Rapid Identification of Officinal Akebiae Caulis and Its Toxic Adulterant Aristolochiae Manshuriensis Caulis (Aristolochia manshuriensis) by Loop-Mediated Isothermal Amplification.

Bo Wang,Lan Wu,Yuhua Shi,Chao Xiong,Peng Zhang,Mingming Zhao,Wei Sun,Shilin Chen,Wei Liu,Ping Wang

doi:10.3389/fpls.2016.00887

Abstract

Mu-tong (Akebiae Caulis) is a traditional Chinese medicine commonly used as a diuretic and antiphlogistic. A common adulterant of Mu-tong is Guan-mu-tong (Aristolochiae Manshuriensis Caulis), which is derived from the stem of Aristolochia manshuriensis Komarov, and contains carcinogenic aristolochic acids. We used an alternative technique, loop-mediated isothermal amplification (LAMP), to differentiate Mu-tong from Guan-mu-tong because LAMP is quick, highly sensitive, and specific. We designed a set of four common primers (G-F3, G-B3, G-FIP, and G-BIP) and a loop primer (G-LB) for LAMP based on the internal transcribed spacer 2 sequence of Ar. manshuriensis. We successfully amplified the LAMP assays and visual detection occurred within 60 min at isothermal conditions of 65°C. The LAMP reaction exhibited a tenfold increase in detection (4.22 pg/μl DNA) over conventional polymerase chain reaction demonstrating that LAMP is a useful technique to detect Guan-mu-tong. We conclude that the LAMP technique is a potentially valuable safety control method for simple and efficient discrimination of Mu-tong from its adulterant Guan-mu-tong.

Highlights

The use of traditional herbal medicine as dietary supplements has become increasingly popular in many countries due to the consumer’s belief that herbal medicine is both natural and harmless
In order to establish proper regulatory oversight for consumer safety, a procedure is needed that identifies the herbal plant in question, and detects possible adulterants. One such case is that of Mu-tong (Akebiae Caulis), a common traditional Chinese medicine used as a diuretic and antiphlogistic (Kawata et al, 2007) which may be adulterated by an aristolochic acid containing herb, Guan-mu-tong (Aristolochiae Manshuriensis Caulis) derived
Guan-mu-tong contains nephrotoxic and carcinogenic aristolochic acids (AAs), which cause aristolochic acid nephropathy (AAN) and upper tract urothelial carcinomas (UTUC) (Grollman et al, 2007; Hoang et al, 2013; Poon et al, 2013), with the inadvertent use of adulterated Guan-mu-tong by consumers leading to possible renal failure (But and Ma, 1999; Lord et al, 1999)

Summary

Introduction

The use of traditional herbal medicine as dietary supplements has become increasingly popular in many countries due to the consumer’s belief that herbal medicine is both natural and harmless. In order to establish proper regulatory oversight for consumer safety, a procedure is needed that identifies the herbal plant in question, and detects possible adulterants One such case is that of Mu-tong (Akebiae Caulis), a common traditional Chinese medicine used as a diuretic and antiphlogistic (Kawata et al, 2007) which may be adulterated by an aristolochic acid containing herb, Guan-mu-tong (Aristolochiae Manshuriensis Caulis) derived. DNA sequencing and DNA barcoding approaches are used for authentication of Aristolochiaceae and non-Aristolochiaceae plant material, but these procedures are time consuming (Han et al, 2005; Li et al, 2014; Wu et al, 2015) Due to these limitations, a rapid and effective identification method is needed for detecting Guan-mu-tong, in adulterated samples of Mutong

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