Advances in pharmacology, biosynthesis, and metabolic engineering of Scutellaria-specialized metabolites.

Scutellaria Linn., which belongs to the family Lamiaceae, is a commonly used medicinal plant for heat clearing and detoxification. In particular, the roots of S. baicalensis and the entire herb of S. barbata have been widely used in traditional medicine for thousands of years. The main active components of Scutellaria, including: baicalein, wogonin, norwogonin, scutellarein, and their glycosides have potential or existing drug usage. However, the wild resources of Scutellaria plants have been overexploited, and degenerated germplasm resources cannot fulfill the requirements of chemical extraction and clinical usage. Metabolic engineering and green production via microorganisms provide alternative strategies for greater efficiency in the production of natural products. Here, we review the progress of: pharmacological investigations, multi-omics, biosynthetic pathways, and metabolic engineering of various Scutellaria species and their active compounds. In addition, based on multi-omics data, we systematically analyze the phylogenetic relationships of Scutellaria and predict candidate transcription factors related to the regulation of active flavonoids. Finally, we propose the prospects of directed evolution of core enzymes and genome-assisted breeding to alleviate the shortage of plant resources of Scutellaria. This review provides important insights into the sustainable utilization and development of Scutellaria resources.

Comparative Genome Analysis of Scutellaria baicalensis and Scutellaria barbata Reveals the Evolution of Active Flavonoid Biosynthesis.

Scutellaria baicalensis (S. baicalensis) and Scutellaria barbata (S. barbata) are common medicinal plants of the Lamiaceae family. Both produce specific flavonoid compounds, including baicalein, scutellarein, norwogonin, and wogonin, as well as their glycosides, which exhibit antioxidant and antitumor activities. Here, we report chromosome-level genome assemblies of S. baicalensis and S. barbata with quantitative chromosomal variation (2n = 18 and 2n = 26, respectively). The divergence of S. baicalensis and S. barbata occurred far earlier than previously reported, and a whole-genome duplication (WGD) event was identified. The insertion of long terminal repeat elements after speciation might be responsible for the observed chromosomal expansion and rearrangement. Comparative genome analysis of the congeneric species revealed the species-specific evolution of chrysin and apigenin biosynthetic genes, such as the S. baicalensis-specific tandem duplication of genes encoding phenylalanine ammonia lyase and chalcone synthase, and the S. barbata-specific duplication of genes encoding 4-CoA ligase. In addition, the paralogous duplication, colinearity, and expression diversity of CYP82D subfamily members revealed the functional divergence of genes encoding flavone hydroxylase between S. baicalensis and S. barbata. Analyzing these Scutellaria genomes reveals the common and species-specific evolution of flavone biosynthetic genes. Thus, these findings would facilitate the development of molecular breeding and studies of biosynthesis and regulation of bioactive compounds.

Chemical and genetic diversity of Astragalus mongholicus grown in different eco-climatic regions.

Astragalus mongholicus Bunge (Fabaceae) is an important plant source of the herbal drug known as Radix Astragali, which is used worldwide as a medicinal ingredient and a component of food supplement. Russian Federation, Mongolia, Kazakhstan, and China are the main natural distribution areas of A. mongholicus in the world. However, the quality of medicinal plant varies among different locations. As for A. mongholicus, limited literature focused on its biodiversity mechanism. Here, we combined the chemometric analysis of chemical components with genetic variation, as well as climatic and edaphic traits, to reveal the biodiversity mechanism of A. mongholicus. Results showed that the detected chemical, genetic and climatic traits comprehensively contributed to the quality diversity of A. mongholicus. The eight main chemical components, as well as the inorganic elements of P, B and Na were all significant chemical factors. The precipitation and sunshine duration were the main distinguishing climatic factors. The inorganic elements As, Mn, P, Se and Pb were the distinguishing edaphic factors. The systematic method was firstly established for this medicinal plant in order to illustrate the formation of diversity in terms of quality, and provide scientific evidence for geographic indications and climatic adaptation in production and in the clinical application of herbal medicinal plants.

Three-Dimensional Evaluation on Ecotypic Diversity of Traditional Chinese Medicine: A Case Study of Artemisia annua L.

Artemisinin is the first-line drug for anti-malaria recommended by the World Health Organization (WHO). As the sole natural plant source of artemisinin, ecotypes of Artemisia annua L. vary widely in artemisinin content between nations, and China is the main producing area of A. annua. Here we present a three-dimensional evaluation on ecotypic diversity of A. annua from 12 main producing areas in China using high-performance liquid chromatography coupled with evaporative light scattering detection (HPLC-ELSD) method, DNA barcoding and ecological analyses. The results indicated that A. annua exhibited high ecotypic diversity. A. annua grown in the South of the Qinling Mountains-Huaihe River Line had a high artemisinin content, whereas the northern ones were low. Similar pattern was noted in the genetic diversity. The southern A. annua had high intraspecific variation in contrast to the northern A. annua. In terms of ecological analyses, humidity and sunshine time could be the major limiting ecological factors that affect the accumulation of artemisinin. This is the first reported three-dimensional evaluation integrating chemical, molecular and ecological analyses of the ecotypic diversity of A. annua. The work will facilitate exploring the genetic basis of chemical variations and developing strategies for the breeding and cultivation of high quality A. annua.

