UHPLC-HRMS based saponins profiling of three morphological regions in American ginseng (Panax quinquefolium L.) and their correlation with the antioxidant activity.

American ginseng (Panax quinquefolium L.) is used as tonic plant and high-grade nourishment. Ultra-high-performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) method was established for identifying the chemical constituent in three morphological regions of American ginseng, including main root (MR), rhizome (RH) and lateral root (LR). The 63 saponins was identified in different morphological regions of 10 American ginseng samples. The chemical maker compounds in corresponding morphological region, while the major compounds of MR (malonyl-ginsenoside Rb1, ginsenoside Rd, Rs2 and pseudo-RC1), LR (stipuleanoside R2, ginsenoside Re and malonyl-ginsenoside Rc), and RH (malonyl-ginsenoside Rd, Rb3, and chikusetsu saponin II) were discovered. Correlation analysis showed that 11 compounds were positively correlated with the antioxidant activity of American ginseng. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-023-01453-4.

Biochar reduces the cadmium content of Panax quinquefolium L. by improving rhizosphere microecology.

Cadmium (Cd) accumulation in American ginseng (Panax quinquefolium L.) can negatively impact its yield and safety. Our previous study found that biochar could reduce cadmium content of P. quinquefolius, however, the mechanism was yet to be elucidated. In the present study, we tested four treatments in order to reveal the mechanism by which this phenomenon occurs: control, Cd, Cd + biochar and biochar. The results showed that the following responses were induced by the addition of biochar under Cd stress. Firstly, the soil physicochemical properties were improved, this is especially true for the soil pH value and soil organic matter content, which were increased by 20.42 % and 15.57 %, respectively. Secondly, the relative abundances of several beneficial microorganism phyla; such as Proteobacteria, Bacteroidota and Actinobacteria; were increased by 10.69 %, 20.11 % and 60.86 %, respectively. Thirdly, treatment with biochar reduced the Cd content by increasing cadmium-chelated metabolites within the soil (e.g., naringenin, caffeic acid, and valine) and increasing detoxification substances in plants (e.g., malic acid, flavonoids, and fumaric acid). Changes in these metabolites were significantly correlated with rhizosphere microecology. In summary, biochar treatment reduced the Cd content in seedlings by improving the soil properties, rhizosphere community, soil metabolites, and plant metabolites.

Panax quinquefolium L. and Salvia miltiorrhiza Bunge. Enhances Angiogenesis by Regulating the miR-155-5p/HIF-1α/VEGF Axis in Acute Myocardial Infarction.

Background: Combination of Panax quinquefolium L and Salvia miltiorrhiza Bunge. (PS) has been widely used in the clinical treatment of ischemic heart disease. The purpose of this study was to explore the therapeutic effect and mechanism of PS on angiogenesis in rats after acute myocardial infarction (AMI). Methods: A rat model of AMI was established by ligating the left anterior descending (LAD) artery. The grouping and administration scheme were as follows: sham group, model group, PS low-dose (PS-L) group, PS high-dose (PS-H) group, PX-478 group and angiotensin converting enzyme inhibitor (ACEI) group. After 28 days of treatment, echocardiography, myocardial infarct size, some angiogenesis markers and the miR-155-5p/HIF-1α/VEGF axis were measured. Results: PS improved cardiac structure and function, reduced infarct size, and alleviated myocardial fibrosis and inflammatory cell infiltration in AMI rats. Mechanistically, PS enhanced the expression of HGF and bFGF in serum, increased the levels of MVD and CD31 in myocardial tissues, and inhibited the activation of the miR-155-5p/HIF-1α/VEGF pathway, which ultimately promoted angiogenesis. In addition, the regulatory effect of PS on angiogenesis was partly abolished by PX-478. Conclusion: PS increased the expression of MVD and CD31 in the myocardium and stimulated angiogenesis. The above effects of PS may be associated with the inhibition of the miR-155-5p/HIF-1α/VEGF axis.

First report of root rot caused by Fusarium commune on American ginseng (Panax quinquefolium L.) in China.

