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Ginseng, a cornerstone of traditional herbal medicine in Asia, garnered significant attention for its therapeutic potential. Central to its pharmacological effects are ginsenosides, the primary active metabolites, many of which fall within the dammarane-type and share protopanaxadiol as a common precursor. Challenges in extracting protopanaxadiol and ginsenosides from ginseng arise due to their low concentrations in the roots. Emerging solutions involve leveraging microbial cell factories employing genetically engineered yeasts. Here, we optimized the fermentation conditions via the Design of Experiment, realizing 1.2 g/L protopanaxadiol in simple shake flask cultivations. Extrapolating the optimized setup to complex ginsenosides, like compound K, achieved 7.3-fold (0.22 g/L) titer improvements. Our adaptable fermentation conditions enable the production of high-value products, such as sustainable triterpenoids synthesis. Through synthetic biology, microbial engineering, and formulation studies, we pave the way for a scalable and sustainable production of bioactive compounds from ginseng., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Ginsenosides are major active components of Panax ginseng , which are generally glycosylated at C3-OH and/or C20-OH of protopanaxadiol (PPD) and C6-OH and/or C20-OH of protopanaxatriol. However, the glucosides of dammarenediol-II (DM), which is the direct precursor of PPD, have scarcely been separated from P. ginseng . Because different positions and numbers of the hydroxyl and glycosyl groups lead to a diversity of structure and function of the ginsenosides, it can be inferred that DM glucosides may have different pharmacological activities compared with natural ginsenosides. Herein, we first constructed the cell factory for de novo biosynthesis of 3- O -(β- D -glucopyranosyl-(1→2)-β- D -glucopyranosyl)-dammar-24-ene-3β,20 S -diol (3β- O -Glc 2 -DM) by introducing the codon-optimized genes encoding dammarenediol-II synthase, two UDP-glycosyltransferases (UGTs) including UGT74AC1-M7 from Siraitia grosvenorii and UGTPg29 from P. ginseng in Saccharomyces cerevisiae via the CRISPR/Cas9 system. The titer of 3β- O -Glc 2 -DM was then increased from 18.9 to 148.0 mg/L by several metabolic engineering strategies including overexpressing the rate-limiting enzymes of triterpenoid biosynthesis, balancing carbon flux of biosynthetic pathways of triterpenoid and ergosterol, and engineering endoplasmic reticulum. Furthermore, the 3β- O -Glc 2 -DM titer of 766.3 mg/L was achieved through fed-batch fermentation in a 3-L bioreactor. Finally, in vitro assays demonstrated that 3β- O -Glc 2 -DM exhibited a protective effect on H/R-induced cardiomyocyte damage. This work provides a feasible approach for production of 3β- O -Glc 2 -DM as a potential cardioprotective drug candidate.

Panax japonicus, a traditional medicinal plant from the Araliaceae family, produces bioactive triterpenes with health benefits. In Traditional Chinese Medicine, its roots have been used, but the chemical basis of its medicinal use is unclear, particularly regarding the metabolism and regulation of triterpene saponin biosynthesis. This study employed an integrative approach using Ultra Performance Liquid Chromatography (UPLC) and transcriptome analysis. Our UPLC analysis showed that ginsenoside Ro and chikusetsusaponin IVa were mainly detected in P. japonicus root. Subsequently, a comparative transcriptome analysis of four P. japonicus tissues (roots, leaves, flowers and fruits) was conducted using Illumina sequencing. As a result, 90,985 unigenes were functionally annotated from a total of 211,650 assembled non-redundant transcripts. Among these, 42,829 unigenes were annotated in NR database. Tissue-specific gene analysis revealed that roots had the highest number of specifically expressed unigenes (11,832). The majority of these unigenes were associated with metabolic processes. Additionally, tissue expression patterns analysis for three common transcription factor families indicated that WRKY family genes showed a significantly root-specific expression pattern, potentially playing a role in triterpene saponin biosynthesis. Notably, we investigated the expression profiles of genes related to the biosynthesis of triterpene saponins and found that four genes, ACCT, HMGS, HMGR and SS, encoding key enzymes in triterpene saponins biosynthesis pathway, were primarily expressed in the root. Overall, our study provides a set of P. japonicus tissue transcriptome data, which will aid in the discovery of triterpene saponin biosynthetic genes and offers valuable genetic information for this medical plant., Competing Interests: Competing interests The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Statement of permission for collecting Panax japonicus Experimental research and field studies on plants (either cultivated or wild), including the collection of plant material have complied with relevant institutional, national, and international guidelines and legislation. The experimental studies and collection of cultivated Panax japonicus plants were conducted in compliance with the collaboration agreements between the Biomedical Research Institute, Hubei University of Medicine (HBMU), and the Longwangzhai Rare Chinese Herbal Medicine Cultivation Farmers’ Cooperative (LRCC). Permission for the collection of P. japonicus was obtained through this collaboration. Voucher specimens of the plants used in this study have been deposited at the Biomedical Research Institute, HBMU, under the code number VofP-ZZS-2022. The identification of the plant material was confirmed by Prof. Yonghong Zhang at HBMU., (© 2024. The Author(s).)

This work was designed for the in-depth characterization and holistic comparison of up to 12 ginseng varieties, which can benefit the development of functional foods and ensure their authenticity in the food industry. An online comprehensive two-dimensional liquid chromatography/quadrupole time-of-flight mass spectrometry (2D-LC/QTOF-MS) approach was established by configurating the XCharge C18 and HSS Cyano columns. Under the optimal conditions, we characterized a total of 1146 ginsenosides (including 876 potentially new compounds) from 12 ginseng varieties by reference to an in-house library of 573 known ginsenosides and 70 reference compounds. The online 2D-LC/QTOF-MS-based untargeted metabolomics workflows were developed, by which 126 potential ginsenoside markers were unveiled and utilized to establish the key identification points for each ginseng species. Compared with the conventional liquid chromatography/mass spectrometry metabolomics, our multidimensional chromatography approach performed better in discriminating multiple ginseng varieties. This work demonstrates a potent and practical methodology to identify easily confused functional plants.

Ginsenoside Ro, as one of the few oleanane-type ginsenosides, is well known for its unique molecular structure and biological activities. Currently, research on the biosynthesis of ginsenoside Ro is still in its early stages. Therefore, the establishment of a new ginsenoside Ro cell factory is of great significance for the in-depth development and utilization of genes related to ginsenoside Ro synthesis, as well as for the exploration of pathways to obtain ginsenoside Ro. In this study, we cloned endogenous constitutive promoters, terminators, and other genetic elements from S. cerevisiae BY4741. These elements were then sequentially assembled with the uridine diphosphate glucuronic acid transferase gene identified in our previously study ( PgUGAT252645 ) and several other reported key enzyme genes, to construct DNA fragments used for integration into the genome of S. cerevisiae BY4741. By sequentially transferring these DNA fragments into chemically competent cells of engineering strains and conducting screening and target product detection, we successfully constructed an engineered S. cerevisiae strain (BY-Ro) for ginsenoside Ro biosynthesis using S. cerevisiae BY4741 as the host cell. Strain BY-Ro produced 253.32 μg/L of ginsenoside Ro under optimal fermentation conditions. According to subsequent measurements and calculations, this equates to 0.033 mg/g DCW, corresponding to approximately 31% of the ginsenoside Ro content found in plant samples. This study not only included a deeper investigation into the function of PgUGAT252645 but also provides a novel engineering platform for ginsenoside Ro biosynthesis.

Panax japonicus C. A. Meyer is renowned for its significant therapeutic effects and is commonly used worldwide. Its active ingredients, triterpenoid saponins, show variation in content among different tissues. The tissue-specific distribution of saponins is potentially related to the expression of vital genes in the biosynthesis pathway. In this study, the contents of five saponins (ginsenoside Ro, chikusetsusaponin IV, chikusetsusaponin IVa, ginsenoside Rg1, and ginsenoside Rb1) in three different tissues were determined by HPLC. Transcriptome sequencing analysis identified differentially expressed genes (DEGs) involved in triterpenoid saponin biosynthesis, highlighting significant correlations between saponin contents and the expression levels of 10 cytochrome p450 monooxygenase (CYP) and 3 UDP-glycosyltransferase (UGT) genes. Cloning, sequencing, and prokaryotic expression of UGT genes confirmed the molecular weights of UGT proteins. Gene sequence alignment and phylogenetic analysis provided preliminary insights into UGT gene functions. Meanwhile, the function of one UGT gene was characterized in the yeast. These findings advance our understanding of the triterpenoid saponin biosynthesis in P. japonicus and support future research in traditional Chinese medicine (TCM) and synthetic biology.

