Transcriptome analysis reveals cadmium exposure enhanced the isoquinoline alkaloid biosynthesis and disease resistance in Coptis chinensis.

Coptis chinensis Franch is a perennial herb from the Ranunculaceae family with a long history of medicinal use. As the medicinal part, the rhizome of coptis often accumulates excessive cadmium (Cd) even at low concentrations in the soil, which not only compromises its medicinal safety but also raises concerns about adverse effects on human health. Therefore, effective strategies are needed to mitigate this accumulation and ensure its safe use in traditional medicine. This study utilized transcriptome profiling and physiological analysis to explore molecular mechanisms associated with ecological significance and the active accumulation of Cd in C. chinensis. The response to Cd in C. chinensis was assessed through RNA sequencing, Cd determination and isoquinoline alkaloid measurement using its roots, stems, and leaves. The transcriptome revealed, a total of 2667, 2998, or 2815 up-regulated deferentially expressed genes in roots, stems or leaves in response to Cd exposure. Furthermore, we identified phenylpropanoid and isoquinoline alkaloid biosynthesis as the key pathways response to Cd exposure, which suggests that C. chinensis may improve its tolerance to Cd through regulating the phenylpropanoid biosynthesis pathway. Under Cd exposure, plant-pathogen interaction in leaves was identified as the key pathway, which indicates that upregulation of genes involved in plant-pathogen interaction could enhance disease resistance in C. chinensis. WGCNA analysis identified WRKY8 (Cluster-55763.31419) and WRKY47 (Cluster-55763.221590) as potential regulators of secondary metabolic synthesis and plant-pathogen interaction pathway in C. chinensis triggered by Cd. The measurement of berberine, coptisine, palmatine, and epiberberine also demonstrated that Cd simulated the four isoquinoline alkaloids in roots. Therefore, our study not only presented a transcriptome expression profiles that revealed significant upregulation of genes involved in metal transport and detoxification pathways but also suggested a possible mechanism to cope with Cd accumulation. This knowledge provides a new insight into gene manipulation for controlling Cd accumulation, enhancing resistance and promoting synthesis of secondary metabolites with potential medicinal properties in other medicinal plant species.

DNA methylation regulates biosynthesis of tanshinones and phenolic acids during growth of Salvia miltiorrhiza.

DNA methylation plays a crucial role in the regulation of plant growth and the biosynthesis of secondary metabolites. Danshen (Salvia miltiorrhiza) is a valuable Chinese herbal medicine commonly used to treat cardiovascular diseases; its active ingredients are tanshinones and phenolic acids, which primarily accumulate in roots. Here, we conducted a targeted metabolic analysis of S. miltiorrhiza roots at 3 distinct growth stages: 40 d old (r40), 60 d old (r60), and 90 d old (r90). The contents of tanshinones (cryptotanshinone, tanshinone I, tanshinone IIA, and rosmariquinone) and phenolic acids (rosmarinic acid and salvianolic acid B) gradually increased during plant development. Whole-genome bisulfite sequencing and transcriptome sequencing of roots at the 3 growth stages revealed an increased level of DNA methylation in the CHH context (H represents A, T, or C) context at r90 compared with r40 and r60. Increased DNA methylation levels were associated with elevated expression of various genes linked to epigenetic regulations, including CHROMOMETHYLASE2 (SmCMT2), Decrease in DNA Methylation 1 (SmDDM1), Argonaute 4 (SmAGO4), and DOMAINS REARRANGED METHYLTRANSFERASE 1 (SmDRM1). Moreover, expression levels of many genes involved in tanshinone and salvianolic acid biosynthesis, such as copalyldiphosphate synthase 5 (SmCPS5), cytochrome P450-related enzyme (SmCYP71D464), geranylgeranyl diphosphate synthase (SmGGPPS1), geranyl diphosphate synthase (SmGPPS), hydroxyphenylpyruvate reductase (SmHPPR), and hydroxyphenylpyruvate dioxygenase (SmHPPD), were altered owing to hyper-methylation, indicating that DNA methylation plays an important role in regulating tanshinone and phenolic acid accumulation. Our data shed light on the epigenetic regulation of root growth and the biosynthesis of active ingredients in S. miltiorrhiza, providing crucial clues for further improvement of active compound production via molecular breeding in S. miltiorrhiza.