American ginseng (Panax quinquefolium L.) is one of the most valuable herb crops because of its unique pharmacological effects. In 2019, American ginseng plants withered and root rot with incidences of 20-45% were observed in about 70000m2 of ginseng production field located in mountainous valley of Benxi city (41º23'32" N, 124º04'27" E), Liaoning Province in China. Disease symptoms included chlorotic leaves with dark brown discoloration extending gradually from the basal to the apical part of the leaves. Water-soaked, irregular lesions appeared on the surface of roots and rotten at later stage. Twenty-five symptomatic roots were surface-sterilized by immersion in 2% sodium hypochlorite (NaOCl) for 3 min, followed by rinsing three times in sterilized water. The sections healthy tissues bordered rotten tissues, i.e. the leading edge, were cut into 4-5 mm pieces with a sterile scalpel and 4 pieces were placed on each PDA plate. After 5 days incubation at 26°C, total of 68 single spores were obtained from the colonies with an inoculation needle under stereomicroscope. Colonies from single conidia were white to greyish white, densely floccose to fluffy, and the reverse grayish yellow with dull violet pigmentation. Single-celled and ovoid microconidia in false heads were borne on aerial monophialidic or polyphialidic conidiophores on Carnation Leaf Agar (CLA) media, and measured 5.0 -14.5 × 3.0 -4.8 µm (n=25). Macroconidia were two to four septa, slightly curved, apical and basal cells were also curved, and they measured 22.5 - 45.5 × 4.5 - 6.3 µm (n=25). Chlamydospores were singly or in pairs, circular or subcircular, smooth, and measuring 5 - 10.5 µm (n=25) in diameter. Morphologically, the isolates were identified as Fusarium commune (Skovgaard et al. 2003; Leslie and Summerell 2006 ). To confirm the identity, the rDNA partial translation elongation factor1 alpha (TEF-a) gene and the internal transcribed spacer (ITS) region of ten isolates were amplified and sequenced (O'Donnell et al. 2015; White et al. 1990). Identical sequences were obtained, and one representative sequence of isolate BGL68 was submitted to GenBank. BLASTn analysis of both the TEF-α (MW589548) and the ITS (MW584396) sequences, revealed 100% and 99.46 % sequence identity to F. commune MZ416741 and KU341322, respectively. The pathogenicity test was conducted under greenhouse conditions. The surface of healthy 2-year-old American ginseng roots was washed and disinfested in 2% NaOCl for 3 min before rinsing in sterilized water. Twenty roots were wounded with a toothpick, resulting in tiny perforations (1.0 × 1.0×3.0 mm), 3 perforations were wounded on each root. Inoculums was prepared from the culture of isolate BGL68 incubate in potato dextrose broth (PD) for 5 days at 26°C,140 rpm. Ten wounded roots were immersed in a conidial suspension (2 × 105 conidia/ml) for four hours in a plastic bucket, and planted in five containers (two roots per container) filled with sterile soil. Another ten wounded roots were immersed in sterilized distilled water and planted in five containers as controls. The containers were incubated for four weeks in a greenhouse at temperature between 23°C and 26°C, under a 12-hr light and dark regime, and irrigate with sterile water every 4 days. Three weeks after inoculation, all inoculated plants exhibited chlorotic leaves, wilting and root rot. The taproot and the fibrous roots showed brown to black root rot and no symptoms in non-inoculated controls. The fungus was reisolated from the inoculated plants, but not from any of the control plants. The experiment was repeated two times with similar results. This is the first report of root rot caused by F. commune on American ginseng in China. The disease might bring a threat to this ginseng production and should be implemented effective control measures to reduce losses.

An efficient isolation and purification of broad partition coefficient range ginsenosides from roots of Panax quinquefolium L. by linear gradient counter-current chromatography coupled with preparative high-performance liquid chromatography.