Panax ginseng is a precious Chinese medicinal plant with a long growth cycle and high medicinal value. Therefore, it is of great significance to explore effective ways to increase its yield and main active substance content to reduce the cost of ginseng, which is widely used in food and clinical applications. Here, we review the key roles of microorganisms in the biological control of ginseng diseases, enhancement of ginseng yield, biotransformation of ginsenosides, and augmentation of ginsenoside bioactivity. The application of microorganisms in P. ginseng faces multiple challenges, including the need for further exploration of efficient microbial strain resources used in the cultivation of ginseng and biotransformation of ginsenosides, lack of microbial application in large-scale field cultivation of ginseng, and unclear mechanism of microbial transformation of ginsenosides. This review provides a deeper understanding of the applications of microorganisms in P. ginseng., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Panax ginseng C.A. Meyer, known as the "King of Herbs," has been used as a nutritional supplement for both food and medicine with the functions of relieving fatigue and improving immunity for thousands of years in China. In agricultural planting, soil environments of different geographical origins lead to obvious differences in the quality of ginseng, but the potential mechanism of the differences remains unclear. In this study, 20 key differential metabolites, including ginsenoside Rb1, glucose 6-phosphate, etc., were found in ginseng from 10 locations in China using an ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS)-untargeted metabolomics approach. The soil properties were analyzed and combined with metagenomics technology to explore the possible relationships among microbial elements in planting soil. Through Spearman correlation analysis, it was found that the top 10 microbial colonies with the highest abundance in the soil were significantly correlated with key metabolites. In addition, the relationship model established by the random forest algorithm and the quantitative relationship between soil microbial abundance and ginseng metabolites were successfully predicted. The XGboost model was used to determine 20(R)-ginseng Rg2 and 2'(R)-ginseng Rg3 as feature labeled metabolites, and the optimal ginseng production area was discovered. These results prove that the accumulation of metabolites in ginseng was influenced by microorganisms in the planting soil, which led to geographical differences in ginseng quality.

Iron plays a crucial role in plant chlorophyll synthesis, respiration, and plant growth. However, excessive iron content can contribute to ginseng poisoning. We previously discovered that the application of silicon (Si) and potassium (K) can mitigate the iron toxicity on ginseng. To elucidate the molecular mechanism of how Si and K alleviate iron toxicity stress in ginseng. We investigated the physiological and transcriptional effects of exogenous Si and K on Panax ginseng. The results suggested that the leaves of ginseng with Si and K addition under iron stress increased antioxidant enzyme activity or secondary metabolite content, such as phenylalanine amino-lyase, polyphenol oxidase, ascorbate peroxidase, total phenols and lignin, by 6.21%-25.94%, 30.12%-309.19%, 32.26%-38.82%, 7.81%-23.66%, and 4.68%-48.42%, respectively. Moreover, Si and K increased the expression of differentially expressed genes (DEGs) associated with resistance to both biotic and abiotic stress, including WRKY (WRKY1, WRKY5, and WRKY65), bHLH (bHLH35, bHLH66, bHLH128, and bHLH149), EREBP, ERF10 and ZIP. Additionally, the amount of DEGs of ginseng by Si and K addition was enriched in metabolic processes, single-organism process pathways, signal transduction, metabolism, synthesis and disease resistance. In conclusion, the utilization of Si and K can potentially reduce the accumulation of iron in ginseng, regulate the expression of iron tolerance genes, and enhance the antioxidant enzyme activity and secondary metabolite production in both leaves and roots, thus alleviating the iron toxicity stress in ginseng., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Masson SAS.)

MicroRNA (miRNA) is a class of non-coding endogenous small-molecule, single-stranded RNAs, and it is involved in post-transcriptional gene expression regulation in plants and plays an important role in plant growth and development. Among them, miRNA156 regulates members of target SPL gene family and thus participates in plant growth and development, hormonal response and adversity stress. However, it has not been reported in ginseng. In this study, based on the previous analysis of the SPL gene family, the age-related and stably expressed SPL gene PgSPL24-09 was obtained in roots. The binding site of miRNA156 to this gene was analyzed using target gene prediction tools, and the interactions between miRNA156 and PgSPL24-09 gene were verified by dual luciferase reporter gene assay and RT-qPCR. At the same time, miRNA156 silencing vector and overexpression vector were constructed and transformed into ginseng adventitious roots and Arabidopsis thaliana to analyze the molecular mechanism of miRNA156-SPL module in regulating the growth of ginseng adventitious roots. This study provides a theoretical basis for the in-depth study of the molecular role of miRNAs in ginseng growth, and also lays the foundation for the study of the role of miRNA156-SPL module in regulating the growth and development of ginseng., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests. Mingzhu Zhao, Kangyu Wang reports financial support was provided by Department of Science and Technology of Jilin Province (20240402046 GH). If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Ginsenoside, the principal active constituent of ginseng, exhibits enhanced bioavailability and medicinal efficacy in rare ginsenosides compared to major ginsenosides. Current research is focused on efficiently and selectively removing sugar groups attached to the major ginsenoside sugar chains to convert them into rare ginsenosides that meet the demands of medical industry and functional foods. The methods for preparing rare ginsenosides encompass chemical, microbial, and enzymatic approaches. Among these, the enzyme conversion method is highly favored by researchers due to its exceptional specificity and robust efficiency. This review summarizes the biological activities of different rare ginsenosides, explores the various glycosidases used in the biotransformation of different major ginsenosides as substrates, and elucidates their respective corresponding biotransformation pathways. These findings will provide valuable references for the development, utilization, and industrial production of ginsenosides., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The accumulation of secondary metabolites in Panax ginseng Meyer (P. ginseng) exhibits significant geographical variation, normally due to environmental factors. The current study aimed at elucidating the key environmental factors modulating the accumulation of secondary metabolites in P. ginseng. Plant and the associated soil samples were collected from ten geographical locations within the latitudinalrange of 27.09°N - 42.39°N and longitudinal range of 99.28°E - 128.19°E. 12 secondary metabolites in P. ginseng toots were measured. And the correlation between secondary metabolites with a series of soil properties and 7 climatic factors were investigated through Pearson's correlation, mantel test, random forest and pathway analysis. The results revealed that climatic factors were stronger drivers of ginseng secondary metabolite profile than soil nutrients. Specifically, temperature seasonality (TS) and soil available phosphorus (AP) were the most effective environments to have significantly and positively influence on the secondary metabolites of ginseng. This findings contribute to identifying optimal cultivation areas for P. ginseng, and hopefully establishing methods for interfering/shaping microclimate for cultivating high-quality P. ginseng., (© 2024. The Author(s).)

Ginseng, from the roots of Panax ginseng C. A. Meyer, is a widely used herbal medicine in Asian countries, known for its excellent therapeutic properties. The growth of P. ginseng is depend on specific and strict environments, with a preference for wetness but intolerance for flooding. Under excessive soil moisture, some irregular rust-like substances are deposited on the root epidermis, causing ginseng rusty symptoms (GRS). This condition leads to a significant reduce in yield and quality, resulting in substantial economic loses. However, there is less knowledge on the cause of GRS and there are no effective treatments available for its treatment once it occurs. Unsuitable environments lead to the generation of large amounts of reactive oxygen species (ROS). We investigated the key indicators associated with the stress response during different physiological stages of GRS development. We observed a significant change in ROS level, MDA contents, antioxidant enzymes activities, and non-enzymatic antioxidants contents prior to the GRS. Through the analysis of soil features with an abundance of moisture, we further determined the source of ROS. The levels of nitrate reductase (NR) and nitric oxide synthase (NOS) activities in the inter-root soil of ginseng with GRS were significantly elevated compared to those of healthy ginseng. These enzymes boost nitric oxide (NO) levels, which in turn showed a favorable correlation with the GRS. The activities of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase first rose and then decreased as GRS developed. Excess soil moisture causes a decrease in oxygen levels. This activated NR and NOS in the soil, resulting in a production of excess NO. The NO then diffused into the ginseng root and triggered a burst of ROS through NADPH located on the cell membrane. Additionally, Fe 2+ in soil was oxidized to red Fe 3+ , and finally led to GRS. This conclusion was also verified by the Sodium Nitroprusside (SNP), a precursor compound producing NO. The presence of NO from NR and NOS in water-saturated soil is responsible for the generation of ROS. Among these, NO is the main component that contribute to the occurrence of GRS., (© 2024. The Author(s).)