Research on Coix seed as a food and medicinal resource, it's chemical components and their pharmacological activities: A review.

ETHNOPHARMACOLOGICAL RELEVANCE: Coix lacryma-jobi var. ma-yuen (Romanet du Caillaud) Stapf is a plant of the genus Coix in the Gramineae family. Coix seed is cultivated in various regions throughout China. In recent years, with the research on the medicinal value of Coix seed, it has received more and more widespread attention from people. Numerous pharmacological effects of Coix seed have been demonstrated through modern pharmacological studies, such as hypoglycemia, improving liver function, anti-tumor, regulating intestinal microbiota, improving spleen function, and anti-inflammatory effects. AIMS OF THE STUDY: This article is a literature review. In recent years, despite the extensive research on Coix seed, there has yet to be a comprehensive review of its traditional usage, medicinal resources, chemical components, and pharmacological effects is still lacking. To fill this gap, the paper provides an overview of the latest research progress on Coix seed, aiming to offer guidance and references for its further development and comprehensive utilization. MATERIAL AND METHODS: To gather information on the traditional usage, phytochemical ingredients, and pharmacological properties of Coix seed, we conducted a literature search using both Chinese and English languages in five databases: PubMed, Web of Science, China National Knowledge Infrastructure (CNKI), and Springer. RESULTS: This article is a literature review. The chemical constituents of Coix seed include various fatty acids, esters, polysaccharides, sterols, alkaloids, triterpenes, tocopherols, lactams, lignans, phenols, flavonoids and other constituents. Modern pharmacological research has indeed shown that Coix seed has many pharmacological effects and is a natural anti-tumor drug. In addition to its anti-tumor effect, it also has pharmacological effects such as hypoglycemia, improving liver function, regulating intestinal microbiota, improving spleen function, and anti-inflammatory effects. CONCLUSIONS: This article provides a brief overview of the traditional uses, biotechnological applications, chemical components, and pharmacological effects of Coix seed. It highlights the importance of establishing quality standards, discovering new active ingredients, and exploring pharmacological mechanisms in Coix seed research. The article also emphasizes the significance of clinical trials, toxicology studies, pharmacokinetics data, and multidisciplinary collaboration for further advancements in this field. Overall, it aims to enhance understanding of Coix seed and its potential in pharmaceutical development and wellness products.

Endophytic fungus Penicillium steckii DF33 promoted tanshinones biosynthesis in Salvia miltiorrhiza by regulating the expression of CYP450 genes.

Salvia miltiorrhiza, a prominent traditional Chinese medicinal resource, has been extensively employed in the management of cardiovascular and cerebrovascular ailments. Ensuring the consistency of S. miltiorrhiza raw materials revolves around the imperative task of maintaining stable tanshinones content and composition. An effective approach in this regard involves the utilization of endophytic fungi as inducers. Within this context, our study spotlights an endophytic fungus, Penicillium steckii DF33, isolated from the roots of S. miltiorrhiza. Remarkably, this fungus has demonstrated a significant capacity to boost the biosynthesis and accumulation of tanshinones. The primary objective of this investigation is to elucidate the underlying regulatory mechanism by which DF33 enhances and regulates the biosynthesis and accumulation of tanshinones. This is achieved through its influence on the differential expression of crucial CYP450 genes within the S. miltiorrhiza hairy roots system. The results revealed that the DF33 elicitor not only promotes the growth of hairy roots but also enhances the accumulation of tanshinones. Notably, the content of cryptotanshinone was reached 1.6452 ± 0.0925 mg g-1, a fourfold increase compared to the control group. Our qRT-PCR results further demonstrate that the DF33 elicitor significantly up-regulates the expression of most key enzyme genes (GGPPS, CPS1, KSL1, CYP76AH1, CYP76AH3, CYP76AK1, CYP71D411) involved in the tanshinone biosynthesis pathway. This effect is particularly pronounced in certain critical CYP450 genes and Tanshinone ⅡA synthase (SmTⅡAS), with their expression levels peaking at 7 days or 14 days, respectively. In summary, endophytic P. steckii DF33 primarily enhances tanshinone biosynthesis by elevating the expression levels of pivotal enzyme genes associated with the modification and transformation stages within the tanshinone biosynthesis pathway. These findings underscore the potential of employing plant probiotics, specifically endophytic and root-associated microbes, to facilitate the biosynthesis and transformation of vital constituents in medicinal plants, and this approach holds promise for enhancing the quality of traditional Chinese medicinal materials.

