Rapid Authentication of the Herbal Medicine Plant Species Aralia continentalis Kitag. and Angelica biserrata C.Q. Yuan and R.H. Shan Using ITS2 Sequences and Multiplex-SCAR Markers

Abstract

Accurate identification of the plant species that are present in herbal medicines is important for quality control. Although the dried roots of Aralia continentalis (Araliae Continentalis Radix) and Angelica biserrata (Angelicae Pubescentis Radix) are used in the same traditional medicine, namely Dok-Hwal in Korean and Du-Huo in Chinese, the medicines are described differently in the national pharmacopeia. Further confusion arises from the distribution of dried Levisticum officinale and Heracleum moellendorffii roots as the same medicine. Medicinal ingredients from all four plants are morphologically similar, and discrimination is difficult using conventional methods. Molecular identification methods offer rapidity and accuracy. The internal transcribed spacer 2 (ITS2) region of the nuclear ribosomal RNA gene (rDNA) was sequenced in all four plant species, and the sequences were used to design species-specific primers. Primers for each species were then combined to allow sample analysis in a single PCR reaction. Commercial herbal medicine samples were obtained from Korea and China and analyzed using the multiplex assay. The assay successfully identified authentic medicines and also identified inauthentic or adulterated samples. The multiplex assay will be a useful tool for identification of authentic Araliae Continentalis Radix and/or Angelicae Pubescentis Radix preparations in Korea and China.

Development of conventional PCR and real‐time PCR assays to discriminate the origins of Chinese pepper oil and herbal materials from Zanthoxylum

“…Although botanical taxonomists recommend analysis of DNA barcodes for accurate species identification, such assays are cumbersome and time‐consuming, involving DNA amplification, electrophoresis, gel rescue and sequence analysis . In recent years, sequence characterized amplified region (SCAR) markers based on species‐specific sequences obtained from DNA barcodes have been used to discriminate species in raw food materials and herbal medicines …”

“…[13][14][15] In recent years, sequence characterized amplified region (SCAR) markers based on species-specific sequences obtained from DNA barcodes have been used to discriminate species in raw food materials and herbal medicines. 16,17 Raw food materials are altered by various manufacturing techniques involving physical, chemical or biochemical treatments. 6,7,18 Therefore, DNA fragmentation (to sizes less than 200 bp) frequently occurs in processed food such as pressed oils, cooked foods and ground material, making it difficult to detect DNA.…”

Differentiating Authentic Adenophorae Radix from Its Adulterants in Commercially-Processed Samples Using Multiplexed ITS Sequence-Based SCAR Markers

“…Judging from our laboratory experience, these additional costs amount to at least $300-$400 USD for the analysis of 10 herbal medicine samples. Therefore, the SCAR method is much more convenient and less expensive for the authentication of herbal medicines and for the identification of the botanical origins of herbal medicines than the DNA barcoding method [22,24,26]. In spite of these disadvantages, DNA barcoding is mightily important for plant species identification with high resolution and for obtaining primary sequence information in species-specific marker development.…”

“…A sequence-characterized amplified region (SCAR) marker assay can reliably and reproducibly distinguish species based on sequence information obtained from DNA fingerprinting or barcoding in diverse plants and herbal medicines. This method amplifies only target-containing samples using specific primers and differentiates positive or negative amplification of target regions, as well as length polymorphisms of target regions by gel electrophoresis of closely related samples [22][23][24]. Therefore, the SCAR method is simple to carry out, its results are simple to interpret, and it produces fewer errors in PCR amplification and sequencing [21].…”

“…Therefore, the SCAR method is simple to carry out, its results are simple to interpret, and it produces fewer errors in PCR amplification and sequencing [21]. Furthermore, the efficiency and ability of this assay improved with the combination of individual SCAR markers and multiplex PCR, namely, multiplexed SCAR marker assays [22,25,26]. These have been abundantly and successfully applied to authenticate medicinal plant species and commercially-processed herbal medicines [22].…”