Culture maturity assessment can be used to control fruiting body flush timing. Culture maturity of sawdust-based substrate was evaluated by using oak mushroom, (Lentinula edodes (Berk.) Pegler). The influence of substrate water potential (${psi}$) on the growth and fruiting of three genotypes of L. edodes was also investigated. Glucosamine content revealed a peak at the fruiting body senescent stage. Glucosamine increased steadily to the sporophore senescent stage, and sharply declined at crop treatment. Lipid phosphate and ergosterol contents peaked at pinning and button break stages, respectively. Therefore lipid phosphate and ergosterol contents would be considered as the convenient measurement for judging culture maturity and fruiting potentials. The substrate pH values before inoculation and on the fruiting stage were varied from 6.3 to 4.0. This pH changes were detected as changes in color from bluish purple to yellow by direct bromphenol blue(BPB) spraying, and shown a good correlation with fruit body yield of the 1 st flush. Concerning water potential of the cultures, a slight reduction of water potential, -0.5MPa, stimulated mycelial and colony growths on liquid, agar and sawdust-based substrates. The water potential of well-colonized matured substrate was -0.7MPa and -4.0MPa, before and after the fruiting, respectively. Excellent water providing capacity (higher ${psi}$) is expected to well-matured cultures with a high density of mycelial colonization. Also, the substrate water potential significantly affected by the interaction between genotypes and spawn run time.