Abstract
With the widespread use of traditional medicine around the world, the safety and efficacy of traditional herbal patent medicine have become an increasing concern to the public. However, it is difficult to supervise the authenticity of herbal materials in mixed herbal products according to the current quality standards, especially for traditional herbal patent medicine, with a distinct variance in the dosage of herbal materials. This study utilized the shotgun metabarcoding approach to analyze the biological ingredients of Fuke Desheng Wan (FKDSW), which is an effective traditional herbal product for the treatment of dysmenorrhea. Six herbal materials were collected, and a lab-made mock FKDSW sample was produced to establish a method for the authentication assessment of biological ingredients in traditional herbal patent medicine based on shotgun metabarcoding. Furthermore, four commercial FKDSW samples were collected to verify the practicality of the shotgun metabarcoding approach. Then, a total of 52.16 Gb raw data for 174 million paired-end reads was generated using the Illumina NovaSeq sequencing platform. Meanwhile, 228, 23, and 14 operational taxonomic units (OTUs) were obtained for the ITS2, matK, and rbcL regions, respectively, after bioinformatic analysis. Moreover, no differences were evident between the assembly sequences obtained via shotgun metabarcoding and their corresponding reference sequences of the same species obtained via Sanger sequencing, except for part of the ITS2 and matK assembly sequences of Paeonia lactiflora Pall., Saussurea costus (Falc.) Lipsch. and Bupleurum chinense DC. with 1–6 different bases. The identification results showed that all six prescribed ingredients were successfully detected and that the non-authentic ingredient of Bupleuri Radix (Chaihu, Bupleurum chinense DC. or Bupleurum scorzonerifolium Willd.) was found in all the commercial samples, namely Bupleurum falcatum L. Here, 25 weed species representing 16 genera of ten families were detected. Moreover, 26 fungal genera belonging to 17 families were found in both lab-made and commercial FKDSW samples. This study demonstrated that the shotgun metabarcoding approach could overcome the biased PCR amplification and authenticate the biological ingredients of traditional herbal patent medicine with a distinct variance in the dosage of the herbal materials. Therefore, this provides an appropriate evaluation method for improving the safety and efficacy of traditional herbal patent medicine.
Highlights
Dysmenorrhea is a common gynecological disease that can harm the health, work status, and quality of life of women
The reads after quality control were aligned to the reference database using basic local alignment search tool (BLAST), the targeted paired-end reads of ITS2, matK, and rbcL were enriched by local python
Xin et al (2018a) applied shotgun metagenomic sequencing for the first time to detect the biological ingredients in Longdan Xiegan Wan, verifying the feasibility of the method for monitoring the species composition in traditional herbal patent medicine
Summary
Dysmenorrhea is a common gynecological disease that can harm the health, work status, and quality of life of women. Fuke Desheng Wan (FKDSW) has been recommended as a gynecological medicine for curing dysmenorrhea caused by liver depression or the stagnancy of both blood and qi (Commission, 1994). FKDSW is a honeyed pill consisting of six herbal materials, including Angelicae Sinensis Radix (Danggui), Paeoniae Radix Alba (Baishao), Aucklandiae Radix (Muxiang), Notopterygii Rhizoma et Radix (Qianghuo), Leonuri Herba (Yimucao), and Bupleuri Radix (Chaihu), at significantly different and very precise dosages. The quality control and biological ingredient assessment of FKDSW are challenging compared to other traditional herbal patent medicines with similar dosage levels of each ingredient. In this case, the characteristics of medicinal materials with low content will be overwhelmed by the characteristics of medicinal materials with high content. There are 7% of herbal products of the Lonicerae japonicae Flos contained both of two adulterants Eucommiae Folium and Lonicerae Flos (Gao et al, 2017)
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