Retention Behaviors and Analytical Methods of High Performance Size-Exclusion Chromatography of Polysaccharides from Duanaliella salina

Several polysaccharides with biological activities including anti-virus and anti-tumor were present in the residue produced by the extraction of β-carotene from Duanaliella salina. In this article, a method of high performance size-exclusion chromatography (HPSEC) coupled with refractive index detection has been developed for the relative molecular mass analysis of the polysaccharides isolated from the residue. The effects of salt and pH values of the mobile phase on the retention behaviors of five polysaccharide fractions were investigated on two HPSEC columns (Waters Ultrahydrogel Linear, 7.8 mm i.d. × 300 mm) connected in series. The results showed that 0.1 mol/L NaAc buffer solution may be utilized as the mobile phase during HPSEC under the conditions of column temperature of 45°C and flow rate of 0.9 mL/min to minimize nonspecific interaction of sulfated polysaccharide fraction (PD4a), complex carbohydrate containing oligonucleic acid (PD4b), and acidic polysaccharide fraction (PD1) with the column matrix. In addition, under the conditions, the association effect of the polysaccharide molecules was eliminated. Thus, the polysaccharide molecules were eluted and separated mainly following the equilibrium exclusion mechanism. The weight mean molecular masses (Mw) of five polysaccharide fractions from D. salina determined under optimum chromatographic conditions were 1548000 for PD1, 33000 for PD2, 67000 for PD3, 424000 for PD4a, and 10000 for PD4b. For sulfated polysaccharide fraction PD4a, the relative standard deviations were 1.7% and 0.88% for Mw and peak area, respectively.