Structure characterization and protective effect against UVB irradiation of polysaccharides isolated from the plateau plant Gentiana dahurica Fisch.

Gentiana dahurica Fisch. (G. dahurica) is one of the legitimate sources of Qinjiao in Traditional Chinese Medicine (TCM) and grows on high-altitude plateaus. Plants develop unique biochemical accumulations to resist plateau conditions, especially the strong UV irradiation. Thus, this study aimed to investigate the polysaccharide of G. dahurica (GDP), its structure and its activity against UVB irradiation. Four GDPs were isolated and two of them were subjected to structural elucidation. The results suggested that GDP-1 has 53.5 % Ara and 30.8 % GalA as its main monosaccharides, with a molecular weight (Mw) of 23 kDa; the GDP-2 has 33.9 % Ara and 48.5 % GalA, with a Mw of 82 kDa. Methylation and NMR spectroscopy analysis revealed that GDP-1 contains →5)-α-Araf-(1 â†’ 5)-α-Araf-(1 â†’ 3,5)-α-Araf-(1 â†’ 3,4)-α-GalpA-(6-OMe)-(1→ as the main chain, the branches of GalA (with esterification), and the terminal Ara; the GDP-2 contains →4)-α-GalpA-(1 â†’ 4)-α-GalpA-(6-OMe)-(1 â†’ 5)-α-Araf-(1 â†’ 3,5)-α-Araf-(1→ as the main chain, the branches of →5)-α-Araf-(1-5)-α-Araf, and the terminal GalA. Both GDP-1 and GDP-2 exhibited concentration-dependent antioxidant activity against DPPH, ABTS and hydroxyl radicals. Moreover, GDPs significantly attenuated the decreases in viability and proliferation of HaCaT cells after UVB irradiation. They can scavenge reactive oxygen species (ROS) and improve the activities of endogenous antioxidant enzymes, including superoxide dismutase (SOD) and glutathione peroxidase (GSH). The potential mechanism explored by flow cytometry assays of cell apoptosis and cell cycle distribution suggested that GDPs exert protective effects against UVB irradiation by reducing ROS and attenuating S phase cell arrest. In brief, the GDP-1 and GDP-2 are α-1,3- and α-1,4- arabinogalacturonan, respectively. The high content of Ara could be attributed to biochemical accumulation in resisting to the plateau environment and to prevent UVB irradiation-related damage in cells. These findings provide insight into authentic medicinal herbs and the development of GDPs in the modern pharmaceutical and cosmetics industry.

Resultant compound from sublimation test for Gentianae Radix in Japanese Pharmacopoeia was 5-(hydroxymethyl)furfural.

Gentianae Radix, an herbal medicine, has been used as a gastrointestinal drug in Japan. In the Japanese Pharmacopoeia 18th Revision, the sublimation test is specified as an identification test for Gentianae Radix. The compound obtained in this sublimation test was believed to be gentisin, a xanthone family compound. However, the compound we identified using liquid chromatography-high-resolution mass spectrometry (LC-HRMS) and 1H- and 13C-NMR was 5-(hydroxymethyl)furfural (5-HMF). The same compound was found to be a sublimate of Gentianae Scabrae Radix and Gentianae Macrophyllae Radix, belonging to the same genus as Gentianae Radix. These results indicate the necessity to revise the identification test for Gentianae Radix to a more unique method.

[A new benzopyran glycoside from Gentiana macrophylla].

Thirteen compounds were isolated and identified from 70% ethanol extract of the roots of Gentiana macrophylla by multi-chromatographic methods, including microporous resin, silica gel, and C_(18) reversed-phase column chromatography, as well as HPLC as follows: macrophylloside G(1), macrophylloside D(2), 5-formyl-2,3-dihydroisocoumarin(3),(+)-medicarpin(4),(+)-syringaresinol(5), liquiritigenin(6),(3R)-sativanone(7),(3R)-3'-O-methylviolanone(8), 4,2',4'-trihydroxychalcone(9), latifolin(10), gentioxepine(11), 6α-hydroxycyclonerolidol(12), and ethyl linoleate(13). Compound 1 was a new benzopyran glycoside. Compounds 4, 6-10, 12, and 13 were isolated for the first time from Gentiana plants. Compounds 1 and 2 showed promising hepatoprotective activity against D-GalN-induced AML12 cell damage at the concentration of 10 µmol·L~(-1), and compound 2 exhibited more significant activity than silybin at the same concentration.