A Review on the Morphology, Cultivation, Identification, Phytochemistry, and Pharmacology of Kitagawia praeruptora (Dunn) Pimenov.

Kitagawia praeruptora (Dunn) Pimenov, commonly known as Qianhu in China, is a widely used folk Chinese herbal medicine. This article reviews its botanical traits, ethnopharmacology, cultivation techniques, identification, phytochemical compositions, and pharmacological effects. Over 70 coumarin compounds, including simple coumarins, pyranocoumarins, and furanocoumarins, have been isolated within this plant. Additionally, K. praeruptora contains other components such as flavonoids, fatty acids, benzoic acids, and sterols. This information highlights the importance of utilizing active ingredients and excavating pharmacological effects. With its remarkable versatility, K. praeruptora exhibits a wide range of pharmacological effects. It has been found to possess expectorant and bronchodilator properties, cardiovascular protection, antimicrobial and antioxidant activities, anti-tumor effects, and even antidiabetic properties. It is recommended to focus on the development of new drugs that leverage the active ingredients of K. praeruptora and explore its potential for new clinical applications and holistic utilization.

Proanthocyanidin Structure-Activity Relationship Analysis by Path Analysis Model.

To fully explore the influence mechanism of interactions between different monomer units of proanthocyanidins (PAs) on biological activity, a path analysis model of the PA structure-activity relationship was proposed. This model subdivides the total correlation between each monomer unit and activity into direct and indirect effects by taking into account not only each monomer unit but also the correlation with its related monomer units. In addition, this method can determine the action mode of each monomer unit affecting the activity by comparing the direct and total indirect effects. Finally, the advantage of this model is demonstrated through an influence mechanism analysis of Rhodiola crenulata PA monomer units on antioxidant and anti-diabetes activities.

Assessment of Genetic Diversity in Bupleurum spp. Basing Agronomic Traits, Medicinal Components and ISSR Markers.

Radix bupleuri is one of the bulk medicinal materials in China and it is widely adopted in clinical applications and drug discovery. The investigation of agronomic traits, active component content and genetic diversity in diverse Radix bupleuri germplasms may provide evidence to promote the selection of better strains. In this research, 13 germplasms from various sources were used to investigate the variations between different Radix bupleuri germplasms. Nine biological characteristics were noted in the field, and the levels of the two primary active ingredients were determined using high performance liquid chromatography (HPLC). Moreover, the molecular marker technique of inter-simple sequence repeat (ISSR) and the unweighted pair group method with arithmetic means (UPGMA) were employed to evaluate the molecular genetic diversity. The findings showed that there was a wide range of variation among the many varieties of Radix bupleuri, with coefficients of variation for agronomic traits and active component content ranging from 7.62% to 41.54% and 36.47% to 53.70%, respectively. Moreover, there are different degrees of relationship between the two. Since there was a significant correlation between root weight and saikosaponin content, it was possible to classify a plant based on its weight and anticipate its saikosaponin content. The 13 species were divided into four groups based on their germplasm by genetic markers-based cluster analysis. This indicated the possibility that the component content would not necessarily be related to germplasm and might easily be influenced by environmental factors. The use of ISSR marker technology made it possible to precisely identify the various Radix bupleuri provenances and its counterfeit products. There may be a way to prevent the misunderstandings caused by the appearance and composition of Chinese medicinal substances. In our study, the germplasm of Radix bupleuri that was widely circulated in the market was comprehensively evaluated in terms of agronomic traits, active components and molecular level, and identified by simple means, to provide a theoretical basis for the evaluation and screening of fine germplasms of Radix bupleuri.

