Development of a low-cost, simple, and rapid identification method for Glycyrrhiza uralensis using PCR-RFLP and evaluation of seeds distributed on the market.

Glycyrrhiza is a crude drug used worldwide, especially in China and Japan. Glycyrrhiza uralensis Fischer and Glycyrrhiza glabra L. are listed as original plants in the 18th revised Japanese Pharmacopoeia; however, G. uralensis is considered the most important Glycyrrhiza species because it has traditionally been used medicinally. Most G. uralensis used in Japan is imported from China, but export restrictions in recent years have raised concerns about stable supply. Therefore, domestic cultivation in Japan is expected to prevent price increases of raw materials and ensure a stable supply. Accurate species identification is an important step in quality control. In recent years, a method for identifying species using molecular markers has been evaluated for the Glycyrrhiza genus; however, in this study, we found that a simpler method, PCR-RFLP, using two types of restriction enzymes, can be used to identify G. uralensis more cheaply and easily than conventional methods. When applied to the ITS1-5.8S-ITS2 region, the analysis of seeds distributed in the market showed that only four of the 190 seeds surveyed were G. uralensis, while most were G. glabra, G. inflata, or their hybrids. Therefore, to appropriately use commercially available G. uralensis seeds, it is important to correctly identify the species at an early stage. Quick identification using PCR-RFLP is thought to be a useful method for ensuring the adequate supply of G. uralensis.