Background: The mycelial fermentation of higher fungi were investigated to posses various bioactivities. Materials and Methods: The mycelial growth and pellet morphology in a 5-L bioreactor were investigated. The mycelial broth containing biomass and extracellular products harvested from the fermentor was tested for antiproliferative activity of colon cancer LoVo cells using 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide assay. Results: The maximum mycelial concentration in a 5-L bioreactor was 12.5 g/L after 8 days cultivation. Further investigation in the mycelial pellets during the fermentation period revealed that the mean diameter of the pellet morphology was positively correlated with mycelial biomass (R2 = 0.82, P < 0.05) and broth viscosity (R2 = 0.90, P < 0.01), significantly. The ethyl acetate extract showed the most significant effects, increasing the inhibition rate up to 87.5% after 48 h at concentration of 1000 μg/mL. Conclusion: The results demonstrated the feasibility of P. vaninii Ljup mycelial fermentation for large-scale production of bioactive and medicinal compounds.