Large Scale Culturing of Actinomycete Secondary Metabolites and Extraction

Aquatic actinomycetes are evolutionarily distinct from their terrestrial counterparts and devote a large portion of their genome to the synthesis of diverse secondary metabolites (Schorn et. al., Microbiology 2016). Actinomycetes flourish in terrestrial, freshwater, and marine ecosystems with aquatic species producing unique natural products. Over half of all approved drugs come from natural products with 55% of all known antibiotics percent coming directly from natural products or inspiring the synthetic warhead (Newman and Cragg, J. Nat. Prod. 2020). The Carlson Lab actinomycete strain collection is derived from unique ecosystems from across North America. Environmental samples are plated on a variety of media to produce diversity plates, and isolates are replated on new A1 agar plates. These are then cultivated in liquid media, A1 or AIDI, and stored cryogenically in glycerol at -80°C. To isolate potential bioactive compounds, XAD16 resin is added to culture broth. The resin is then rinsed with acetone to release the compounds. Acetone is removed through evaporation in vacuo to yield a crude extract that is further partitioned using ethyl acetate. This material is then fractionated using silica gel chromatography to produce 4 fractions. This step allows to separate the extract into different fractions based on their polarity. These fractions are evaluated for various biological activities including antibiotic, antifungal, and anticancer activity.