Hydroethanolic extract of Gentiana kurroo Royle rhizome ameliorates ethanol-induced liver injury by reducing oxidative stress, inflammation and fibrogenesis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Gentiana kurroo Royle is a medicinal plant mentioned as Traymana in Ayurveda. In the folklore, it is used to cure fever, stomach ache, skin diseases and liver disorders. However, limited reports are available on the therapeutic potential of Gentiana kurroo Royle against alcohol-induced liver damage. AIM OF THE STUDY: To assess the effectiveness of the hydroethanolic extract of Gentiana kurroo Royle rhizome (GKRE) against alcohol-induced liver injury and explore the mechanism of action. MATERIALS AND METHODS: GKRE was characterized using UHPLC-QTOF-MS/MS. The binding affinity of the identified compound was studied in silico. In vitro studies were performed in the Huh-7 cell line. An acute oral toxicity study (2 g/kg BW) of GKRE was done in rats following OECD 420 guidelines. In the efficacy study, rats were treated with 50% ethanol (5 mL/kg BW, orally) for 4 weeks, followed by a single intraperitoneal dose of CCl4 (30%; 1 mL/kg BW) to induce liver injury. After 4th week, the rats were treated with GKRE at 100, 200 and 400 mg/kg BW doses for the next fifteen days. The biochemical and antioxidant parameters were analyzed using commercial kits and a biochemistry analyzer. Histopathology, gene and protein expressions were studied using qRT PCR and western blotting. RESULTS: Thirteen compounds were detected in GKRE. Few compounds showed a strong interaction with the fibrotic and inflammatory proteins in silico. GKRE reduced (p < 0.05) the ethanol-induced ROS production and inflammation in Huh-7 cells. The acute oral toxicity study revealed no adverse effect of GKRE in rats at 2 g/kg BW. GKRE improved (p < 0.05) the body and liver weights in ethanol-treated rats. GKRE improved (p < 0.05) the mRNA levels of ADH, SREBP1c and mitochondrial biogenesis genes in the liver tissues. GKRE also improved (p < 0.05) the liver damage markers, lipid peroxidation and levels of antioxidant enzymes in the liver. A reduced severity (p < 0.05) of pathological changes, fibrotic tissue deposition and caspase 3/7 activity were observed in the liver tissues of GKRE-treated rats. Further, GKRE downregulated (p < 0.05) the expression of fibrotic (TGFß, αSMA and SMADs) and inflammatory markers (TNFα, IL6, IL1ß and NFκB) in the liver. CONCLUSION: GKRE showed efficacy against alcohol-induced liver damage by inhibiting oxidative stress, apoptosis, inflammation and fibrogenesis in the liver.

Comparative analysis of phytochemical profile and antioxidant and anti-inflammatory activity of four Gentiana species from the Qinghai-Tibet Plateau.

