Microorganisms associated with sea sponges have proven to be good natural product resources that are biologically active and pharmaceutically important. This research aimed to identify actinomycetes related to a sponge from Bar-ranglompo Island Makassar and the antibacterial compounds. Identification of actinomycetes was based on molecular characterization of sequence gen16S rRNA. The antibacterial compound was separated using vacuum liquid chromatog-raphy and preparative Thin Layer Chromatography (TLC). The structure determination was done based on spectroscopy 1H-NMR, 13C-NMR, 2D NMR, and mass spectra. Molecular characterization showed that actinomycetes strain BLP 20 had the closest relationship with Streptomyces parvulus and Uncultured Streptomyces sp. with a similarity value of 83%. The results obtained from the characterization of antibacterial compounds based on spectroscopic data indicate that these compounds lead to Actinomycin D. Characterization and identification of Strain 20 / BLP by molecular phylogenetic analysis of 16S rRNA sequences revealed the closest relationship with Uncultured Streptomy-ces sp and S. parvulus with a similarity value of 83 %, which indicated a new species. The structure of the active compound isolated from actinomycetes strain 20 / BLP leads to Actino-mycin D.
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