Swertiamarin from Enicostemma littorale, counteracts PD associated neurotoxicity via enhancement α-synuclein suppressive genes and SKN-1/NRF-2 activation through MAPK pathway.

The elusive targets and the multifactorial etiology of Parkinson's disease (PD) have hampered the discovery of a potent drug for PD. Furthermore, the presently available medications provide only symptomatic relief and have failed to mitigate the pathogenesis associated with PD. Therefore, the current study was aimed to evaluate the prospective of swertiamarin (SW), a secoiridoid glycoside isolated from a traditional medicinal plant, Enicostemma littorale Blume to ameliorate the characteristic features of PD in Caenorhabditis elegans. SW (25 µM) administration decreased the α-synuclein (α-syn) deposition, inhibited apoptosis and increased dopamine level mediated through upregulating the expression of genes linked to ceramide synthesis, mitochondrial morphology and function regulation, fatty acid desaturase genes along with stress responsive MAPK (mitogen-activated protein kinase) pathway genes. The neuroprotective effect of SW was evident from the robust reduction of 6-hydroxydopamine (6-OHDA) induced dopaminergic neurodegeneration independent of dopamine transporter (dat-1). SW mediated translational regulation of MAPK pathway genes was observed through increase expression of SKN-1 and GST-4. Further, in-silico molecular docking analysis of SW with C. elegans MEK-1 showed a promising binding affinity affirming the in-vivo results. Overall, these novel finding supports that SW is a possible lead for drug development against the multi- factorial PD pathologies.

Competitive immunochromatographic test strips for the rapid semi-quantitative analysis of the biologically active bitter glycoside, amarogentin.

Amarogentin (AG), a biologically active secoiridoid glycoside, is responsible for the efficacy of Gentianaceae based medications. Thus, qualitative and quantitative analyses of AG are of significance for batch to batch quality control purposes. By conjugating colloidal gold nanoparticles with the AG-specific monoclonal antibody, MAb 1E9, we were able to develop a single-step competitive immunochromatographic assay (ICA) for simple quantification of the AG content in plant samples. With a limit of detection of 250 ng/mL, the analytical results were obtained after immersing the ICA test strip in the detection mixture for 15 min. This new ICA is superior to conventional ICAs as it is considerably faster due to the speed with which the test strips can be produced and the omission of the time-consuming preparation phase that was previously required to make the fiber pad. Moreover, our ICA only needs a small amount of analyte (20 µL).The reliability of the reported test strip was confirmed by comparing its semi-quantitative results with those obtained via an indirect competitive enzyme-linked immunosorbent assay (icELISA). The positive correlation between these methods (R2 = 0.984) indicated that this new ICA could be applied for the semi-quantitative analysis of the AG content in plant samples.

Molecular docking, antiproliferative and anticonvulsant activities of swertiamarin isolated from Enicostemma axillare.

Present study aimed for molecular docking, antiproliferative and anticonvulsant activities of swertiamarin isolated from the successive methanol extract of Enicostemma axillare. Molecular docking of swertiamarin on telomerase targets (PDB ID: 5UGW, 3DU6 and 4ERD), followed by antiproliferative activity on HEp2 and HT-29 cells by MTT and SRB assays. Also tested for anticonvulsant activity by pentylenetetrazole (PTZ, 80 mg/kg bw) induced convulsant. Molecular docking study predicted good total score of the swertiamarin with the selected targets. Swertiamarin possesses antiproliferative activity on HEp-2 and HT-29 cells with lower CTC50 values. It also served as significant anticonvulsant agent with prolonged onset and reduced duration of the seizures. These results confirm that swertiamarin exhibited potential antiproliferative and anticonvulsant activities.

Effects of Anthocleista djalonensis root extracts on reproductive hormones and testicular marker enzymes in adult male Wistar rats.

