Table_1_Identifying the Species of Seeds in Traditional Chinese Medicine Using DNA Barcoding.DOCX

Seed is not only the main reproductive organ of most of herbal plants but also an important part of Traditional Chinese Medicine (TCM). Seed TCMs possess important medicinal properties and have been widely used as components of pharmaceutical products. In parallel with the increasing popularity and accessibility of seeds as medicinal products in recent years, numerous substitutes and adulterants have also appeared on the market. Due to the small volume and similar appearances of many seed TCMs, they are very difficult to accurately identify the constituent plant species through organoleptic methods. Usage of the wrong herb may be ineffective or may worsen the condition and even cause death. Correct identification of seed herbal medicines is therefore essential for their safe use. Here, we acquired 177 ITS2 sequences and 15 psbA-trnH sequences from 51 kinds of seed TCMs belonging to 64 species that have been described in the Chinese Pharmacopoeia. Tree-building analysis showed that the ITS2 sequences of 48 seed TCMs can be differentiated from each other, and they formed distinct, non-overlapping groups in the maximum-likelihood tree. Furthermore, all of the sequences acquired in this study have been submitted to the public DNA barcoding system for herbal medicine, and this integrated database was used to identify 400 seed TCM samples purchased from medicinal markets, drug stores, and the Internet, enabling the identification of 7.5% of the samples as containing non-declared species. This study provides a brief operating procedure for the identification of seed TCMs found in herbal medicine. In the future, researchers and traditional herbal medicine enterprises can use this system to test their herbal materials.

种子不仅是大多数草本植物的主要繁殖器官，亦是传统中医药（TCM）的重要组成部分。种子类TCM蕴含着重要的药用特性，并广泛作为药品的组成部分被应用。近年来，随着种子作为药用产品的人气和可获得性的提升，市场上亦出现了众多替代品和掺杂物。由于许多种子类TCM体积微小且外观相似，通过感官鉴定方法难以准确识别其构成植物种类。误用草药不仅可能无效，甚至可能加剧病情，甚至导致死亡。因此，准确识别种子类草药对于其安全使用至关重要。本研究中，我们从中国药典中描述的64种属于51种种子TCM中获取了177条ITS2序列和15条psbA-trnH序列。树状图构建分析表明，48种种子TCM的ITS2序列可以相互区分，并在最大似然树中形成了独特的、非重叠的组群。此外，本研究中获取的所有序列均已提交至公共草药DNA条形码系统，并利用该综合数据库识别了从药材市场、药店和互联网购买的400份种子TCM样本，从而识别出7.5%的样本含有未申报的物种。本研究为草药中种子类TCM的鉴定提供了简明操作流程。未来，研究人员和传统草药企业可利用此系统对其草药材料进行检测。