[Applications of ion chromatography for the analysis of Chinese herbal medicine components].

Ion chromatography (IC) is a novel high performance liquid chromatographic technique that is suitable for the separation and analysis of ionic substances in different matrix samples. Since 1975, it has been widely used in many fields, such as the environment, energy, food, and medicine. IC compensates for the separation limitations of traditional gas chromatography and high performance liquid chromatography and can realize the qualitative analysis and quantitative detection of strongly polar components. This chromatographic technique features not only simple operations but also rapid analysis. The sensors used in IC are characterized by high sensitivity and selectivity, and the technique can simultaneously separate and determine multiple components. Several advances in IC instrumentation and chromatographic theories have been developed in recent years. IC can analyze various types of samples, including ions, sugars, amino acids, and organic acids (bases). Chinese herbal medicines are typically characterized by highly complex chemical compositions and may contain carbohydrates, proteins, alkaloids, and other active components. They also contain toxic residues such as sulfur dioxide, which may be produced during the processing of medicinal materials. Therefore, the analysis and elucidation of the precise chemical constituents of Chinese herbal medicines present key problems that must be resolved in modern Chinese herbal medicine research. In this context, IC has become an important method for analyzing and identifying the complex components of Chinese herbal medicines because this method is suitable for detecting a single active ingredients among complex components. This paper introduces the different types and principles of IC as well as research progress in this technique. As the applications of IC-based methods in pharmaceutical science, cell biology, and microbiology increase, further development is necessary to expand the applications of this technique. The development of innovative techniques has enabled IC technologies to achieve higher analytical sensitivity, better selectivity, and wider application. The components of Chinese herbal medicines can be divided into endogenous and exogenous components according to their source: endogenous components include glycosides, amino acids, and organic acids, while exogenous components include toxic residues such as sulfur dioxide. Next, the applications of IC to the complex components of Chinese herbal medicines in recent decades are summarized. The most commonly used IC technologies and methods include ion exchange chromatography and conductivity detection. The advantages of IC for the analysis of alkaloids have been demonstrated. This method exhibits better characteristics than traditional analytical methods. However, the applications of IC for the speciation analysis of inorganic anions are limited. Moreover, few reports on the direct application of the technique for the determination of the main active substances in Chinese herbal medicines, including flavonoids, phenylpropanoids, and steroids, have been reported. Finally, this paper reviews new IC technologies and their application progress in Chinese herbal medicine, focusing on their prospects for the effective separation and analysis of complex components. In particular, we discuss the available sample (on-line) pretreatment technologies and explore possible technologies for the selective and efficient enrichment and separation of different components. Next, we assess innovative research on solid-phase materials that can improve the separation effect and analytical sensitivity of IC. We also describe the features of multidimensional chromatography, which combines the advantages of various chromatographic techniques. This review provides a theoretical reference for the further development of IC technology for the analysis of the complex chemical components of Chinese herbal medicines.

A novel bHLH gene responsive to low nitrogen positively regulates the biosynthesis of medicinal tropane alkaloids in Atropa belladonna.

Medicinal tropane alkaloids (TAs), including hyoscyamine, anisodamine and scopolamine, are essential anticholinergic drugs specifically produced in several solanaceous plants. Atropa belladonna is one of the most important medicinal plants that produces TAs. Therefore, it is necessary to cultivate new A. belladonna germplasm with the high content of TAs. Here, we found that the levels of TAs were elevated under low nitrogen (LN) condition, and identified a LN-responsive bHLH transcription factor (TF) of A. belladonna (named LNIR) regulating the biosynthesis of TAs. The expression level of LNIR was highest in secondary roots where TAs are synthesized specifically, and was significantly induced by LN. Further research revealed that LNIR directly activated the transcription of hyoscyamine 6ß-hydroxylase gene (H6H) by binding to its promoter, which converts hyoscyamine into anisodamine and subsequently epoxidizes anisodamine to form scopolamine. Overexpression of LNIR upregulated the expression levels of TA biosynthesis genes and consequently led to the increased production of TAs. In summary, we functionally identified a LN-responsive bHLH gene that facilitated the development of A. belladonna with high-yield TAs under the decreased usage of nitrogen fertilizer.

The impact of thermal pretreatment on phytochemical profiles and bioactivity of freeze-dried lily bulbs (Lilium lancifolium).