Distinguishing Astragalus mongholicus and Its Planting Soil Samples from Different Regions by ICP-AES.

"Daodi herb" enjoys a good reputation for its quality and clinical effects. As one of the most popular daodi herbs, Astragalus membranaceus (Fisch.) Bge var. mongholicus (Bge.) Hsiao (A. membranaceus) is popularly used for its anti-oxidant, anti-inflammatory and immune-enhancing properties. In this study, we used inductively coupled plasma atomic emission spectrometry (ICP-AES) technique to investigate the inorganic elements contents in A. mongholicu and its soil samples from daodi area (Shanxi) and non-daodi areas (Inner Mongolia and Gansu). A total of 21 inorganic elements (Pb, Cd, As, Hg, Cu, P, K, Zn, Mn, Ca, Mg, Fe, Se, B, Al, Na, Cr, Ni, Ba, Ti and Sr) were simultaneously determined. Principal component analysis (PCA) was performed to differentiate A. mongholicu and soil samples from the three main producing areas. It was found that the inorganic element characteristics as well as the uptake and accumulation behavior of the three kinds of samples were significantly different. The high contents of Fe, B, Al, Na, Cr and Ni could be used as a standard in the elements fingerprint to identify daodi and non-daodi A. Mongholicus. As the main effective compounds were closely related to the pharmacodynamics activities, the inter-relationships between selected elements and components could reflect that the quality of A. Mongholicus from Shanxi were superior to others to a certain degree. This finding highlighted the usefulness of ICP-AES elemental analysis and evidenced that the inorganic element profile can be employed to evaluate the genuineness of A. mongholicus.

Geoherbalism evaluation of Radix Angelica sinensis based on electronic nose.

Radix Angelica sinensis (Danggui, DG), derived from the dry root of Angelicae sinensis, is popularly used for its antioxidant, hematinic and immuno-enhancement. However, DG from different origins possess different quality, and difficult to identity. In this study, we used electronic nose technique to investigate DG from different producing areas for monitoring the correlation of origin and quality. The electronic nose was employed to establish classification model of DG originated from four main producing areas of Gansu, Yunnan, Sichuan and Hubei in China. Principal component analysis (PCA) and discriminant function analysis (DFA) were performed to differentiate DG samples from four main producing areas. The content of phthalides of DG were determined to confirm the quality changes and investigate its correlation with the odor response values by Gas Chromatography-Mass Spectrometer (GC-MS). The results of PCA and DFA analysis showed that the electronic nose could accurately distinguish DG from four main producing areas. The method of electronic nose for identification could be verified by GC-MS technology, and the main ingredient content was consistent with its odor of DG. In conclusion, electronic nose could effectively identify different origins of DG, and could be applied for rapid identification and quality control of genuine Angelica herbs.

Integrated analysis for identifying radix astragali and its adulterants based on DNA barcoding.

Radix Astragali is a popular herb used in traditional Chinese medicine for its proimmune and antidiabetic properties. However, methods are needed to help distinguish Radix Astragali from its varied adulterants. DNA barcoding is a widely applicable molecular method used to identify medicinal plants. Yet, its use has been hampered by genetic distance, base variation, and limitations of the bio-NJ tree. Herein, we report the validation of an integrated analysis method for plant species identification using DNA barcoding that focuses on genetic distance, identification efficiency, inter- and intraspecific variation, and barcoding gap. We collected 478 sequences from six candidate DNA barcodes (ITS2, ITS, psbA-trnH, rbcL, matK, and COI) from 29 species of Radix Astragali and adulterants. The internal transcribed spacer (ITS) sequence was demonstrated as the optimal barcode for identifying Radix Astragali and its adulterants. This new analysis method is helpful in identifying Radix Astragali and expedites the utilization and data mining of DNA barcoding.

Review of the botanical characteristics, phytochemistry, and pharmacology of Astragalus membranaceus (Huangqi).

Astragalus membranaceus is one of the most widely used traditional Chinese herbal medicines. It is used as immune stimulant, tonic, antioxidant, hepatoprotectant, diuretic, antidiabetic, anticancer, and expectorant. The current paper reviews the botanical characteristics, phytochemistry, and pharmacology of Astragali Radix. Information on Astragali Radix was gathered via the Internet (using Google Scholar, Baidu Scholar, Elsevier, ACS, Medline Plus, CNKI, and Web of Science) as well as from libraries and local books. More than 100 compounds, including flavonoids, saponins, polysaccharides, and amino acids, have so far been identified, and the various biological activities of the compounds have been reported. As an important traditional Chinese medicine, further studies on Astragali Radix can lead to the development of new drugs and therapies for various diseases. The improvement of its utilization should be studied further.

