The bright blue colouration of Centaurea cyanus (cornflower) is attributed to a supramolecular pigment formed via metal ion chelation with accumulated cyanidin (Cy) and co-pigments, yet the underlying genetic regulatory mechanisms remain incompletely understood. Elucidating this mechanism will facilitate the molecular breeding of cyanidin-based blue flowers, thereby enhancing the ornamental value of flowering plants. Initially, in vitro blue pigment reconstruction experiments identified that Fe3+ is the primary metal ion enabling the blue transformation of Cy, and the molar ratio of Cy to Fe3+ was found to be critical. Genomic analysis identified 11 VIT/VTL genes and four CcFers genes in cornflower. Expression profiling revealed an inverse expression relationship between CcVIT1a and CcFers across various tissues. Notably, CcVIT1a specifically and highly expressed in blue-coloured ray florets. Subcellular localization and yeast mutant complementation assay showed CcVIT1a in tonoplast, whereas the CcFers reside in plastids. This spatial separation suggests their coordinated role in regulating intracellular iron homeostasis. Virus-induced gene silencing of CcVIT1a caused a colour shift from blue to violet, indicating its critical role in blue colouration of cornflower. Concurrently, the expression of the CcFer1a was significantly upregulated, while the other three CcFers were downregulated, further supporting the idea that CcVIT1a and the CcFers collaboratively regulate intracellular iron homeostasis. Transient overexpression of CcVIT1a in chrysanthemum CB (Cy-determined blue flowers) sample 'Dante purple' induced a colour transition from red-purple to violet, accompanied by the downregulation of both CmVIT and CmFer1. These findings imply a similar synergistic mechanism governing iron homeostasis in chrysanthemum. This study elucidates the cooperative roles of VIT1 and Fer in regulating intracellular iron partitioning. We demonstrate that upregulation of CcVIT1a enhances iron sequestration into the vacuole, thereby promoting the cyanidin-based blue colouration. Collectively, our findings provide novel molecular insights and a potential strategy for breeding blue flowers through the manipulation of cyanidin and iron metabolism.

Soil salinization severely constrains the growth and sustainable production of Chrysanthemum morifolium. However, the molecular mechanisms underlying salt tolerance in chrysanthemum remain largely uncharacterized. To identify suitable materials for dissecting the molecular mechanisms of salt tolerance, four chrysanthemum cultivars were evaluated under salt stress. Among them, ‘Hangbaiju’ was identified as the most salt-tolerant cultivar as reflected by enhanced osmotic adjustment capacity, reduced membrane damage, and improved maintenance of photosynthetic performance. Based on this screening, comparative transcriptome sequencing was performed to identify candidate genes associated with salt stress response, leading to the identification of CmNHX1–4. CmNHX1–4 were localized to the tonoplast and exhibited sustained transcriptional activation during prolonged salt stress. Notably, CmNHX1–4 displayed distinct temporal and spatial expression patterns across roots and leaves. Functional assays further demonstrated that heterologous expression of CmNHX1–4 in yeast and Arabidopsis significantly enhanced salt tolerance. Moreover, K-means clustering and WGCNA analysis combined with promoter analysis of CmNHXs indicated the potential regulation of CmNHXs by WRKY transcription factors via W-box elements. Further experiments demonstrated that CmWRKY54 directly bound to the promoter of CmNHX1 and activated its transcription. Collectively, our findings reveal a novel regulatory module whereby CmWRKY54 enhances salt tolerance through activating NHX1 expression, which is potentially implicated in mediating the vacuolar sequestration of cytosolic Na+. These results expand our mechanistic understanding of salt tolerance mechanisms in chrysanthemum and provide potential candidate genes for salt tolerance improvement in horticultural crops.