Lily bulbs are susceptible to deterioration during storage if improperly handled. To resolve this problem, it is necessary to investigate suitable processing techniques. The aim of this study is to evaluate the effects of steaming, blanching and microwave pretreatment on freeze-dried lily bulbs in terms of color, phenolic content and bioactivity. Results showed that appropriate steaming and blanching pretreatment could contribute to product characteristics similar to those of freeze-dried lily bulbs, with the maximum L* value reduced by only 7.57% and 0.55% respectively. Thermal pretreatment affected the retention, degradation and transformation of polyphenol, especially for regalosides. The polyphenol was closely associated with the browning of lily bulbs. Thermal processing caused the decline of regaloside A and the increase of regaloside B, which were the major phenolic monomers that can effectively inhibit the browning of lily bulbs. The antioxidant activity of freeze-dried lily pretreated with blanching for 6 min was the highest (4.39 ± 0.32 µmol TE/g DW), with an improvement of nearly 25.39% compared to that of untreated freeze-dried lily. Thus, the combination of freeze-dried with steaming or blanching pretreatment could be proposed as a sustainable strategy to improve the quality of lily bulbs for industrial application.

Salidroside protects pulmonary artery endothelial cells against hypoxia-induced apoptosis via the AhR/NF-κB and Nrf2/HO-1 pathways.

BACKGROUND: The apoptosis of pulmonary artery endothelial cells (PAECs) is an important factor contributing to the development of pulmonary hypertension (PH), a serious cardio-pulmonary vascular disorder. Salidroside (SAL) is a bioactive compound derived from an herb Rhodiola, but the potential protective effects of SAL on PAECs and the underlying mechanisms remain elusive. PURPOSE: The objective of this study was to determine the role of SAL in the hypoxia-induced apoptosis of PAECs and to dissect the underlying mechanisms. STUDY DESIGN: Male Sprague-Dawley (SD) rats were subjected to hypoxia (10% O2) for 4 weeks to establish a model of PH. Rats were intraperitoneally injected daily with SAL (2, 8, and 32 mg/kg/d) or vehicle. To define the molecular mechanisms of SAL in PAECs, an in vitro model of hypoxic cell injury was also generated by exposed PAECs to 1% O2 for 48 h. METHODS: Various techniques including hematoxylin and eosin (HE) staining, immunofluorescence, flow cytometry, CCK-8, Western blot, qPCR, molecular docking, and surface plasmon resonance (SPR) were used to determine the role of SAL in rats and in PAECs in vitro. RESULTS: Hypoxia stimulation increases AhR nuclear translocation and activates the NF-κB signaling pathway, as evidenced by upregulated expression of CYP1A1, CYP1B1, IL-1ß, and IL-6, resulting in oxidative stress and inflammatory response and ultimately apoptosis of PAECs. SAL inhibited the activation of AhR and NF-κB, while promoted the nuclear translocation of Nrf2 and increased the expression of its downstream antioxidant proteins HO-1 and NQO1 in PAECs, ameliorating the hypoxia-induced oxidative stress in PAECs. Furthermore, SAL lowered right ventricular systolic pressure, and decreased pulmonary vascular remodeling and right ventricular hypertrophy in hypoxia-exposed rats. CONCLUSIONS: SAL may attenuate the apoptosis of PAECs by suppressing NF-κB and activating Nrf2/HO-1 pathways, thereby delaying the progressive pathology of PH.

Functional divergence of two arginine decarboxylase genes in tropane alkaloid biosynthesis and root growth in Atropa belladonna.

Putrescine, produced via the arginine decarboxylase (ADC)/ornithine decarboxylase (ODC)-mediated pathway, is an initial precursor for polyamines metabolism and the root-specific biosynthesis of medicinal tropane alkaloids (TAs). These alkaloids are widely used as muscarinic acetylcholine antagonists in clinics. Although the functions of ODC in biosynthesis of polyamines and TAs have been well investigated, the role of ADC is still poorly understood. In this study, enzyme inhibitor treatment showed that ADC was involved in the biosynthesis of putrescine-derived metabolites and root growth in Atropa belladonna. Further analysis found that there were six ADC unigenes in the A. belladonna transcriptome, with two of them, AbADC1 and AbADC2, exhibiting high expression in the roots. To investigate their roles in TAs/polyamines metabolism and root growth, RNA interference (RNAi) was used to suppress either AbADC1 or AbADC2 expression in A. belladonna hairy roots. Suppression of the AbADC1 expression resulted in a significant reduction in the putrescine content and hairy root biomass. However, it had no noticeable effect on the levels of N-methylputrescine and the TAs hyoscyamine, anisodamine, and scopolamine. On the other hand, suppression of AbADC2 expression markedly reduced the levels of putrescine, N-methylputrescine, and TAs, but had no significant effect on hairy root biomass. According to ß-glucuronidase (GUS) staining assays, AbADC1 was mainly expressed in the root elongation and division region while AbADC2 was mainly expressed in the cylinder of the root maturation region. These differences in expression led to functional divergence, with AbADC1 primarily regulating root growth and AbADC2 contributing to TA biosynthesis.