This study evaluated the effect of methanol and aqueous ethanol root extracts (200 mg/kg body weight) of Anthocleista djalonensis on sex hormone concentrations and testicular marker enzymes of adult rats after 60 days of administration followed by 60 days of treatment withdrawal. The results showed no significant changes in testosterone and follicle-stimulating hormone (FSH) levels during the 60 days of extract treatment. Interestingly, 60 days after treatment withdrawal, there was an increase in intratesticular and serum testosterone and serum FSH in the methanol but not aqueous ethanol extract post-treatment groups. Intratesticular 3ß-hydroxysteroid dehydrogenase (3ß-HSD) activity remained unaffected while that of 17ß-HSD increased slightly during treatment of both extracts and the increase reached a statistical significance level (p < .05) during post-treatment. Gamma-glutamyltranspeptidase activity in the testis of the methanol but not aqueous ethanol extract-treated animals remained high during post-treatment compared to initial treatment values. Phytochemical analysis of the plant extracts showed that phenol and flavonoid constituents were higher in the methanol than the aqueous ethanol extract and has higher antioxidant activity. Altogether, post-treatment effect of the extract on the testis was more effective than treatment-related effect and the methanol extract appears to have better and consistent effects on the investigated parameters probably due to higher antioxidant activity conferred to it by its phenolic and flavonoid contents.

Anti-inflammatory lupane triterpenoids from Menyanthes trifoliata.

A new lupane triterpenoid, 23-O-trans-feruloylcylicodiscic acid (1), as well as four known analogues (2‒5), was isolated from the EtOAc fraction of Menyanthes trefoliata. The structure of compound 1 was elucidated on the basis of extensive spectroscopic analysis, including 2D NMR data. The structures of the known compounds were established by comparison of their spectroscopic data with that in the literature. Compounds 1, 2, and 4 exhibited certain anti-NO production activities.

Green synthesis and characterization of silver nanoparticles using Enicostemma axillare (Lam.) leaf extract.

In the present article, the facile green synthesis of silver nanoparticles (AgNPs) using aqueous leaf extract of Enicostemma axillare (Lam.) has reported. This is a simple, cost-effective, stable for a long time and reproducible aqueous synthesis method to obtain a self-assembly Ag nanoparticles. The size and shape of Ag nanoparticles were characterized by XRD, TEM, and SEM-EDS. The formation and stability of the reduced silver nanoparticles in the colloidal solution were monitored by UV-Vis spectrophotometer analysis. Zeta potential was confirmed by DLS study. The mean particle diameter of silver nanoparticles was calculated from the TEM, SEM and the size of the particles was measured between 15 and 20 nm. TEM analysis revealed the spherical shape of the particles. Crystalline nature of the nanoparticles in the face-centred cubic structure are confirmed by the peaks in the XRD pattern corresponding to (111), (200), (220) and (311) planes. This study showed the biogenic, environmentally friendly and cost-effective synthesis and characterization of the silver nanoparticles.

[A new naphthaldehyde derivative from Comastoma pulmonarium and its anti-tobacco mosaic virus (anti-TMV) activity].

A new naphthaldehyde derivative has been isolated from Comastoma pulmonarium by using various chromatographic techniques, including silica gel, Sephadex LH-20, MCI-gel resin and RP-HPLC. This compounds was determined as 5-methoxy-2-methyl-7-(2-oxopropyl)naphthalene-1-carbaldehyde(1) by NMR, MS, IR and UV spectra. This compound was also evaluated for its anti-tobacco mosaic virus (anti-TMV) activity. The result showed that it showed high anti-TMV activity with inhibition rate of 32.8%. The inhibition rate is close to that of positive control (ningnanmycin).

Interaction effects on cytochrome P450 both in vitro and in vivo studies by two major bioactive xanthones from Halenia elliptica D. Don.

