In the present study, we focused on the molecular identification of Gracilaria tenuistipitata (a red sea algae), screening of its effective bioactive compounds and appraisal of their therapeutic potency. To confirm its identity, we conducted the first molecular analysis, utilizing the plastid-encoded rbcL gene and mitochondrial Cytochrome c Oxidase Subunit I 5′ region (COI-5P) to accurately determine the species. This study aimed to profile and evaluate the biological potential of G. tenuistipitata by comparing its ethanolic and isopropanolic extracts using HPLC, GC-MS, and FTIR. Phytochemical screening revealed a diverse biochemical composition, including alkaloids, phenolic compounds, and flavonoids. The ethanolic extract exhibited higher total phenolic (18.94 mg GAE/g) and flavonoid (42.375 mg QE/g) content compared to the isopropanolic extract. This corresponded with its greater antioxidant potential, as shown by a lower DPPH IC50 value (770.88 µg/mL vs. 821 µg/mL for isopropanolic) and higher total antioxidant capacity. HPLC analysis quantified 11 polyphenols, with gallic acid being most abundant in the ethanolic extract (653.06 µg/g), while GC-MS identified 9 key volatile compounds, with hexadecanoic acid, methyl ester predominating in the isopropanolic extract (67.357 µg/mL). FTIR analysis confirmed the presence of hydroxyl, carboxyl, and carbonyl functional groups and supported the seaweed’s safety by indicating the absence of toxic cyanide groups. Collectively, this study provides a foundational, comparative chemical profile of G. tenuistipitata, highlighting how solvent choice impacts bioactive yield and underscoring its potential as a source of natural antioxidants.