Water is crucial for microbial growth, heat transfer and substrate hydrolysis, and dynamically changes with time in solid-state fermentation. However, water dynamics in the solid substrate is difficult to define and measure. Here, nuclear magnetic resonance was used to monitor water dynamics during the pure culture of Aspergillus oryzae YH6 on wheat in a model system to mimic solid starter (Qu or Koji) preparation. During fermentation, overall water content gradually decreased from 0.84 to 0.36 g/g, and water activity decreased from 0.99 to 0.93. Water content in different state (bound, immobilized and free) changed differently and all moved to more "bound" direction. The internal water distribution over the substrate matrix also showed a faster reduction inward both in the radical and axial direction. Our findings provide the prerequisites for optimal processes where water dynamics in solid-state fermentation can be monitored and controlled.