Sequence Analysis of Chinese and Japanese Curcuma Drugs on the 18S rRNA Gene and trnK Gene and the Application of Amplification-Refractory Mutation System Analysis for Their Authentication.

Abstract

Herbal drugs derived from rhizomes of Curcuma plants including Curcuma longa, C. zedoaria, etc. have attracted attention because of their antiinflammatory, 1) antihepatotoxic, 2) and neuroprotective activity.3) These drugs were originally used in traditional Asian medicines, and in Chinese medicine they are mainly used for treating various syndromes due to obstruction of blood circulation and retention of blood stasis such as arthralgia, psychataxia, and dysmenorrhea. 4) In China and Japan, more than 10 Curcuma species are distributed or cultivated, and the underground parts of six species have been used medicinally. In the Chinese herbal literature, 5) each Curcuma drug is prescribed for a specific ailment, although at present their clinical application is complicated because of the morphological similarity of the drugs as well as the plants. To develop the ultimate identification of Curcuma plants, we previously carried out molecular analysis of six Curcuma species based on 18S ribosomal RNA gene and trnK gene sequences. 6) As a result, two types of 18S rRNA gene sequences and five types of trnK gene sequences were demonstrated, and the identical sequence was observed between C. phaeocaulis and the Chinese population of C. zedoaria, and the Japanese population of C. zedoaria and C. kwangsiensis with a purple-colored band in the leaf blades (purple-cloud type), respectively. We report here the sequence analysis of Curcuma drugs available in Chinese and Japanese markets in both gene regions to identify their botanical origins. Moreover, to develop a more convenient method for identification of Curcuma plants and drugs, amplification-refractory mutation system (ARMS) analysis of both gene regions was investigated using species-specific primers designed by nucleotide differences among Curcuma species. Since five new species have been recently recorded in Chinese botanical papers 7,8) based on minute morphological differences from existing species, their relationship with other species was investigated at the molecular taxonomic to adjust the genus Curcuma in China. MATERIALS AND METHODS MaterialsThe following 11 Curcuma species comprising 20 specimens were examined: C. longa, C. phaeocaulis, C. zedoaria (Chinese and Japanese populations), C. kwangsiensis (purple-cloud and pubescent types), C. wenyujin, C. aromatica, C. sichuanensis, C. chuanezhu, C. chuanyujin, C. chuanhuangjiang, and C. yunnanensis (Table 1). Twenty-five Curcuma drugs were purchased from Chinese markets near the cultivation areas, the National Institute for Control of Pharmaceutical and Biological Products (Beijing, China), Tochimoto Tenkaido Co., Ltd. (Osaka, Japan), and Uchida Wakanyaku Co., Ltd. (Tokyo, Japan) ( Table 2). The authors identified the plant specimens with the help of Professor D. Fang and Dr. X. H. Xiao. All materials were stored in the Museum of Materia Medica, Institute of Natural Medicine, Toyama Medical and Pharmaceutical University (TMPW), Japan.Extraction and Purification of Total DNA and PCR Amplification Total DNA was ex...

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SNP, ARMS and SSH Authentication of Medicinal Dendrobium officinale KIMURA et MIGO and Application for Identification of Fengdou Drugs

“…DNA sequences carry lots of genetic information that is special for each plant. 17) Molecular methods such as SNP analysis, 18) ARMS 19) and SSH 16) have been widely used for authentication of medicinal herbs based on DNA sequences.…”

Rapid Identification of Curcuma longa and C. aromatica by LAMP

“…These species-specific regions made it possible to identify Curcuma plants and drugs by sequence analysis. In our previous study, we developed the amplification-refractory mutation system (ARMS) 6) and singlenucleotide polymorphism (SNP) analysis 7) for Curcuma species. In the present study, we used LAMP to identify Curcuma plants.…”