4.2. Production of Bioactive Culture Fluids

Ganoderma lucidum cultures were grown in stationary 2000 mL Erlenmeyer flasks containing 200 mL of potato dextrose broth. Flasks were inoculated with 5 pieces (cut with a 1 cm diameter No. 8 core borer) of actively growing mycelia from 14 day-old RSM cultures and incubated at 30 °C for 21 days. Mycelial growth was removed by centrifugation (14,000 rpm, 15 min, 5 °C). Culture fluids were collected and sterilized under vacuum with 0.22 μm membrane filters (Membrane Filters, Isopore^TM, Wicklow, Ireland), freeze-dried, and stored at room temperature in a desiccator until use.