Chrysanthemum (Chrysanthemum × morifolium Ramat.) is a globally popular ornamental plant, but most cultivars lack efficient petal-based transient transformation systems, limiting floral trait molecular mechanism exploration. Protoplasts are versatile tools for gene localization, interaction, and functional characterization. Here, we established a petal protoplast isolation and transient transformation system for C. morifolium ‘Wandai Fengguang’ via L9(34) orthogonal design: optimal isolation (0.6 M mannitol, 8 h enzymatic digestion time, 0.4% macerozyme R-10, 4% cellulase R-10) and transformation (40% PEG4000, 12 μg plasmid, 10 min transfection, a protoplast density of 1 × 106 protoplasts mL−1). Under these conditions, protoplast yield was 5.14 × 106 protoplasts g−1·FW, viability 87.41%, and transformation efficiency 51.50%, with good applicability for six additional germplasms. We further analyzed CmVIT1 protein localization. Compared with the previous system, this system significantly improved protoplast yield and transformation efficiency, facilitating the transient transformation of genes related to floral traits in chrysanthemum and providing a methodological framework for other horticultural crops.

Anoectochilus roxburghii, a valuable medicinal orchid, is severely threatened by drought stress. This study investigated its drought tolerance mechanisms focusing on late embryogenesis abundant (LEA) proteins and WRKY transcription factors (TFs). Physiological analyses showed that SLs treatment significantly reduced relative water content (RWC) and relative electrical conductivity (REC) compared to drought-stressed controls. Transcriptomic screening and qRT-PCR analyses identified ArLEA5/9 and ArWRKY57/70 as key drought-responsive genes, whose expression was upregulated under drought but suppressed by SLs. Overexpression of these genes in tobacco leaves and A. roxburghii protocorm-like bodies (PLBs) resulted in reduced water loss and enhanced drought resistance, with increased soluble proteins, proline, and decreased malondialdehyde (MDA) content. Yeast two-hybrid (Y2H), bimolecular fluorescence complementation (BIFC), and yeast one-hybrid (Y1H) assays confirmed that ArWRKY57 forms a plasma membrane complex with ArWRKY70 and directly regulates ArLEA5 transcription. This study represents the first to identify candidate drought-resistant genes (ArLEA5, ArLEA9, ArWRKY57, and ArWRKY70) and characterize the ArWRKY57-ArWRKY70-ArLEA5 regulatory module as a key mechanism underlying drought tolerance in A. roxburghii, thereby facilitating molecular breeding for drought-tolerant medicinal orchids.

Anoectochilus roxburghii (Wall.) Lindl., a rare and precious traditional Chinese medicinal plant, faces the challenge of seed developmental failure due to incomplete endosperm development. The miR166-HD-Zip III module has been reported to be involved in the regulation of reproductive development. However, the function of miR166-HD-Zip III module in A. roxburghii remains unclear. In this study, 38 ArHD-Zip TFs were identified from the full-length transcriptome of A. roxburghii. Among them, ArHDZ22 was found to promote flowering and seed development through phylogenetic analysis, sequence analysis and overexpression functional verification experiments. Subsequently, aro-miR166b/c that may regulate ArHDZ22 were screened by Small RNA sequencing, K-means analysis, differential expression analysis and qRT-PCR. The Dual-luciferase assay and tobacco transient transformation demonstrated that aro-miR166b/c could degrade ArHDZ22 directly. In addition, the over-expression experiments in Arabidopsis thaliana showed that aro-miR166b/c was able to regulate reproductive development by targeting ArHDZ22. Overall, this study revealed the regulatory role of the aro-miR166b/c-ArHDZ22 module in reproductive development, further elucidating the molecular mechanism of the reproductive development of A. roxburghii, and also provides a solid theoretical basis for its efficient breeding and germplasm innovation of A. roxburghii.

Anoectochilus roxburghii, as a horticultural plant, is renowned for its distinctive characteristics and widespread cultivation across various Asian regions. The beautiful golden veins make it of great ornamental value. At the same time, it is often used in medicine and food. Due to the immature development of the embryos and seed coat cells, the seeds of A. roxburghii were aborted. Only under the condition of symbiosis with fungi, the germination process of seeds can be promoted. Coupled with the dependence of cross-pollination on natural factors such as wind and insects, the reproduction of A.roxburghii is more complicated and difficult. In this study, we found that exogenous spermidine (Spd) treatment up-regulates the expression of ArWRKY6. Tissue-specific expression analysis showed that ArWRKY6 expressed most at the early stage of flower buds period and gradually decreased with the opening of flowers. ArWRKY6 overexpression plant demonstrated that it could promote flowering, and positively affected pistil and seed development. Our results provide a foundation for further analysis of the function of WRKY transcription factors in reproductive development of A. roxburghii, and provided a theoretical basis for hybridization breeding.

