Exopolysaccharide-producing cyanobacterial strains in biological soil crusts are described, in addition to their chemical properties and antioxidant and flocculation activities. The EPSs from Pudukkottai blackish biological soil crusts (PBBSCs) showed significant amounts of total soluble proteins (0.1687 mg/mL) and carbohydrates (0.8056 mg/mL) compared with the Ariyalur blackish biological soil crusts (ABBSCs). LC-MS analysis of the cyanobacterial polysaccharides revealed the presence of natural sugars such as ribose and glucose/mannose, and uronic acids. The FTIR spectrum showed specific peak for OH and -NH stretching, C-H stretching, and carboxylic acids as the dominant groups in EPS. The in vitro DPPH assay of EPSs from PBBSCs showed 74.3% scavenging activity. Furthermore, the reducing power was determined to be 0.59 ata 500 mg/mL concentration, respectively. The extracted EPSs from the biological soil crust flocculated Kaolin clay suspension maximum at 500 mg/mL. Consequently, the cyanobacterial strain and exopolysaccharide characterization from the sacred forest's biological soil crust were analyzed for their bioactive potential, bio-crust diversity, and distribution.