Nucleosil 5 ods column

The α-ketol (Me) preparations were purified firstly by RP-HPLC on Macherey–Nagel Nucleosil 5 ODS column (250 × 4 mm, 5 μm) using the solvent mixture methanol–water (linear gradient from 76:24 to 96:4, by volume) at a flow rate of 0.4 mL/min. Final purification was carried out by NP-HPLC on Macherey–Nagel Nucleodur 100–3 column (250 × 4.6 mm, 3 μm) eluted with hexane/isopropanol (99:1, by volume), flow rate 0.4 mL/min. Enantiomers of purified α-ketol methyl ester were separated on Chiralcel OB-H column (250 × 0.46 mm, 5 μm) with hexane/isopropanol 94:6 (by volume), flow rate 0.4 mL/min.

The products (Me esters) were separated using RP-HPLC on a Macherey–Nagel Nucleosil 5 ODS column (250 × 4 mm, 5 μm) using the solvent mixture methanol/water (linear gradient from 60:40 to 96:4, by volume) at a flow rate of 0.4 mL/min. The peaks of the products were collected and purified using NP-HPLC, as described in the previous section. All HPLC analyses were performed with a Shimadzu LC-20AB solvent delivery pump with UV spectral monitoring (190–370 nm) using a Shimadzu SPD-M20A diode array detector (Shimadzu, Kyoto, Japan). Separate products were collected after NP-HPLC separation, redissolved in [²H₆]benzene, and then, the NMR spectra were recorded. For additional qualitative information, the products (Me/TMS) were re-analyzed using GC-MS after hydrogenation over PtO₂ or after sequential NaBH₄ reduction, hydrogenation, methylation, and trimethylsilylation.
Product 1 collected after the NP-HPLC purification was dissolved in [²H₆]benzene and subjected to the NMR spectral records. Then, product 1 was reduced with NaBH₄ to compound 2, which was finally purified using NP-HPLC, as described in the previous section. Then, product 2 was dissolved in [²H₆]benzene, and its NMR spectra were recorded.

The products were purified first using RP-HPLC on a Macherey–Nagel Nucleosil 5 ODS column (250 × 4 mm, 5 µm) using the solvent mixture methanol/water (linear gradient from 76:24 to 96:4, by volume) at a flow rate of 0.4 mL/min. The peaks of the products were collected and purified using NP-HPLC on a Macherey–Nagel Nucleodur 100–3 column (250 × 4.6 mm, 3 μm) using the solvent mixture hexane/isopropanol (98.5:1.5, by volume) at a flow rate of 0.4 mL/min. After NaBH₄ reduction, the final purification of the products was performed using NP-HPLC as described above. Product enantiomers (methyl esters) were separated on a Chiralcel OD-H column (250 × 4.6 mm, 5 µm) with hexane/isopropanol (97:3, by volume) at a flow rate of 0.4 mL/min.