ETHNOPHARMACOLOGICAL RELEVANCE: Gentiana species, known as the traditional Tibetan medicine "Bangjian," have been integral to clinical practice for millennia. Despite their longstanding use, our understanding of the variation in chemical constituents and bioactive effects among different species is limited. AIM OF THE STUDY: In the present study, we aimed to assess the differences in chemical profiles and bioactivities among four Gentiana species (G. veitchiorum, G. trichotoma, G. crassuloides, and G. squarrosa) and explore potential bioactive markers. MATERIALS AND METHODS: The chemical composition of the four Gentiana species was analyzed using UPLC-QE-Orbitrap-MS. The antioxidant activity of the extracts was compared through DPPH, ABTS, and reducing power assays. The anti-inflammatory activity was evaluated by measuring the inhibitory effects on lipopolysaccharide-induced secretion of nitric oxide (NO), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α) by RAW264.7 macrophages. Additionally, compounds strongly correlated with anti-inflammatory and antioxidant activities were identified through spectrum-effect relationship analysis. RESULTS: A total of 50 compounds were identified across the four Gentiana species. In vitro antioxidant assays demonstrated DPPH and ABTS scavenging abilities and reducing power within the concentration range of 62.5-2000 µg/mL. All four species inhibited the production of NO, IL-6, and TNF-α in RAW264.7 cells. Spectrum-effect relationship analysis revealed that gentiascabraside A, gentiatibetine, tachioside, lutonarin, and isotachioside were associated with the highest antioxidant activity; and swertiamarin, tarennoside, eleganoside C, and alpigenoside were associated with the highest anti-inflammatory activity. CONCLUSIONS: This study presents, for the first time, the chemical profiles and bioactivities of G. trichotoma, G. crassuloides, and G. squarrosa, which were comprehensively compared with those of G. veitchiorum. The findings provide novel insights to understand the traditional use and/or expand the current use of Gentiana species. Additionally, this research highlights the potential of Gentiana species as natural sources of antioxidants and anti-inflammatory agents, suggesting promising applications in tea production or medicinal contexts in the near future.

ResD-Net: A model for rapid prediction of antioxidant activity in gentian root using FT-IR spectroscopy.

Gentian, an herb resource known for its antioxidant properties, has garnered significant attention. However, existing methods are time-consuming and destructive for assessing the antioxidant activity in gentian root samples. In this study, we propose a method for swiftly predicting the antioxidant activity of gentian root using FT-IR spectroscopy combined with chemometrics. We employed machine learning and deep learning models to establish the relationship between FT-IR spectra and DPPH free radical scavenging activity. The results of model fitting reveal that the deep learning model outperforms the machine learning model. The model's performance was enhanced by incorporating the Double-Net and residual connection strategy. The enhanced model, named ResD-Net, excels in feature extraction and also avoids gradient vanishing. The ResD-Net model achieves an R2 of 0.933, an RMSE of 0.02, and an RPD of 3.856. These results support the accuracy and applicability of this method for rapidly predicting antioxidant activity in gentian root samples.

Genomic characterization of WRKY transcription factors related to secoiridoid biosynthesis in Gentiana macrophylla.

Gentiana macrophylla is one of Chinese herbal medicines in which 4 kinds of iridoids or secoiridoids, such as loganic acid, sweroside, swertiamarin, and gentiopicroside, are identified as the dominant medicinal secondary metabolites. WRKY, as a large family of transcription factors (TFs), plays an important role in the synthesis of secondary metabolites in plants. Therefore, WRKY genes involved in the biosynthesis of secoiridoids in G. macrophylla were systematically studied. First, a comprehensive genome-wide analysis was performed, and 42 GmWRKY genes were identified, which were unevenly distributed in 12 chromosomes. Accordingly, gene structure, collinearity, sequence alignment, phylogenetic, conserved motif and promoter analyses were performed, and the GmWRKY proteins were divided into three subfamilies based on phylogenetic and multiple sequence alignment analyses. Moreover, the enzyme-encoding genes of the secoiridoid biosynthesis pathway and their promoters were then analysed, and the contents of the four secoiridoids were determined in different tissues. Accordingly, correlation analysis was performed using Pearson's correlation coefficient to construct WRKY gene-enzyme-encoding genes and WRKY gene-metabolite networks. Meanwhile, G. macrophylla seedlings were treated with methyl jasmonate (MeJA) to detect the dynamic change trend of GmWRKYs, biosynthetic genes, and medicinal ingredient accumulation. Thus, a total of 12 GmWRKYs were identified to be involved in the biosynthesis of secoiridoids, of which 8 (GmWRKY1, 6, 12, 17, 33, 34, 38 and 39) were found to regulate the synthesis of gentiopicroside, and 4 (GmWRKY7, 14, 26 and 41) were found to regulate the synthesis of loganic acid. Taken together, this study systematically identified WRKY transcription factors related to the biosynthesis of secoiridoids in G. macrophylla, which could be used as a cue for further investigation of WRKY gene functions in secondary metabolite accumulation.

