Agilent dd2 500 m nmr spectrometer

Carboxyl reduction in SPm was carried out according to the method of Taylor and Conrad [45 (link)], and the reduction product was designated as SPm-R. Desulfations of the polysaccharides SPm and SPm-R were carried out as previous described [46 (link)], and the effectiveness of desulfation procedure was confirmed by determination of residual sulfate in the polysaccharide. The desulfation polysaccharide products of SPm and SPm-R were designated as SPm-Ds and SPm-RDs, respectively. Methylation analysis was performed for the polysaccharides SPm, SPm-Ds, SPm-R and SPm-RDs [47 (link),48 (link)]. FTIR spectrum was measured on a Nicolet Nexus 470 spectrometer (Thermo Fisher Scientific, Waltham, MA, USA). NMR spectra were recorded on an Agilent DD2 500 M NMR spectrometer (Agilent Technologies Co. Ltd., Palo Alto, CA, USA). Acetone was used as internal standard (2.225 ppm for ¹H and 31.07 ppm for ¹³C).

FTIR spectrum of the polysaccharide was measured on a Nicolet Nexus 470 spectrometer (Thermo Fisher Scientific, Waltham, MA). The polysaccharide was mixed with KBr powder, ground and pressed into a 1 mm pellet for FTIR measurements in the frequency range of 4000–400 cm⁻¹ [55 (link)]. ¹H and ¹³C NMR spectra were performed at 23 °C on an Agilent DD2 500 M NMR spectrometer (Agilent Technologies CO., LTD., USA). Approximately 70 mg of polysaccharide was dissolved in 1 mL of D₂O (99.97%) followed by freeze-drying for three times, and the final sample was dissolved in 0.5 mL of 99.97% D₂O. ¹H–¹H COSY, ¹H–¹³C HSQC, and ¹H–¹H NOESY experiments were also carried out. Chemical shifts are expressed in ppm using acetone as an internal standard at 2.23 ppm for ¹H and 31.07 ppm for ¹³C.

The FTIR spectrum of the polysaccharide was measured on a Nicolet Nexus 470 spectrometer (Thermo Fisher Scientific, Waltham, MA, USA) according to the method of Shingel [58 (link)]. The polysaccharide was mixed with KBr powder, ground, and pressed into a 1 mm pellet for FTIR measurements in the frequency range of 4000–400 cm⁻¹. NMR spectra, including ¹H and ¹³C NMR, ¹H–¹H COSY, ¹H–¹³C HSQC, and ¹H–¹H NOESY, were performed at 23 °C through an Agilent DD2 500M NMR spectrometer (Agilent Technologies, Santa Clara, CA, USA). Briefly, polysaccharide (50 mg) was dissolved in deuterium and lyophilized, the procedure was repeated three times, and then it was dissolved in 0.5 mL of 99.97% D₂O (120 mg/mL). Chemical shifts were expressed in ppm using acetone as an internal standard at 2.225 ppm for ¹H and 31.07 ppm for ¹³C.