As a famous health food, roots of Panax quinquefolium L. possessed immune regulation and enhancement of the central nervous system, in which ginsenosides are the main active component with different numbers and positions of sugars, causing different chemical polarities with a challenge for the separation and isolation. In this study, a fast and effective bilinear gradient counter-current chromatography was proposed for preparative isolation ginsenosides with a broad partition coefficient range from roots of Panax quinquefolium L. In terms of the established method, the mobile phases comprising n-butanol and ethyl acetate were achieved by adjusting the proportion. Coupled with the preparative HPLC, eleven main ginsenosides were successfully separated, including ginsenoside Rg1 (1), Re (2), acetyl ginsenoside Rg1 (3), Rb1 (4), Rc (5), Rg2 (6), Rb3 (7), quinquefolium R1 (8), Rd (9), gypenoside X VII (10) and notoginsenoside Fd (11), with purities exceeding 95% according to the HPLC results. Tandem mass spectrometry and electrospray ionization mass spectrometry were adopted for recognizing the isolated compound architectures. Our study suggests that linear gradient counter-current chromatography effectively separates the broad partition coefficient range of ginsenosides compounds from the roots of Panax quinquefolium L. In addition, it can apply to active compound isolation from other complicated natural products.

Microbial inoculants and garbage fermentation liquid reduced root-knot nematode disease and As uptake in Panax quinquefolium cultivation by modulating rhizosphere microbiota community.

Objective: To find a suitable ecological cultivation measure to solve the problem of root-knot nematode disease of Panax quinquefolium (Panacis Quinquefolii Radix) and the heavy metals accumulating in its roots. Methods: Three-year-old P. quinquefolium was treated with four different combinations of microbial inoculant (MI) and garbage fermentation liquid (GFL) [the joint application of 'TuXiu' MI and Fifty potassium MI (TF), the combination use of 'No. 1' MI and Fifty potassium MI (NF), 'Gulefeng' poly-γ-glutamic acid MI (PGA), GFL], and the untreated control (CK). Here, high-throughput sequencing, ICP-MS and UPLC were employed to systematically characterize changes of microbial diversity and structure composition, heavy metals (As, Cd and Pb) content and ginsenoside content among different treatments. Results: The results revealed that different MIs and GFL could increase the root dry weight of P. quinquefolium, PGA enhanced it by 83.24%, followed by GFL (49.93%), meanwhile, PGA and GFL were able to lessen root-knot nematode disease incidence by 57.25% and 64.35%. The treatment of PGA and GFL can also effectively reduce heavy metals in roots. The As content in GFL and PGA was decreased by 52.17% and 43.48% respectively, while the Cd and Pb contents of GFL and PGA was decreased somewhat. Additionally, the content of total ginsenosides was increased by 42.14% and 42.07%, in response to TF and NF, respectively. Our metagenomic analysis showed that the relative abundance of particular soil microbial community members related to the biocontrol of root-knot nematode disease and plant pathogen (i.e., Chaetomium in NF, Xylari in GFL, and Microascus in PGA), heavy metal bioremediation (Hyphomacrobium in PGA and Xylaria in GFL), and nitrogen fixation (Nordella and Nitrospira in TF) was significantly increased; notably, potential harmful microflora, such as Plectosaphaerella and Rhizobacter, were more abundant in the control group. Conclusion: MI and GFL could improve the quality of P. quinquefolium by modifying its rhizosphere microbial community structure and composition, both of them are beneficial to the development of ecological cultivation of P. quinquefolium.

Transformation Mechanism of Rare Ginsenosides in American Ginseng by Different Processing Methods and Antitumour Effects.