Gentiana macrophylla flavonoids from Tibetan medicine decreases circ_0059665 to alleviate the progression of non-small cell lung cancer under hypoxia.

BACKGROUND: Gentiana macrophylla Pall. is a traditional Tibetan medicinal herb possessing antinociceptive and anti-inflammatory activities. Circular RNAs (circRNAs) have been identified to be involved in the tumorigenesis of non-small cell lung cancer (NSCLC). Here, this study focused on investigating the function and mechanism of Gentiana macrophylla flavonoids (GF) and circ_0059665 in NSCLC progression. METHODS: The contents of mRNA and protein were detected using qRT-PCR and western blotting analysis. Cell proliferative and invasive abilities were evaluated by cell counting kit-8, EdU, colony formation and transwell assays, respectively. M2 macrophage polarization was analyzed by flow cytometry. RESULTS: GF treatment suppressed NSCLC cell proliferation, invasion and M2 macrophage polarization under hypoxic conditions. Circ_0059665 was highly expressed in NSCLC tissues and cells. Its expression was increased under hypoxic conditions but was reduced following GF treatment. Furthermore, circ_0059665 overexpression reversed the anticancer effects of GF on NSCLC cells under hypoxic conditions. Mechanistically, circ_0059665 acted as a sponge for miR-512-5p to regulate NOVA2 expression. Hypoxia decreased miR-512-5p levels, and increased NOVA2 levels in NSCLC cells, while these tendencies were abolished after GF treatment. Circ_0059665 silencing inhibited NSCLC cell proliferation, invasion and M2 macrophage polarization in hypoxic environments, which were counteracted by NOVA2 overexpression. Moreover, NOVA2 upregulation reversed the suppressive effects of GF on NSCLC cells with hypoxia treatment. In addition, GF impeded NSCLC tumor growth in vivo via suppressing circ_0059665. CONCLUSION: GF treatment in hypoxic environments suppressed NSCLC cell proliferation, invasion and M2 macrophage polarization via the circ_0059665/miR-512-5p/NOVA2 axis.

Regulatory mechanisms of Gentiopicroside on human diseases: a brief review.

Gentiopicroside (GPS), a single compound isolated from Gentiana lutea L. and the crucial representative of secoiridoid constituent, has been permitted for centuries in traditional Chinese medicine. GPS and its metabolites have been increasingly used in the search for clinical management with therapeutic properties and fewer side effects. The objective of this review was to provide a comprehensive overview of the involvement of molecular pathways in the therapeutic effects of GPS on human diseases and chronic conditions. This study presents a meticulously conducted comprehensive search of the PubMed and Google Scholar databases (from 1983 to 2023), aimed at identifying articles relating to regulatory mechanisms of GPS on human diseases and the pharmacokinetics of GPS. The inclusion criteria were meticulously and precisely defined to encompass original research papers that explicitly focused on elucidating the regulatory mechanisms of GPS in various human diseases through in vitro and animal studies. Notably, these studies were mandated to integrate specific genetic markers or pathways as essential components of their research inquiries. The evaluated pharmacokinetic parameters included maximum plasma concentration (Cmax), time to reach maximum plasma concentration (Tmax), area under the curve (AUC), clearance, and plasma half-life (t1/2). Subsequently, through a rigorous screening process of titles and abstracts, studies conducted in vitro or on animals, as well as those reporting pharmacokinetic data related to drugs other than GPS or language barriers, were systematically excluded. Drawing from the data and studies pertaining to this review, we conducted a thorough and informative analysis of the pharmacological characteristics and biological functions of GPS. These encompassed a wide range of effects, including hepatoprotective, anti-inflammatory, antifibrotic, antioxidant, analgesic, antitumor, and immunomodulatory properties. The analysis provided a comprehensive and insightful understanding of GPS's pharmacological profile and its diverse activities. Enhancing theoretical and experimental methodologies could prove advantageous in expanding the clinical applications of GPS. This could involve optimizing the bioavailability and pharmacokinetics of GPS, uncovering additional biomarkers and potential biotransformation pathways, and investigating its combined effects with standard-of-care medications.

