DNA Metabarcoding Authentication of Ayurvedic Herbal Products on the European Market Raises Concerns of Quality and Fidelity

Ayurveda is one of the oldest systems of medicine in the world, but the growing commercial interest in Ayurveda based products has increased the incentive for adulteration and substitution within this herbal market. Fraudulent practices such as the use of undeclared fillers and use of other species of inferior quality is driven both by the increased as well as insufficient supply capacity of especially wild plant species. Developing novel strategies to exhaustively assess and monitor both the quality of raw materials and final marketed herbal products is a challenge in herbal pharmacovigilance. Seventy-nine Ayurvedic herbal products sold as tablets, capsules, powders, and extracts were randomly purchased via e-commerce and pharmacies across Europe, and DNA metabarcoding was used to assess the ability of this method to authenticate these products. Our analysis reveals that only two out of 12 single ingredient products contained only one species as labeled, eight out of 27 multiple ingredient products contained none of the species listed on the label, and the remaining 19 products contained 1 to 5 of the species listed on the label along with many other species not specified on the label. The fidelity for single ingredient products was 67%, the overall ingredient fidelity for multi ingredient products was 21%, and for all products 24%. The low level of fidelity raises concerns about the reliability of the products, and detection of threatened species raises further concerns about illegal plant trade. The study highlights the necessity for quality control of the marketed herbal products and shows that DNA metabarcoding is an effective analytical approach to authenticate complex multi ingredient herbal products. However, effort needs to be done to standardize the protocols for DNA metabarcoding before this approach can be implemented as routine analytical approaches for plant identification, and approved for use in regulated procedures

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 Gui Zhi Fu Ling Wan (FL) and Ge Xia Zhu Yu Tang (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 a resolution for all species indirectly through the 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 are assigned to the same species. Our study emphasizes that integrating DNA metabarcoding with phytochemical analysis brings valuable data for the comprehensive authentication of Traditional Chinese Medicines ensuring their quality and safe use

Authentication of Garcinia fruits and food supplements using DNA barcoding and NMR spectroscopy

Garcinia L. (Clusiaceae) fruits are a rich source of (−)-hydroxycitric acid, and this has gained considerable attention as an anti-obesity agent and a popular weight loss food supplement. In this study, we assessed adulteration of morphologically similar samples of Garcinia using DNA barcoding, and used NMR to quantify the content of (−)-hydroxycitric acid and (−)-hydroxycitric acid lactone in raw herbal drugs and Garcinia food supplements. DNA barcoding revealed that mostly G. gummi-gutta (previously known as G. cambogia) and G. indica were traded in Indian herbal markets, and there was no adulteration. The content of (−)-hydroxycitric acid and (−)-hydroxycitric acid lactone in the two species varied from 1.7% to 16.3%, and 3.5% to 20.7% respectively. Analysis of ten Garcinia food supplements revealed a large variation in the content of (−)-hydroxycitric acid, from 29 mg (4.6%) to 289 mg (50.6%) content per capsule or tablet. Only one product contained quantifiable amounts of (−)-hydroxycitric acid lactone. Furthermore the study demonstrates that DNA barcoding and NMR could be effectively used as a regulatory tool to authenticate Garcinia fruit rinds and food supplements

Elderberry and Elderflower Extracts, Phenolic Compounds, and Metabolites and Their Effect on Complement, RAW 264.7 Macrophages and Dendritic Cells

Modulation of complement activity and inhibition of nitric oxide (NO) production by macrophages and dendritic cells may have therapeutic value in inflammatory diseases. Elderberry and elderflower extracts, constituents, and metabolites were investigated for their effects on the complement system, and on NO production in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages and murine dendritic D2SC/I cells. The EtOH crude extracts from elderberry and elderflower and the isolated anthocyanins and procyanidins possessed strong complement fixating activity and strong inhibitory activity on NO production in RAW cells and dendritic cells. Phenolic compounds in the range of 0.1–100 µM showed a dose-dependent inhibition of NO production, with quercetin, rutin, and kaempferol as the most potent ones. Among the metabolites, caffeic acid and 3,4-dihydroxyphenylacetic acid showed the strongest inhibitory effects on NO production in both cell lines, without having cytotoxic effect. Only 4-methylcatechol was cytotoxic at the highest tested concentration (100 µM). Elderberry and elderflower constituents may possess inflammatory modulating activity, which increases their nutritional value

