An integrated system for identifying the hidden assassins in traditional medicines containing aristolochic acids.

Lan Wu,Yingjie Zhu,Shilin Chen,Bo Wang,Yaoli Li,Jingyuan Song,Yung-Chi Cheng,Haiyu Zhao,Li Xiang,Shaoqing Cai,Wei Sun,Hui Yao

doi:10.1038/srep11318

Abstract

Traditional herbal medicines adulterated and contaminated with plant materials from the Aristolochiaceae family, which contain aristolochic acids (AAs), cause aristolochic acid nephropathy. Approximately 256 traditional Chinese patent medicines, containing Aristolochiaceous materials, are still being sold in Chinese markets today. In order to protect consumers from health risks due to AAs, the hidden assassins, efficient methods to differentiate Aristolochiaceous herbs from their putative substitutes need to be established. In this study, 158 Aristolochiaceous samples representing 46 species and four genera as well as 131 non-Aristolochiaceous samples representing 33 species, 20 genera and 12 families were analyzed using DNA barcodes based on the ITS2 and psbA-trnH sequences. Aristolochiaceous materials and their non-Aristolochiaceous substitutes were successfully identified using BLAST1, the nearest distance method and the neighbor-joining (NJ) tree. In addition, based on sequence information of ITS2, we developed a Real-Time PCR assay which successfully identified herbal material from the Aristolochiaceae family. Using Ultra High Performance Liquid Chromatography-Mass Spectrometer (UHPLC-HR-MS), we demonstrated that most representatives from the Aristolochiaceae family contain toxic AAs. Therefore, integrated DNA barcodes, Real-Time PCR assays using TaqMan probes and UHPLC-HR-MS system provides an efficient and reliable authentication system to protect consumers from health risks due to the hidden assassins (AAs).

Highlights

For Aristolochiaceous herbs, the results showed that 39 species contained aristolochic acid I (AA I) (Fig. 3; additional details in Supplementary Table S6 and Fig. S2 online) and 14 species contained aristolochic acid II (AA II) (Fig. 3; additional details in Supplementary Table S6 and Fig. S3 online), and no aristolochic acids (AAs) were found in two Aristolochia species (Ar. kwangsiensis and Ar. elegans) together with four Asarum species (As. caudigerum, As. caulescens, As. pulchellum, and As. campaniflorum)
The internal transcribed spacer 2 (ITS2) region is more applicable than psbA-trnH for the identification of Aristolochiaceous plant materials
The NJ tree demonstrated that the ITS2 region can be used to accurately distinguish Aristolochiaceous species from their putative substitutes

Summary

Introduction

1990s, an epidemic of urothelial carcinomas related to AAs occurred in Belgium, where a number of women were prescribed the Chinese herb Fangji The women developed renal interstitial fibrosis and upper tract urothelial carcinomas-aristolochic acid nephropathy (UTUC-AAN)[4]. In another case, patients prescribed herbal preparations for the treatment of eczema suffered from interstitial nephritis, when preparations contained AAs. The main reason for this was the incorrect substitution of adulterants such as Guanmutong (Ar. manshuriensis Kom.) instead of Mutong (Akebia quinata (Houtt.) Decne.) and Guangfangji (Ar. fangchi) instead of Fangji (Stephania tetrandra)[5]. There has been a report of chronic interstitial fibrosis related to the ingestion of Indian herbal medicines that contain AAs9. AAN has been reported in many countries but its true incidence is unknown and what is currently known is probably only the tip of the iceberg[10,11,12]

Methods

Results

Conclusion