Dracotangusions A and B, two new sesquiterpenes from Dracocephalum tanguticum Maxim. with anti-inflammatory activity.

Two new guaiane sesquiterpenoids were isolated from the dried aerial parts of Dracocephalum tanguticum Maxim., named as dracotangusions A (1) and B (2), together with four known sesquiterpenoids, which were identified as Curcumenone (3), (4Z,7Z,9Z)-11-Hydroxy-4,7,9-germacratriene-1,6-dione (4), Kobusone (5), and (1S,10S), (4S, 5S)-(+)-germacrone-1(10)-4-diepoxide (6). The structures of isolates were determined by UV, IR, HR-ESI-MS, and NMR analysis. What is noteworthy is that four known sesquiterpenoids were isolated for the first time from the genus of Dracocephalum L. All compounds inhibited the extremely significant difference (p < 0.01) in anti-inflammatory activity, suggesting that these compounds may be promising candidates as an anti-inflammatory agent.

Endophytic bacterial community structure and diversity of the medicinal plant Mirabilis himalaica from different locations.

Endophytic bacteria play important roles in medicinal plant growth, abiotic stress, and metabolism. Mirabilis himalaica (Edgew.) Heimerl is known for its medicinal value as Tibetan traditional plant; however, little is known about the endophytic bacteria associated with this plant in different geographic conditions and vegetal tissues. To compare the endophytic bacterial community associated with this plant in different geographic conditions and vegetal tissues, we collected the leaves, stems, and roots of M. himalaica from five locations, Nongmu college (NM), Gongbujiangda (GB), Zhanang County (ZL), Lang County (LX), and Sangri County (SR), and sequenced the 16S rRNA V5-V7 region with the Illumina sequencing method. A total of 522,450 high-quality sequences and 4970 operational taxonomic units (OTUs) were obtained. The different tissues from different locations harbored unique bacterial assemblages. Proteobacteria and Actinobacteria were the dominant phyla in all the samples, while the dominant genera changed based on the different tissues. The endophytic bacterial structures in the leaf and stem tissues were different compared to root tissues. Redundancy analysis (RDA) showed that the endophytic bacterial community was significantly correlated with pH, available phosphorus (AP), total phosphorus (TP), total nitrogen (TN), and soil organic matter (SOM). These findings suggested that the geographic conditions, climate type, ecosystem type, and tissues determined the endophytic bacterial composition and relative abundances. This conclusion could facilitate an understanding of the relationship and ecological function of the endophytic bacteria associated with M. himalaica and provide valuable information for artificial planting of M. himalaica and identifying and applying functional endophytic bacteria.

Herpetrione, a New Type of PPARα Ligand as a Therapeutic Strategy Against Nonalcoholic Steatohepatitis.

Non-alcoholic fatty liver disease, especially nonalcoholic steatohepatitis (NASH), is a leading cause of cirrhosis and liver cancer worldwide; nevertheless, there are no Food and Drug Administration-approved drugs for treating NASH until now. Peroxisome proliferator-activated receptor alpha (PPARα) is an interesting therapeutic target for treating metabolic disorders in the clinic, including NASH. Herpetrione, a natural lignan compound isolated from Tibetan medicine Herpetospermum caudigerum, exerts various hepatoprotective effects, but its efficacy and molecular mechanism in treating NASH have not yet been elucidated. Here, we discovered that herpetrione lessened lipid accumulation and inflammation in hepatocytes stimulated with oleic acid and lipopolysaccharide, and effectively alleviated NASH caused by a high-fat diet or methionine-choline-deficient diet by regulating glucolipid metabolism, insulin resistance, and inflammation. Mechanistically, RNA-sequencing analyses further showed that herpetrione activated PPAR signaling, which was validated by protein expression. Furthermore, the analysis of molecular interactions illustrated that herpetrione bound directly to the PPARα protein, with binding sites extending to the Arm III domain. PPARα deficiency also abrogated the protective effects of herpetrione against NASH, suggesting that herpetrione protects against hepatic steatosis and inflammation by activation of PPARα signaling, thereby alleviating NASH. Our findings shed light on the efficacy of a natural product for treating NASH, as well as the broader prospects for NASH treatment by targeting PPARα.