Improving the cultivation mode and technology for traditional Chinese medicine has become important for its sustainable development. Monoculture enhances plant diseases, which decreases yield and quality. Intercropping is an effective measure to counterbalance that negative effect. In this study, we focused on Panax quinquefolium L. (ginseng) and four treatments were set up: the control without intercropping, P. quinquefolius + ryegrass (Lolium perenne L.), P. quinquefolius + red clover (Trifolium pratense L.), and P. quinquefolius + ryegrass + red clover. An LC-MS/MS system was used to detect the changes in the P. quinquefolius secondary metabolites, and high-throughput sequencing technology was used to determine the changes in the P. quinquefolius' rhizosphere soil microorganisms. Ginsenoside content, soil enzyme activities, and arbuscular mycorrhizal infection rate of P. quinquefolius were also measured using HPLC, ELISA kits, and microscopy, respectively. Co-intertia and Pearson's analysis were performed to explore the relationship between the metabolites and the P. quinquefolius microorganisms. Intercropping significantly increased the content of ginsenoside metabolites and recruited a large number of beneficial bacteria to the P. quinquefolius rhizosphere. The P. quinquefolius secondary metabolites were associated with the rhizosphere microbial community. For example, the dominant microorganisms, such as Acidobacteriota and Chloroflexi, played a key role in promoting the synthesis of ginsenoside Rd and (20R) ginsenoside Rg3 by P. quinquefolius. Intercropping led to changes in the P. quinquefolius secondary metabolites by driving and reshaping the rhizosphere microorganisms. These findings revealed the potential application of intercropping for improving the quality of P. quinquefolius., (© 2024 Scandinavian Plant Physiology Society.)

Proanthocyanidins (PAs) are flavonoid compounds with important defensive roles in plants. The application of PAs in industries such as the pharmaceutical industry has led to increased interest in enhancing their biosynthesis. In Arabidopsis thaliana, PAs are biosynthesized under the regulation of an R2R3-MYB transcription factor TRANSPARENT TESTA 2 (TT2), which can interact with other proteins, including TRANSPARENT TESTA GLABRA 1 (TTG1), while also regulating a plant's response to abiotic stressors. However, the regulation of PA biosynthesis in the high-value medicinal plant Panax ginseng (ginseng) has not yet been studied. Understanding the mechanism of PAs biosynthesis regulation in ginseng may be helpful in increasing the plant's range of pharmacological applications. This study found that the overexpression of PgTT2 increased PA biosynthesis by an average of 67.3% in ginseng adventitious roots and 50.5% in arabidopsis seeds. Furthermore, transgenic arabidopsis plants overexpressing PgTT2 produced increased reactive oxygen species (ROS) scavenging ability by influencing abscisic acid synthesis and signaling. However, under high salinity stress, seed germination and growth rate of seedlings were decreased. An expression analysis of plants facing salt stress revealed increased transcripts of an ABA biosynthetic gene, NCED3, and ABA signaling genes ABI5 and ABI3. Moreover, the PgTT2 protein showed a direct interaction with PgTTG1 in yeast two-hybrid assays. This study therefore reveals novel information on the transcriptional regulation of PA production in ginseng and shows how PgTT2 influences the ABA response pathway to regulate responses to ROS and salt stress., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ok Ran Lee reports financial support was provided by National Research Foundation of Korea, Rural Development Administration, and Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Objective: This study aims to investigate the effect of red ginseng polysaccharide (RGP) on gastric cancer (GC) development and explore its mechanism., Methods: GC cell lines AGS were treated with varying concentrations of RGP (50, 100, and 200 μg/mL). AGS cells treated with 200 μg/mL RGP were transfected with aquaporin 3 (AQP3) overexpression vector. Cell proliferation, viability, and apoptosis were evaluated by MTT, colony formation assay, and flow cytometry, respectively. Real-time quantitative reverse transcription PCR (qRT-PCR) was used to detect the expression of AQP3. The levels of Fe2+, malondialdehyde, and lactate dehydrogenase were measured using their respective detection kits, and the reactive oxygen species levels was determined by probe 2',7'-dichlorodihydrofluorescein diacetate. The expression of ferroptosis-related protein and PI3K/Akt pathway-related protein were assessed by western blot. In vivo experiments in nude mice were performed and the mice were divided into four groups ( n  = 5/group) which gavage administrated with 150 mg/kg normal saline, and 75, 150, 300 mg/kg RGP, respectively. Their tumor weight and volume were recorded., Results: RGP treatment effectively inhibited the proliferation and viability of AGS cells in a dosage-dependent manner and induced apoptosis. It induced ferroptosis in AGS cells, as well as inhibiting the expression of PI3K/Akt-related proteins. AQP3 overexpression could reversed the effect of RGP treatment on ferroptosis. Confirmatory in vivo experiments showed that RGP could reduce the growth of implanted tumor, with increased RGP concentration resulting in greater tumor inhibitory effects., Conclusion: RGP might have therapeutic potential against GC, effectively inhibiting the proliferation and viability of AGS cells.

Introduction: Ginseng berry (GB) has previously been demonstrated to improve systemic insulin resistance and regulate hepatic glucose metabolism and steatosis in mice with diet-induced obesity (DIO)., Objectives: In this study, the role of GB in metabolism was assessed using metabolomics analysis on the total liver metabolites of DIO mice., Methods: Metabolomic profiling was performed using capillary electrophoresis time-of-flight mass spectrometry (CE-TOF/MS) of liver tissue from mice on a 12-wk normal chow diet (NC), high-fat diet (HFD), and HFD supplemented with 0.1% GB (HFD + GB). The detected metabolites, its pathways, and functions were analyzed through partial least square discriminant analysis (PLS-DA), the small molecular pathway database (SMPDB), and MetaboAnalyst 5.0., Results: The liver metabolite profiles of NC, HFD, and GB-fed mice (HFD + GB) were highly compartmentalized. Metabolites involved in major liver functions, such as mitochondrial function, gluconeogenesis/glycolysis, fatty acid metabolism, and primary bile acid biosynthesis, showed differences after GB intake. The metabolites that showed significant correlations with fasting blood glucose (FBG), insulin, and homeostatic model assessment for insulin resistance (HOMA-IR) were highly associated with mitochondrial membrane function, energy homeostasis, and glucose metabolism. Ginseng berry intake increased the levels of metabolites involved in mitochondrial membrane function, decreased the levels of metabolites related to glucose metabolism, and was highly correlated with metabolic phenotypes., Conclusion: This study demonstrated that long-term intake of GB changed the metabolite of hepatosteatotic livers in DIO mice, normalizing global liver metabolites involved in mitochondrial function and glucose metabolism and indicating the potential mechanism of GB in ameliorating hyperglycemia in DIO mice., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

To convert ginsenosides Rb1, Rb2, Rb3, and Rc into Rd by a single enzyme, a putative β-glycosidase (Pxbgl) from the xylan-degrading bacterium Petroclostridium xylanilyticum was identified and used. The k cat / K m value of Pxbgl for Rb3 was 18.18 ± 0.07 mM -1 /s, which was significantly higher than those of Pxbgl for other ginsenosides. Pxbgl converted almost all Rb3 to Rd with a productivity of 5884 μM/h, which was 346-fold higher than that of only β-xylosidase from Thermoascus aurantiacus . The productivity of Rd from the Panax ginseng root and Panax notoginseng leaf was 146 and 995 μM/h, respectively. Mutants N293 K and I447L from site-directed mutagenesis based on bioinformatics analysis showed an increase in specific activity of 29 and 7% toward Rb3, respectively. This is the first report of a β-glycosidase that can simultaneously remove four different glycosyls at the C-20 position of natural PPD-type ginsenosides and produce Rd as the sole product from P. notoginseng leaf extracts with the highest productivity.

Panax ginseng C.A. Mey., known for its medicinal and dietary supplement properties, primarily contains pharmacologically active ginsenosides. However, the regulatory mechanisms linking ginseng root development with ginsenoside biosynthesis are still unclear. Root meristem growth factors (RGFs) are crucial for regulating plant root growth. In our study, we identified five ginseng RGF peptide sequences from the ginseng genome and transcriptome libraries. We treated Arabidopsis and ginseng adventitious roots with exogenous Panax ginseng RGFs (PgRGFs) to assess their activities. Our results demonstrate that PgRGF1 influences gravitropic responses and reduces lateral root formation in Arabidopsis. PgRGF1 has been found to restrict the number and length of ginseng adventitious root branches in ginseng. Given the medicinal properties of ginseng, We determined the ginsenoside content and performed transcriptomic analysis of PgRGF1-treated ginseng adventitious roots. Specifically, the total ginsenoside content in ginseng adventitious roots decreased by 19.98 % and 63.71 % following treatments with 1 μM and 10 μM PgRGF1, respectively, compared to the control. The results revealed that PgRGF1 affects the accumulation of ginsenosides by regulating the expression of genes associated with auxin transportation and ginsenoside biosynthesis. These findings suggest that PgRGF1, as a peptide hormone regulator in ginseng, can modulate adventitious root growth and ginsenoside accumulation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Ginseng saponins (ginsenosides), bioactive compounds derived from ginseng, are widely used natural products with potent therapeutic properties in the management of various ailments, particularly tumors, cardiovascular and cerebrovascular diseases, and immune system disorders. Autophagy, a highly regulated and multistep process involving the breakdown of impaired organelles and macromolecules by autophagolysosomes and autophagy-related genes (ATGs), has gained increasing attention as a potential target for ginsenoside-mediated disease treatment. This review aims to provide a comprehensive overview of recent research advances in the understanding of autophagy-related signaling pathways and the role of ginsenoside-mediated autophagy regulation. By delving into the intricate autophagy signaling pathways underpinning the pharmacological properties of ginsenosides, we highlight their therapeutic potential in addressing various conditions. Our findings serve as a comprehensive reference for further investigation into the medicinal properties of ginseng or ginseng-related products., (© 2024 Wiley-VHCA AG, Zurich, Switzerland.)

