Applied Barcoding: The Practicalities of DNA Testing for Herbals

Abstract

DNA barcoding is a widely accepted technique for the identification of plant materials, and its application to the authentication of commercial medicinal plants has attracted significant attention. The incorporation of DNA-based technologies into the quality testing protocols of international pharmacopoeias represents a step-change in status, requiring the establishment of standardized, reliable and reproducible methods. The process by which this can be achieved for any herbal medicine is described, using Hypericum perforatum L. (St John’s Wort) and potential adulterant Hypericum species as a case study. A range of practical issues are considered including quality control of DNA sequences from public repositories and the construction of individual curated databases, choice of DNA barcode region(s) and the identification of informative polymorphic nucleotide sequences. A decision tree informs the structure of the manuscript and provides a template to guide the development of future DNA barcode tests for herbals.

DNA-Based Authentication and Metabolomics Analysis of Medicinal Plants Samples by DNA Barcoding and Ultra-High-Performance Liquid Chromatography/Triple Quadrupole Mass Spectrometry (UHPLC-MS)

“…Herbal products are sold as ground or powdered form of a raw herb, which makes correct botanical identification difficult [ 22 ]. DNA barcoding constitutes a very useful tool for quality control and, consequently, for clinical safety [ 23 , 24 ]. Molecular analyses is crucial for accurate and fast identification of medicinal plants, since the plant fragments sold in the market is difficult to identify using traditional methods especially due to lack of morphological features.…”

DNA Barcoding of St. Johnʼs wort (Hypericum spp.) Growing Wild in North-Eastern Greece

“…Although rbcL is widely used because of its easy amplification and high sequence quality, the region shows low intraspecific variation due to its slow evolutionary rate; thus it can be useful as a barcode only above genus level or in combination with other loci [15,16]. Howard et al [33] reported that despite being able to discriminate Hypericum species at the genus and section level, rbcL cannot differentiate the closely related species H. perforatum and H. maculatum. Our silico analysis results show that this region cannot be used effectively for the discrimination of Hypericum species, so we examined only matK and trnH-psbA for the identification of Hypericum species from NE Greece.…”

“…matK region is often difficult to amplify and sequence because of its relatively large size (about 1000 bp) and the lack of universal primers [36]. In Hypericum species, poor PCR amplification of matK has been reported by Howard et al [33] using published primers. Likewise, in our study, the 2 pairs of primers that have been previously used for the amplification of Hypericum species [37] failed to amplify the matK region.…”

“…Costa et al [27] supported that ITS exhibits big intraspecific variation when compared with matK, which is a sufficient barcode alone. Howard et al also reported that using different genomic regions could improve the species identification and recently included matK, rbcL, and trnH-psbA in a study, showing that ITS cannot differentiate H. perforatum from H. maculatum [33].…”

“…matK and trnH-psbA have been previously shown to be able to serve as single-region barcodes for wild medicinal plants contributing to the conservation and trade control of valuable plant resources [20][21]. DNA barcoding studies of the genus Hypericum have focused on the ITS region for phylogenetic analysis [22][23][24][25][26], detection of Hypericum species in herbal products [27][28][29][30][31], and species identification [32][33][34]. However, the low discrimination power and the multi-copy nature of this nuclear region complicate the analysis, so the use of plastid regions with a fast evolutionary rate is proposed [22,23,25,27,33].…”