The mechanism by which ginsenosides from Panax quinquefolium L. transform into rare saponins by different processing methods and their antitumour effects have yet to be fully elucidated. Our study aimed to detect the effect of amino acids and processing methods on the conversion of ginsenosides in American ginseng to rare ginsenosides, using 8 monomeric ginsenosides as substrates to discuss the reaction pathway and mechanism. S180 tumour-bearing mice were established to study the antitumour effects of American ginseng total saponins (AGS-Q) or American ginseng total saponins after transformation (AGS-H) synergistic CTX. The results showed that aspartic acid was the best catalyst, and the thermal extraction method had the best effect. Under the optimal conditions, including a reaction temperature of 110°C, an aspartic acid concentration of 5%, a reaction time of 2.5 h and a liquid-solid ratio of 30 mL/g, the highest conversion of Rk1 and Rg5 was 6.58 ± 0.11 mg/g and 3.74 ± 0.05 mg/g, respectively. In the reaction pathway, the diol group saponins participated in the transformation process, and the triol group saponins basically did not participate in the transformation process. AGS-Q or AGS-H synergistic CTX, or AGS-H synergistic CTX/2 could significantly increase the tumour inhibition rate, spleen index and white blood cell count, had a significant upregulation effect on IL-2 and IL-10 immune cytokines; significantly restored the ratio of CD4+/CD8+; and significantly inhibited the level of CD4+CD25+. AGS-Q or AGS-H synergistic with CTX or CTX/2 can significantly upregulate the expression of Bax and cleaved-Caspase-3 and inhibit the expression of antiapoptotic protein Bcl-2. AGS synergistic CTX in the treatment of S180 tumour-bearing mice can improve the efficacy and reduce toxicity.

Microbial inoculants and garbage fermentation liquid reduced root-knot nematode disease and As uptake in Panax quinquefolium cultivation by modulating rhizosphere microbiota community / 中草药·英文版

Objective: To find a suitable ecological cultivation measure to solve the problem of root-knot nematode disease of Panax quinquefolium (Panacis Quinquefolii Radix) and the heavy metals accumulating in its roots. Methods: Three-year-old P. quinquefolium was treated with four different combinations of microbial inoculant (MI) and garbage fermentation liquid (GFL) [the joint application of ‘TuXiu’ MI and Fifty potassium MI (TF), the combination use of ‘No. 1′ MI and Fifty potassium MI (NF), ‘Gulefeng’ poly-γ-glutamic acid MI (PGA), GFL], and the untreated control (CK). Here, high-throughput sequencing, ICP-MS and UPLC were employed to systematically characterize changes of microbial diversity and structure composition, heavy metals (As, Cd and Pb) content and ginsenoside content among different treatments. Results: The results revealed that different MIs and GFL could increase the root dry weight of P. quinquefolium, PGA enhanced it by 83.24%, followed by GFL (49.93%), meanwhile, PGA and GFL were able to lessen root-knot nematode disease incidence by 57.25% and 64.35%. The treatment of PGA and GFL can also effectively reduce heavy metals in roots. The As content in GFL and PGA was decreased by 52.17% and 43.48% respectively, while the Cd and Pb contents of GFL and PGA was decreased somewhat. Additionally, the content of total ginsenosides was increased by 42.14% and 42.07%, in response to TF and NF, respectively. Our metagenomic analysis showed that the relative abundance of particular soil microbial community members related to the biocontrol of root-knot nematode disease and plant pathogen (i.e., Chaetomium in NF, Xylari in GFL, and Microascus in PGA), heavy metal bioremediation (Hyphomacrobium in PGA and Xylaria in GFL), and nitrogen fixation (Nordella and Nitrospira in TF) was significantly increased; notably, potential harmful microflora, such as Plectosaphaerella and Rhizobacter, were more abundant in the control group. Conclusion: MI and GFL could improve the quality of P. quinquefolium by modifying its rhizosphere microbial community structure and composition, both of them are beneficial to the development of ecological cultivation of P. quinquefolium.

Machine learning methods to predict the cultivation age of Panacis Quinquefolii Radix.

