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.

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.

[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.

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.

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.

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.

(±)-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.

Rapid Identification of Wild Gentiana Genus in Different Geographical Locations Based on FT-IR and an Improved Neural Network Structure Double-Net.

Gentiana Genus, a herb mainly distributed in Asia and Europe, has been used to treat the damp heat disease of the liver for over 2000 years in China. Previous studies have shown significant differences in the compositional contents of wild Gentiana Genus samples from different geographical origins. Therefore, the traceable geographic locations of the wild Gentiana Genus samples are essential to ensure practical medicinal value. Over the last few years, the developments in chemometrics have facilitated the analysis of the composition of medicinal herbs via spectroscopy. Notably, FT-IR spectroscopy is widely used because of its benefit of allowing rapid, nondestructive measurements. In this paper, we collected wild Gentiana Genus samples from seven different provinces (222 samples in total). Twenty-one different FT-IR spectral pre-processing methods that were used in our experiments. Meanwhile, we also designed a neural network, Double-Net, to predict the geographical locations of wild Gentiana Genus plants via FT-IR spectroscopy. The experiments showed that the accuracy of the neural network structure Double-Net we designed can reach 100%, and the F1_score can reach 1.0.

The chromosome-level genome assembly of Gentiana dahurica (Gentianaceae) provides insights into gentiopicroside biosynthesis.

Gentiana dahurica Fisch. is a perennial herb of the family Gentianaceae. This species is used as a traditional Tibetan medicine because of its rich gentiopicroside constituents. Here, we generate a high-quality, chromosome-level genome of G. dahurica with a total length of 1,416.54 Mb. Comparative genomic analyses showed that G. dahurica shared one whole-genome duplication (WGD) event with Gelsemium sempervirens of the family Gelsemiaceaei and had one additional species-specific WGD after the ancient whole-genome triplication with other eudicots. Further transcriptome analyses identified numerous enzyme coding genes and the transcription factors related to gentiopicroside biosynthesis. A set of candidate cytochrome P450 genes were identified for being involved in biosynthetic shifts from swertiamarin to gentiopicroside. Both gene expressions and the contents measured by high-performance liquid chromatography indicated that the gentiopicrosides were mainly synthesized in the rhizomes with the highest contents. In addition, we found that two above-mentioned WGDs, contributed greatly to the identified candidate genes involving in gentiopicroside biosynthesis. The first reference genome of Gentianaceae we generated here will definitely accelerate evolutionary, ecological, and pharmaceutical studies of this family.

Quantitative analysis of the profiles of twelve major compounds in Gentiana straminea Maxim. Roots by LC-MS/MS in an extensive germplasm survey in the Qinghai-Tibetan plateau.

ETHNOBOTANICAL RELEVANCE: Gentiana straminea Maxim. is a well-known Tibetan traditional herb, which has been used to treat rheumatic arthritis, iceteric hepatitis, and other diseases for thousands years. However, there is still lack of comprehensive active constituents profiling of this species throughout the Qinghai-Tibet Plateau (QTP). AIM OF STUDY: This study was designed to provide a comprehensive quality map of G.straminea germplasm based on twelve active constituents (loganic acid, gentiopicroside, swertiamarin, sweroside, 6-O-ß-D-glucosylgentiopicroside, oleanic acid, morroniside, trilobatin, isoorientin, isovite, Shanzhisidemethylester and quercetin) on the QTP. MATERIALS AND METHODS: G.straminea root samples collected throughout QTP in the flowering period were analyzed by the LC-MS/MS. Statistics analysis methods PCA, clustering and ecological regions analysis for G.straminea constituents differentiation was demonstrated. RESULTS: The active constituents varied greatly across the QTP; the majority of constituents were secoiridoid derivatives, with gentiopicroside being the most abundant compound. Most constituents were significantly affected by the latitudes and altitudes other than longitudes. PCA and hierarchical clustering analysis showed that all samples could be separated into six distinct groups, and 15 populations showed the highest constituent abundances. Further, geographical region analysis showed that the highest quality populations mainly located near the source region of Yellow River, especially in the Qinghai and Sichuan areas. Additionally, correlation analysis showed that there were relationships among genetiopicroside, loganic acid, and other compounds, which might be related to the enzymatic pathways involved in the metabolism of these constituents. CONCLUSION: LC-MS/MS method allowed separation of quality profiling of G.straminea on the QTP, 15 populations showed the highest constituent abundances. In six geographical groups, the highest quality populations mainly located near the source region of Yellow River, especially in the Qinghai and Sichuan areas, which may be due to the climate caused by the westerlies and Indian Ocean monsoons in the QTP.

In-vitro cytotoxicity in relation to chemotypic diversity in diploid and tetraploid populations of Gentiana kurroo Royle.