Objective Anoectochilus roxburghii is a valuable medicinal and ornamental plant. The aim of this study is to investigate the morphological and biochemical responses during the flower development stages of A. roxburghii, and to assess the effects of exogenous polyamines (PAs) on bud differentiation and metabolism, thereby providing a theoretical basis for understanding the flowering form and physiology of A. roxburghii. Methods In this study, morphological and biochemical responses in flower development stages of A. roxburghii were investigated using paraffin sections and stereomicroscope. A. roxburghii was divided into five periods, including vegetative growth period, flower bud period, flowering period, late flowering period and fruiting period. During the flowering phase, specific biochemical parameters were measured, including soluble sugar content, superoxide dismutase (SOD) activity, soluble protein content, peroxidase (POD) activity, and catalase (CAT) activity. These measurements were conducted to understand the biochemical changes occurring within A. roxburghii during its flowering process. Furthermore, the effects of PAs on bud differentiation were examined. Additionally, the activities of S-adenosylmethionine decarboxylase (SAMDC) and polyamine oxidase (PAO), as well as the content of polyphenols, polysaccharides, and flavonoids in A. roxburghii, were measured after PA treatment to evaluate the metabolic changes induced by exogenous PAs. Results During the flowering phase of A. roxburghii, soluble sugar content and SOD activity were steadily declining. Soluble protein content was initially increasing and then reducing, and POD and CAT activities showed opposite pattern. In addition, the effects of exogenous PAs on bud differentiation were investigated. Results showed that 3 mmol/L putrescine or 0.3 mmol/L spermidine significantly promoted the bud differentiation of A. roxburghii and advanced the flowering. The activities of SAMDC, PAO, and the content of polyphenols, polysaccharides and flavonoids in A. roxburghii significantly increased after PA treatment, demonstrating that exogenous PA can accelerate metabolism and improved the active ingredients content. Conclusion The flower development of A. roxburghii was divided into five stages, with significant changes in soluble sugar, protein, POD, SOD, CAT, MDA, and PRO levels. Exogenous putrescine and spermidine enhanced bud differentiation and accelerated flowering, increasing SAMDC and PAO activities, suggesting accelerated PA metabolism. PAs also improved active component content. These findings provide a theoretical basis for studying flower morphology and PA-induced flowering regulation of A. roxburghii.

The MYB and bHLH transcription factors could participate in the synthesis of anthocyanins through interaction or upstream-downstream regulation. Anthocyanins, as one of the most important active substances in plants, have high nutritional value of human beings. At present, anthocyanin biosynthesis has been widely studied, however, the regulatory mechanism of anthocyanins in Dendrobium officinale (D. officinale) remain unknow. Here, the anthocyanin components of two different colored cultivars from D. officinale were analyzed by UPLC-MS/MS. We have found that delphinidin and its derivatives were the main anthocyanin components in D. officinale, among which Malvidin-3-O-glucoside and Malvidin-3-O-galactoside were the main active substances. Subsequently, the key structural genes DoCHS1/2, DoF3H, DoF3’H, DoDFR1/2, DoANS, and transcription factors DoMYB5–1, DoMYB7, DobHLH5–1 were identified based on transcriptomic analysis, which were mainly involved in the anthocyanin biosynthesis. Among that, both DoMYB5–1 and DoMYB7 could interact with DobHLH5–1 to form complexes (DoMYB5–1-DobHLH5–1, DoMYB7-DobHLH5–1), and then regulated the expression of DoDFR1 and DoF3’H to promote anthocyanin synthesis through the protein-protein interaction, transient overexpression experiments and Y1H assay. Furthermore, we also found that the interaction of DoMYB5–1 and DobHLH5–1 could cascade-rergulate the complex DoMYB7-DobHLH5–1, thereby regulating the expression of DoDFR1 and then participating in the regulation of anthocyanin metabolism in D. officinale. Overall, this study clarified the regulatory mechanism of anthocyanin accumulation in D. officinale, which was helpful to reveal the biological mechanism of secondary metabolism in plants and provided a theoretical reference for the quality improvement of D. officinale.