Antioxidative and Cytoprotective Effects of Ganoderma applanatum and Fomitopsis pinicola in PC12 Adrenal Phaeochromocytoma Cells.

Considering the impact of oxidative stress on the development of many diseases, together with the role of natural antioxidants in maintaining physiological balance in humans, medicinal mushrooms are potential sources of bioactive compounds against many diseases. In the present work, in vitro evaluation of the biological activities of the alcoholic extracts of two wild tree mushrooms, namely, Ganoderma applanatum and Fomitopsis pinicola, has been performed. Extraction of G. applanatum (GAE) and F. pinicola (FPE) was conducted with 60% ethanol and 100% ethanol sequentially. UPLC-MS/MS identification was conducted on the two mushrooms extracts. A total of 15 substances were identified in GAE, including 3 spiro meroterpenoids and 12 triterpenoids; a total of 14 chemical constituents were iden¬tified in FPE, including 8 triterpenoids, 4 triterpene glycosides, 1 lanosterol, and 1 lanostanoid. The resulting extracts were examined for their in vitro antioxidative and cytoprotective effects against AAPH-induced oxidative damage. Our results demonstrated that both extracts have potent antioxidative activities, when GAE was 0.2 mg/mL, the clearance rates of DPPH and ABTS have reached 93.34% and 99.93%, respectively. When FPE was 1.4 mg/mL and 0.6 mg/mL, the scavenging rates of DPPH and ABTS have reached 91.76% and 100%, respectively. Both the alcoholic extracts of G. applanatum and F. pinicola were able to protect the AAPH-induced damage and could effectively inhibit cell aging via ß-galactosidase (SA ß-gal) staining activity test and scanning electron microscopy analysis.

Dynamic analysis of physiological indices and transcriptome profiling revealing the mechanisms of the allelopathic effects of phenolic acids on Pinellia ternata.

Pinellia ternata (Thunb.) is a famous traditional Chinese medicine with high medicinal value, but its culture is strongly hindered by the continuous cropping obstacles (CCO) which are tightly associated with allelopathic effects. Deciphering the response mechanisms of P. ternata to allelochemicals is critical for overcoming the CCO. Here, we elucidate the response of P. ternata to phenolic acids treatment via physiological indices, cellular approaches, and transcriptome analysis. Phenolic acids showed a significant effect on the growth of P. ternata seedlings, similar to the phenotype of continuous cropping. Cellular analysis demonstrated that phenolic acids remarkably induced root cell death. Physiological analysis revealed that phenolic acids induced the overaccumulated of H2O2 and O 2 - in root cells. However, two exogenous antioxidants (L-ascorbic acid and ß-gentiobiose) aid in the scavenging of over-accumulated H2O2 and O 2 - by promoting the antioxidant enzyme activity such as superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT). Transcriptome analysis demonstrated that differentially expressed genes (DEGs) related to the cell wall degeneration and reactive oxygen species (ROS) metabolism were upregulated by phenolic acid treatment. In addition, downregulated DEGs involved in sucrose and starch metabolism and phenylpropanoid biosynthesis pathways decreased the key metabolites contents. Taken together, phenolic acids caused root cell death by inducing the overaccumulation of H2O2 and O 2 - , and L-ascorbic acid and ß-gentiobiose effectively alleviated ROS stress. The present study elucidates the underlying mechanism of the allelopathic effect of phenolic acids, offers valuable information for further understanding the mechanism of CCO, and could contribute to improving guidance for further P. ternata production.

Systematic Identification and Validation of Suitable Reference Genes for the Normalization of Gene Expression in Prunella vulgaris under Different Organs and Spike Development Stages.