Lignan glucosides from Gentiana macrophylla with potential anti-arthritis and hepatoprotective activities.

Ten lignans, including six previously undescribed phenolic ester glycosyl lignans (1-6), were isolated from a well-known traditional Chinese medicine, Qin-Jiao, which is the dry root of Gentiana macrophylla Pall. (Gentianaceae). Their structures were determined by spectroscopic and chemical methods, especially 2D NMR techniques. Quantum chemical calculations of theoretical ECD spectra allowed the determination of their absolute configurations. Refer to its traditional applications for the treatment of rheumatic arthralgia and hepatopathy, these compounds were evaluated on a TNF-α induced MH7A human synoviocyte inflammation model and a D-GalN induced AML12 hepatocyte injury model. Compounds 1, 2, 5, and 6 significantly reduced the release of proinflammatory cytokine IL-1ß in MH7A cells at 15 µM and they also could strongly protect AML12 cells against D-GalN injury at 30 µM. Flow cytometry and Western blot analysis showed that compound 5 ameliorated D-GalN induced AML12 cell apoptosis by upregulating the expression of anti-apoptotic Bcl-2 protein and down-regulating the expression of pro-apoptotic Bax protein.

Gentiana Scabra Bge Extract (GSE) Protects Against Alcoholic Liver Disease by Regulating the TLR4/NF-κB Pathway in Mice.

BACKGROUND: Alcohol abuse leads to alcoholic liver disease (ALD), for which no effective treatment is yet known. Gentiana Scabra Bge is a traditional Chinese medicine; its extract has a significant liver protection effect, but its effects on the mechanism of improving alcohol-induced toxicity remain unclear. Therefore, this study used cell and mouse models to investigate how Gentiana Scabra Bge extract (GSE) might affect the TLT4/NF-κB inflammation pathway in ALD. METHODS: In mice, we induced the alcoholic liver injury model by applying alcohol and induced the inflammatory cell model by lipopolysaccharide (LPS)-induced macrophages. Using an enzyme-linked immunosorbent assay (ELISA) kit, aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and interleukin 1ß (IL-1ß), interleukin 6 (IL-6), and tumor necrosis factor-α (TNF-α) levels were measured in liver tissue; we also performed histological analysis of liver tissue sections to assess the hepatoprotective effect of GSE on alcohol. Using real-time fluorescence quantification, we determined the expression of toll-like receptor 4 (TLR4) and nuclear factor κB (NF-κB) mRNA levels; we used Western blotting to detect the expression of TLR4/NF-κB signaling pathway-related proteins. RESULTS: We demonstrate that GSE decreased AST and ALT activity, ameliorated liver dysfunction, decreased cytokine levels, and reduced LPS-induced cellular inflammation. In addition, GSE protected mouse liver cells from the inflammatory response by reducing alcohol-induced liver pathological damage and downregulating genes and proteins such as nuclear factors. CONCLUSIONS: GSE can attenuate liver injury in mice through the TLR4/NF-κB pathway by inhibiting the activation of nuclear factors.

Beauvericin Immunotoxicity Prevention by Gentiana lutea L. Flower In Vitro.

Beauvericin (BEA) is an emerging mycotoxin produced by some species of Fusarium genera that widely contaminates food and feed. Gentiana lutea is a protected medicinal plant known for its antioxidant and anti-inflammatory properties, which are attributed to its rich content of bioactive compounds. In order to evaluate the beneficial effects of G. lutea flower against BEA cytotoxicity, the aim of this study is to evaluate changes in protein expression after Jurkat cell exposure through a proteomics approach. To carry out the experiment, cells were exposed to intestinally digested G. lutea flower alone or in combination with the BEA standard (100 nM) over 7 days. Differentially expressed proteins were statistically evaluated (p < 0.05), revealing a total of 172 proteins with respect to the control in cells exposed to the BEA standard, 145 proteins for G. lutea alone, and 139 proteins when exposing the cells to the combined exposure. Bioinformatic analysis revealed processes implicated in mitochondria, ATP-related activity, and RNA binding. After careful analysis of differentially expressed proteins, it was evident that G. lutea attenuated, in most cases, the negative effects of BEA. Furthermore, it decreased the presence of major oncoproteins involved in the modulation of immune function.

