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

Abstract

There are many polysaccharides with biological activities including anti-virus and anti-tumor in the residue produced by the extraction of beta-carotene from Duanaliella salina. In this paper, a method of high performance size-exclusion chromatography 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 mobile phase on retention behaviors of five polysaccharides fractions were investigated on two HPSEC columns (Waters Ultrahydragel Linear, 7.8 mm i. d. x 300 mm) connected in series. The results showed that 0. 1 mol/L NaAc buffer solution may be utilized as mobile phase during HPSEC under the conditions of column temperature of 45 degrees C and flow rate of 0.9 mL/min to minimize nonspecific interaction of sulfated polysaccharides 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. Thereby, the polysaccharide molecules were eluted and separated following equilibrium exclusion mechanism mainly. The weight mean molecular masses (M(w)) of five polysaccharide fractions from D. salina determined under optimum chromatographic conditions were 1 548 000 for PD1, 33 000 for PD2, 67 000 for PD3, 424 000 for PD4a and 10 000 for PD4b. For sulfated polysaccharide fraction PD4a, the relative standard deviations were 1.7% and 0.88% for M(w) and peak area, respectively.