The major components, 1-hydroxy-2,3,5-trimethoxy-xanthone (HM-1) and 1,5-dihydroxy-2,3-dimethoxy-xanthone (HM-5) isolated from Halenia elliptica D. Don (Gentianaceae), could cause vasodilatation in rat coronary artery with different mechanisms. In this work, high-performance liquid chromatography coupled to ion trap time-of-flight mass spectrometry (LCMS-IT-TOF) was used to clarify the metabolic pathways, and CYP450 isoform involvement of HM-1 and HM-5 were also studied in rat. At the same time, in vivo inhibition effects of HM-1 and ethyl acetate extracts from origin herb were studied. Three metabolites of HM-5 were found in rat liver microsomes (RLMs); demethylation and hydroxylation were the major phase I metabolic reactions for HM-5. Multiple CYP450s were involved in metabolism of HM-1 and HM-5. The inhibition study showed that HM-5 inhibited Cyp1a2, 2c6 and 2d2 in RLMs. HM-1 inhibited activities of Cyp1a2, Cyp2c6 and Cyp3a2. In vivo experiment demonstrated that both HM-1 and ethyl acetate extracts could inhibit Cyp3a2 in rats. In conclusion, the metabolism of xanthones from the origin herb involved multiple CYP450 isoforms; in vitro, metabolism of HM-5 was similar to that of its parent drug HM-1, but their inhibition effects upon CYP450s were different; in vivo, Cyp3a2 could be inhibited by HM-1 and ethyl acetate extracts.

Chemotaxonomic Studies of Nine Gentianaceae Species from Western China Based on Liquid Chromatography Tandem Mass Spectrometry and Fourier Transform Infrared Spectroscopy.

INTRODUCTION: Gentianaceae species which widely occur all over the world are used as folk medicine and raw food material with bitter properties. Although comparative analysis on metabolites in several Gentianaceae species has been reported, metabolic similarities used for chemotaxonomic studies are not yet clear. OBJECTIVE: To systematically characterise the variations of holistic metabolome and characteristic metabolites (iridoid glycosides and phenols) in nine Gentianaceae species from western China. METHODOLOGY: Fourier transform infrared (FT-IR) spectroscopy was applied to determine the variations of holistic metabolome. A targeted metabolic profiling using liquid chromatography with tandem mass spectrometry (LC-MS/MS) was established for determination of seven characteristic metabolites and identification of their derivatives. Both FT-IR and LC-MS/MS data were subjected to chemometrics analysis for exploring variations in iridoid glycosides and phenols within these species. RESULTS: Holistic metabolome in genera Gentiana and Swertia was largely different. Diversity of the biosynthetic pathway of iridoid glycosides was also observed in these species. Principal component analysis (PCA) showed a clear separation according to infrageneric classifications of genus Gentiana. Some secondary metabolites, such as mangiferin, rhodenthoside A-C, isoorientin, isovitexin, amarogentin, and swertianolin would serve as potential chemotaxonomic markers to differentiate Gentianaceae species. Furthermore, the accumulation of the six major metabolites seems to depend on geographical regions in Sect. Monopodiae and Sect. Cruciata. CONCLUSIONS: The combination of LC-MS/MS and FT-IR would provide some potential evidence on chemotaxonomic studies of Gentianaceae. Copyright © 2016 John Wiley & Sons, Ltd.

Anti-apoptotic and cytoprotective effect of Enicostemma littorale against oxidative stress in Islets of Langerhans.

Context Oxidative stress induces apoptosis within Islets of Langerhans in diabetes mellitus (DM). Enicostemma littorale blume, herb of the Gentianaceae family is used as an anti-diabetic agent across rural India. Objective This report demonstrates potent anti-apoptotic and cyto-protective activity of Enicostemma littorale MeOH extract (EL MeOH ext.) against 50 µM H2O2 in isolated rat Islets. Materials and methods In this study, the whole plant methanolic extract of EL with doses 0.25-4 mg/mL each for the preincubation duration of 0.5-4 h against 50 µM H2O2 were tested for optimum protective dose and time by Trypan blue dye exclusion assay. Islet intracellular reactive oxygen species (ROS) was quantified by DCFDA staining and cell death using PS/PI & FDA/PI staining. Further, comet assay, biochemical assessment of caspase-3 and antioxidant enzyme activities along with immunoblotting of PARP-1, caspase-3, TNF-α activation and p-P38 MapK (stress kinase) induction was performed. Results The optimized dose of EL MeOH ext. 2 mg/mL for 2 h was used throughout the study, which significantly decreased total Intracellular ROS and cell death. Further, caspase-3 activity, PARP-1 cleavage, p-P38 MapK (stress kinase) activation and TNF-α levels, which had been significantly elevated, were normalized. Antioxidant enzymes like catalase, superoxide dismutase, reduced glutathione and glutathione peroxidase, along with Comet assay, demonstrated that pretreatment with EL MeOH ext. can augment antioxidant enzyme activities and protect from DNA damage. Discussion and conclusions Significant anti-apoptotic and cyto-protective effects were mediated by EL with Islets of Langerhans subjected to oxidative stress-induced cell death.