Bletilla striata is widely used as healthy food and herbal medicine for its hemostasis, anti-ulcer, promoting wound healing, antibacterial, anti-inflammatory, and immune regulatory functions. Drying methods affect the quality of herbal medicine, including appearance and more importantly the content of active components, which play a decisive role in the medical and tonic functions. In this study, hot air drying (HD), infrared drying (ID), microwave drying (MD), vacuum drying (VD), and freeze drying (FD) were used to evaluate the effect of drying methods on the quality of B. striata tubers. Significant effects of drying methods on the appearance, color, microstructure, water activity, water content, and active component content of B. striata tubers were observed. FD-treated samples retained the majority of the appearance, color, and microstructure, while samples processed with other methods exhibiting various deformation and discoloration. In addition, a significant decline in water content and water activity was detected in FD-treated samples, yet, the content of active components, such as B. striata polysaccharides, total polyphenols, and militarine, was relatively high. After comprehensive comparison, freeze-drying is proposed to be the most suitable method for B. striata drying, which is also recommendable for other tuber drying.

Cornflower is favored by its exquisite capitulum with diverse color variations, espessially for the pure blue color, which are widely applied in food decoration, garden design as well as pharmaceutical drugs for its abundant secondary metabolites. Recently, a vast ocean of gene resources have been easily obtained by muti-omics technology in cornflower. However, the functional gene remains hard to identify due to a lack of efficient technical systems in cornflower capitula. Virus-induced gene silencing (VIGS), a tissue culture-independent method, shows great potential in confirming gene function of non-model species. This study established a VIGS method based on tobacco rattle virus (TRV) to effectively silence the anthocyanidin synthase gene (CcANS) in cornflower capitulum. To optimize silencing efficiency, we systematically screened various infiltration parameters, including infection method, flower developmental stage, and bacterial concentration. The results showed that the best silencing efficiency was obtained by co-injection of the bacterial suspension into both bud and pedicel at stage 1 with the concentration of OD600=1.2-1.8, in which the best silent phenotype can be obtained as high as 20 %-23 %. The insertion fragment could be as short as 84 bp, and the frequency of silencing signal transfer could reach 30 % without decreasing the silencing efficiency. This study provides an important technical means for verifying the gene function at the reproductive stage in cornflower.

Light, as an important environmental signal for plant growth and development, has been reported to be involved in the metabolism of phenylpropane. However, the light-induced phenylpropane biosynthesis mechanism in Chrysanthemum morifolium Ramat., especially flavonoid and chlorogenic acid (CGA) metabolism, is not clear. In this study, we found the flower phenotype of chrysanthemum 'HangBaiJu' became lighter after bagging. Besides, the content of main active ingredients, such as Luteolin-7-O-glucoside (Lu-7-O-G), Diosmetin-7-O-glucoside (Di-7-O-G), Apigenin-7-O-glucoside, Apigenin and CGA, was significantly decreased. Comparative transcriptome analysis (pairwise comparison, WGCNA, and k-means clustering) revealed that the expression of structural genes, such as CmPAL, CmCHS1/2, CmF3’H, CmFNS and CmHQT, was notably declined under dark conditions. Meanwhile, ‘bridge proteins’ CmMYB3/6/16 and signal transduction factors CmBBX20/22.2/22.3 were identified as key regulatory factors in the light-mediated synthesis of flavonoids and CGA in chrysanthemum. Among that, CmBBX20 could promote the accumulation of flavonoids and CGA by directly binding to the G-box core sequences of downstream target genes based on transient overexpression and Y1H assays. Overall, the preliminary molecular mechanism of BBXs and MYBs coordinately regulating the accumulation of flavonoids and CGA in chrysanthemum under different light inductions has been clarified, which provided a theoretical basis for the molecular breeding and quality improvement of chrysanthemum.