Ginsenosides, the primary bioactive constituents in ginseng (Panax ginseng), possess substantial pharmacological potential and are in high demand in the market. The plant hormone methyl jasmonate (MeJA) effectively elicits ginsenoside biosynthesis in P. ginseng, though the regulatory mechanism remains largely unexplored. NAC transcription factors are critical in intricate plant regulatory networks and participate in numerous plant physiological activities. In this study, we identified a MeJA-responsive NAC transcription factor gene, PgNAC72, from a transcriptome library produced from MeJA-treated P. ginseng callus. Predominantly expressed in P. ginseng flowers, PgNAC72 localizes to the nucleus. Overexpressing PgNAC72 (OE-PgNAC72) in P. ginseng callus notably elevated total saponin levels, particularly dammarane-type ginsenosides, by upregulating dammarenediol synthase (PgDDS), encoding a key enzyme in the ginsenoside biosynthesis pathway. Electrophoretic mobility shift assays and dual-luciferase assays confirmed that PgNAC72 binds to the NAC-binding elements in the PgDDS promoter, thereby activating its transcription. Further RNA-seq and terpenoid metabolomic data in the OE-PgNAC72 line confirmed that PgNAC72 enhances ginsenoside biosynthesis. These findings uncover a regulatory role of PgNAC72 in MeJA-mediated ginsenoside biosynthesis, providing insights into the ginsenoside regulatory network and presenting a valuable target gene for metabolic engineering., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Panax ginseng is a perennial herb with the main active compounds of ginsenosides. Among the reported ginsenosides, ginsenoside Rg_1 not only has a wide range of medicinal functions and abundant content but also is one of the major ginsenoside for the quality evaluation of this herb in the Chinese Pharmacopoeia. The main biosynthesis pathway of ginsenoside Rg_1 in P. ginseng has been clarified, which lays a foundation for the comprehensive and in-depth analysis of the biosynthesis and regulatory mechanism of ginseno-side Rg_1. However, the biosynthesis of ginsenoside Rg_1 is associated with other complex processes involving a variety of regulatory genes and catalyzing enzyme genes, which remain to be studied comprehensively. With the transcriptome data of 344 root samples from 4-year-old P. ginseng plants and their corresponding ginsenoside Rg_1 content obtained in the previous study, this study screened out 217 differentially expressed genes(DEGs) with Rg_1 content changes by DEseq2 analysis in R language. Furthermore, the weighted gene co-expression network analysis(WGCNA) revealed 40 hub genes among the DEGs.Pearsoncorrelation analysis was further perforned to yield 20 candidate genes significantly correlated with ginsenoside Rg_1 content, and these genes were annotated to multiple metabolic processes including primary metabolism and secondary metabolism. Finally, the treatment of P. ginseng adventitious roots with methyl jasmonate indicated that 16 of these genes promoted the biosynthesis of ginsenoside Rg_1 in response to methyl jasmonate induction. Finally, one of the 16 genes was randomly selected to verify the function of the gene by genetic transformation and qRT-PCR and to confirm the rationality of the methodology of this study. The above results lay a foundation for studying the mechanism for regulation on the synthesis of ginsenoside Rg_1 and provide genetic resources for the industrial production of ginsenoside Rg_1.

Ginseng frequently encounters environmental stress during its growth and development. Late Embryogenesis Abundant (LEA) proteins play a crucial role in combating adversity stress, particularly against abiotic challenges In this study, 107 LEA genes from ginseng, spanning eight subfamilies, were identified, demonstrating significant evolutionary conservation, with the LEA2 subfamily being most prominent. Gene duplication events, primarily segmental duplications, have played a major role in the expansion of the LEA gene family, which has undergone strong purifying selection. PgLEAs were unevenly distributed across 22 chromosomes, with each subfamily featuring unique structural domains and conserved motifs. PgLEAs were expressed in various tissues, exhibiting distinct variations in abundance and tissue specificity. Numerous regulatory cis-elements, related to abiotic stress and hormones, were identified in the promoter region. Additionally, PgLEAs were regulated by a diverse array of abiotic stress-related transcription factors. A total of 35 PgLEAs were differentially expressed following treatments with ABA, GA, and IAA. Twenty-three PgLEAs showed significant but varied responses to drought, extreme temperatures, and salinity stress. The transformation of tobacco with the key gene PgLEA2-50 enhanced osmoregulation and antioxidant levels in transgenic lines, improving their resistance to abiotic stress. This study offers insights into functional gene analysis, focusing on LEA proteins, and establishes a foundational framework for research on ginseng's resilience to abiotic stress., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Ginseng ( Panax ginseng C. A. Meyer) is an ancient and valuable Chinese herbal medicine, and ginsenoside, as the main active ingredient of ginseng, has received wide attention because of its various pharmacological active effects. Cytochrome P450 is the largest family of enzymes in plant metabolism and is involved in the biosynthesis of terpenoids, alkaloids, lipids, and other primary and secondary plant metabolites. It is significant to explore more PgCYP450 genes with unknown functions and reveal their roles in ginsenoside synthesis. In this study, based on the five PgCYP450 genes screened in the pre-laboratory, through the correlation analysis with the content of ginsenosides and the analysis of the interactions network of the key enzyme genes for ginsenoside synthesis, we screened out those highly correlated with ginsenosides, PgCYP309 , as the target gene from among the five PgCYP450 genes. Methyl jasmonate-induced treatment of ginseng adventitious roots showed that the PgCYP309 gene responded to methyl jasmonate induction and was involved in the synthesis of ginsenosides. The PgCYP309 gene was cloned and the overexpression vector pBI121-PgCYP309 and the interference vector pART27-PgCYP309 were constructed. Transformation of ginseng adventitious roots by the Agrobacterium fermentum -mediated method and successful induction of transgenic ginseng hairy roots were achieved. The transformation rate of ginseng hairy roots with overexpression of the PgCYP309 gene was 22.7%, and the transformation rate of ginseng hairy roots with interference of the PgCYP309 gene was 40%. Analysis of ginseng saponin content and relative gene expression levels in positive ginseng hairy root asexual lines revealed a significant increase in PPD, PPT, and PPT-type monomeric saponins Re and Rg2. The relative expression levels of PgCYP309 and PgCYP716A53v2 genes were also significantly increased. PgCYP309 gene promotes the synthesis of ginsenosides, and it was preliminarily verified that PgCYP309 gene can promote the synthesis of dammarane-type ginsenosides.

Ginseng (Panax ginseng C. A. Mey.) is an important and valuable medicinal plant species used in traditional Chinese medicine, and its metabolite ginsenoside is the primary active ingredient. The FAR1/FHY3 gene family members play critical roles in plant growth and development as well as participate in a variety of physiological processes, including plant development and signaling of hormones. Studies have indicated that methyl jasmonate treatment of ginseng adventitious roots resulted in a significant increase in the content of protopanaxadiol ginsenosides. Therefore, it is highly significant to screen the FAR1/FHY3 gene family members in ginseng and preliminarily investigate their expression patterns in response to methyl jasmonic acid signaling. In this study, we screened and identified the FAR1/FHY3 family genes in the ginseng transcriptome databases. And then, we analyzed their gene structure and phylogeny, chromosomal localization and expression patterns, and promoter cis-acting elements, and made GO functional annotations on the members of this family. After that, we treated the ginseng adventitious roots with 200 mM methyl jasmonate and investigated the trend of the expression of four genes containing the largest number of methyl jasmonate cis-acting elements at different treatment times. All four genes were able to respond to methyl jasmonate, the most significant change was in the PgFAR40 gene. This study provides data support for subsequent studies of this family member in ginseng and provides experimental reference for subsequent validation of the function of this family member under methyl jasmonic acid signaling., (© 2024. The Author(s).)

