Dg 2080 53

Thin layer chromatography (TLC) was conducted by using precoated silica gel 600 F254 plates (Art. 5715, Merck, Darmstadt, Germany) or reverse phase C18 F254 plates (Art. 15389, Merck, Darmstadt, Germany). Flash column chromatography was carried out on silica gel by a medium-pressure gradient system equipped with a Pump Module C-605 and a Pump Manager C-615 (BÜCHI, Flawil, Switzerland). High resolution ESIMS were recorded on a Mariner Biospectrometry Workstation (Applied Biosystems of Thermo Fisher Scientific, Waltham, MA, USA) equipped with an electrospray ion source in the positive mode. High performance liquid chromatography (HPLC) was performed on a high pressure gradient system that is composed of pumps PU-2087, a degasser DG-2080-53, a mixer MX-2080-32, and a detector UV-2075 (JASCO, Tokyo, Japan). FT-IR spectra were recorded on an FT/IR-400 spectrometer instrument (JASCO). Specific rotations were observed on a DIP-370 polarimeter (JASCO). NMR spectra were investigated on an Avance ARX400 (400 MHz for ¹H) or Avance III HD 600 MHz Cryo-probe spectrometer (600 MHz for ¹H) (Bruker Bio Spin, Yokohama, Japan). The chemical shifts (ppm) were referenced to the solvent residual peak at δ_H 7.26 ppm (CDCl₃), δ_H 3.30/δ_C 49.0 ppm (CD₃OD), or δ_H 2.06/δ_C 29.9 ppm (acetone-d₆).

Size Exclusion Chromatography (SEC) has been used to determinate the molecular weights of polymer samples. A SEC system from JASCO was used including a Degasser DG-2080-53, a pump PU-980, an autosampler AS-2051 Plus, a column oven, a UV-detector UV-975, and a refractive index detector RI-2031 Plus. NaNO₃ (0.1 M) containing NaN₃ was used as an eluent at 30 °C. A SUPREMA columns guard/1000/30 Å from Polymer Standards Service was used. Pullulan standards were used for calibration.

The HPLC-FD system consists of a Model Ternary Gradient Unit LG2080-02, a Model 3-Line Degasser DG-2080-53, and a Model FB-1520S fluorescence detector (excitation wavelength, 280 nm; emission wavelength, 340 nm) together with a Model Intelligent HPLC Pump PU-2080-Plus (JASCO, Tokyo, Japan). A Shodex Asahipak ES-502N 7C column (7.5 mm ID × 100 mm, Showa Denko Co., Tokyo, Japan) was used for the anion-exchange HPLC with an ethanol gradient on 0.05 M sodium acetate, 0.40 M Na₂SO₄ (acetate-sulfate buffer), pH 4.85, at the flow-rate of 1.00 mL/min^{15 (link),19 (link)}. The following ethanol concentrations were used for the analysis: 0–1 min, 0%; 1–50 min, linear increase from 0 to 10%; 50–55 min, linear decrease from 10 to 0%; 55–60 min, 0%. All solvents were filtered through a filter unit (0.22 μm, pore size, TPP) and the samples were filtered through filter units (Millex-GV 0.22 μm Filter Unit, Millipore, Bedford, MA, USA) before use.