ETHNOPHARMACOLOGICAL RELEVANCE: Gentiana kurroo is a multipurpose critically endangered medicinal herb prescribed as medicine in Ayurveda in India and exhibits various pharmacological properties including anti-cancer activity. The species is rich repository of pharmacologically active secondary metabolites together with secoiridoidal glycosides. AIM OF THE STUDY: The study aimed to investigate the chemical diversity in different populations/cytotypes prevailing in G. kurroo to identify elite genetic stocks in terms of optimum accumulation/biosynthesis of desired metabolites and having higher in-vitro cytotoxicity potential in relation to chemotypic diversity. MATERIAL AND METHODS: The wild plants of the species were collected from different ranges of altitudes from the Kashmir Himalayas. For cytological evaluation, the standard meiotic analysis was performed. The standard LC-MS/MS technique was employed for phytochemical analysis based on different marker compounds viz. sweroside, swertiamarin, and gentiopicroside. Different tissues such as root-stock, aerial parts, and flowers were used for chemo-profiling. Further, the methanolic extracts of diploid and tetraploid cytotypes were assessed for cytotoxic activity by using MTT assay against four different human cancer cell lines. RESULTS: The quantification of major bioactive compounds based on tissue- and location-specific comparison, as well as in-vitro cytotoxic potential among extant cytotypes, was evaluated. The comprehensive cytomorphological studies of the populations from NW Himalayas revealed the occurrence of different chromosomal races viz. n = 13, 26. The tetraploid cytotype was hitherto unreported. The tissue-specific chemo-profiling revealed relative dominance of different phytoconstituents in root-stock. There was a noticeable increase in the quantity of the analyzed compounds in relation to increasing ploidy status along the increasing altitudes. The MTT assay of methanolic extracts of diploid and tetraploid cytotypes displayed significant cytotoxicity potential in tetraploids. The root-stock extracts of tetraploids were highly active extracts with IC50 value ranges from 5.65 to 8.53 µg/mL against HCT-116 colon cancer. CONCLUSION: The chemical evaluation of major bioactive compounds in diverse cytotypes from different plant parts along different altitudes presented an appreciable variability in sweroside, swertiamarin, and gentiopicroside contents. Additionally, the concentrations of these phytoconstituents varied for cytotoxicity potential among different screened cytotypes. This quantitative difference of active bio-constituents was in correspondence with the growth inhibition percentage of different tested cancer cell lines. Thus, the present investigation strongly alludes towards a prognostic approach for the identification of elite cytotypes/chemotypes with significant pharmacological potential.

A new and effective evaluation method for Radix Gentianae Macrophyllae herbs based on 2-phenylethyl ß-d-glucopyranoside, 2-methoxyanofinic acid and gentioxepine by UPLC-QTOF-MS exploring and HPLC-DAD quantification.

Radix Gentianae Macrophyllae (RGM) is a traditional Chinese medicine belonging to the Gentiana genus and including four species of herbs. Owing to the lack of characteristic constituents, it is difficult to discriminate RGMs of different origins or even differentiate between herbs in the same genus. The current research aimed to explore the characteristic aromatic compounds, verify their significance in distinguishing different origins of RGM herbs and provide a simple and effective quality evaluation method. A selective extraction method was developed for noniridoid compounds and the extract was then subjected to UPLC-QTOF-MS analysis to obtain the RGM-MS ion pair database for noniridoid components. An HPLC-DAD quantitative analysis method was further developed based on characteristic aromatic compounds (2-phenylethyl ß-d-glucopyranoside, 2-methoxyanofinic acid and gentioxepine) after the ion screening in the MS database. By means of principal component analysis and hierarchical clustering analysis analysis, the significant relationship between aromatic compounds contents and different species of RGM was revealed. As a result, the significance of 2-phenylethyl ß-d-glucopyranoside, 2-methoxyanofinic acid and gentioxepine in distinguishing four species of RGM herbs was verified and a sensitive and reproducible HPLC-DAD method was established using these markers, which can be used for the classification and quantitative analysis of RGMs.

Osteoprotective Effects of Loganic Acid on Osteoblastic and Osteoclastic Cells and Osteoporosis-Induced Mice.

Osteoporosis is a common disease caused by an imbalance of processes between bone resorption by osteoclasts and bone formation by osteoblasts in postmenopausal women. The roots of Gentiana lutea L. (GL) are reported to have beneficial effects on various human diseases related to liver functions and gastrointestinal motility, as well as on arthritis. Here, we fractionated and isolated bioactive constituent(s) responsible for anti-osteoporotic effects of GL root extract. A single phytochemical compound, loganic acid, was identified as a candidate osteoprotective agent. Its anti-osteoporotic effects were examined in vitro and in vivo. Treatment with loganic acid significantly increased osteoblastic differentiation in preosteoblast MC3T3-E1 cells by promoting alkaline phosphatase activity and increasing mRNA expression levels of bone metabolic markers such as Alpl, Bglap, and Sp7. However, loganic acid inhibited osteoclast differentiation of primary-cultured monocytes derived from mouse bone marrow. For in vivo experiments, the effect of loganic acid on ovariectomized (OVX) mice was examined for 12 weeks. Loganic acid prevented OVX-induced bone mineral density loss and improved bone structural properties in osteoporotic model mice. These results suggest that loganic acid may be a potential therapeutic candidate for treatment of osteoporosis.

