Highly active antiretroviral therapy (HAART) is the only available remedial measure to treat HIV infected patients, as recognized by the WHO. However, it is associated with toxicity (nephrotoxicity), high cost and most preferably drug resistance in the first-line treatment. Wherefore, potential and novel natural source is the only option for the modern world to challenge this global issue. In recent years, sulfated polysaccharide from marine macroalgae shown to be biologically active as anti-inflammatory, anticoagulant, antitumor, immunomodulatory and antiviral agents. As a direct inhibitor of HIV including other retroviruses, it is considered as a “new generation antiretroviral drug”. In our present study, Fucoidan, a sulfated polysaccharide has been extracted from two different macroalgae Dictyota bartayesiana (DD) and Turbinaria decurrens (TD) based on hot water extraction method and further confirmed by FT-IR and RP-HPLC methods. Both the crude and purified fucoidan samples were evaluated for anti-HIV activity after ion exchange chromatography purification. The maximum inhibitory activity of crude and purified fucoidan samples are 90.5% and 89% in the fucoidan extracts of DD. Whereas, it was 89.7% and 92% in the fucoidan extracts of TD. Simultaneously, the IC 50 values were determined and recorded as 1.56 µg/ml and 57.6 ng/ml in both the crude and purified fucoidan extracts of DD respectively. Similarly, for TD, it was 3 µg/ml and 131.7 ng/ml in the fucoidan extracts of TD. Therefore, further extensive research work is the most needful to fill the gaps to develop this sulfated polysaccharide as a potential drug for the treatment of HIV patients.