Terpenoid Glucosides from Gentiana macrophylla That Attenuate TNF-α Induced Pulmonary Inflammation in A549 Cells.

Four previously undescribed terpenoid glucosides, including one sesquiterpenoid di-glucoside (1), two new iridoid glucosides (2, 3), and a new triterpenoid tri-glucoside (4), were isolated from a 70% ethanol extract of the root of Gentiana macrophylla (Gentianaceae), along with eight known terpenoids. Their structures were determined by spectroscopic techniques, including 1D, 2D NMR, and HRMS (ESI), as well as chemical methods. The absolute configuration of compound 1 was determined by quantum chemical calculation of its theoretical electronic circular dichroism (ECD) spectrum. The sugar moieties of all the new compounds were confirmed to be D-glucose by GC analysis after acid hydrolysis and acetylation. Anti-pulmonary inflammation activity of the iridoids were evaluated on a TNF-α induced inflammation model in A549 cells. Compound 2 could significantly alleviate the release of proinflammatory cytokines IL-1ß and IL-8 and increase the expression of anti-inflammatory cytokine IL-10.

Identification of Gentian-Related Species Based on Two-Dimensional Correlation Spectroscopy (2D-COS) Combined with Residual Neural Network (ResNet).

Gentian is a traditional Chinese herb with heat-clearing, damp-drying, inflammation-alleviating and digestion-promoting effects, which is widely used in clinical practice. However, there are many species of gentian. According to the pharmacopoeia, Gentiana manshurica Kitag, Gentiana scabra Bge, Gentiana triflora Pall and Gentianarigescens Franch are included. Therefore, accurately identifying the species of gentian is important in clinical use. In recent years, with the advantages of low cost, convenience, fast analysis and high sensitivity, infrared spectroscopy (IR) has been extensively used in herbal identification. Unlike one-dimensional spectroscopy, a two-dimensional correlation spectrum (2D-COS) can improve the resolution of the spectrum and better highlight the details that are difficult to detect. In addition, the residual neural network (ResNet) is an important breakthrough in convolutional neural networks (CNNs) for significant advantages related to image recognition. Herein, we propose a new method for identifying gentian-related species using 2D-COS combined with ResNet. A total of 173 gentian samples from seven different species are collected in this study. In order to eliminate a large amount of redundant information and improve the efficiency of machine learning, the extracted feature band method was used to optimize the model. Four feature bands were selected from the infrared spectrum, namely 3500-3000 cm-1, 3000-2750 cm-1, 1750-1100 cm-1 and 1100-400 cm-1, respectively. The one-dimensional spectral data were converted into synchronous 2D-COS images, asynchronous 2D-COS images, and integrative 2D-COS images using Matlab (R2022a). The identification strategy for these three 2D-COS images was based on ResNet, which analyzes 2D-COS images based on single feature bands and full bands as well as fused feature bands. According to the results, (1) compared with the other two 2D-COS images, synchronous 2D-COS images are more suitable for the ResNet model, and (2) after extracting a single feature band 1750-1100 cm-1 to optimize ResNet, the model has the best convergence performance, the accuracy of training, test and external validation is 1 and the loss value is only 0.155. In summary, 2D-COS combined with ResNet is an effective and accurate method to identify gentian-related species.

Quality by design-based capillary electrophoresis method development for the quantitative analysis of four iridoid compounds in Gentiana macrophylla Radix.

In this study, the capillary electrophoresis-photodiode array detector was employed for the analysis of four iridoid compounds in Gentiana macrophylla Radix (RGM), and the method was optimized based on the concept of analytical quality by design (AQbD). The peak areas relative standard deviation (n = 3) and resolution of the four analytes were selected as critical method attributes. Fractional factorial design test combined with Pareto analysis were employed to screen critical method parameters (buffer concentration, pH, sodium dodecyl sulfate [SDS] micelle concentration, temperature, and voltage). Subsequently, three main factors (buffer concentration, buffer pH, and SDS concentration) were selected by central composite design test for constructing the design space. The optimal separation conditions as follows: capillary column (50.2 cm × 50 µm, detection length 40 cm). Working background electrolyte consisted of 51 mmol/L borax solution (pH = 9.47) and 40 mmol/L SDS. The samples were injected by pressure (5 s at 0.5 psi) and the detection was performed at 254 nm. Applied voltage was 20 kV and column temperature was 23°C. The developed method is rapid and reliable for the quantitative analysis of four iridoid compounds in RGM, providing a reference for the application of AQbD concept in the analysis of natural products.

