Ogata Jun, Kawamura Maiko, Hakamatsuka Takashi, Kikura-Hanajiri Ruri
Division of Pharmacognosy, Phytochemistry and Narcotics, National Institute of Health Sciences.
Yakugaku Zasshi. 2020;140(12):1501-1508. doi: 10.1248/yakushi.20-00170.
In Japan, mitragynine, 7-hydroxymitragynine and Mitragyna speciosa Korth. (M. speciosa, "Kratom") were controlled as Designated Substances under the Pharmaceutical and Medical Device Act from March 2016. In this study, the origins of 16 Kratom products obtained from the illegal drug market in Japan were investigated by DNA analyses and LC-MS analyses. When the PCR-restriction fragment length polymorphism (RFLP) was performed using the restriction enzyme XmaI (as reported by Sukrong et al. to be able to distinguish M. speciosa), the same DNA fragment patterns were obtained from all 16 products. On the other hand, as a result of the identification of the plant species of each product by nucleotide sequence analyses, the sequences of M. speciosa were detected in only 14 products. Despite the facts that mitragynine and 7-hydroxymitragynine were detected also in the other two products by the LC-MS analyses, M. speciosa DNAs were not amplified from these products by the PCR. Moreover, the DNA amplicons of the other psychotropic plant (Mesembryanthemum sp., e.g. "Kanna") were detected. This plant PCR amplicon has the restriction site for the XmaI at the same position of the M. speciosa PCR amplicon and it is difficult to distinguish "Kratom" and "Kanna" by the conventional PCR-RFLP. When the restriction enzyme XhoI was used simultaneously with the Xmal, the specific DNA fragment was only observed from the M. speciosa amplicon and it was possible to distinguish both species using this improved PCR-RFLP method. This method is useful to identify the origin of Kratom products distributed in the illegal drug market.
在日本，自2016年3月起，帽柱木碱、7-羟基帽柱木碱以及柯迪 kratom（Mitragyna speciosa Korth.，“kratom”）根据《药品和医疗器械法》被列为指定物质。在本研究中，通过DNA分析和液相色谱-质谱联用（LC-MS）分析，对从日本非法毒品市场获取的16种kratom产品的来源进行了调查。当使用限制性内切酶XmaI进行聚合酶链反应-限制性片段长度多态性（PCR-RFLP）分析时（如Sukrong等人所报道，该酶能够区分柯迪 kratom），从所有16种产品中获得了相同的DNA片段模式。另一方面，通过核苷酸序列分析鉴定各产品的植物种类后，仅在14种产品中检测到了柯迪 kratom的序列。尽管通过LC-MS分析在另外两种产品中也检测到了帽柱木碱和7-羟基帽柱木碱，但通过PCR未能从这些产品中扩增出柯迪 kratom的DNA。此外，还检测到了其他精神活性植物（Mesembryanthemum sp.，例如“卡纳”）的DNA扩增子。这种植物的PCR扩增子在柯迪 kratom PCR扩增子的相同位置具有XmaI的限制性位点，因此难以通过传统的PCR-RFLP区分“kratom”和“卡纳”。当同时使用限制性内切酶XhoI和Xmal时，仅在柯迪 kratom的扩增子中观察到特定的DNA片段，使用这种改进的PCR-RFLP方法可以区分这两种植物。该方法有助于鉴定非法毒品市场上流通的kratom产品的来源。