Ultra-performance liquid chromatography-quadrupole\time-of- flight mass spectrometry with multivariate statistical analysis for exploring potential chemical markers to distinguish between raw and processed Rheum palmatum.

BACKGROUND: The long term use of Rheum palmatum for the treatment of diseases associated with chronic hepatitis and renal failure can lead to liver and kidney damage. To reduce the toxicity of R. palmatum and alleviate any symptoms of decanta and celialgia, the raw material has been subjected to a specific process prior to its use for hundreds of years. Despite its extensive use in medicine, very little is currently known about the nature of the components present in this material in terms of their efficacy and overall toxicity, and the effect that processing has on the levels of these components in the processed material. The aim of this investigation was to explore potential differences in the chemical markers between batches of raw and processed R. palmatum and to develop a deeper understanding of the underlying mechanisms responsible for the enhanced efficacy and reduced toxicity of the processed material. METHODS: Raw and processed R. palmatum samples were analyzed by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF-MS) coupled with multivariate statistical analysis using principal component analysis (PCA) and orthogonal partial least square discriminant analysis (OPLS-DA). RESULTS: The emodin-8-O-glucoside, emodin-O-glucoside, catechin-glucopyranoside, gallic acid-3-O-glucoside, torachrysone, and chrysophanol dimethyl ether were rapidly explored as representative markers to distinguish for the first time between the raw and processed R. palmatum material. Among the potential chemical markers, Emodin-8-O-glucoside and gallic acid-3-O-glucoside were determined to be the best markers for the raw and processed R. palmatum. CONCLUSION: UPLC/Q-TOF-MS with multivariate statistical analysis represents an efficient method for exploring the chemical markers in the raw and processed R. palmatum material, as well as investigating the mechanisms associated with the processing, quality control, and safe application of R. palmatum.

[Identification of radix et rhizoma clematidis and its adulterants using DNA barcoding].

This study provides the candidate sequences in the identification of Radix et Rhizoma Clematidis and its adulterants using DNA barcoding. We amplified and sequenced the region psbA-trnH, with the data of 284 sequences from GenBank, the differential intra- and inter-specific divergences, genetic distance, barcoding gap were used to evaluate five barcodes, and the identification efficiency was assessed using BLAST1 and Nearest Distance methods. The results showed that psbA-trnH barcodes performed high identification efficiency and inter-specific divergences among the five different DNA barcodes. Analysis of the barcoding gap and NJ tree showed psbA-trnH was superior to other barcodes. Based on the identification and PCR amplification efficiency, psbA-trnH can be the ideal barcode to identify Radix et Rhizoma Clematidis and its adulterants accurately.

Sulfur fumigation, a better or worse choice in preservation of Traditional Chinese Medicine?

Sulfur fumigation (SF) in Traditional Chinese Medicine (TCM) is a highly efficient and important traditional preservation method in China. This method has generated a great deal of concern and has been disputed in the last few years because of its uncertain safety. SF can alter the quality of TCMs by damaging the bioactive compounds, changing chemical profiles, and generating detrimental exogenous materials. However, SF is still widely used in the herbal medicinal industry because of its various benefits, such as its pesticidal and anti-bacterial effects, easy operation, and low-cost. This review contains the current situation, chemical mechanism and reactions during SF, the pharmacological and pharmacokinetic research, and the influence of quality caused by SF. In addition, a quantification-operation sulfur fumigation device (QOSFD), which can maintain the quality of TCMs by controlling the SF processing parameters, has been designed and introduced. The key technologies of this device involve controlling the O(2) content and the temperature of SO(2) as well as the quantification of sulfur in SF. This device can reduce the possibility of reaction between bioactive compounds and sulfur/sulfurous acid, as well as control the limitation of SO(2) residues. The QOSFD is regarded as a promising preservation technique in the field of TCM, medicinal materials, agriculture, and fruit industry.

[Identification of gentianae macrophyllae radix using the ITS2 barcodes].

DNA barcoding is a rapidly developing frontier technology in the world and will be useful in promoting the quality control and standardization of traditional Chinese medicine. Until now, many studies concerning DNA barcoding have focused on leaf samples but rarely on Chinese herbal medicine. There are three issues involved in DNA barcoding for traditional Chinese medicinal materials: (1) the extraction methods for total DNA of the rhizomes of the medicinal materials; (2) intra-specific variation among samples from different places of origin; (3) accuracy and stability of this method. In this study, Gentianae Macrophyllae Radix was used to verify the stability and accuracy of DNA barcoding technology. Five regions (ITS2, psbA-trnH, matK, rbcL, and ITS) were tested for their ability to identify 86 samples of Gentianae Macrophyllae Radix and their adulterants. After improving the DNA extraction method, genomic DNA from all samples was successfully obtained. To evaluate each barcode's utility for species authentication, PCR amplification efficiency, genetic divergence, and species authentication were assessed. Among all tested regions only ITS2 locus showed 100% of PCR amplification and identification efficiencies. Based on the established method, we successfully identified two samples of Gentianae Macrophyllae Radix bought in pharmacy to the original species.