Studies On The Relationship Between SO₄^2- Content Of Polysaccharide From Gracilaria Lemaneiformis And Anti-influenza Virus Activities

In this paper the relationship between SO42- content of polysaccharide from Gracilaria lemaneiformis(PGL) and anti-influenza virus activities was studied, its initial antiviral mechanism was also approached. Extraction conditions of polysaccharide were optimized and suitable desulfuration and sulfation methods were selected at the same time. The main study results were as follows:Extraction conditions of polysaccharide from Gracilaria lemaneiformis were obtained by the L933orthogonal design, while temperature of 100℃, time of 5 hours, dry alage to water ratio of 1:100 was the optimum, the ratio of polysaccharide to dry alage was 22.97% in this condition; Purified PGL's(PPGL) polysaccharide content was 94.48%, protein content was 0.95%, SO42- content was 13.25%; Three fractions called GL-1,GL-2 and GL-3 were obtained by DEAE-celluose column chromatograph, it showed that PPGL was composed of three components at lease; Results of ultraviolet analysis and elution from Sephadex G-100 of GL-1, GL-2 and GL-3 showed that three of them were purified well. The average molecular weight and SO42- content of each fraction were estimated at 5.1kD, 52kD, 67kD and 0.00%, 12.63%, 22.61%.Using hydrochloric acid- methanol, dilute hydrochloric acid and dissolvent methods to deal with PPGL to get desthio-PGL, results showed that SO42- contents of three methods were 1.75%, 0.00% and 2.25%; And the degree of substitution(DS) were 0.09, 0.00 and 0.12 separately. In general, hydrochloric acid- methanol method was the best of the three.Using reforming Wolfrom method to deal with desthio-PGL from hydrochloric acid- methanol method to obtain sulfated PGL. Taking SO42- content and DS as index, the sulfation condition the volume ratio of chlorosulfonic acid to pyridine of 1:4, temperature of 65℃, time of 4 hours was the optimum; The SO42- content and DS of sulfated PGL was 17.53% and 2.01 separately in this condition.Antiviral results showed that the inhibiting effect of PPGL to influenza virus H1-364 was better than to influenza virus H3-616; Putting PPGL, desthio-PGL, different SO42- content of sufation samples (S1, S2, S3) and purified ploysacchrides (GL-1, GL-2 and GL-3) on MDCK which was affected by H1-364, results demonstrated that PPGL's inhibiting effect was the best and it's max virus inhibition ratio was 88.82%; Polysacchride samples'antiviral ability was better when SO42- content was higher within limits; While SO42- content and DS were near 13% and 1.10, the polysacchride samples got the best antiviral ability, while SO42- content was higher or lower than 13%, the antviral ability reduced.Antiviral mechanism initial approach showed that PPGL had no direct effect on H1-364, it could not effect virus to release, but it could interfere H1-364 to adsorb on host cell and it also had effect on virus replication.Study on antiviral activities of polysaccharide from Gracilaria lemaneiformis was firstly approached in this paper, it revealed the relationship between SO42- content and antiviral activities. Study results had significant theory and academic value to physiological activities study of Gracilaria lemaneiformis, they also provided significant instructions for exploitating polysaccharide from Gracilaria lemaneiformis reasonably.