Insights into the mechanism underlying UV-B induced flavonoid metabolism in callus of a Tibetan medicinal plant Mirabilis himalaica.

Mirabilis himalaica is an important Tibetan medicinal plant in China. However, it has become a rare and class I endangered Tibetan medicine plant. Therefore, the use of callus to propagate germplasm resources is of great significance. We found that the flavonoid content of M. himalaica callus increased continuously with the extension of UV-B treatment. Multi-omics profiles were used to reveal the co-expression patterns of gene networks of flavonoid metabolism in M. himalaica callus during UV-B radiation. Results showed that five medicinal metabolics, including geranin, eriodictyol, astragalin, isoquercetin, pyrotechnic acid, and one anthocyanin malvide-3-O-glucoside were identified. The transcriptome data were divided into 46 modules according to the expression pattern by WGCNA (weighted gene co-expression network analysis), of which the module Turquoise had the strongest correlation with six target metabolites. We found that seven structural genes and twenty-five transcription factors were related to the metabolism of flavonoid synthesis, among which the structural genes CHI, C4H and UGT79B6 had strong co-expression relationships with the 6 target metabolites. WRKY42, WRKY7, bHLH128 and other transcription factors had strong co-expression relationships with multiple structural genes. Consequently, these findings suggest callus grown under UV-B treatment could be an effective alternative medical resource of M. himalaica, which is valuable for conservation and usage of this wild and endangered plant.

Hyperuricemia: A key contributor to endothelial dysfunction in cardiovascular diseases.

As an end product of purine metabolism, uric acid (UA) is a major endogenous antioxidant in humans. However, impaired UA synthesis and excretion can lead to hyperuricemia (HUA), which may in turn induce endothelial dysfunction (ED) and contribute to the pathogenesis of cardiovascular diseases (CVDs; e.g., atherosclerosis and hypertension). In this review, we discuss recent advances and novel insights into the effects exerted by HUA conditions in ED and related underlying mechanisms focusing on impaired UA metabolism, reduction in the synthesis and bioavailability of nitric oxide, endothelial cell injury, the endothelial-to-mesenchymal transition, insulin resistance, procoagulant activity, and acquisition of an inflammatory phenotype. We additionally discuss intervention strategies for HUA-induced ED and the paradoxical roles of UA in endothelial function. We summarize major conclusions and perspectives: the deleterious effects of HUA contribute to the initiation and progression of CVD-related ED. However, the treatment strategies (in addition to urate-lowering therapy) for increasing endothelial function are limited because the majority of literature on pharmacological and pathophysiological mechanisms underlying HUA-induced ED solely describes in vitro models. Therefore, a better understanding of the mechanisms involved in HUA-induced ED is critical to the development of novel therapies for preventing and treating CVD-HUA comorbidities.

Effect of Microwave Treatments Combined with Hot-Air Drying on Phytochemical Profiles and Antioxidant Activities in Lily Bulbs (Lilium lancifolium).

Lily bulbs (Lilium lancifolium Thunb.) are rich in phytochemicals and have many potential biological activities which could be deep-processed for food or medicine purposes. This study investigated the effects of microwaves combined with hot-air drying on phytochemical profiles and antioxidant activities in lily bulbs. The results showed that six characteristic phytochemicals were identified in lily bulbs. They also showed that with an increase in microwave power and treatment time, regaloside A, regaloside B, regaloside E, and chlorogenic acid increased dramatically in lily bulbs. The 900 W (2 min) and the 500 W (5 min) groups could significantly suppress the browning of lily bulbs, with total color difference values of 28.97 ± 4.05 and 28.58 ± 3.31, respectively, and increase the content of detected phytochemicals. The highest oxygen radical absorbance activity was found in the 500 W, 5 min group, a 1.6-fold increase as compared with the control (57.16 ± 1.07 µmol TE/g DW), which was significantly relevant to the group's phytochemical composition. Microwaves enhanced the phytochemicals and antioxidant capacity of lily bulbs, which could be an efficient and environmentally friendly strategy for improving the nutrition quality of lily bulbs during dehydration processing.