The quantitative real-time PCR (qRT-PCR) is an efficient and sensitive method for determining gene expression levels, but the accuracy of the results substantially depends on the stability of the reference gene (RG). Therefore, choosing an appropriate reference gene is a critical step in normalizing qRT-PCR data. Prunella vulgaris L. is a traditional Chinese medicine herb widely used in China. Its main medicinal part is the fruiting spike which is termed Spica Prunellae. However, thus far, few studies have been conducted on the mechanism of Spica Prunellae development. Meanwhile, no reliable RGs have been reported in P. vulgaris. The expression levels of 14 candidate RGs were analyzed in this study in various organs and at different stages of Spica Prunellae development. Four statistical algorithms (Delta Ct, BestKeeper, NormFinder, and geNorm) were utilized to identify the RGs' stability, and an integrated stability rating was generated via the RefFinder website online. The final ranking results revealed that eIF-2 was the most stable RG, whereas VAB2 was the least suitable as an RG. Furthermore, eIF-2 + Histon3.3 was identified as the best RG combination in different periods and the total samples. Finally, the expressions of the PvTAT and Pv4CL2 genes related to the regulation of rosmarinic acid synthesis in different organs were used to verify the stable and unstable RGs. The stable RGs in P. vulgaris were originally identified and verified in this work. This achievement provides strong support for obtaining a reliable qPCR analysis and lays the foundation for in-depth research on the developmental mechanism of Spica Prunellae.

Characterization and Comparative Analysis of Chloroplast Genomes in Five Uncaria Species Endemic to China.

Uncaria, a perennial vine from the Rubiaceae family, is a typical Chinese traditional medicine. Currently, uncertainty exists over the Uncaria genus' evolutionary relationships and germplasm identification. The complete chloroplast genomes of four Uncaria species mentioned in the Chinese Pharmacopoeia and Uncaria scandens (an easily confused counterfeit) were sequenced and annotated. The findings demonstrated that the whole chloroplast genome of Uncaria genus is 153,780-155,138 bp in full length, encoding a total of 128-131 genes, containing 83-86 protein-coding genes, eight rRNAs and 37 tRNAs. These regions, which include eleven highly variable loci and 31-49 SSRs, can be used to create significant molecular markers for the Uncaria genus. The phylogenetic tree was constructed according to protein-coding genes and the whole chloroplast genome sequences of five Uncaria species using four methods. The topology of the two phylogenetic trees showed no difference. The sequences of U. rhynchophylla and U. scandens are clustered in one group, while the U. hirsuta and U. macrophylla are clustered in another group. U. sessilifructus is clustered together with the above two small clades. New insights on the relationship were revealed via phylogenetic research in five Uncaria species. This study will provide a theoretical basis for identifying U. rhynchophylla and its counterfeits, as well as the species of the Uncaria genus. This research provides the initial chloroplast genome report of Uncaria, contributes to elucidating the chloroplast genome evolution of Uncaria in China.

Antiviral activities of Polygonum perfoliatum L. extract and related phenolic acid constituents against hepatitis B virus.

Chronic hepatitis B virus (HBV) infection is an important public health problem. Polygonum perfoliatum L. is a traditional medicinal herb and has been reported to have pharmacological activities such as anti-inflammatory, antibacterial, and antiviral. In this study, the antiviral activities and mechanisms of Polygonum perfoliatum L. extract against HBV and the effective components were investigated. The results showed that the total extract of Polygonum perfoliatum L. reduced the levels of HBV e antigen (HBeAg) secretion and the viral covalently closed circular DNA (CCC DNA) formation, but had little or no negative effects on viral capsid assembly and pregenomic RNA packaging. Further fractionation showed that the water extract (WE) fraction exerted comparable anti-HBV activities with the total extract, especially in inhibiting the CCC DNA formation and HBeAg production, indicating that the effective antiviral components are mainly distributed in this fraction. Further study showed that the phenolic acids constituents, protocatechuic acid, and gallic acid, but not ethyl caffeate, which is reported enriched in the WE fraction, showed strong anti-HBV activities in inhibiting viral core DNA synthesis, CCC DNA formation, and HBeAg production. These results suggested that the Polygonum perfoliatum L. total extract and the related phenolic acids like protocatechuic acid and gallic acid could inhibit HBV replication and also indicated the potential utility of Polygonum perfoliatum L. and related constituents as sources of novel antivirals against HBV.

Multi-algorithm cooperation comprehensive research of bZIP genes under nitrogen stress in Panax notoginseng.