[Research on origin, property and efficacy of Tibetan medicine "Dida"].

Tibetan medicine "Dida" isoccasionally misused due to its complex origins, which ultimately affects its clinical efficacy. The accurate name, origin, property, and efficacy of "Dida"are highly important for its further research and development. In the present study, by viewing the classic Tibetan medicine and modern literature, and combining the clinical practice of Tibetan medicine, the origins, properties and the clinic effects of "Dida" were defined. "Dida" originated from multiple plant species of Swertia, Gentianopsis, Halenia, Lomatogonium, Comastoma(Gentianaceae), Hedyotis (Saxifragaceae) and Erysimum (Cruciferae). The medicinal properties of "Dida" is mainly bitter and cold. It has been commonly used to treat febrile diseases and hepatic and gall diseases. This study suggested that the relevant herbalogical study, species identification and pharmacological effects of "Dida" should be taken based on the Tibetan medicine theories and clinical practice. Thus the medicine can be better used and ensure its safety and quality simultaneously.

Rapid preparative isolation of erythrocentaurin from Enicostemma littorale by medium-pressure liquid chromatography, its estimation by high-pressure thin-layer chromatography, and its α-amylase inhibitory activity.

Erythrocentaurin is a relatively simple natural product present among the members of Gentianaceae. A preparative method for the isolation of erythrocentaurin from the ethyl acetate fraction of Enicostemma littorale using medium-pressure liquid chromatography has been reported. The method consisted of a simple step gradient from 10 to 20% ethyl acetate in n-hexane. Using a 70 × 460 mm Si60 column, this method is capable of processing 20 g of material in <3 h (purity ≈ 97%). The recovery of erythrocentaurin was 87.77%. Estimation of erythrocentaurin in extracts and fractions based on high-pressure thin-layer chromatography was carried out on silica gel 60 F(254) plates with toluene/ethyl acetate/formic acid (80:18:2 v/v/v) as the mobile phase. The densitometric analysis was performed at 230 nm. A well-separated compact band of erythrocentaurin appeared at R(f )0.54 ± 0.04. The analytical method showed good linearity in the concentration range of 200-1500 ng/band with a correlation coefficient of 0.99417. The limits of detection and quantification were found to be ≈60 and ≈180 ng/band, respectively. Erythrocentaurin exhibited a concentration-dependent α-amylase inhibition (IC(50) 1.67 ± 0.28 mg/mL). The outcome of the study should be considered for pharmacokinetic and biotransformation studies involving E. littorale.

Nonprenylated Xanthones from Gentiana lutea, Frasera caroliniensis, and Centaurium erythraea as Novel Inhibitors of Vascular Smooth Muscle Cell Proliferation.

Aberrant proliferation of vascular smooth muscle cells (VSMC) plays a major role in restenosis, the pathological renarrowing of the blood vessel lumen after surgical treatment of stenosis. Since available anti-proliferative pharmaceuticals produce unfavorable side effects, there is high demand for the identification of novel VSMC proliferation inhibitors. A natural product screening approach using a resazurin conversion assay enabled the identification of gentisin (1) from Gentiana lutea as a novel inhibitor of VSMC proliferation with an IC50 value of 7.84 µM. Aiming to identify further anti-proliferative compounds, 13 additional nonprenylated xanthones, isolated from different plant species, were also tested. While some compounds showed no or moderate activity at 30 µM, 1-hydroxy-2,3,4,5-tetramethoxyxanthone (4), swerchirin (6), and methylswertianin (7) showed IC50 values between 10.2 and 12.5 µM. The anti-proliferative effect of 1, 4, 6, and 7 was confirmed by the quantification of DNA synthesis (BrdU incorporation) in VSMC. Cell death quantification (determined by LDH release in the culture medium) revealed that the compounds are not cytotoxic in the investigated concentration range. In conclusion, nonprenylated xanthones are identified as novel, non-toxic VSMC proliferation inhibitors, which might contribute to the development of new therapeutic applications to combat restenosis.