Revealing evolution of tropane alkaloid biosynthesis by analyzing two genomes in the Solanaceae family.

Tropane alkaloids (TAs) are widely distributed in the Solanaceae, while some important medicinal tropane alkaloids (mTAs), such as hyoscyamine and scopolamine, are restricted to certain species/tribes in this family. Little is known about the genomic basis and evolution of TAs biosynthesis and specialization in the Solanaceae. Here, we present chromosome-level genomes of two representative mTAs-producing species: Atropa belladonna and Datura stramonium. Our results reveal that the two species employ a conserved biosynthetic pathway to produce mTAs despite being distantly related within the nightshade family. A conserved gene cluster combined with gene duplication underlies the wide distribution of TAs in this family. We also provide evidence that branching genes leading to mTAs likely have evolved in early ancestral Solanaceae species but have been lost in most of the lineages, with A. belladonna and D. stramonium being exceptions. Furthermore, we identify a cytochrome P450 that modifies hyoscyamine into norhyoscyamine. Our results provide a genomic basis for evolutionary insights into the biosynthesis of TAs in the Solanaceae and will be useful for biotechnological production of mTAs via synthetic biology approaches.

Oxybaphus himalaicus Mitigates Lipopolysaccharide-Induced Acute Kidney Injury by Inhibiting TLR4/MD2 Complex Formation.

Acute kidney injury (AKI) is described as the abrupt decrease in kidney function always accompanied by inflammation. The roots of Oxybaphus himalaicus Edgew. have long been used in Tibetan folk medicine for the treatment of nephritis. Nevertheless, modern pharmacological studies, especially about the underlying mechanism of O. himalaicus medications, are still lacking. Here, in lipopolysaccharide (LPS)-induced RAW264.7 macrophages, the O. himalaicus extract (OE) showed significant anti-inflammatory activity with the dose dependently reducing the LPS-stimulated release of nitric oxide and the mRNA level and protein expression of inflammatory cytokines and reversed the activation of nuclear factor kappa B (NF-κB). Co-immunoprecipitation assay indicated that OE inhibited Toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD2) complex formation and further suppressed both myeloid differentiation factor 88 (MyD88)-dependent and TIR-domain-containing adapter-inducing interferon-ß (TRIF)-dependent cascades activation. In addition, OE could restrain NADPH oxidase 2 (NOX2) endocytosis by blocking TLR4/MD2 complex formation to prevent reactive oxygen species production. In LPS-induced AKI mice, OE treatment mitigated renal injury and inflammatory infiltration by inhibiting TLR4/MD2 complex formation. UPLC-MS/MS analysis tentatively identified 41 components in OE. Our results indicated that OE presented significant anti-inflammatory activity by inhibiting TLR4/MD2 complex formation, which alleviated LPS-induced AKI in mice.

Ornithine decarboxylase regulates putrescine-related metabolism and pollen development in Atropa belladonna.

Polyamines, including putrescine, spermidine, and spermine, play critical roles in cell physiology by different forms. As a rate-limiting enzyme that converts ornithine to putrescine, ornithine decarboxylase (ODC, EC 1.1.1.37) has been studied in detail in animals and microorganisms, but its specific functions are poorly understood in plants. In this study, the metabolic and developmental roles of the ODC gene were studied through RNAi-mediated suppression of the ODC gene (AbODC) in A. belladonna. Suppression of AbODC reduced the production of precursors of medicinal tropane alkaloids, including putrescine and N-methylputrescine, as well as hyoscyamine and scopolamine. In AbODC-RNAi roots, the production of putrescine and spermidine in free form was reduced, but in the AbODC-RNAi leaves, the content of free polyamines was not altered. In the roots/leaves of AbODC-RNAi plants, the production of conjugated and bound polyamines was reduced. In addition, suppression of the ODC gene resulted in reduction of polyamines and pollen sterility in AbODC-RNAi flowers. In floral organs, GUS-staining results indicated that AbODC was domainantly expressed in pollen. In summary, ornithine decarboxylase not only plays a key role in regulating the biosynthesis of diverse forms of polyamines and medicinal tropane alkaloids, but also participates in pollen development.