Basic leucine zipper (bZIP) transcription factors play an irreplaceable position in the regulation of plant secondary metabolism, growth and development, and resistance to abiotic stress. Panax notoginseng is a traditional medicinal plant in China, but the systematic identification and the resistance of Panax notoginseng bZIP (PnbZIP) family under nitrogen stress have not been reported before, considering the excessive application of N fertilizers. In this study, we conducted a genome-wide identification of the PnbZIP family and analyzed its phylogeny, tissue selectivity, and abiotic resistence. 74 PnbZIPs were distributed on 12 chromosomes and 8 were not successfully located. Through phylogenetic analysis of Arabidopsis and Panax notoginseng, we divided them into 14 subgroups. In the same subgroup, bZIPs had similiar intron/exon structure and conserved motifs. In the analysis of chromosome structure, two PnbZIP genes were duplicated in tandem on chromosome 3. Intraspecific collinearity analysis showed that 28 PnbZIPs participated in segmental replication. Each PnbZIP promoter contained at least one stress response element or stress-related hormone response element. RNA-seq and qRT-PCR methods were used to analyze the expression patterns of the PnbZIP gene in different tissues (roots, flowers, and leaves) and under different nitrogen stresses. The results showed that the PnbZIP gene had the highest expression level in flowers and reflected tissue-specific expressions. Meanwhile, under the stress of ammonium nitrogen fertilizer and nitrate nitrogen fertilizer, PnbZIPs in roots were differently expressed. 10 PnbZIP stress-responsive genes were screened for significant expression, among which PnbZIP46 was significantly up-regulated, which could be a candidate gene for resistance to Nitrogen stress. This study laid the foundation for functional identification of PnbZIPs and improved the cultivation of Panax notoginseng.

[Three-dimensional multi-component quality evaluation of Chinese medicine based on proportion consistency of active components: a study of Salvia miltiorrhiza].

To reveal the law of the proportion consistency of the active components in Chinese medicine and explore a new way to evaluate the quality of Chinese medicine, the present study investigated the content of tanshinone Ⅱ_A, cryptotanshinone, tanshinone Ⅰ, salvianolic acid B, rosmarinic acid, caffeic acid, and lithospermic acid in 895 samples of Salvia miltiorrhiza from 87 literature articles. The samples with salvianolic acid B and tanshinones(total content of tanshinone Ⅱ_A, tanshinone Ⅰ, and cryptotanshinone) meeting the requirements of Chinese Pharmacopoeia were analyzed for proportion consistency of components, and the proportion distribution of components was compared. The results showed that the proportions in and between tanshinones and salvianolic acids were stable. The content ratios of cryptotanshinone to tanshinone Ⅱ_A, rosmarinic acid to salvianolic acid B, and salvianolic acid B to tanshinone Ⅱ_A fluctuated from 0.1 to 1.7, from 0.02 to 0.14, and from 10 to 60, respectively. It indicated that the content proportions of different components were not fixed, but fluctuated in a certain range. The fluctuation range of components in the same group was small, but that in different groups was large. The content proportions of active components were stable for Chinese medicinal materials with fixed varieties or origins. The samples whose content proportions were out of the fluctuation ranges were generally unqualified medicinal materials. We proposed a "three-dimensional multi-component" quality evaluation model of Chinese medicine based on the above findings. Specifically, "three-dimensional" means to accurately evaluate the quality of Chinese medicine from varieties, proportions, and content of active components, and "multi-component" means to highlight the information of multiple components in Chinese medicine, especially the quality markers and equivalent component groups. The proportion of components in Chinese medicine can be used as an important index for the quality evaluation of Chinese medicine. The proportions of active components are stable in Chinese medicinal materials with fixed varieties or places of origin.

Structural and interactions analysis of a transcription factor PnMYB2 in Panax notoginseng.