Chrysanthemum morifolium ‘HangBaiJu’, a popular medicinal and edible plant, exerts its biological activities primarily through the presence of flavones and caffeoylquinic acids (CQAs). However, the regulatory mechanism of flavone and CQA biosynthesis in the chrysanthemum capitulum remains unclear. In this study, the content of flavones and CQAs during the development of chrysanthemum capitulum was determined by HPLC, revealing an accumulation pattern with higher levels at S1 and S2 and a gradual decrease at S3 to S5. Transcriptomic analysis revealed that CmPAL1/2, CmCHS1/2, CmFNS, CmHQT, and CmHCT were key structural genes in flavones and CQAs biosynthesis. Furthermore, weighted gene co-expression correlation network analysis (WGCNA), k-means clustering, correlation analysis and protein interaction prediction were carried out in this study to identify transcription factors (TFs) associated with flavone and CQA biosynthesis, including MYB, bHLH, AP2/ERF, and MADS-box families. The TFs CmERF/PTI6 and CmCMD77 were proposed to act as upstream regulators of CmMYB3 and CmbHLH143, while CmMYB3 and CmbHLH143 might form a complex to directly regulate the structural genes CmPAL1/2, CmCHS1/2, CmFNS, CmHQT, and CmHCT, thereby controlling flavone and CQA biosynthesis. Overall, these findings provide initial insights into the TF regulatory network underlying flavones and CQAs accumulation in the chrysanthemum capitulum, which laid a theoretical foundation for the quality improvement of C. morifolium ‘HangBaiJu’ and the high-quality development of the industry.

Cold stress seriously affects plant development and secondary metabolism. The basic region/leucine zipper (bZIP) is one of the largest transcription factor (TFs) family and widely involved in plant cold stress response. However, the function of bZIP in Dendrobium catenatum has not been well-documented. Cold inhibited the growth of D. catenatum and increased total polysaccharide and alkaloid contents in stems. Here, 62 DcbZIP genes were identified in D. catenatum, which were divided into 13 subfamilies. Among them, 58 DcbZIPs responded to cold stress, which were selected based on the transcriptome database produced from cold-treated D. catenatum seedlings. Specifically, the expression of DcbZIP3/6/28 was highly induced by cold treatment in leaves or stems. Gene sequence analysis indicated that DcbZIP3/6/28 contains the bZIP conserved domain and is localized to the cell nucleus. Co-expression networks showed that DcbZIP6 was significantly negatively correlated with PAL2 (palmitoyl-CoA), which is involved in flavonoid metabolism. Moreover, DcbZIP28 has significant negative correlations with various metabolism-related genes in the polysaccharide metabolic pathway, including PFKA1 (6-phosphofructokinase), ALDO2 (aldose-6-phosphate reductase) and SCRK5 (fructokinase). These results implied that DcbZIP6 or DcbZIP28 are mainly involved in flavonoid or polysaccharide metabolism. Overall, these findings provide new insights into the roles of the DcbZIP gene family in secondary metabolism in D. catenatum under cold stress.

Ethnopharmacological relevanceIn recent years, Chinese herbal medicine has gained more and more recognition in disease prevention and control due to its low toxicity and comprehensive treatment. C. morifolium (Chrysanthemum morifolium Ramat.), as the medicine food homology plant with the bioactivity of anti-oxidation, anti-inflammatory, neuroprotection and cardiovascular protection, has important therapeutic effects and health benefits for colds, inflammation, cardiovascular diseases and various chronic diseases.Aim of the studyBy reviewing the historical development, classification and distribution of germplasm resources, phytochemistry, pharmacology, and modern application of C. morifolium, the paper provides a reliable basis for the further research and application of chrysanthemum as therapeutic agents and functional additives.Materials and methodsThe literature and information about C. morifolium published in the last ten years were collected from various platforms, including Google Scholar, PubMed, ScienceDirect, Web of Science and China Knowledge Network.ResultsA comprehensive analysis confirmed that C. morifolium originated in China, and it went through the development process from food and tea to medicine for more than 3,000 years. During this period, different cultivars emerged through several breeding techniques and were distributed throughout the world. Moreover, A variety of chemical components such as flavonoids, phenolic acids, volatile oils, and terpenes in chrysanthemum have been proven they possess various pharmacology of anti-inflammatory, anti-oxidant, and prevention of chronic diseases by regulating inflammatory cytokines, oxidative stress responses and signaling pathways, which are the essential conditions to play a role in TCM, nutraceuticals and diet.ConclusionThis paper provides a comprehensive review of historical development, classification, phytochemistry, pharmacology, and modern application of C. morifolium. However, future studies should continue to focus on the bioactive compounds and the synergistic mechanism of the "multi-component, multi-target, and multi-pathway" of chrysanthemum, and it is necessary to develop more innovative products with therapeutic effects.

