Judul: Aktivitas Antioksidan dan Sitotoksisitas Ekstrak Pigmen yang Berasal dari Bakteri yang Berasosisasi dengan Spons Laut
Abstrak: Poor lifestyle and environmental factors, including free radicals exposure, can cause degenerative diseases. Free radicals such as Reactive Oxygen Species (ROS) are naturally generated in the cells during metabolism. ROS is stabilized by indigenous antioxidants that are naturally present in the cells. Besides indigenous antioxidants, dietary antioxidant intake protects the body from free radicals that can trigger degenerative diseases. One of the antioxidant sources is bio-pigment, which can be produced by plants and microbes, including bacteria. The production of pigment from bacteria has advantages such as faster production and being environmentally friendly. The pigmented bacteria can live in various habitats, including in the oceans. Marine bacteria can form a symbiotic mutualism relationship with other organisms, such as sponges. Bacteria associated with sponges produce secondary metabolites similar to the hosts. The colored body of the sponge is thought to be caused by the presence of the associated pigmented bacteria. Several previous studies have reported various biological activities of pigments produced by bacteria associated with sponges, such as antimicrobial, anticancer, and antioxidant. In this study, isolate AGS112 and SAB E-64 produce yellow-red and bright yellow intracellular pigments, respectively, that might have antioxidant activity. Thus, this study aimed to determine the antioxidant and cytotoxicity activity of pigments derived from marine sponge-associated bacteria, followed by its function in the antioxidative genes expression regulation in the model organism Schizosaccharomyces pombe. This study was initiated by the rejuvenation of 2 bacterial isolates, AGS112 and SAB E-64. AGS112 was identified based on 16S rRNA. Meanwhile, SAB E64 had been identified in previous studies as Kocuria marina, which belongs to the actinobacteria group. Furthermore, the production and extraction of pigment produced by B. haikouensis AGS112 and K. marina SAB E-64 were carried out using chloroform and methanol, respectively. The crude pigment extracts were then analyzed using UV-Vis to determine the maximum wavelength absorption of each pigment, followed by the total phenolic and flavonoid measurements. In addition, the cytotoxicity activity test was carried out using the Brine Shrimp Lethality Test (BSLT) method. The two crude pigment extracts were then fractionated using the Thin Layer Chromatography (TLC) method, followed by bio-autography TLC analysis. The selected active fraction was isolated using the TLC-preparative method. Then, both crude pigment extracts and the active fraction extracts were tested for their antioxidant activity against DPPH radicals. The pigment extract with more potent antioxidant activity was then selected to be analyzed in vivo using model organism S. pombe through oxidative stress response assay (spot assay and TPC method), followed by relative gene expression analysis of sod1 and ctt1 genes using qRT-PCR. The metabolite profiling of the selected active fraction extract was analyzed using Liquid Chromatography Mass-Spectrophotometry (LC-MS). Based on the 16S rRNA gene, AGS112 has the closest similarity to Bacillus haikouensis C-89 (99%). The yields of crude pigment extracts of B. haikouensis AGS112 and K. marina SAB E-64 were 12.57% and 19.7% (w/v), respectively. The BSLT results showed that the LC50 values of each crude pigment extract were <1000 µg/mL that indicated the crude pigment extracts at certain concentrations is classified as cytotoxic compound and may have
Keyword: Antioxidant, DPPH, Pigment, Schizosaccharomyces pombe, Sponge-associated bacteria