s Serial Extraction Technique of Rich Antibacterial Compounds in Sargassum cristaefolium Using Different Solvents and Testing their Activity

Background: Sargassum cristaefolium, as one of the brown seaweeds locally found in Indonesia, is extracted using the serial technique employing different solvents. Methods: S. cristaefolium powder (50 mesh) was extracted with three different solvents, including hexane, ethyl acetate, and methanol. S. cristaefolium powder residue was dried prior to serial re-extraction using different solvents. Three serial extracts were obtained and named as 1-stage extract, 2-stage extract, and 3-stage extract. Besides, a single-step extract (i.e., extraction using only methanol) was produced to be compared with three serial extracts in antibacterial activity tests (against E. coli and S. aureus). The three serial extracts were detected for their antibacterial compounds using GC-MS, LC-HRMS, and FT-IR. Results: The 3-stage extract exhibited the highest extraction yield. On S. aureus, the inhibition zone in all extracts was not significantly different. On E.coli, the highest inhibition zone (5.42±0.14 mm) was of the 3-stage extract; indeed, it was higher than both antibiotic and a single- step extract. Antibacterial compounds, such as phenol, 9-Tricosene(Z)-, palmitic acid, and oleamide, were present in all extracts. Other antibacterial compound types, both the 1-stage and 2-stage extracts, contained 7 types, whilst the 3-stage extract contained the most types (11 types). Particularly, hexyl cinnamic aldehyde, betaine and several cinnamic aldehyde groups were detected only in the 3-stage extract comprising the dominant area. The carboxylic acid groups were detected in all extracts to confirm the fatty acid structure. Conclusion: The serial extraction technique could produce the 3-stage extract which exhibited the strongest antibacterial activity and contained the richest antibacterial compounds.