Chronic Heart Failure (CHF) is a significant global public health issue, with high mortality and morbidity rates and associated costs. Disease modules, which are collections of disease-related genes, offer an effective approach to understanding diseases from a biological network perspective. We employed the multi-Steiner tree algorithm within the NeDRex platform to extract CHF disease modules, and subsequently utilized the Trustrank algorithm to rank potential drugs for repurposing. The constructed disease module was then used to investigate the mechanism by which Panax ginseng ameliorates CHF. The active constituents of Panax ginseng were identified through a comprehensive review of the TCMSP database and relevant literature. The Swiss target prediction database was utilized to determine the action targets of these components. These targets were then cross-referenced with the CHF disease module in the STRING database to establish protein-protein interaction (PPI) relationships. Potential action pathways were uncovered through Gene Ontology (GO) and KEGG pathway enrichment analyses on the DAVID platform. Molecular docking, the determination of the interaction of biological macromolecules with their ligands, and visualization were conducted using Autodock Vina, PLIP, and PyMOL, respectively. The findings suggest that drugs such as dasatinib and mitoxantrone, which have low docking scores with key disease proteins and are reported in the literature as effective against CHF, could be promising. Key components of Panax ginseng, including ginsenoside rh4 and ginsenoside rg5, may exert their effects by targeting key proteins such as AKT1, TNF, NFKB1, among others, thereby influencing the PI3K-Akt and calcium signaling pathways. In conclusion, drugs like dasatinib and midostaurin may be suitable for CHF treatment, and Panax ginseng could potentially mitigate the progression of CHF through a multi-component-multi-target-multi-pathway approach. Disease module analysis emerges as an effective strategy for exploring drug repurposing and the mechanisms of traditional Chinese medicine in disease treatment., (© 2024. The Author(s).)

Dysbiosis of gut microbiota is believed to be associated with inflammatory bowel disease (IBD). Ginsenoside compound K (CK), the main metabolite of Panax ginseng ginsenoside, has proven effective as an anti-inflammatory agent in IBD. However, the mechanisms by which CK modulates gut microbiota to ameliorate IBD remain poorly understood. Herein, CK demonstrated the potential to suppress the release of proinflammatory cytokines by gut microbiota modulation. Notably, supplementation with CK promoted the restoration of a harmonious balance in gut microbiota, primarily by enhancing the populations of Lactobacillus and Akkermansia . Furthermore, CK considerably elevated the concentrations of tryptophan metabolites derived from Lactobacillus that could activate the aryl hydrocarbon receptor. Overall, the promising alleviative efficacy of CK primarily stemmed from the promotion of Lactobacillus growth and production of tryptophan metabolites, suggesting that CK should be regarded as a prospective prebiotic agent for IBD in the future.

The allelopathic autotoxicity of ginsenosides is an important cause of continuous cropping obstacles in ginseng planting. There is no report on the potential molecular mechanism of the correlation between polarity of ginsenoside components and their allelopathic autotoxicity. This study applied a combination of metabolomics and transcriptomics analysis techniques, combined with apparent morphology, physiological indexes, and cell vitality detection of the ginseng hairy roots, through which the molecular mechanism of correlation between polarity and allelopathic autotoxicity of ginsenosides were comprehensively studied. The hairy roots of ginseng presented more severe cell apoptosis under the stress of low-polarity ginsenoside components (ZG70). ZG70 exerted allelopathic autotoxicity by regulating the key enzyme genes of cis-zeatin (cZ) synthesis pathway, indole-3-acetic acid (IAA) synthesis pathway, and jasmonates (JAs) signaling transduction pathway. The common pathway for high-polarity ginsenoside components (ZG50) and ZG70 to induce the development of allelopathic autotoxicity was through the expression of key enzymes in the gibberellin (GA) signal transduction pathway, thereby inhibiting the growth of ginseng hairy roots. cZ, indole-3-acetamid (IAM), gibberellin A1 (GA1), and jasmonoyl-L-isoleucine (JA-ILE) were the key response factors in this process. It could be concluded that the polarity of ginsenoside components were negatively correlated with their allelopathic autotoxicity.

Ginseng is widely recognized for its diverse health benefits and serves as a functional food ingredient with global popularity. Ginsenosides with a broad range of pharmacological effects are the most crucial active ingredients in ginseng. This study aimed to derive ginseng glucosyl oleanolate (GGO) from ginsenoside Ro through enzymatic conversion and evaluate its impact on liver cancer in vitro and in vivo. GGO exhibited concentration-dependent HepG2 cell death and markedly inhibited cell proliferation via the MAPK signaling pathway. It also attenuated tumor growth in immunocompromised mice undergoing heterograft transplantation. Furthermore, GGO intervention caused a modulation of gut microbiota composition by specific bacterial populations, including Lactobacillus, Bacteroides, Clostridium, Enterococcus , etc., and ameliorated SCFA metabolism and colonic inflammation. These findings offer promising evidence for the potential use of GGO as a natural functional food ingredient in the prevention and treatment of cancer.

Tau is a microtubule-associated protein that plays a critical role in the stabilization and modulation of neuronal axons. Tau pathology is stronger associated with cognitive decline in patients with Alzheimer's disease (AD) than amyloid beta (Aβ) pathology. Hence, tau targeting is a promising approach for the treatment of AD. Previous studies have demonstrated that the non-saponin fraction with rich polysaccharide (NFP) from Korean red ginseng (KRG) can modulate tau aggregation and exert a therapeutic effect on AD. Therefore, we investigated the efficacy of NFP isolated from KRG on tau pathology in experimental models of AD. Our results showed that NFP from KRG ameliorated deposition and hyperphosphorylation of tau in the brain of 3xTg mice. Moreover, NFP from KRG modulated the aggregation and dissociation of tau K18 in vitro. We demonstrated the alleviatory effects of NFP from KRG on hyperphosphorylated tau and tau kinase in okadaic acid-treated HT22 cells. Furthermore, NFP from KRG mitigated Aβ deposition, neurodegeneration, and neuroinflammation in 3xTg mice. We revealed the neuroprotective effects of NFP from KRG on tau-induced neuronal loss in HT22 cells. Our results indicate that NFP extracted from KRG is a novel therapeutic agent for the treatment of AD associated with tau pathology., Competing Interests: Declaration of competing interest The authors have declared that no competing interest exists., (Copyright © 2024. Published by Elsevier B.V.)

Pq3-O-UGT2, derived from Panax quinquefolius, functions as a ginsenoside glucosyltransferase, utilizing UDP-glucose (UDPG) as the sugar donor to catalyze the glycosylation of Rh2 and F2. An essential step in comprehending its catalytic mechanism involves structural analysis. In preparation for structural analysis, we expressed Pq3-O-UGT2 in the Escherichia coli (E. coli) strain Rosetta (DE3). The recombinant Pq3-O-UGT2 was purified through Ni-NTA affinity purification, a two-step ion exchange chromatography, and subsequently size-exclusion chromatography (SEC). Notably, the purified Pq3-O-UGT2 showed substantial activity toward Rh2 and F2, catalyzing the formation of Rg3 and Rd, respectively. This activity was discernible within a pH range of 4.0-9.0 and temperature range of 30-55 °C, with optimal conditions observed at pH 7.0-8.0 and 37 °C. The catalytic efficiency of Pq3-O-UGT2 toward Rh2 and F2 was 31.43 s -1  mΜ -1 and 169.31 s -1  mΜ -1 , respectively. We further crystalized Pq3-O-UGT2 in both its apo form and co-crystalized forms with UDPG, Rh2 and F2, respectively. High-quality crystals were obtained and X-ray diffraction data was collected for all co-crystalized samples. Analysis of the diffraction data revealed that the crystal of Pq3-O-UGT2 co-crystalized with UDP-Glc belonged to space group P1, while the other two crystals belonged to space group P2 1 2 1 2 1 . Together, this study has laid a robust foundation for subsequent structural analysis of Pq3-O-UGT2., Competing Interests: Declaration of competing interest The authors declare that there is no conflicts of interest or financial bias in this work., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Panax ginseng C. A. Meyer ( Ginseng ) is one of the most used traditional Chinese herbal medicines, with its roots being used as the main common medicinal parts; its therapeutic potential has garnered significant attention. AUXIN/INDOLE-3-ACETIC ACID ( Aux/IAA ) is a family of early auxin-responsive genes capable of regulating root development in plants through the auxin signaling pathway. In the present study, 84 Aux/IAA genes were identified from the ginseng genome and their complexity and diversity were determined through their protein domains, phylogenetic relationships, gene structures, and cis-acting element predictions. Phylogenetic analyses classified PgIAA into six subgroups, with members in the same group showing greater sequence similarity. Analyses of interspecific collinearity suggest that segmental duplications likely drove the evolution of PgIAA genes, followed by purifying selection. An analysis of cis-regulatory elements suggested that PgIAA family genes may be involved in the regulation of plant hormones. RNA-seq data show that the expression pattern of Aux/IAA genes in Ginseng is tissue-specific, and PgIAA02 and PgIAA36 are specifically highly expressed in lateral, fibrous, and arm roots, suggesting their potential function in root development. The PgIAA02 overexpression lines exhibited an inhibition of lateral root growth in Ginseng . In addition, yeast two-hybrid and subcellular localization experiments showed that PgIAA02 interacted with PgARF22/PgARF36 (ARF: auxin response factor) in the nucleus and participated in the biological process of root development. The above results lay the foundation for an in-depth study of Aux/IAA and provide preliminary information for further research on the role of the Aux/IAA gene family in the root development of Ginseng .

