Pu 1580 intelligent pump

Optical rotations were determined on a P-1030 digital polarimeter (JASCO, Tokyo, Japan). IR spectra were measured on a FT-710 Fourier transform infrared spectrophotometer (Horiba, Kyoto, Japan), while UV spectra were taken on a V-520 UV/Vis spectrophotometer (JASCO, Tokyo, Japan). NMR data were obtained by an Ultrashield 600 spectrometer (Bruker, Ettlingen, Germany) with tetramethylsilane (TMS) as an internal standard. Positive ion HR-ESI-MS measurement was performed with the LTQ Orbitrap XL mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA). Column chromatography (CC) was carried out on silica gel 60 (E. Merck, Darmstadt, Germany), and octadecyl silica (ODS) gel (Cosmosil 75C18-OPN (NacalaiTesque, Kyoto, Japan; Φ = 35 mm, L = 350 mm). HPLC was performed on ODS gel (Inertsil ODS-3, GL-science, Φ = 6 mm, 250 mm, flow rate = 1.6 mL/min, GL Sciences, Tokyo, Japan), and the eluate was observed by refractive index detector with the RI-930 intelligent detector (JASCO, Tokyo, Japan) and a PU-1580 intelligent pump (JASCO, Tokyo, Japan). All chemicals were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan) and TCI (Tokyo, Japan).

Glucose and ethanol were quantified by HPLC. The HPLC system included a Jasco AS-2055 Intelligent auto sampler (Jasco, Tokyo, Japan) and a Jasco PU-1580 Intelligent pump (Jasco). The chromatographic separation was performed on a Rezex ROA H⁺ organic acid column, 5 μm, 7.8mm x 300 mm, (Phenomenex, Macclesfield, UK) at ambient temperature. The mobile phase was 0.005N H₂SO₄ with a flow rate of 0.5 mL/min. For detection a Jasco RI-2031 Intelligent refractive index detector (Jasco) was employed. Data acquisition was via the Azur software (version 4.6.0.0, Datalys, St Martin D’heres, France) and concentrations were determined by peak area comparison with injections of authentic standards. The injected volume was 10 μL and analysis was completed in 40 minutes. All chemicals used were analytical grade (>95% purity, Sigma-Aldrich, UK).

Glucose, ethanol, acetic acid and glycerol were quantified by HPLC. The HPLC system included a Jasco AS-2055 Intelligent auto sampler (Jasco, Tokyo, Japan) and a Jasco PU-1580 Intelligent pump (Jasco). The chromatographic separation was performed on a Rezex ROA H⁺ organic acid column, 5 μm, 7.8 mm × 300 mm, (Phenomenex, Macclesfield, UK) at ambient temperature. The mobile phase was 0.005 N H₂SO₄ with a flow rate of 0.5 mL/min. For detection a Jasco RI-2031 Intelligent refractive index detector (Jasco) was employed. Data acquisition was via the Azur software (version 4.6.0.0, Datalys, St Martin D’heres, France) and concentrations were determined by peak area comparison with injections of authentic standards. The injected volume was 10 μL and analysis was completed in 40 min. All chemicals used were analytical grade (>95% purity, Sigma-Aldrich, UK).