Comparison of plastid genomes and ITS of two sister species in Gentiana and a discussion on potential threats for the endangered species from hybridization.

BACKGROUND: Gentiana rigescens Franchet is an endangered medicinal herb from the family Gentianaceae with medicinal values. Gentiana cephalantha Franchet is a sister species to G. rigescens possessing similar morphology and wider distribution. To explore the phylogeny of the two species and reveal potential hybridization, we adopted next-generation sequencing technology to acquire their complete chloroplast genomes from sympatric and allopatric distributions, as along with Sanger sequencing to produce the nrDNA ITS sequences. RESULTS: The plastid genomes were highly similar between G. rigescens and G. cephalantha. The lengths of the genomes ranged from 146,795 to 147,001 bp in G. rigescens and from 146,856 to 147,016 bp in G. cephalantha. All genomes consisted of 116 genes, including 78 protein-coding genes, 30 tRNA genes, four rRNA genes and four pseudogenes. The total length of the ITS sequence was 626 bp, including six informative sites. Heterozygotes occurred intensively in individuals from sympatric distribution. Phylogenetic analysis was performed based on chloroplast genomes, coding sequences (CDS), hypervariable sequences (HVR), and nrDNA ITS. Analysis based on all the datasets showed that G. rigescens and G. cephalantha formed a monophyly. The two species were well separated in phylogenetic trees using ITS, except for potential hybrids, but were mixed based on plastid genomes. This study supports that G. rigescens and G. cephalantha are closely related, but independent species. However, hybridization was confirmed to occur frequently between G. rigescens and G. cephalantha in sympatric distribution owing to the lack of stable reproductive barriers. Asymmetric introgression, along with hybridization and backcrossing, may probably lead to genetic swamping and even extinction of G. rigescens. CONCLUSION: G. rigescens and G. cephalantha are recently diverged species which might not have undergone stable post-zygotic isolation. Though plastid genome shows obvious advantage in exploring phylogenetic relationships of some complicated genera, the intrinsic phylogeny was not revealed because of matrilineal inheritance here; nuclear genomes or regions are hence crucial for uncovering the truth. As an endangered species, G. rigescens faces serious threats from both natural hybridization and human activities; therefore, a balance between conservation and utilization of the species is extremely critical in formulating conservation strategies.

Comparative analysis of mitochondrial genomes of two alpine medicinal plants of Gentiana (Gentianaceae).

Gentiana crassicaulis and G. straminea are alpine plants of Gentiana with important medicinal value and complex genetic backgrounds. In this study, the mitochondrial genomes (mtDNAs) of these two species were sequenced. The mtDNAs of G. crassicaulis and G. straminea are 368,808 and 410,086 bp long, respectively, 52 and 49 unique genes are annotated in the two species, and the gene arrangement varies widely. Compared to G. crassicaulis, G. straminea loses three effective genes, namely atp6, trnG-GCC and trnV-GAC. As a pseudogene, the atp6 gene of G. straminea is incomplete, which is rare in higher plants. We detected 1696 and 1858 pairs of long repeats and 213 SSRs and 250 SSs in the mtDNAs of G. crassicaulis and G. straminea, respectively. There are 392 SNPs and 18 InDels between the two genomes, and syntenic sequence and structural variation analysis show low collinearity between the two genomes. Chloroplast DNA transferring to mtDNA is observed in both species, and 46,511 and 55,043 bp transferred segments containing three tRNA genes are identified, respectively. Comparative analysis of mtDNAs of G. crassicaulis, G. straminea and four species of Gentianales determined 18 core genes, and there is no specific gene in G. crassicaulis and G. straminea. The phylogenetic tree based on mtDNAs places Gentianaceae in a branch of Gentianales. This study is the first to analyze the mtDNAs of Gentianaceae, which could provide information for analysis of the structure of mtDNAs of higher plants and phylogenetic research of Gentianaceae and Gentianales.