During ageing, the permeability of the intestinal barrier increases, the integrity of the intestinal barrier decreases, and the physiology of intestinal cells changes. Furthermore, intestinal inflammation and excessive oxidative stress are both likely to cause systemic diseases. Ginseng oligopeptides have a positive significant effect in terms of improving human health and delaying ageing, but their role in the ageing of the intestine has not been studied much. In our experiment, we constructed a gut-on-a-chip model and induced senescence of the chip with H 2 O 2 so as to explore the effects of ginseng oligopeptides on the senescent intestine. The experimental results showed that ginseng oligopeptides had no obvious effects on the integrity of the intestine, including the TEER value and the expression of tight junction proteins. However, ginseng oligopeptides might have other positive effects, such as inhibiting excessive cell proliferation, promoting mucin secretion, and increasing the antioxidant capacity of the intestine, to improve intestinal health.

Ginseng ( Panax ginseng C.A. Meyer) is a perennial herb belonging to the family Araliaceae and has been used for thousands of years in East Asia as an essential traditional medicine with a wide range of pharmacological activities of its main active ingredient, ginsenosides. The AP2/ERF gene family, widely present in plants, is a class of transcription factors capable of responding to ethylene regulation that has an influential role in regulating the synthesis of major active ingredients in medicinal plants and in response to biotic and abiotic stresses, which have not been reported in Panax ginseng . In this study, the AP2/ERF gene was localized on the ginseng chromosome, and an AP2/ERF gene duplication event was also discovered in Panax ginseng . The expression of seven ERF genes and three key enzyme genes related to saponin synthesis was measured by fluorescence quantitative PCR using ethylene treatment of ginseng hairy roots, and it was observed that ethylene promoted the expression of genes related to the synthesis of ginsenosides, among which the PgERF120 gene was the most sensitive to ethylene. We analyzed the sequence features and expression patterns of the PgERF120 gene and found that the expression of the PgERF120 gene was specific in time and space. The PgERF120 gene was subsequently cloned, and plant overexpression and RNA interference vectors were constructed. Ginseng adventitious roots were transformed using the Agrobacterium tumefaciens -mediated method to obtain transgenic ginseng hairy roots, and the gene expression, ginsenoside content and malondialdehyde content in overexpression-positive hairy roots were also analyzed. This study preliminarily verified that the PgERF120 gene can be involved in the regulation of ginsenoside synthesis, which provides a theoretical basis for the study of functional genes in ginseng and a genetic resource for the subsequent use of synthetic biology methods to improve the yield of ginsenosides.

In this study, a new polysaccharide (PSPJ) with specific molecular weight and monosaccharide compositions was isolated and purified from the water extract of Panacis Japonici Rhizoma (PJR). 16S rRNA analysis and untargeted metabolomic analysis were used to assess PSPJ's efficacy in averting non-alcoholic fatty liver disease (NAFLD). This study indicated that PSPJ significantly reduced liver fat accumulation, the increase in blood lipids and ALT caused by HFD, indicating that PSPJ can prevent NAFLD. We demonstrated through cell experiments that PSPJ does not directly affect liver cells. The gut microbiota disorder and alterations in short-chain fatty acids (SCFAs) induced by the high-fat diet (HFD) were ameliorated by PSPJ, as evidenced by the analysis of 16S rRNA. In particular, supplementing PSPJ reduced the abundance of Turicibacter, Dubosiella, and Staphylococcus, and increased the abundance of Bacteroides, Blautia, and Lactobacillus. Untargeted metabolomic analysis shows that PSPJ improves liver metabolic disorders by regulating arachidonic acid metabolism, carbohydrate digestion and absorption, fatty acid biosynthesis, fatty acid metabolism and retinol metabolism. The findings of our investigation indicate that PSPJ has the potential to modulate liver metabolism through alterations in the composition of intestinal bacteria, hence preventing NAFLD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The author would like to express our sincere gratitude to Nanjing Agricultural University for providing reasonable experimental conditions and equipment. At the same time, thanks for the support of the above fundings., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Di(2-ethylhexyl) phthalate (DEHP) is regarded as a priority environmental pollutant. This study explored the adsorption and accumulation of DEHP within the ginseng-soil system and the mechanism of DEHP toxicity to ginseng (Panax ginseng C.A. Meyer). Under exposure to 22.10 mg/kg DEHP in soil, DEHP mainly accumulated in ginseng leaves (20.28 mg/kg), stems (4.84 mg/kg) and roots (2.00 mg/kg) after 42 days. The oxidative damage, metabolism, protein express of ginseng were comprehensively measured and analyzed. The results revealed that MDA presented an activation trend in ginseng stems and leaves after 42 days of DEHP exposure, while the opposite trend was observed for POD. Levels of ginsenoside metabolites Rg2, Rg3, Rg5, Rd, Rf and CK decreased in the ginseng rhizosphere exudates under DEHP stress. Further investigations revealed that DEHP disrupts ginsenoside synthesis by inducing glycosyltransferase (GS) and squalene synthase (SS) protein interactions. Molecular docking indicated that DEHP could stably bind to GS and SS by intermolecular forces. These findings provide new information on the ecotoxicological effect of DEHP on ginseng root., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Panax ginseng has been widely applied as an important herb in traditional medicine to treat numerous human disorders. However, the inflammatory regulation effect of P. ginseng distillate (GSD) has not yet been fully assessed. To determine whether GSD can ameliorate inflammatory processes, a GSD was prepared using the vacuum distillation process for the first time, and the regulation effect on lipopolysaccharide-induced macrophages was assessed. The results showed that GSD effectively inhibited nitric oxide (NO) formation and activation of inducible nitric oxide synthase (iNOS) mRNA in murine macrophage cell, but not cyclooxygenase-2 production. The mRNA expression pattern of tumor necrosis factor alpha and IL-6 were also reduced by GSD. Furthermore, we confirmed that GSD exerted its anti-inflammatory effects by downregulating c-Jun NH 2 -terminal kinase (JNK) phosphorylation, the extracellular signal-regulated kinase phosphorylation, and signaling pathway of nuclear factor kappa B (NF-κB). Our findings revealed that the inflammatory regulation activity of GSD could be induced by iNOS and NO formation inhibition mediated by regulation of nuclear factor kappa B and p38/JNK MAPK pathways.

Objective: To investigate the protective effect of total saponins of Panax japonicus (TSPJ) against CCl 4 -induced acute liver injury (ALI) in rats and explore the underlying pharmacological mechanisms., Methods: Male SD rat models of CCl 4 -induced ALI were given intraperitoneal injections of distilled water, 100 mg/kg biphenyl bisabololol, or 50, 100, and 200 mg/kg TSPJ during modeling ( n =8). Liver functions (AST, ALT, TBil and ALP) of the rats were assessed and liver pathologies were observed with HE staining. Immunohistochemistry was used to detect the expressions of PI3K/Akt/NF-κB signaling pathway molecules in liver tissue; ELISA was used to determine the levels of T-SOD, GSH-P x , and MDA. Western blotting was performed to detect the expression levels of PI3K-Akt and SIRT6-NF-κB pathways in the liver tissue., Results: Network pharmacological analysis indicated that the key pathways including PI3K/Akt mediated the therapeutic effect of TSPJ on ALI. In the rat models of ALI, treatments with biphenyl bisabololol and TSPJ significantly ameliorated CCl 4 -induced increase of serum levels AST, ALT, ALP, TBil and MDA and decrease of T-SOD and GSH-P x levels (all P < 0.01). The rat models of ALI showed significantly increased expression of p-NF-κB ( P < 0.01), decreased expressions of PI3K, p-Akt and SIRT6 proteins, and elevated expression levels of p-NF-κB, TNF-α and IL-6 proteins in the liver, which were all significantly improved in the treatment groups ( P < 0.05 or 0.01)., Conclusion: TSPJ can effectively alleviate CCl 4 -induced ALI in rats by suppressing inflammatory responses and oxidative stress in the liver via regulating the PI3K/Akt and SIRT6/NF-κB pathways.

