What's in the box? Authentication of Echinacea herbal products using DNA metabarcoding and HPTLC

Background: Differences in regulatory policies between countries as well as a lack of appropriate standardized methods for the authentication and quality control of herbal products directly impact their quality and safety. Echinacea products are among the top-selling herbal products in Europe and the United States with indications for a broad range of ailments. The increased use of Echinacea species has led to concerns about adulterated products resulting from challenges in morphology-based identification, due to overlapping morphological variation, frequent hybridization between species, and deliberate adulteration. Purpose: This study addressed the need for a novel analytical strategy in the authentication of herbal products. Methods: A combination of high performance thin layer chromatography (HPTLC) and DNA metabarcoding was employed. Fifty-three Echinacea herbal products marketed across Europe were tested to evaluate the accuracy of these methods in plant identification and their potential for detecting substitutes, adulterants and other unreported plant constituents. Results: HPTLC provides high resolution in the detection of Echinacea phytochemical target compounds, but does not offer information on the other species within the product. Alternatively, we showed that the limitation of HPTLC in detecting non-targeted species can be overcome by the complementary use of DNA metabarcoding. Using DNA metabarcoding, Echinacea species were detected in 34 out of the 38 retained products (89%), but with a lack of discriminatory resolution at the species level due to the low level of molecular divergence within the Echinacea genus. All of the tested herbal products showed considerable discrepancies between ingredients listed on the label and the ones detected using DNA metabarcoding, registering an overall ingredient fidelity of only 43%. Conclusion: The results confirm that DNA metabarcoding can be used to test for the presence of Echinacea species and simultaneously to detect other species present in even highly processed and multi-ingredient herbal products.