The main active ingredients of the traditional Chinese medicinal plant, Panax notoginseng, are the Panax notoginseng saponins (PNS). They can be synthesized via the mevalonate pathway; PnSS and PnSE1 are the key rate-limiting enzymes in this pathway. In this study, an interaction between PnMYB2 and the key enzymes was identified and characterized from the P. notoginseng cDNA library using the Y1H technique. Subsequently, X-α-gal color reaction confirmed the interaction between PnMYB2 and the upstream sequences of PnSS and PnSE1 promoters. Full-length cDNA sequence of PnMYB2 was isolated and characterized. PnMYB2 has an open reading frame of 864 bp, encoding 287 amino acids. 3D structural analysis of PnMYB2 indicated that its structure was similar to that of the template. Phylogenetic analysis revealed that PnMYB2 and PgMYB2 are highly homologous and belong to the R2R3 MYB transcription factor (TF). Subcellular localization analysis showed that PnMYB2 was localized in the nucleus. The recombinant protein PnMYB2 was successfully obtained through prokaryotic expression and was confirmed to be an inclusion body protein. Furthermore, electrophoretic mobility shift assay (EMSA) experiments demonstrated that PnMYB2 specifically binds to MYB core and AC-rich elements. This study provides a theoretical basis for transcriptional regulation of saponin biosynthesis in P. notoginseng.

The complete chloroplast genome sequence of traditional Chinese medicine Uncaria macrophylla (Rubiaceae).

Uncaria macrophylla (Rubiaceae) is a medicinal vine plant of the Rubiaceae family that was distributed in East Asia and Southeast Asia. The first complete chloroplast genome of Uncaria macrophylla was sequenced and assembled in this study. The genome is 155,138 bp in length and contained 129 encoded genes in total, including 79 protein-coding genes, eight ribosomal RNA genes, and 37 transfer RNA genes. The phylogenomic analysis showed that U. macrophylla was closely related to Uncaria rhynchophylla according to the current sampling extent.

Inhibitory activities of Ranunculus japonicus Thunb. ethanol extract against hepatitis B virus.

The chronic hepatitis B virus (HBV) infection is a worldwide public health problem, which cannot be cured by current therapeutics due to the persistence of viral CCC DNA in the infected hepatocytes. Screening from medicinal herbs for anti-HBV activities showed that the ethanol extract from Ranunculus japonicus Thunb. could decrease the production of HBV e antigen (HBeAg). Further study showed that the extract had no effect on core protein expression but significantly reduced the efficiency of viral capsid assembly. The levels of viral pgRNA and total core DNA were not affected significantly. However, the ratio of RC DNA/SS DNA decreased, indicating that the conversion of RC DNA from SS DNA was delayed by the extract. More interestingly, though similar levels of RC DNA were accumulated, the CCC DNA level and its formation efficiency were reduced significantly, which was also consistent with the decreased level of HBeAg, indicating that R. japonicus Thunb. extract could inhibit the CCC DNA formation. Together, this study found that R. japonicus Thunb. extract could inhibit HBV replication at multiple steps, especially showed significant inhibitory effects on capsid assembly and CCC DNA formation.

Transcriptome and HPLC Analysis Reveal the Regulatory Mechanisms of Aurantio-Obtusin in Space Environment-Induced Senna obtusifolia Lines.

Senna obtusifolia is a famous medicinal plant that is widely used in Asian countries. Its seed plays an important role in the treatment of many diseases because it contains various anthraquinones and flavonoids. Our previous studies have indicated that three space environment-induced S. obtusifolia lines (SP-lines) i.e., QC10, QC29, and QC46, have higher seed yield and aurantio-obtusin (AO) content. However, the underlying mechanism of higher AO content in SP-lines is still unknown. Herein, transcriptome sequencing and HPLC were employed to analyze the differences between SP-lines and ground control (GC3) and elucidate the regulatory mechanisms of AO accumulation in SP-lines. The results show that 4002 differentially expressed genes (DEGs) were identified in SP-lines versus (vs.) GC3. DEGs in the QC10 vs. GC3, QC29 vs. GC3, and QC46 vs. GC3 comparisons were classified into 28, 36, and 81 GO terms and involved in 63, 74, and 107 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. KEGG pathway and gene expression analysis revealed that DEGs involved in anthraquinone pathways were significantly elevated in QC10 and QC46. Integrating the results of GO annotation, KEGG enrichment, and gene expression analysis, we propose that the elevated genes such as DAHPS, DHQS, and MenB enhance the metabolic flux in the anthraquinone pathway and promote AO content in QC10 and QC46. Taken together, this study elucidated the mechanism of AO content in SP-lines and provides valuable genetic information for S. obtusifolia. In addition, to the best of our knowledge, this study presents the first transcriptome analysis of environment-induced medicinal plants and paves the way to select elite S. obtusifolia varieties in the future.