BACKGROUND: American ginseng (AG) is a valuable medicine widely consumed as a herbal remedy throughout the world. Huge price difference among AG with different growth years leads to intentional adulteration for higher profits. Thus, developing reliable approaches to authenticate the cultivation ages of AG products is of great use in preventing age falsification. METHODS: A total of 106 batches of AG samples along with their 9 physicochemical features were collected and measured from experiments, which was then split into a training set and two test sets (test set 1 and 2) according to the cultivation regions. Principle component analysis (PCA) was carried out to examine the distribution of the three data sets. Four machine learning (ML) algorithms, namely elastic net, k-nearest neighbors, support vector machine and multi-layer perception (MLP) were employed to construct predictive models using the features as inputs and their growth years as outputs. In addition, a similarity-based applicability domain (AD) was defined for these models to ensure the reliability of the predictive results for AG samples produced in different regions. RESULTS: A positive correlation was observed between the several features and the growth years. PCA revealed diverse distributions among different cultivation regions. The most accurate model derived from MLP shows good prediction power for the fivefold cross validation and the test set 1 with mean square error (MSE) of 0.017 and 0.016 respectively, but a higher MSE value of 1.260 for the test set 2. After applying the AD, all models showed much lower prediction errors for the test samples within AD (IDs) than those outside the AD (ODs). MLP remains the best predictive model with an MSE value of 0.030 for the IDs. CONCLUSION: Cultivation years have a close relationship with bioactive components of AG. The constructed models and AD are also able to predict the cultivation years and discriminate samples that have inaccurate prediction results. The AD-equipped models used in this study provide useful tools for determining the age of AG in the market and are freely available at https://github.com/dreadlesss/Panax_age_predictor .

Panax quinquefolium L. Ginsenosides from Hairy Root Cultures and Their Clones Exert Cytotoxic, Genotoxic and Pro-Apoptotic Activity towards Human Colon Adenocarcinoma Cell Line Caco-2.

American ginseng, Panax quinquefolium (L.), is traditionally used in folk medicine. It exhibits a range of anti-inflammatory, hepatoprotective, anti-diabetic, anti-obesity, anti-hyperlipidemic and anti-carcinogenic effects. Its main components are ginsenosides, also known as panaxosides or triterpene saponins. In order to obtain high yields of ginsenosides, different methods of controlled production are involved, i.e., with hairy root cultures. However, they are still employed under in vitro conditions. Our studies revealed that hairy root cultures subjected to an elicitation process can be considered as a potent source of ginsenosides. The present study examines the biological activity of ginseng hairy root cultures against the Caco-2 human adenocarcinoma cell line. Among our six different clones of P. quinquefolium hairy roots, extracts B and Be (treated with elicitor) were the strongest inhibitors of the cellular metabolic activity. While all extracts induced DNA damage, B and Be also generated reactive oxygen species (ROS) in a concentration-dependent manner, which was correlated with the depletion of the mitochondrial membrane potential and induction of apoptosis. These findings indicate that further research concerning P. quinquefolium hairy root cultures should focus on the activity of rare ginsenosides and other biologically active compound profiles (i.e., phenolic compounds).

Comparison of Ginsenoside Components of Various Tissues of New Zealand Forest-Grown Asian Ginseng (Panax Ginseng) and American Ginseng (Panax Quinquefolium L.).

Asian ginseng (Panax ginseng) and American ginseng (Panax quinquefolium L.) are the two most important ginseng species for their medicinal properties. Ginseng is not only popular to consume, but is also increasingly popular to cultivate. In the North Island of New Zealand, Asian ginseng and American ginseng have been grown in Taupo and Rotorua for more than 15 years. There are no publications comparing the chemical constituents between New Zealand-grown Asian ginseng (NZPG) and New Zealand-grown American ginseng (NZPQ). In this study, fourteen ginsenoside reference standards and LC-MS2 technology were employed to analyze the ginsenoside components of various parts (fine root, rhizome, main root, stem, and leaf) from NZPG and NZPQ. Fifty and 43 ginsenosides were identified from various parts of NZPG and NZPQ, respectively, and 29 ginsenosides were found in both ginseng species. Ginsenoside concentrations in different parts of ginsengs were varied. Compared to other tissues, the fine roots contained the most abundant ginsenosides, not only in NZPG (142.49 ± 1.14 mg/g) but also in NZPQ (115.69 ± 3.51 mg/g). For the individual ginsenosides of both NZPG and NZPQ, concentration of Rb1 was highest in the underground parts (fine root, rhizome, and main root), and ginsenoside Re was highest in the aboveground parts (stem and leaf).