Although some species that accumulate only cyanidin (Cy) in nature can produce blue flowers through iron ions, there has been no evidence of blue chrysanthemums being generated in this manner. This study revealed that flavonoid extracts from the ray florets of the chrysanthemum cultivar ‘Wandai Fengguang’ turned blue when exposed to Fe3+. Samples that could turn blue were labeled as CB (Cy-determined blue flowers), while samples that did not turn blue were labeled as CN (Cy-determined non-blue flowers). After a series of experiments, a stable screening system was established using flavonoid extracts containing NaAc buffer at pH 5.5 and a total anthocyanin concentration (TAC) of 30 μmol · L-1, and the addition of Fe3+ from 0 to 0.25 μmol · L-1 allowed for the selection of five CB samples from 39 chrysanthemum cultivars. All five CB samples exhibited flower color phenotypes that belonged to Cluster-Ⅰ with redness (a*) values ranging from 29.03 to 45.99, yellowness (b*) values from −11.31 to 3.77, and brightness (L*) values from 29.07 to 45.99. Additionally, the ratio of TAC to total luteolin content (TLC) was found to be a critical factor for distinguishing between CB and CN samples. To realize the desired blue hue in the flavonoid extracts with the participation of Fe3+, a TAC to TLC ratio of 2.25 and above is required. Moreover, the protoplasts and ray florets of CB samples that turned blue with the involvement of Fe2+ showed great potential for cultivating blue chrysanthemums through ferric–anthocyanin chelate. Overall, this study reveals that blue flowers can be cultivated through the increase in the iron ion content, combined with the accumulation of Cy.

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Dendrobium catenatum is a well-known ornamental and medicinal plant producing polysaccharide as the main active ingredient. Cellulose synthase-like (Csl) genes encode glycosyltransferases that are involved in polysaccharide synthases, influencing plant growth development. However, the Csl gene family has not yet been systematically cataloged in D. catenatum. Here, a total of 59 Csl genes were identified in D. catenatum and classified into A-J subgroups. Transcriptomic analysis revealed that nine DcCsl genes were differentially expressed in two varieties (‘Green’ and ‘Red’). The DcCslG3b showed high expression in D. catenatum ‘Green’, which was positively correlated with the polysaccharide content. DcCslG3b, localized in the endoplasmic reticulum, was significantly increased in leaves or stems of D. catenatum under drought and low-temperature treatment, respectively. Co-expression network and promoter cis-element analysis indicated that the ERF2 is potential transcription factor of DcCslG3b. Furthermore, the transient expression of DcCslG3b remarkably reduced water loss rate of tobacco leaves, thus might result in drought tolerance of plants. This study will lead to a better understanding of Csl-mediated polysaccharide synthesis and abiotic stress response in D. catenatum.

Anoectochilus roxburghii is a precious Chinese herb with excellent pharmacological effects. When meet stress, plants produced polyamines (PAs) which are identified in active tissues and play crucial roles in stress responses. Nevertheless, the molecular mechanism of PAs responding to stress in A. roxburghii were largely unknown. Here, for the first time, three PAOs in A. roxburghii were identified from transcriptome data and functionally analyzed. Expression analysis was performed using fluorescence quantitative PCR and eventually, transgenic Arabidopsis thaliana were created to confirm the biological function of ArPAOs. It was found that the highest expression was in young flower buds and the expression levels of three ArPAO genes in various tissues were also different, which indicated their different catalytic preferences and functions. Under the treatment of exogenous spermine and spermidine, the expression of three ArPAOs fluctuated and the overall expression level increased. Overexpression (OE) of ArPAOs could enhance water stress tolerance through mediating polyamine catabolism. It might because the excessive expression of ArPAOs reduce the content of spermine and spermidine, creating a negative feedback regulation upstream and causes plants manufacturing PAs which reinforce water stress tolerance. Taken together, the study firstly reported the overexpression of ArPAOs enhance plant water stress tolerance and regulate the content of polyamines. This work will provide a vital theoretical foundation for uncovering the operation and regulatory mechanism of ArPAOs in promoting plant development and managing stress.

