H5β, t, JH5β,CH2=3 Hz), 3.29 (H2β, dd, JH1β,H2β=8 Hz, JH2β,H3β=10 Hz), 1.4-1.51 (CH2, α and β), 0.77 (CH3, α and β). 13C-NMR (100 MHZ, D2O) δ (ppm): 96.42 (C1β), 92.23 (C1α), 76.43, 73.15, 72.04, 69.72 (β anomer), 71.81, 70.22, 69.51, 68.45 (α anomer), 23.01 (CH2, α and β), 9.41 (CH3, α and β). HRMS Calcd for C7H14O5+H+ 179.0919, found 179.0914.