Role of MicroRNA-Like RNAs in the Regulation of Spore Morphological Differences in the Entomopathogenic Fungus Metarhizium acridum.

Metarhizium acridum is an important microbial pesticide. Conidia (CO) and blastospores (BS) are two types of spores that occur in different patterns in the M. acridum life cycle and exhibit significant differences in cell morphology, structure, and activity. It may suggest that the fungus has a complex gene regulation mechanism. While previous studies on the differences between CO and BS have mainly focused on cell structure and application, little is known regarding the differences between CO and BS in fungi on the transcriptome levels. MicroRNAs (miRNAs) are small noncoding RNAs crucial to gene regulation and cell function. Understanding the miRNA-like RNAs (milRNA) and mRNA expression profiles related to cell growth and cellular morphological changes would elucidate the roles of miRNAs in spore morphological differences. In this study, 4,646 differentially expressed genes (DEGs) were identified and mainly classified in the GO terms cell, cell part, biological process, and catalytic activity. The KEGG annotation suggested that they were enriched in amino acid biosynthesis, carbohydrate metabolism, ribosome, and oxidative phosphorylation and might be involved in cell activity and structure. There were 113 differentially expressed milRNAs (DEMs), targeting 493 DEGs. Target gene functional analysis revealed that the target genes were mainly enriched in RNA transport, purine metabolism, and the cell cycle. In addition, we identified essential genes from milRNA-mRNA pairs that might participate in cell budding growth and cell membrane and wall integrity, including adenosine deaminase, glycosyl hydrolase, and G-patch domain protein (dno-miR-328-3p), WD repeat-containing protein pop1 (age-miR-127), and GPI-anchored wall transfer protein (cgr-miR-598). MilRNAs might therefore play a crucial role in cell growth and cellular morphological changes as transcriptional and post-transcriptional regulators.

AabHLH112, a bHLH transcription factor, positively regulates sesquiterpenes biosynthesis in Artemisia annua.

The bHLH transcription factors play important roles in the regulation of plant growth, development, and secondary metabolism. ß-Caryophyllene, epi-cedrol, and ß-farnesene, three kinds of sesquiterpenes mainly found in plants, are widely used as spice in the food industry and biological pesticides in agricultural production. Furthermore, they also have a significant value in the pharmaceutical industry. However, there is currently a lack of knowledge on the function of bHLH family TFs in ß-caryophyllene, epi-cedrol, and ß-farnesene biosynthesis. Here, we found that AabHLH112 transcription factor had a novel function to positively regulate ß-carophyllene, epi-cedrol, and ß-farnesene biosynthesis in Artemisia annua. Exogenous MeJA enhanced the expression of AabHLH112 and genes of ß-caryophyllene synthase (CPS), epi-cedrol synthase (ECS), and ß-farnesene synthase (BFS), as well as sesquiterpenes content. Dual-LUC assay showed the activation of AaCPS, AaECS, and AaBFS promoters were enhanced by AabHLH112. Yeast one-hybrid assay showed AabHLH112 could bind to the G-box (CANNTG) cis-element in promoters of both AaCPS and AaECS. In addition, overexpression of AabHLH112 in A. annua significantly elevated the expression levels of AaCPS, AaECS, and AaBFS as well as the contents of ß-caryophyllene, epi-cedrol, and ß-farnesene, while suppressing AabHLH112 expression by RNAi reduced the expression of the three genes and the contents of the three sesquiterpenes. These results suggested that AabHLH112 is a positive regulator of ß-caryophyllene, epi-cedrol, and ß-farnesene biosynthesis in A. annua.

Transcriptome and Metabolome Analyses of Codonopsis convolvulacea Kurz Tuber, Stem, and Leaf Reveal the Presence of Important Metabolites and Key Pathways Controlling Their Biosynthesis.