A new ocotillol-type ginsenoside from stems and leaves of Panax quinquefolium L. and its anti-oxidative effect on hydrogen peroxide exposed A549 cells.

A new ocotillol-type ginsenoside, namely 12-one-pseudoginsenoside F11 (12-one-PF11), was isolated from stems and leaves of Panax quinquefolium, whose structure was elucidated 6-O-[α-L-rhamnopyranosyl-(1-2)-ß-D-glucopyranosyl]-dammar-12-one-20S,24R-epoxy-3ß,6α,25-triol. 12-one-PF11 significantly suppressed hydrogen peroxide induced oxidative stress in human lung carcinoma A549 cells. As compared with model group, 12-one-PF11 improved cell viability of A549 cells in a dose-dependent manner, and significantly decreased the generation of malondialdehyde (MDA) and increased production of superoxide dismutase (SOD) and glutathione (GSH) and protein expression levels of nuclear related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in A549 cells.

Molecular identification of Sanqi Tablets using DNA barcoding and its methodological evaluation / 中草药

Objective: In order to study the application of DNA barcoding in the authentication of Chinese patent medicines, Sanqi Tablets were used as the object to investigate the applicability, specificity and precision of this method. Methods: Fifteen batches of commercially available Sanqi Tablet samples were collected. The conditions of DNA extraction for Sanqi Tablet had been first investigated, and the DNA was used for testing the applicability of the methods such as PCR amplification, sequence acquisition, and species authentication in the principles for molecular identification of traditional Chinese materia medica using DNA barcoding. The specificity and reproducibility of DNA barcoding in identification of Sanqi Tablets and its adulterations from the roots of Panax notoginseng, P. ginseng and P. quinquefolius were also studied. Results: The Sanqi Tablet sample with an amount of sampling to be 100 mg and a water bath at 56 ℃ for 8 h gave an average concentration of 60.7 ng/μL and then the PCR amplification, sequence acquisition and species assignment were all successful. The ITS2 sequences of P. notoginseng, P. ginseng and P. quinquefolius were all 230 bp in length, and there were seven stable SNP loci between P. notoginseng and P. ginseng, P. notoginseng and P. quinquefolius. ITS2 sequences could be successfully obtained from lab-made and the adulterated Sanqi Tablets, and the Sanger sequencing chromatograms of different ratios of P. notoginseng and P. ginseng mixtures, P. notoginseng and P. quinquefolius mixtures had heterozygous peaks with corresponding peak height ratio at SNP positions. The repeatability, intermediate precision and reproducibility were all in line with the requirements of “General Regulation 9101” in the Chinese Pharmacopoeia. Conclusion: The ITS2 sequence can stably and accurately authenticate the raw materials of Sanqi Tablets with substantial specificity and precision. The DNA barcoding identification method of Sanqi Tablets will provide a new technical tool for ensuring the safety of Sanqi Tablets in clinical medications, and provide reference for the identification of other single-herb products documented in the Chinese Pharmacopoeia.

American Ginseng (Panax quinquefolium L.) as a Source of Bioactive Phytochemicals with Pro-Health Properties.

Panax quinquefolium L. (American Ginseng, AG) is an herb characteristic for regions of North America and Asia. Due to its beneficial properties it has been extensively investigated for decades. Nowadays, it is one of the most commonly applied medical herbs worldwide. Active compounds of AG are ginsenosides, saponins of the glycosides group that are abundant in roots, leaves, stem, and fruits of the plant. Ginsenosides are suggested to be primarily responsible for health-beneficial effects of AG. AG acts on the nervous system; it was reported to improve the cognitive function in a mouse model of Alzheimer's disease, display anxiolytic activity, and neuroprotective effects against neuronal damage resulting from ischemic stroke in animals, demonstrate anxiolytic activity, and induce neuroprotective effects against neuronal damage in ischemic stroke in animals. Administration of AG leads to inhibition of hypertrophy in heart failure by regulation of reactive oxygen species (ROS) in mice as well as depletion of cardiac contractile function in rats. It also has an anti-diabetic and anti-obesity potential as it increases insulin sensitivity and inhibits formation of adipose tissue. AG displays anti-cancer effect by induction of apoptosis of cancer cells and reducing local inflammation. It exerts antimicrobial effects against several pathogenic strains of bacteria. Therefore, AG presents a high potential to induce beneficial health effects in humans and should be further explored to formulate precise nutritional recommendations, as well as to assess its value in prevention and therapy of some disorders, including cancer.