Systematic investigation on the distribution of four hidden toxic Aconitum alkaloids in commonly used Aconitum herbs and their acute toxicity.

Yunaconitine (YAC), crassicauline A (CCA), 8-deacetylyunaconitine (DYA), and 8-deacetylcrassicauline A (DCA), as hidden toxic Aconitum alkaloids, are detected in some products of processed Aconitum carmichaelii lateral root and poisoning cases. The distribution and toxicity of these four components in Aconitum herbs should be further systematically studied for medication safety. This study developed a new UHPLC-QQQ-MS/MS method to determine ten Aconitum alkaloids, including aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, benzoylhypaconine, YAC, CCA, DYA, and DCA, for Aconitum herbs simultaneously. YAC and CCA were founded in some samples of unprocessed A. carmichaelii lateral root (7.04%), A. carmichaelii root (9.43%), A. brachypodum root (6.00%), and A. ouvrardianum root (100%). Four hidden toxic Aconitum alkaloids were detected in processed A. carmichaelii lateral root (2.56%) and A. vilmorinianum root (100%). Four hidden toxic Aconitum alkaloids played significant roles in the classification of Aconitum herbs by OPLS-DA analysis. The acute toxicity test was performed by up-and-down procedure (UDP). The oral administration of the half lethal dose (LD50) of YAC, CCA, DYA, and DCA to female ICR mice was 2.37 mg/kg, 5.60 mg/kg, 60.0 mg/kg, and 753 mg/kg, respectively. The LD50 by intravenous injection was 0.200 mg/kg, 0.980 mg/kg, 7.60 mg/kg, and 34.0 mg/kg, respectively. The LD50 of unprocessed A. carmichaelii lateral root, A. vilmorinianum root, and A. brachypodum root to mice orally was 1.89 g/kg, 0.950 g/kg, and 0.380 g/kg, respectively. Symptoms of Aconitum alkaloid poisoning in mice were decreased activity, fur erect, palpebral edema, vomiting, polypnea, and convulsions. The main change of organs was flatulence. No poisoning or death occurred in mice at the maximum dosage (27.0 g/kg) of A. ouvrardianum root orally. To better control the quality and safety of Aconitum herbs, this study provides favorable support for improving the existing standards to strengthen the supervision of the four hidden toxic Aconitum alkaloids.

Endophytic fungus Cladosporium tenuissimum DF11, an efficient inducer of tanshinone biosynthesis in Salvia miltiorrhiza roots.

Salvia miltiorrhiza is a traditional medicinal plant mainly used for cardiovascular and cerebrovascular disease treatment. Tanshinones are the main bioactive constituents of S. miltiorrhiza, which mainly accumulate around its root periderm tissue. Endophytic fungi are important bioelicitors or probiotics that can promote the accumulation of secondary metabolites and sustainable cultivation of medicinal plants. Among them, endophytic Cladosporium spp., possessing a variety of biotransformation and metabolic abilities, is an ideal elicitor source. Here, we used a gnotobiotic system to investigate the effects of the endophytic fungus Cladosporium tenuissimum DF11 on tanshinone biosynthesis in S. miltiorrhiza roots. The results showed that C. tenuissimum DF11 mainly colonizes the intercellular space of the root tissues and promotes tanshinone biosynthesis and accumulation in S. miltiorrhiza roots by upregulating the expression of the genes encoding for key enzymes HMGR, DXS, DXR, GGPPS, CPS, KSL and CYP76AH1 of the tanshinone biosynthesis pathway. The expression levels of almost all genes encoding for key enzymes reached the response peak in the first or second week after DF11 colonization. Taken together, the endophytic fungus C. tenuissimum DF11 could promote secondary metabolite accumulation in S. miltiorrhiza roots. These results indicate that DF11 will be a potential biofertilizer fungus to regulate and stabilize the quality of cultivated S. miltiorrhiza medicinal materials.