Codonopsis convolvulacea Kurz. var. vinciflora (Kom.) L.T. Shen is a member of Campanulaceae, which is used in traditional Chinese medicine. However, apart from a few Codonopsis species, no detailed knowledge is available on the metabolite composition and respective transcriptome signatures. We performed a combined transcriptome and metabolome analysis of the tuber, stem, and leaf of C. convolvulacea and found 1,144 metabolites and 231,840 unigenes in three experimental groups. The analysis revealed considerable variations in the three tissues. Tubers were rich in amino acids and derivatives, flavonoids, and organic acids, whereas the stems and leaves were rich in alkaloids and flavonoids, respectively. Transcriptome sequencing revealed candidate genes being involved in flavonoid, tryptophan, and alkaloid biosyntheses. In particular, we indicated that the variation in the isoflavone content is linked to the expressions of CHI, CYP73A, C3'H, F3H, CYP75B1, anthocyanidin synthase, and FLS. In a similar way, the levels of indole, L-tyrosine, and tryptamine were also consistent with the expressions of TDC/DDCs in the respective tissues. In addition, the expression levels of ASP5, ARO8, GOT, and AOC3 indicated that L-tryptophan is being converted to downstream metabolites. Overall, our datasets present a useful resource for future research on the uses of this medicinal plant and put forward many research questions.

Comparative analyses of chloroplast genomes from Six Rhodiola species: variable DNA markers identification and phylogenetic relationships within the genus.

BACKGROUND: As a valuable medicinal plant, Rhodiola has a very long history of folk medicine used as an important adaptogen, tonic, and hemostatic. However, our knowledge of the chloroplast genome level of Rhodiola is limited. This drawback has limited studies on the identification, evolution, genetic diversity and other relevant studies on Rhodiola. RESULTS: Six Rhodiola complete chloroplast genomes were determined and compared to another Rhodiola cp genome at the genome scale. The results revealed a cp genome with a typical quadripartite and circular structure that ranged in size from 150,771 to 151,891 base pairs. High similarity of genome organization, gene number, gene order, and GC content were found among the chloroplast genomes of Rhodiola. 186 (R. wallichiana) to 200 (R. gelida) SSRs and 144 pairs of repeats were detected in the 6 Rhodiola cp genomes. Thirteen mutational hotspots for genome divergence were determined and could be used as candidate markers for phylogenetic analyses and Rhodiola species identification. The phylogenetic relationships inferred by members of Rhodiola cluster into two clades: dioecious and hermaphrodite. Our findings are helpful for understanding Rhodiola's taxonomic, phylogenetic, and evolutionary relationships. CONCLUSIONS: Comparative analysis of chloroplast genomes of Rhodiola facilitates medicinal resource conservation, phylogenetic reconstruction and biogeographical research of Rhodiola.

Biotechnological Approaches on Engineering Medicinal Tropane Alkaloid Production in Plants.

Hyoscyamine and scopolamine, belonging to medicinal tropane alkaloids (MTAs), are potent anticholinergic drugs. Their industrial production relies on medicinal plants, but the levels of the two alkaloids are very low in planta. Engineering the MTA's production is an everlasting hot topic for pharmaceutical industry. With understanding the MTA's biosynthesis, biotechnological approaches are established to produce hyoscyamine and scopolamine in an efficient manner. Great advances have been obtained in engineering MTA's production in planta. In this review, we summarize the advances on the biosynthesis of MTAs and engineering the MTA's production in hairy root cultures, as well in plants. The problems and perspectives on engineering the MTA's production are also discussed.

Molecular insights into AabZIP1-mediated regulation on artemisinin biosynthesis and drought tolerance in Artemisia annua.

Artemisia annua is the main natural source of artemisinin production. In A. annua, extended drought stress severely reduces its biomass and artemisinin production while short-term water-withholding or abscisic acid (ABA) treatment can increase artemisinin biosynthesis. ABA-responsive transcription factor AabZIP1 and JA signaling AaMYC2 have been shown in separate studies to promote artemisinin production by targeting several artemisinin biosynthesis genes. Here, we found AabZIP1 promote the expression of multiple artemisinin biosynthesis genes including AaDBR2 and AaALDH1, which AabZIP1 does not directly activate. Subsequently, it was found that AabZIP1 up-regulates AaMYC2 expression through direct binding to its promoter, and that AaMYC2 binds to the promoter of AaALDH1 to activate its transcription. In addition, AabZIP1 directly transactivates wax biosynthesis genes AaCER1 and AaCYP86A1. The biosynthesis of artemisinin and cuticular wax and the tolerance of drought stress were significantly increased by AabZIP1 overexpression, whereas they were significantly decreased in RNAi-AabZIP1 plants. Collectively, we have uncovered the AabZIP1-AaMYC2 transcriptional module as a point of cross-talk between ABA and JA signaling in artemisinin biosynthesis, which may have general implications. We have also identified AabZIP1 as a promising candidate gene for the development of A. annua plants with high artemisinin content and drought tolerance in metabolic engineering breeding.