Metabolic Profiling and Mass Spectrometry Imaging of Rice Koji During the Extended Koji -Making Process

During sake brewing, rice koji produces saccharification enzymes that play crucial roles in flavor development. In this study, we analyzed the metabolic changes in rice koji across different koji-making durations and clarified the characteristics of hine koji prepared via a longer koji-making period than usual. Profiling of hydrophilic low-molecular-weight components using gas chromatography/mass spectrometry (GC/MS) revealed that several components, including erythritol, increased with an extended koji-making time. Quantitative analyses via high-performance liquid chromatography (HPLC) showed that glucose levels plateaued after a certain point, whereas erythritol levels increased continuously. Furthermore, mass spectrometry imaging (MSI) revealed a uniform distribution of glucose throughout rice koji, whereas erythritol was localized at the periphery and slightly inside the rice koji. This study provides significant insights into the koji-making process, offering a valuable tool for optimizing koji-making procedures and improving the sake quality.