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In order to explore the effect of Chlorella polysaccharide sulfation modification on biological activity, Chlorella sp.22 was used as experimental material in this study. The purified polysaccharide HDB-1 was obtained by extracting intracellular polysaccharides and separated and purified by DEAE-52 cellulose anion exchange column. The purified polysaccharide HDB-1 was sulfated to obtain SHDB-1. The antioxidant activity and antitumor activity of polysaccharides before and after sulfation modification were studied. The results showed that the purified component HDB-1 was mainly α-furanose. SHDB-1 was obtained by sulfation modification of HDB-1 by sulfur trioxide-pyridine method. Its degree of substitution was 1.046. The antioxidant activity results showed that the IC50 (semi-inhibitory concentration) values of HDB-1 and SHDB-1 DPPH radical clearance were 29.28 mg/mL and 14.49 mg/mL, respectively. The IC50 values of hydroxyl radical scavenging were 36.75 mg/mL and 28.59 mg/mL, respectively, indicating that the antioxidant effect of SHDB-1 was better than that of HDB-1. The antitumor results showed that HDB-1 and SHDB-1 were not toxic to cells at 1 mg/mL concentrations. The IC50 values of HDB-1 and SHDB-1 were 960.16 μg/mL and 658.19 μg/mL, respectively, indicating that SHDB-1 inhibited the proliferation of Hela in cervical cancer cells better than HDB-1. The above results show that compared with HDB-1, the antioxidant activity and in vitro inhibition of cancer cell proliferation of the sulfation-modified Chlorella intracellular polysaccharides SHDB-1 are significantly improved.