Heleos 2 multi angle light scattering

The total polysaccharide content was determined using the phenol-sulfuric acid protocol described previously [35 (link)]. The monosaccharide composition was verified by high-performance anion exchange chromatography using a CarboPac^TM PA1 column (4.5 mm × 50 mm; Thermo Fisher Scientific, Waltham, MA, USA). The samples were acidified with 4 M trifluoroacetic acid (TFA). Subsequently, the monosaccharide composition was identified and verified by comparing its retention time with that of the standard mixture. The functional groups were determined using a Bruker FTIR, Alpha II (Bruker, Karlsruhe, Germany) instrument with a wavenumber range of 400–4000 cm⁻¹. The molecular weight was determined using a DAWN Heleos II multi-angle light scattering and Optilab T-rEX refractive index detector (MALS-RI, Wyatt Technology, Santa Barbara, CA, USA) system equipped with a Shimadzu HPLC system connected to a PL Aquagel-OH MIXED-H (7.5 × 300 mm, Agilent Technologies, Santa Clara, CA, USA). The sample was filtered through a membrane filter of a pore diameter of 0.22 µm before being eluted with 0.5 mol/L NaCl at a flow rate of 0.5 mL/min. Data analysis was carried out using ASTRA 6 software (Wyatt Technology) with a refractive index increment (dn/dc) of 0.140 mL/g. The structure of the commercial-grade fucoidan was analyzed for comparison.