Metabolic and performance effects of Yerba Mate on well-trained cyclists

INTRODUCTION: Yerba Mate (YM) is a South American plant, rich in polyphenols, saponins, and xanthines, of growing scientific interest because of its metabolic effects. YM has been shown to increase fat utilization during exercise in untrained humans, but its effects on well-trained individuals during exercise are unknown. METHODS: We characterized metabolic and physical performance effects of YM in 11 well-trained male cyclists. In a double-blind crossover design, participants ingested 5 g of YM or placebo (PL; maltodextrin) daily for 5 d and 1 h before experimental trials. RESULTS: Ergometer-based tests included a submaximal step test (SST) at 30%-80% of V ̇O2max (6 × 5-min stages), followed by a cycloergometer-based time trial (TT) test to complete mechanical work (~30 min; n = 9). Before and during tests, blood and respiratory gas samples were collected. YM increased resting plasma adrenaline concentration (P = 0.002), and fat utilization by 23% at 30%-50% V ̇O2max versus PL (Glass effect sizes (ES) ± 95% confidence interval (CI), 0.8 ± 0.55) correlating strongly with post-SST plasma (glycerol; r = 0.758). Treatment effects on rates of perceived exertion, heart rate, and gross efficiency were unclear during SST. Respiratory exchange ratio during TT indicated carbohydrate dependence and did not differ between treatments (PL, 0.95 ± 0.03 (SD); YM, 0.95 ± 0.02). TT performance showed a small (ES = 0.38 ± 0.33) but significant (P = 0.0278) improvement with YM (PL, 30.1 ± 1.8 min (SD); YM, 29.4 ± 1.4 min; 2.2% ± 2% (95% CI)), with an average increase of 7-W power output (ES = 0.2 ± 0.19; P = 0.0418; 2.3% ± 2% (95% CI)) and 2.8% V ̇O2 (P = 0.019). Pacing displayed lower power output after 30% of total TT workload in PL vs YM. CONCLUSIONS: YM increased fat utilization during submaximal exercise and improved TT performance, but performance-enhancement effect was unrelated to measures of substrate metabolism during maximal exercise

Effect of Phenolic Compounds from Elderflowers on Glucose- and Fatty Acid Uptake in Human Myotubes and HepG2-Cells

Type 2 diabetes (T2D) is manifested by progressive metabolic impairments in tissues such as skeletal muscle and liver, and these tissues become less responsive to insulin, leading to hyperglycemia. In the present study, stimulation of glucose and oleic acid uptake by elderflower extracts, constituents and metabolites were tested in vitro using the HepG2 hepatocellular liver carcinoma cell line and human skeletal muscle cells. Among the crude extracts, the 96% EtOH extract showed the highest increase in glucose and oleic acid uptake in human skeletal muscle cells and HepG2-cells. The flavonoids and phenolic acids contained therein were potent stimulators of glucose and fatty acid uptake in a dose-dependent manner. Most of the phenolic constituents and several of the metabolites showed high antioxidant activity and showed considerably higher α-amylase and α-glucosidase inhibition than acarbose. Elderflower might therefore be valuable as a functional food against diabetes

Extracts, Anthocyanins and Procyanidins from Aronia melanocarpa as Radical Scavengers and Enzyme Inhibitors

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Ethnobotany of dioecious species: Traditional knowledge on dioecious plants in India

Ethnopharmacological relevance: More than 15,000 angiosperm species are dioecious, i.e., having distinct male and female individual plants. The allocation of resources between male and female plants is different, and also variation in secondary metabolites and sex-biased herbivory is reported among dioecious plants. However, little is known about the ethnobotany of dioecious species and whether preferences exist for a specific gender, e.g., in food, medicine or timber. Aim of the study: The aim of this study was: 1) to study whether Indian folk healers have preference for plant genders, and to document their knowledge and use of dioecious species; 2) to understand the concept of plant gender in Indian systems of medicine and folk medicine, and whether Ayurvedic literature includes any references to gender preference. Materials and methods: Lists of dioecious plants used in Indian systems of medicine and folk medicine were compiled. Ethnobotanical data was collected on perceptions and awareness of dioecious plants, and preferences of use for specific genders of dioecious species using semi-structured interviews with folk healers in Tamil Nadu, India. In addition, twenty Ayurvedic doctors were interviewed to gain insight into the concept of plant gender in Ayurveda. Results: Indian systems of medicine contain 5–7% dioecious species, and this estimate is congruent with the number of dioecious species in flowering plants in general. Informants recognized the phenomenon of dioecy in 31 out of 40 species, and reported gender preferences for 13 species with respect to uses as timber, food and medicine. Among informants different plant traits such as plant size, fruit size, and visibility of fruits determines the perception of a plant being a male or female. Ayurvedic classical literature provides no straightforward evidence on gender preferences in preparation of medicines or treatment of illness, however it contains details about reproductive morphology and sexual differentiation of plants. Conclusions: A knowledge gap exists in ethnobotanical and ethnopharmacological literature on traditional knowledge of dioecious plants. From this explorative study it is evident that people have traditional knowledge on plant gender and preferential usages towards one gender. Based on this, we propose that researchers conducting ethnobotanical and ethnopharmacological studies should consider documenting traditional knowledge on sexual systems of plants, and test the existence of gender specific usages in their conceptual framework and hypothesis testing. Incorporating such concepts could provide new dimensions of scientific knowledge with potential implications to conservation biology, chemical ecology, ethnoecology and drug discovery