Complementary authentication of Chinese herbal products to treat endometriosis using DNA metabarcoding and HPTLC shows a high level of variability

Traditional Chinese Medicine (TCM) is popular for the treatment of endometriosis, a complex gynecological disease that affects 10% of women globally. The growing market for TCMs has yielded a significant incentive for product adulteration, and although emerging technologies show promise to improve their quality control, many challenges remain. We tested the authenticity of two traditional Chinese herbal formulae used in women’s healthcare for the treatment of endometriosis, known as <em>Gui Zhi Fu Ling Wan</em> (FL) and <em>Ge Xia Zhu Yu Tang</em> (GX). Dual-locus DNA metabarcoding analysis coupled with high-performance thin-layer chromatography (HPTLC) were used to authenticate 19 FL and six GX commercial herbal products, as well as three ad hoc prepared artificial mixtures. HPTLC was able to detect most of the expected ingredients via comparative component analysis. DNA metabarcoding was able to detect an unexpected species diversity in the products, including 38 unexpected taxa. Chromatography has resolution for all species indirectly through identification of marker compounds for the different species ingredients. Metabarcoding on the other hand yields an overview of species diversity in each sample, but interpretation of the results can be challenging. Detected species might not be present in quantities that matter, and without validated quantification some detected species can be hard to interpret. Comparative analysis of the two analytical approaches also reveals that DNA for species might be absent or too fragmented to amplify as the relevant chemical marker compounds can be detected but no amplicons assigned to the same species. Our study emphasizes that integrating DNA metabarcoding with phytochemical analysis brings valuable data for comprehensive authentication of Traditional Chinese Medicines ensuring their quality and safe use.