Activation of NOD-like receptor protein 3 (NLRP3) inflammasome exacerbates liver inflammation and fibrosis in nonalcoholic steatohepatitis (NASH), suggesting that development of inflammasome inhibitor can become leading candidate to ameliorate NASH. Panax ginseng (P. ginseng) contains numerous bioactive natural components to reduce inflammation. This study aims to identify inhibitory components of P. ginseng for NLRP3 inflammasome activation. We separated polar and non-polar fractions of P. ginseng and tested modulation of NLRP3 inflammasome, and then identified pure component for inflammasome inhibitor which ameliorates diet-induced NASH. Non-polar P. ginseng fractions obtained from ethyl acetate solvent attenuated IL-1β secretion and expression of active caspase-1. We revealed that panaxydol (PND) is pure component to inhibit NLRP3 inflammasome activation. PND blocked inflammasome cytokines release, pyroptotic cell death, caspase-1 activation and specking of inflammasome complex. Inhibitory effect of PND was specific to NLRP3-dependent pathway via potential interaction with ATP binding motif of NLRP3. Moreover, in vivo studies showed that PND plays beneficial roles to reduce tissue inflammations through disruption of NLRP3 inflammasome and to ameliorate the development of NASH. These results provide new insight of natural products, panaxydol, for NLRP3 inflammasome inhibitor and could offer potential therapeutic candidate for reliving NASH., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Panax ginseng , a traditional Chinese medicine with a history spanning thousands of years, faces overexploitation and challenges related to extended growth periods. Tissue-cultured adventitious roots and stem cells are alternatives to wild and field-cultivated ginseng. In this study, we assessed the in vitro xanthine oxidase and α-glucosidase inhibitory activities of saponin extracts among cultured cambial meristematic cells (CMC), adventitious ginseng roots (AGR), and field-cultivated ginseng roots (CGR). The xanthine oxidase (XO) and α-glucosidase inhibitory activities were determined by uric acid estimation and the p -NPG method, respectively. Spectrophotometry and the Folin-Ciocalteu, aluminum nitrate, and Bradford methods were employed to ascertain the total saponins and phenolic, flavonoid, and protein contents. The calculated IC 50 values for total saponin extracts against XO and α-glucosidase were 0.665, 0.844, and >1.6 mg/mL and 0.332, 0.745, and 0.042 mg/mL for AGR, CMC, CGR, respectively. Comparing the total saponin, crude protein, and total phenolic contents revealed that AGR > CMC > CGR. To the best of our knowledge, this study presents the first report on the in vitro comparison of xanthine oxidase and α-glucosidase inhibitory activities among AGR, CMC, and CGR. The findings offer valuable insights into the development of hypoglycemic and antihyperuricemic medicinal, nutraceutical, and functional products utilizing AGR and CMC.

Inflammatory bowel disease (IBD) is closely linked to the homeostasis of the intestinal environment, and exosomes can be used to treat IBD due to their high biocompatibility and ability to be effectively absorbed by the intestinal tract. However, Ginseng-derived nanoparticles (GDNPs) have not been studied in this context and their mechanism of action remains unclear. Here, we investigated GDNPs ability to mediate intercellular communication in a complex inflammatory microenvironment in order to treat IBD. We found that GDNPs scavenge reactive oxygen species from immune cells and intestinal epithelial cells, inhibit the expression of pro-inflammatory factors, promote the proliferation and differentiation of intestinal stem cells, as well as enhancing the diversity of the intestinal flora. GDNPs significantly stabilise the intestinal barrier thereby promoting tissue repair. Overall, we proved that GDNPs can ameliorate inflammation and oxidative stress in vivo and in vitro, acting on the TLR4/MAPK and p62/Keap1/Nrf2 pathways, and exerting an anti-inflammatory and antioxidant effect. GDNPs mitigated IBD in mice by reducing inflammatory factors and improving the intestinal environment. This study offers new evidence of the potential therapeutic effects of GDNPs in the context of IBD, providing the conceptual ground for an alternative therapeutic strategy., (© 2024. The Author(s).)

Panax ginseng Meyer and Inula japonica Thunb. are well established in traditional medicine and are known for their therapeutic properties in managing a range of ailments such as diabetes, asthma, and cancer. Although P. ginseng and I. japonica can alleviate pulmonary fibrosis (PF), the anti-fibrosis effect on PF by the combination of two herbal medicines remains unexplored. Therefore, this study explores this combined effect. In conditions that were not cytotoxic, MRC-5 cells underwent treatment using the formula combining P. ginseng and I. japonica (ISE081), followed by stimulation with transforming growth factor (TGF)-β1, to explore the fibroblast-to-myofibroblast transition (FMT). After harvesting the cells, mRNA levels and protein expressions associated with inflammation and FMT-related markers were determined to evaluate the antiinflammation activities and antifibrosis effect of ISE081. Additionally, the anti-migratory effects of ISE081 were validated through a wound-healing assay. ISE081 remarkably reduced the mRNA levels of interleukin (IL)-6, IL-8, α-smooth muscle actin (SMA), and TGF-β1 in MRC-5 cells and suppressed the α-SMA and fibronectin expressions, respectively. Furthermore, ISE081 inhibited Smad2/3 phosphorylation and wound migration of MRC-5 cells. Under the same conditions, comparing those of ISE081, P. ginseng did not affect the expression of α-SMA, fibronectin, and Smad2/3 phosphorylation, whereas I. japonica significantly inhibited them but with cytotoxicity. The results indicate that the synergistic application of P. ginseng and I. japonica enhances the anti-fibrotic properties in pulmonary fibroblasts and concurrently diminishes toxicity. Therefore, ISE081 has the potential as a prevention and treatment herbal medicine for PF.

Plants usually produce defence metabolites in non-active forms to minimize the risk of harm to themselves and spatiotemporally activate these defence metabolites upon pathogen attack. This so-called two-component system plays a decisive role in the chemical defence of various plants. Here, we discovered that Panax notoginseng, a valuable medicinal plant, has evolved a two-component chemical defence system composed of a chloroplast-localized β-glucosidase, denominated PnGH1, and its substrates 20(S)-protopanaxadiol ginsenosides. The β-glucosidase and its substrates are spatially separated in cells under physiological conditions, and ginsenoside hydrolysis is therefore activated only upon chloroplast disruption, which is caused by the induced exoenzymes of pathogenic fungi upon exposure to plant leaves. This activation of PnGH1-mediated hydrolysis results in the production of a series of less-polar ginsenosides by selective hydrolysis of an outer glucose at the C-3 site, with a broader spectrum and more potent antifungal activity in vitro and in vivo than the precursor molecules. Furthermore, such β-glucosidase-mediated hydrolysis upon fungal infection was also found in the congeneric species P. quinquefolium and P. ginseng. Our findings reveal a two-component chemical defence system in Panax species and offer insights for developing botanical pesticides for disease management in Panax species., (© 2024. The Author(s).)

Panax ginseng is an important medicinal plant, and ginsenosides are the main bioactive molecules of ginseng. The TCP (TBI, CYC, PCF) family is a group of transcription factors (TFs) that play an important role in plant growth and development, hormone signalling and synthesis of secondary metabolites. In our study, 78 PgTCP transcripts were identified from the established ginseng transcriptome database. A phylogenetic tree analysis showed that the 67 PgTCP transcripts with complete open reading frames were classified into three subfamilies, including CIN, PCF, and CYC/TB1. Protein structure analysis showed that PgTCP genes had bHLH structures. Chromosomal localization analysis showed that 63 PgTCP genes were localized on 17 of the 24 chromosomes of the Chinese ginseng genome. Expression pattern analysis showed that PgTCP genes differed among different lineages and were spatiotemporally specific. Coexpression network analysis indicated that PgTCP genes were coexpressed and involved in plant activities or metabolic regulation in ginseng. The expression levels of PgTCP genes from class I (PCF) were significantly downregulated, while the expression levels of PgTCP genes from class II (CIN and CYC/TB1) were upregulated, suggesting that TCP genes may be involved in the regulation of secondary metabolism in ginseng. As the PgTCP26-02 gene was found to be related to ginsenoside synthesis, its predicted protein structure and expression pattern were further analysed. Our results provide new insights into the origin, differentiation, evolution and function of the PgTCP gene family in ginseng, as well as the regulation of plant secondary metabolism., (© 2024. The Author(s).)