Multicomponent assessment and ginsenoside conversions of Panax quinquefolium L. roots before and after steaming by HPLC-MSn.

BACKGROUND: The structural conversions in ginsenosides induced by steaming or heating or acidic condition could improve red ginseng bioactivities significantly. In this paper, the chemical transformations of red American ginseng from fresh Panax quinquefolium L. under steaming were investigated, and the possible mechanisms were discussed. METHODS: A method with reversed-phase high-performance liquid chromatography coupled with linear ion trap mass spectrometry (HPLC-MSn)-equipped electrospray ionization ion source was developed for structural analysis and quantitation of ginsenosides in dried and red American ginseng. RESULTS: In total, 59 ginsenosides of protopanaxadiol, protopanaxatriol, oleanane, and ocotillol types were identified in American ginseng before and after steaming process by matching the molecular weight and/or comparing MSn fragmentation with that of standards and/or known published compounds, and some of them were determined to be disappeared or newly generated under different steaming time and temperature. The specific fragments of each aglycone-type ginsenosides were determined as well as aglycone hydrated and dehydrated ones. The mechanisms were deduced as hydrolysis, hydration, dehydration, and isomerization of neutral and acidic ginsenosides. Furthermore, the relative peak areas of detected compounds were calculated based on peak areas ratio. CONCLUSION: The multicomponent assessment of American ginseng was conducted by HPLC-MSn. The result is expected to provide possibility for holistic evaluation of the processing procedures of red American ginseng and a scientific basis for the usage of American ginseng in prescription.

Seeds identification of Panax ginseng and Panax quinquefolium using ITS2 DNA barcordes / 中草药

Objective Traditional identification methods of pharmacognosy is difficult to distinguish the seeds of Panax ginseng and Panax quinquefolius. In order to improve the efficacy and accuracy of identification and provide the scientific foundation for the establishment of Chinese herbal medicine seed quality standards, molecular identification methods of the seeds were established by DNA barcoding technology. Methods The pharmacognostical identification method was used to study the morphological identification and microscopic characters of different seeds. DNA barcodes and Chinese Pharmacopoeia species standard barcode database were employed to identify the seeds by ITS2 sequence comparison, genetic distance comparison and systematic NJ tree construction. Results Intraspecific genetic distances of individuals participating were smaller than interspecific genetic distances. Phylogenetic tree map showed that two species were repectively clustered into one. A total of 42 samples of seeds from Panax ginseng and Panax quinquefolius produced by nine areas were all top-quality and easy to distinguish. Conclusion ITS2 DNA barcodes can identify and differentiate the seeds of P. ginseng and P. quinquefolius germplasm resources quickly, accurately and efficiently.

Quality ecotype of Panax quinquefolium L. based on heredity-chemistry-ecology characteristics / 药学学报

Medicinal materials in China differ in quality by different ecological types. Our research group found that there were two ecotypes of domestic Panax quinquefolium L. according to the characteristics of ginsenosides, inside versus outside Shanhaiguan. The genetic and ecological mechanisms of quality variation of Panax quinquefolium L. is unknown. Based on the genetic-chemical-ecological strategy, transcriptome and HPLC technology were used for comprehensive correlation analyses of transcriptomic data, ginsenoside content and environmental climate ecological factors. The transcriptomic results showed that key genes of ginsenoside biosynthesis, such as HMGR, AS and FPS, were significantly down-regulated in the inside Shanhaiguan ecotype. HPLC results showed that the quality of outside Shanhaiguan ecotype Panax quinquefolium L. was higher than that of the inside ecotype, with the content of ginsenosides in outside Panax quinquefolium L. was higher than that of inside ecotype except Rb2. Correlation analyses revealed that content of Panax quinquefolium L. ginsenoside is positively related to the expression levels of ginsenoside biosynthesis key genes (MK, HMGS, HMGR, and AS), and negatively related to the expression of glycosyl transferase (GT). The content of ginsenosides is negative related with climate factors, such as temperature, sunshine, and is positively related with moisture in both ecological environments. This study has provided a new mechanistic insight into the quality variations of two ecotypes for Panax quinquefolium L. and established a scientific basis for studying the ecological factors for the quality of traditional Chinese medicine.