New insight into the antigenotoxic activity of Gentiana lutea extracts - Protective effect against food borne mutagens.

Food mutagens formed from amino acids during heating of meat have the potential to induce serious consequences on human health. As a result, the identification of naturally occurring, genoprotective agents, is of great importance. The aim of this study was to chemically characterize a root and leaf extracts of Gentiana lutea and to investigate the antigenotoxic effects of extracts and pure constituents (gentiopicroside and mangiferin). Antigenotoxic effects were shown for combinations with the food borne mutagens IQ and PhIP using hepatoma HepG2 cells. Furthermore, their antioxidant activity and their capacity to modulate Nrf2 expression and affect the glutathione redox status were tested. Chemical analyses showed that the most abundant constituents found in root extract are gentiopicroside and sweroside. On the other hand, homoorientin and isovitexin were the dominant ones in leaf extract. Strong genoprotective activities of all tested compounds against both mutagens were observed in alkaline comet assays (up to 77% of tail intensity inhibition, p < 0.001). The protection against glutathione depletion was partially due to the radical scavenging activity and up-regulation of Nrf2 expression by the substances. The results of this study strongly encourage further investigations of the antimutagenic properties of G. lutea.

Identification of Antifungal Bisphosphocholines from Medicinal Gentiana Species.

Gentiana species including G. crassicaulis, G. macrophylla, G. dahurica, and G. straminea are used in traditional Chinese medicine as "Qinjiao" for the treatment of rheumatism, hepatitis, and pain. Four antifungal bisphosphocholines [irlbacholine (2) and three new analogues, gentianalines A-C (1, 3, and 4)] were identified from G. crassicaulis by a bioassay-guided fractionation and structure elucidation approach. Subsequent chemical analysis of 56 "Qinjiao" samples (45 from G. crassicaulis, five from G. macrophylla, three from G. dahurica, and three from G. straminea) showed that bisphosphocholines were present in all four Gentiana species, with irlbacholine as the major compound ranging from 2.0 to 6.2 mg per gram of dried material. Irlbacholine exhibited potent in vitro antifungal activity against Cryptococcus neoformans, Aspergillus fumigatus, Candida albicans, and Candida glabrata with minimum inhibitory concentration (MIC) values of 0.63, 1.25, 10.0, and 5.0 µg/mL, respectively. Identification of the bisphosphocholines, a rare class of antifungal natural products, in these medicinal plants provides scientific evidence to complement their medicinal use. The bisphosphocholines carrying a long aliphatic chain possess amphiphilic molecule-like properties with a tendency of retention in both normal and reversed-phase silica gel column chromatography and thereby may be neglected in natural products discovery. This report may stimulate interest in this class of compounds, which warrant the further study of other biological activities as well.

Amarogentin from Gentiana rigescens Franch Exhibits Antiaging and Neuroprotective Effects through Antioxidative Stress.

In the present study, the replicative lifespan assay of yeast was used to guide the isolation of antiaging substance from Gentiana rigescens Franch, a traditional Chinese medicine. A compound with antiaging effect was isolated, and the chemical structure of this molecule as amarogentin was identified by spectral analysis and compared with the reported data. It significantly extended the replicative lifespan of K6001 yeast at doses of 1, 3, and 10 µM. Furthermore, amarogentin improved the survival rate of yeast under oxidative stress by increasing the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx), and these enzymes' gene expression. In addition, this compound did not extend the replicative lifespan of sod1, sod2, uth1, and skn7 mutants with K6001 background. These results suggested that amarogentin exhibited antiaging effect on yeast via increase of SOD2, CAT, GPx gene expression, enzyme activity, and antioxidative stress. Moreover, we evaluated antioxidant activity of this natural products using PC12 cell system, a useful model for studying the nervous system at the cellular level. Amarogentin significantly improved the survival rate of PC12 cells under H2O2-induced oxidative stress and increased the activities of SOD and SOD2, and gene expression of SOD2, CAT, GPx, Nrf2, and Bcl-x1. Meanwhile, the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) of PC12 cells were significantly reduced after treatment of the amarogentin. These results indicated that antioxidative stress play an important role for antiaging and neuroprotection of amarogentin. Interestingly, amarogentin exhibited neuritogenic activity in PC12 cells. Therefore, the natural products, amarogentin from G. rigescens with antioxidant activity could be a good candidate molecule to develop drug for treating neurodegenerative diseases.