High-Yield Production of Cytotoxic Actinomycin D from Endophytic Streptomyces parvulus AL036 through Nutritional Optimization

Endophytic actinomycetes are bacteria that live inside plant tissues without causing harm to the plant, producing various bioactive compounds. This study aimed to isolate and characterize an endophytic actinomycete from Alpinia galanga roots, and to investigate its bioactive compounds. A Streptomyces strain designated AL036 was isolated and identified as Streptomyces parvulus through morphological, biochemical, and 16S rDNA sequence analysis. Similar to Streptomyces parvulus Tc022, this strain exhibited actinomycin D as its major bioactive compound. Media optimization experiments and cytotoxic activity against non-cancerous and cancer cell lines were carried out. The starch casein medium was identified as a medium supporting significantly higher actinomycin D production (103.67 mg/L) compared to the initial medium (ISP-2). Further optimization revealed that a specific carbon-to-nitrogen ratio (20 g/L soluble starch and 2 g/L casein) maximized production (199.33 mg/L). Sugar supplementation did not enhance production but influenced mycelial growth. The purified actinomycin D exhibited cytotoxicity against HeLa cells and LLC-MK2 cells, with IC ₅₀ values of 10.22 ± 2.53 µg/ml and 17.78 ± 3.97 µg/ml, respectively. This study isolated a new Streptomyces parvulus AL036, with efficient actinomycin D production similar to Tc022. Media optimization significantly improved production, highlighting the importance of carbon and nitrogen sources. Both crude extract and purified actinomycin D showed cytotoxicity and selectivity for some cancer cells.