Objective: To discover the material substances of the numbing characteristics in Polygonatum Rhizoma. Methods: Polygonati Rhizoma from eight regions were processed by the "nine-steam-nine-bask" method, and its content changes of saponin, polyphenol and calcium oxalate crystal at different steaming times were determined. The degree change of numbing was determined by the cell hemolysis test and sensory properties, and a correlation analysis of numbing was conducted. Results: With the increase of steaming times, the contents of polyphenol and saponin in Polygonati Rhizoma from different regions were initially increased and then decreased, while the content of calcium oxalate crystal and hemolysis rate and the sensory score of numbing decreased gradually. The correlation analysis results indicated that there was a significant positive correlation (P＜0.05) or a highly significant positive correlation (P＜0.01) between the content of calcium oxalate crystal and the numbing degree. SEM results showed that the structure of calcium oxalate crystal was gradually destroyed after steaming, which coincided with the change of numbing. Conclusion: This research proves that calcium oxalate crystal may be the key substance for the numbing in Polygonatum Rhizoma, which can provide a scientific basis for the study of the numbing elimination technology of Polygonatum Rhizoma.