Background: Intestinal stem cells (ISCs) are the reservoir source of various types of intestinal cells, and the decline of stem cell function in the gut may be a potential factor for aging-related disease. The present study aimed to explore the regulatory mechanisms of Panax ginseng C.A.Meyer (Araliaceae, Panax genus) that could restore gut aging by enhancing intestinal function and regulating ISCs in aging mice based on the Wnt/β-catenin signaling pathway., Methods: A total of 60 ICR male mice were randomly divided into control, model, metformin, and ginseng water decoction (GWD) 3.6, 1.8, and 0.9 g/kg groups. The aging model was induced by 1 % D-galactose (s.c. 0.1 mL/10 g) for 28 days. Moreover, GWD was given to aging mice intragastrically (i.g.) once a day for 28 successive days. The learning memory ability, pathological status, and function in the ileum tissue, the activity of digestive enzymes, and short-chain fatty acid (SCFA) content in the colon were evaluated, and the related mechanism was investigated., Results: Ginseng can decrease the escape latency time and increase the swimming speed and the number of crossing platforms in aging mice. Moreover, the pathology of ileum tissue improved, the length of the intestinal villi increased, and the width of the villi and the depth of the crypts decreased. The activities of trypsin, α-amylase, and lipase increased in duodenal content and intestinal mucosa. In the colon, the content of SCFA, such as acetic acid, propionic acid and butyric acid, increased, indicating that ginseng significantly improves intestinal function impairment. The mRNA expressions and protein levels of β-catenin, C-myc, GSK-3β, Lgr5, and Olfm4 were upregulated in the ginseng group., Conclusions: Ginseng improves intestinal function and regulates the function of ISCs in order to protect intestinal health by activating the Wnt/β-catenin signaling pathway in aging mice., Competing Interests: Declaration of competing interest None., (Copyright © 2023. Published by Elsevier Inc.)

Objective: This study aimed to explore the mechanism of total saponin of black ginseng (TSBG) in treating heart failure (HF) in DOX-induced HF model rats., Methods: Rats with HF induced by the intraperitoneal injection of DOX were treated with TSBG (low dose, 30 mg/kg/day; medium dose, 60 mg/kg/day; high dose, 120 mg/kg/day) and shakubar trivalsartan (80 mg/kg/day, positive control) for four weeks. Serum BNP and ANP levels were tested by ELISA, and pathological tissue sections were examined. Serum metabolites were measured using nontargeted metabolomic techniques. The expression of Akt/mTOR autophagy-associated proteins in heart tissue was detected using Western blot, including Beclin1, p62, LCII and LC3I., Results: Compared with the model group, rats in the TSBG-H group had a significantly lower heart index (p < 0.05), significantly lower serum levels of BNP (p < 0.01) and ANP (p < 0.01) and significantly fewer cardiac histopathological changes. Metabolomic results showed that TSBG significantly back-regulated 12 metabolites (p < 0.05), including cholesterol, histamine, sphinganine, putrescine, arachidonic acid, 3-sulfinoalanine, hypotaurine, gluconic acid and lysoPC (18:0:0). These metabolite changes were involved in taurine and hypotaurine metabolism, arachidonic acid metabolism, sphingolipid metabolism, etc. The protein expression level of p-Akt/Akt and p-mTOR/mTOR was significantly up-regulated (p < 0.001), whereas that of Beclin1, p62 (p < 0.001) and LCII/LC3I was down-regulated (p < 0.05)., Conclusion: TSBG has an excellent therapeutic effect on DOX-induced HF in rats, probably by regulating the Akt/mTOR autophagy signalling pathway, resulting in the improvement of taurine and hypotaurine metabolism, arachidonic acid metabolism and sphingolipid metabolism, which may provide a reference for elucidating the potential mechanism of action of TSBG against HF., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Polysaccharides isolated from Korean ginseng berries (GBPs) have shown beneficial effects such as immunomodulatory, anti-inflammatory, anti-cancer, and anti-diabetic properties. However, little is known about anti-inflammatory effects of GBPs. Thus, the purpose of this study was to investigate anti-inflammatory properties of four fractions of GBPs, namely GBP-C, GBP-F1, GBP-F2, and GBP-F3, in macrophages. Their toxicities and effects on NO production in RAW264.7 cells were assessed by culturing cells with various concentrations of GBPs and stimulating cells with LPS. Furthermore, expression levels of inflammatory mediators, cytokines, cell surface molecules, and immune signaling pathways were evaluated in LPS-stimulated macrophages using different fractions of GBPs at 450 μg/mL. These GBPs activated LPS-stimulated RAW264.7 cells to significantly reduce NO production. They suppressed the expression of mRNA and cell surface molecules via MAPK and NF-κB pathways. Collectively, results revealed that all four GBP fractions showed anti-inflammatory effects, with GBP-F1 having a more efficient anti-inflammatory effect than GBP-C, GBP-F2, and GBP-F3. The structure of GBP-F1 mainly consists of 1 → 3)- Araf, 1 → 4)- Glcp, and 1 → 6)-Galp glycosidic linkages. These results demonstrate that GBPs can be employed as alternative natural sources of anti-inflammatory agents., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Rod-in et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

In recent years, the phenomenon of acute poisoning and organ damage caused by organophosphorus pesticides (OPs) has been a frequent occurrence. Chlorpyrifos (CPF) is one of the most widely used organophosphorus pesticides. The main active components of ginseng stems and leaves are total ginseng stem-and-leaf saponins (GSLSs), which have various biological effects, including anti-inflammatory, antioxidant and anti-tumor activities. We speculate that these could have great potential in the treatment of severe diseases and the relief of organophosphorus-pesticide-induced side effects; however, their mechanism of action is still unknown. At present, our work aims to evaluate the effects of GSLSs on the antioxidation of CPF in vivo and in vitro and their potential pharmacological mechanisms. Mice treated with CPF (5 mg/kg) showed severe intestinal mucosal injury, an elevated diamine oxidase (DAO) index, the decreased expression of occlusive protein-1 (ZO-1) and occlusive protein, an impaired intestinal mucosal oxidation system and intestinal villi relaxation. In addition, chlorpyrifos exposure significantly increased the contents of the inflammatory factor TNF-α and the oxidative-stress-related indicators superoxide dismutase (SOD), catalase (CAT), glutathione SH (GSH), glutathione peroxidase (GSH-PX), reactive oxygen species (ROS) and total antioxidant capacity (T-AOC); elevated the level of lipid peroxide malondialdehyde (MDA); reversed the expression of Bax and caspase; and activated NF-κB-related proteins. Interestingly, GSLS supplementation at doses of 100 and 200 mg/kg significantly reversed these changes after treatment. Similar results were observed in cultured RAW264.7 cells. Using flow cytometry, Hoechst staining showed that GSLSs (30 μg/mL, 60 μg/mL) could improve the cell injury and apoptosis caused by CPF and reduce the accumulation of ROS in cells. In conclusion, GSLSs play a protective role against CPF-induced enterotoxicity by inhibiting NF-κB-mediated apoptosis and alleviating oxidative stress and inflammation.

Ginsenoside Rg5, a relatively uncommon secondary ginsenoside, exhibits notable pharmacological activity and is commonly hypothesized to originate from the dehydration of Rg3. In this work, we compared different conversion pathways using Rb1, R -Rg3 and S -Rg3 as the raw material under simple acid catalysis. Interestingly, the results indicate that the conversion follows this reaction activity order Rb1 > S -Rg3 > R -Rg3, which is contrary to the common understanding of Rg5 obtained from Rg3 by dehydration. Our experimental results have been fully confirmed by theoretical calculations and a NOESY analysis. The DFT analysis reveals that the free energies of S-Rg3 and R-Rg3 in generating carbocation are 7.56 mol/L and 7.57 mol/L, respectively, which are significantly higher than the free energy of 1.81 mol/L when Rb1 generates the same carbocation. This finding aligns with experimental evidence suggesting that Rb1 is more prone to generating Rg5 than Rg3. The findings from the nuclear magnetic resonance (NMR) analysis suggest that the fatty chains (C22-C27) in R-Rg3 and S-Rg3 adopt a Gauche conformation and an anti conformation with C16-C17 and C13-C17, respectively, due to the relatively weak repulsive van der Waals force. Therefore, the configuration of R-Rg3 is more conducive to the formation of intramolecular hydrogen bonds between 20C-OH and 12C-OH, whereas S-Rg3 lacks this capability. Consequently, this also explains the fact that S-Rg3 is more prone to dehydration to generate Rg5 than R-Rg3. Additionally, our research reveals that the synthetic route of Rg5 derived from protopanaxadiol (PPD)-type ginsenosides (including Rb1, Rb2, Rb3, Rc and Rd) exhibits notable advantages in terms of efficacy, purity and yield when compared to the pathway originating from Rg3. Moreover, this study presents a highly effective and practical approach for the extensive synthesis of Rg5, thereby facilitating the exploration of its pharmacological properties and potential application in drug discovery.