ETHNOPHARMACOLOGICAL RELEVANCE:Bamboos are perennial evergreen plants that belong to the subfamily Bambusoideae of the true grass family Poaceae, with more than thousands of species distributed around the world. They are used as a traditional medicine with demonstrated effects of anti-oxidation, free radical scavenging, anti-inflammatory, liver protection and ameliorating cognitive deficits. Bamboo leaf is mainly used for the treatment of atherosclerotic, diabetic and nervous system diseases.AIM OF THE STUDY:This review aims to provide up-to-date information on the traditional medicinal properties, phytochemistry, pharmacology, and purification technologies of bamboo leaf.MATERIALS AND METHODS:Relevant information on bamboo leaf was obtained by an online search of worldwide accepted scientific databases (Web of Science, ScienceDirect, Elsevier, SpringerLink, ACS Publications, Wiley Online Library and CNKI).RESULTS:More than 100 chemical compounds, including flavonoids and flavonoid glycosides, volatile components, phenolic acids, polysaccharide, coenzyme Q10, phenylpropanoid and amino acids have been reported to be present. These compounds were usually extracted by column chromatography and membrane separation technologies. Preparative high performance liquid chromatography (PHPLC), high-speed counter-current chromatography (HSCCC), simulated moving bed chromatography (SMB) and dynamic axial compression chromatography (DAC) were the advanced separation technologies have been used to isolate C-glycosides from bamboo leaf flavonoid, the main bioactive ingredient of bamboo leaf. Currently, bamboo leaf is mainly used for the treatment of atherosclerotic, diabetic, hepatic diseases and nervous system related symptoms, which are attributed to the presence of bioactive components of bamboo leaf.CONCLUSIONS:Phytochemical and pharmacological analyses of bamboo leaf have been revealed in recent studies. However, most of the pharmacological studies on bamboo leaf have focused on bamboo leaf flavonoids. Further studies need to pay more attention to other phytochemical components of bamboo leaf. In addition, there is lack of sufficient clinical data and toxicity studies on bamboo leaf. Therefore, more clinical and toxicity researches on this plant and constituents are recommended.

Carotenoids are one of the most important pigments for the coloring of many plants, fruits, and flowers. Recently, significant progress has been made in carotenoid metabolism. However, our specific understanding of the transcriptional regulation that controls the expression of carotenoid metabolic genes remains extremely limited. Anemone-type chrysanthemums, a special group of chrysanthemum cultivars, contain elongated disc florets in the capitulum that usually differ in color from the ray florets because of their different carotenoid contents. In this study, the carotenoid composition and content of ray and disc florets from the anemone-type chrysanthemum cultivar "Dong Li Fen Gui" were analyzed by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and the key structural gene CmCCD4a-2, whose differential expression resulted in different carotenoid contents in these two types of florets, was identified. The promoter sequence of CmCCD4a-2 was then used as bait to screen a chrysanthemum flower cDNA library, and the transcription factors (TFs) CmAP3 and CmUIF1 were identified. Y2H, BiFC, and Y3H experiments demonstrated that these two TFs were connected by CmPI to form a CmAP3-CmPI-CmUIF1 TF complex. This TF complex regulated carotenoid metabolism by directly activating the expression of CmCCD4a-2. A large number of target genes regulated directly by the CmAP3-CmPI-CmUIF1 TF complex, including carotenoid biosynthetic genes, flavonoid biosynthetic genes, and flower development-related genes, were identified by DNA-affinity purification sequencing (DAP-seq). This result indicated that the CmAP3-CmPI-CmUIF1 TF complex may participate in multiple processes. These findings expand our knowledge of the transcriptional regulation of carotenoid metabolism in plants and will be helpful for manipulating carotenoid accumulation in chrysanthemum.