(±)-Gentiovarisin A and gentiovarisin B, unusual secoiridoid dimer skeletons from gentiopicroside.

Gentiana scabra, a famous traditional Chinese medicine (TCM), has been documented in Chinese Pharmacopoeia for the treatment of hepatitis. Its index component gentiopicroside could not be detected in the decoction, which suggested that the quality control of the TCM with this ingredient needs attention. The transformed products were obtained from gentiopicroside, mimicking the traditional process of G. scabra. Further investigation of the heat-transformed products yielded two secoiridoid dimers, gentiovarisin A (1) and B (2), with an unprecedented 6/6/6/6/6-fused pentacyclic skeletons. Their structures were elucidated by extensive spectroscopic analyses and single-crystal X-ray diffraction analysis, and the absolute configurations of 1 were confirmed as (+)-1 and (-)-1 by ECD method. Plausible transformation pathways of the isolates were also proposed. Compounds 1 and 2 exhibited in vitro hepatoprotective activity similar to gentiopicroside, while (+)-1 displayed a more potent hepatoprotective activity than N-Acetyl-L-cysteine.

The healing bitterness of Gentiana lutea L., phytochemistry and biological activities: A systematic review.

Over many years, natural products have been a source of healing agents and have exhibited beneficial uses for treating human diseases. The Gentiana genus is the biggest genus in the Gentianaceae, with over 400 species distributed mainly in alpine zones of temperate countries around the world. Plants in the Gentiana genus have historically been used to treat a wide range of diseases. Still, only in the last years has particular attention been paid to the biological activities of Gentiana lutea Linn., also known as yellow Gentian or bitterwort. Several in vitro/vivo investigations and human interventional trials have demonstrated the promising activity of G. lutea extracts against oxidative stress, microbial infections, inflammation, obesity, atherosclerosis, etc.. A systematic approach was performed using Pubmed and Scopus databases to update G. lutea chemistry and activity. Specifically, this systematic review synthesized the major specialized bitter metabolites and the biological activity data obtained from different cell lines, animal models, and human interventional trials. This review aims to the exaltation of G. lutea as a source of bioactive compounds that can prevent and treat several human illnesses.

Phytochemical characterization and anti-inflammatory activity of a water extract of Gentiana purpurea roots.

ETHNOPHARMACOLOGICAL RELEVANCE: Gentiana purpurea was one of the most important medicinal plants in Norway during the 18th and 19th centuries, and the roots were used against different types of gastrointestinal and airway diseases. AIM OF THE STUDY: To explore the content of bioactive compounds in a water extract from the roots, a preparation commonly used in traditional medicine in Norway, to assess the anti-inflammatory potential, and furthermore to quantify the major bitter compounds in both roots and leaves. MATERIALS AND METHODS: G. purpurea roots were boiled in water, the water extract applied on a Diaion HP20 column and further fractionated with Sephadex LH20, reverse phase C18 and normal phase silica gel to obtain the low molecular compounds. 1D NMR, 2D NMR, and ESI-MS were used for structure elucidation. HPLC-DAD analysis was used for quantification. The inhibition of TNF-α secretion in ConA stimulated peripheral blood mononuclear cells (PBMCs) was investigated. RESULTS: Eleven compounds were isolated and identified from the hot water extract of G. purpurea roots. Gentiopicrin, amarogentin, erythrocentaurin and gentiogenal showed dose-dependent inhibition of TNF-α secretion. Gentiopicrin is the major secondary metabolite in the roots, while sweroside dominates in the leaves. CONCLUSIONS: The present work gives a comprehensive overview of the major low-molecular weight compounds in the water extracts of G. purpurea, including metabolites produced during the decoction process, and show new anti-inflammatory activities for the native bitter compounds as well as the metabolites produced during preparation of the crude drug.