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      Differentiation of Cyanthillium cinereum, a smoking cessation herb, from its adulterant Emilia sonchifolia using macroscopic and microscopic examination, HPTLC profiles and DNA barcodes

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          Abstract

          Cyanthillium cinereum (L.) H.Rob. is one of the most popular herbal smoking cessation aids currently used in Thailand, and its adulteration with Emilia sonchifolia (L.) DC. is often found in the herbal market. Therefore, the quality of the raw material must be considered. This work aimed to integrate macro- and microscopic, chemical and genetic authentication strategies to differentiate C. cinereum raw material from its adulterant. Different morphological features between C. cinereum and E. sonchifolia were simply recognized at the leaf base. For microscopic characteristics, trichome and pappus features were different between the two plants. HPTLC profiles showed a distinct band that could be used to unambiguously differentiate C. cinereum from E. sonchifolia. Four triterpenoid compounds, β-amyrin, taraxasterol, lupeol, and betulin, were identified from the distinct HPTLC band of C. cinereum. The use of core DNA barcode regions; rbcL, matK, ITS and psbA- trnH provided species-level resolution to differentiate the two plants. Taken together, the integration of macroscopic and microscopic characterization, phytochemical analysis by HPTLC and DNA barcoding distinguished C. cinereum from E. sonchifolia. The signatures of C. cinereum obtained here can help manufacturers to increase the quality control of C. cinereum raw material in commercialized smoking cessation products.

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          Integrative taxonomy: a multisource approach to exploring biodiversity.

          Good alpha taxonomy is central to biology. On the basis of a survey of arthropod studies that used multiple disciplines for species delimitation, we evaluated the performance of single disciplines. All included disciplines had a considerable failure rate. Rigor in species delimitation can thus be increased when several disciplines chosen for complementarity are used. We present a flexible procedure and stopping rule for integrative taxonomy that uses the information from different disciplines separately. Disagreement among disciplines over the number and demarcation of species is resolved by elucidating and invoking evolutionary explanations for disagreement. With the identification of further promising study organisms and of new questions for in-depth analysis, evolutionary biology should profit from integrative taxonomy. An important rationale is clarity in researcher bias in the decision-making process. The success of integrative taxonomy will further increase through methodological progress, taxonomic training of evolutionary biologists, and balanced resource allocation.
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            Comparative analysis of a large dataset indicates that internal transcribed spacer (ITS) should be incorporated into the core barcode for seed plants.

            A two-marker combination of plastid rbcL and matK has previously been recommended as the core plant barcode, to be supplemented with additional markers such as plastid trnH-psbA and nuclear ribosomal internal transcribed spacer (ITS). To assess the effectiveness and universality of these barcode markers in seed plants, we sampled 6,286 individuals representing 1,757 species in 141 genera of 75 families (42 orders) by using four different methods of data analysis. These analyses indicate that (i) the three plastid markers showed high levels of universality (87.1-92.7%), whereas ITS performed relatively well (79%) in angiosperms but not so well in gymnosperms; (ii) in taxonomic groups for which direct sequencing of the marker is possible, ITS showed the highest discriminatory power of the four markers, and a combination of ITS and any plastid DNA marker was able to discriminate 69.9-79.1% of species, compared with only 49.7% with rbcL + matK; and (iii) where multiple individuals of a single species were tested, ascriptions based on ITS and plastid DNA barcodes were incongruent in some samples for 45.2% of the sampled genera (for genera with more than one species sampled). This finding highlights the importance of both sampling multiple individuals and using markers with different modes of inheritance. In cases where it is difficult to amplify and directly sequence ITS in its entirety, just using ITS2 is a useful backup because it is easier to amplify and sequence this subset of the marker. We therefore propose that ITS/ITS2 should be incorporated into the core barcode for seed plants.
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              Molecular Identification of Commercialized Medicinal Plants in Southern Morocco

              Background Medicinal plant trade is important for local livelihoods. However, many medicinal plants are difficult to identify when they are sold as roots, powders or bark. DNA barcoding involves using a short, agreed-upon region of a genome as a unique identifier for species– ideally, as a global standard. Research Question What is the functionality, efficacy and accuracy of the use of barcoding for identifying root material, using medicinal plant roots sold by herbalists in Marrakech, Morocco, as a test dataset. Methodology In total, 111 root samples were sequenced for four proposed barcode regions rpoC1, psbA-trnH, matK and ITS. Sequences were searched against a tailored reference database of Moroccan medicinal plants and their closest relatives using BLAST and Blastclust, and through inference of RAxML phylograms of the aligned market and reference samples. Principal Findings Sequencing success was high for rpoC1, psbA-trnH, and ITS, but low for matK. Searches using rpoC1 alone resulted in a number of ambiguous identifications, indicating insufficient DNA variation for accurate species-level identification. Combining rpoC1, psbA-trnH and ITS allowed the majority of the market samples to be identified to genus level. For a minority of the market samples, the barcoding identification differed significantly from previous hypotheses based on the vernacular names. Conclusions/Significance Endemic plant species are commercialized in Marrakech. Adulteration is common and this may indicate that the products are becoming locally endangered. Nevertheless the majority of the traded roots belong to species that are common and not known to be endangered. A significant conclusion from our results is that unknown samples are more difficult to identify than earlier suggested, especially if the reference sequences were obtained from different populations. A global barcoding database should therefore contain sequences from different populations of the same species to assure the reference sequences characterize the species throughout its distributional range.
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                Author and article information

                Contributors
                suchada.su@chula.ac.th
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                8 September 2020
                8 September 2020
                2020
                : 10
                : 14753
                Affiliations
                [1 ]GRID grid.7922.e, ISNI 0000 0001 0244 7875, Research Unit of DNA Barcoding of Thai Medicinal Plants, Chulalongkorn University Drug and Health Products Innovation Promotion Center, Department of Pharmacognosy and Pharmaceutical Botany, Faculty of Pharmaceutical Sciences, , Chulalongkorn University, ; Bangkok, 10330 Thailand
                [2 ]GRID grid.412665.2, ISNI 0000 0000 9427 298X, College of Pharmacy, , Rangsit University, ; Pathumthani, 12000 Thailand
                [3 ]GRID grid.7922.e, ISNI 0000 0001 0244 7875, Department of Pharmacy Practice, Faculty of Pharmaceutical Sciences, , Chulalongkorn University, ; Bangkok, 10330 Thailand
                [4 ]GRID grid.7922.e, ISNI 0000 0001 0244 7875, Department of Pharmaceutics and Industrial Pharmacy, Faculty of Pharmaceutical Sciences, , Chulalongkorn University, ; Bangkok, 10330 Thailand
                [5 ]GRID grid.444207.1, ISNI 0000 0004 0635 0976, School of Pharmacy, , Eastern Asia University, ; Bangkok, 12110 Thailand
                Article
                71702
                10.1038/s41598-020-71702-7
                7479599
                31913322
                e0db3d5c-391d-4438-bc2c-aaa987c8eb12
                © The Author(s) 2020

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 2 February 2020
                : 17 August 2020
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                © The Author(s) 2020

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                biological techniques,plant sciences
                Uncategorized
                biological techniques, plant sciences

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