Determination of ginsenoside Re, pseudo-ginsenoside F11, ginsenoside Rb3, and ginsenoside Rd from flower buds of Panax quinquefolium / 中草药

Objective To establish the determination method of four ginsenosides from flower buds of Panax quinquefolium by Waters Acquity UPLC H-Class with Xevo TQD. Methods The chromatographic separation was performed on a Waters Acquity UPLC BEH C18 column (50 mm × 2.1mm, 1.7 μm). The mobile phase was a mixture of acetonitrile and water containing 0.05% ammonium hydroxide with gradient elution; The flow rate was 0.45 mL/min, and the column temperature was 35 ℃; Multiple reaction monitoring (MRM) acquisition under negative ion scan mode by ESI was also performed. Results The method was established with good precision, stability, repeatability, and accuracy. Ginsenoside Re, pseudo-ginsenoside F11, ginsenoside Rb3, and ginsenoside Rd showed a good linearity in the range of corresponding concentrations. Conclusion The UPLC-MS/MS method is rapid, simple, accurate, reliable, which can be used for the analysis of ginsenosides from flower buds of P. quinquefolium.

Extraction of Eight Ginsenosides from Leaves of Panax quinquefolium L. By Ultrasoinc Fountain-based Ultrasonic-assisted Nebulization Extraction Coupled with Solid Phase Extraction / 分析化学

A new ultrasonic-assisted extraction (UANE) method coupled with solid phase extraction (SPE) using ultrasonic fountain was established for the extraction of eight common ginsenosides from leaves of Panax quinquefolium L. The extraction system has been designed and several experimental parameters,including the type and volume of extraction solvent,pH value and salt concentration of extraction solvent,type and volume of elution solvent,and amount of C18, extraction time were examined and optimized. Under the optimal conditions,the recoveries of ginsenosides were in the range of 96. 3% -110. 6%, the relative standard deviations (RSDs) were in the range of 2.8%-4.3%,indicating that the method has a good performance for the extraction of these ginsenosides. Compared with traditional UANE-SPE method, the modified method simplified the extraction device,shortened the extraction time and improved the extraction efficiency.

Tissue-specific metabolite profiling and quantitative analysis of ginsenosides in Panax quinquefolium using laser microdissection and liquid chromatography-quadrupole/time of flight-mass spectrometry.

BACKGROUND: The root of Panax quinquefolium L., famous as American ginseng all over the world, is one of the most widely-used medicinal or edible materials. Ginsenosides are recognized as the main bioactive chemical components responsible for various functions of American ginseng. In this study, tissue-specific chemicals of P. quinquefolium were analyzed by laser microdissection and ultra-high performance liquid chromatography- quadrupole/time-of-flight-mass spectrometry (UHPLC-Q/TOF-MS) to elucidate the distribution pattern of ginsenosides in tissues. The contents of ginsenosides in various tissues were also compared. RESULTS: A total of 34 peaks were identified or temporarily identified in the chromatograms of tissue extractions. The cork, primary xylem or cortex contained higher contents of ginsenosides than phloem, secondary xylem and cambium. Thus, it would be reasonable to deduce that the ratio of total areas of cork, primary xylem and the cortex to the area of the whole transection could help to judge the quality of American ginseng by microscopic characteristics. CONCLUSION: This study sheds new light on the role of microscopic research in quality evaluation, and provides useful information for probing the biochemical pathways of ginsenosides. Graphical abstract.