Medicinal Plants Recommended by the World Health Organization: DNA Barcode Identification Associated with Chemical Analyses Guarantees Their Quality

<div><p>Medicinal plants are used throughout the world, and the regulations defining their proper use, such as identification of the correct species and verification of the presence, purity and concentration of the required chemical compounds, are widely recognized. Herbal medicines are made from vegetal drugs, the processed products of medicinal species. These processed materials present a number of challenges in terms of botanical identification, and according to the World Health Organization (WHO), the use of incorrect species is a threat to consumer safety. The samples used in this study consisted of the dried leaves, flowers and roots of 257 samples from 8 distinct species approved by the WHO for the production of medicinal herbs and sold in Brazilian markets. Identification of the samples in this study using DNA barcoding (<i>matK</i>, <i>rbcL</i> and <i>ITS2</i> regions) revealed that the level of substitutions may be as high as 71%. Using qualitative and quantitative chemical analyses, this study identified situations in which the correct species was being sold, but the chemical compounds were not present. Even more troubling, some samples identified as substitutions using DNA barcoding contained the chemical compounds from the correct species at the minimum required concentration. This last situation may lead to the use of unknown species or species whose safety for human consumption remains unknown. This study concludes that DNA barcoding should be used in a complementary manner for species identification with chemical analyses to detect and quantify the required chemical compounds, thus improving the quality of this class of medicines.</p></div

Identification levels for the analyzed samples when using each or a combination of the chosen markers.

<p>No sequence: samples for which the DNA barcoding protocol did not work. Unidentified: samples that could not be identified. The sequences from these samples did not show similarity levels above 98% to any of the sequences within the databases. Family: samples that could be identified at the family level. The sequences from these samples showed equal similarity levels to database sequences from multiple species belonging to the same family. Genus: samples that could be identified to the genus level. The sequences from these samples showed equal similarity levels to database sequences from multiple species belonging to the same genus. Species: samples that could be identified to the species level. The sequences from these samples showed similarity levels above 98% to database sequences from a unique species.</p

Percentage of samples analyzed according to species and genetic marker.

<p>Percentage of samples analyzed according to species and genetic marker.</p

Barcode gap analyses.

<p>The numbers represent the maximum intra-specific divergence. Values above this number were considered as a different species.</p><p>X—The Barcode Gap was not calculated because there was no clear division between intra- and interspecific genetic divergence.</p><p>*The genus <i>Peumus</i> possess only one specie, which makes the Barcode Gap not applicable.</p><p>Barcode gap analyses.</p

Comparison between the DNA barcode and TLC findings.

<p>ID: sample number. Green: samples that were identified as the expected medicinal species using DNA barcoding and that contained the expected chemical marker from the medicinal species according to TLC. Yellow: samples that were not identified within the genus of the medicinal species using DNA barcoding. Red: samples that were identified using DNA barcoding as a genus or family that varied from the expected one and that did not contain the chemical marker according to TLC. X: samples that did not generate any sequence using DNA barcoding or that could not be tested using TLC. -: absent samples.</p

Species analyzed in this study and their therapeutical recommendations.

<p>Species analyzed in this study and their therapeutical recommendations.</p

Conditions used for the chemical analyses.

<p>Conditions used for the chemical analyses.</p

DNA final barcode identification of the analyzed samples.

<p>DNA final barcode identification of the analyzed samples.</p