Factorial design of essential oil extraction from Fagraea fragrans Roxb. flowers and evaluation of its biological activities for perfumery and cosmetic applications.

OBJECTIVE: To optimize the extraction yields of essential oil from Fagraea fragrans Roxb. flowers in hydro-distillation using a central composite design (CCD) and to evaluate its biological activities for perfumery and cosmetic applications. METHODS: Central composite design was applied to study the influences of operational parameters [water to flower weight (X(1)) and distillation time (X(2))] on the yields of essential oil (Y). Chemical compositions of the essential oil extracted from the optimized condition were identified by gas chromatography-mass spectrometry. Antioxidant activities of the essential oil were determined against ABTS(•+) and DPPH(•) radicals, and the cytotoxic effects were assessed on human embryonic kidney (HEK293) cells by the use of the MTT assay. Also, the aromatic properties of the essential oil were evaluated by five healthy trained volunteers. RESULTS: The best conditions to obtain the maximum essential oil yield were 7.5 mL g(-1) (X(1)) and 215 min (X(2)). The experimental yield of the essential oil (0.35 ± 0.02% v/w) was close to the value predicted by a mathematical model (0.35 ± 0.01% v/w). 3-Octadecyne, Z,Z,Z-7,10,13-hexadecatrienal, E-nerolidol, pentadecanal and linalool were the major constituents of the essential oil. The essential oil showed moderate antioxidant capacities with no toxic effects on HEK293 cells at 1-250 µg mL(-1). Also, the essential oil exhibited a very strong aroma and was classified to be top- to middle-notes. CONCLUSION: The results offer the effectively operational conditions in the extraction of essential oil from F. fragrans using hydro-distillation. The essential oil could be used as a natural fragrance, having antioxidant activity with slight cytotoxicity, for perfumery and cosmetic applications.

[Study on identification of four kinds of Gentianaceae Mongolian medicine Digeda with spectroscopy techniques].

To study the identification of Gentianaceae Mongolian medicine Digeda with spectroscopy techniques, near infrared spectroscopy and differential scanning calorimetry techniques were applied to study on the identification of 4 kinds of Gentianaceae Mongolian medicine Digeda, and characteristic spectrums obtained were systematically analyzed. In NIR study, the four species of Digeda exist some differences in 4 250-4 400 cm(-1) and 5 650-5 800 cm(-1) of one-dimensional spectra, and show significant differences in 4 100- 4 400 cm(-1), 4 401-4 900 cm(-1) and 5 400-5 800 cm(-1) of the second derivative spectra. DSC curves of them present distinct topological pattern, characteristic peak and peak temperature. Using near infrared spectroscopy and differential scanning calorimetry analysis can realize efficient and accurate identification of four kinds of Mongolian medicine Digeda, and provide scientific basis for the efficient and accurate identification of other Gentianaceae Mongolian medicine Digeda.

[Preliminary comparative study of swertiamarin and swertisin on three kinds of Digeda-species Mongolian medicinal materials].

Lomatogonium rotatum (L.) Fries, Gentianopsis barbata (Froel) Ma, and Gentianella acuta (Michx.) Hulten, the three kinds of Digeda-species Mongolian medicinal materials belonging to the family Gentianaceae, bad been widely used for the treatment of liver diseases. To analyze comparatively the content of swertiamarin and swertisin among these three kinds of Digeda-species Mongolian medicinal materials. HPLC method was applied for qualitative and quantitative analysis of swertiamarin and swertisin. The Phenomenex C18 (4.6 mm x 250 mm, 5 µm) was used, chromatographic methanol and water as mobile phase, the flow rate was 1.5 mL x min(-1) with UV detected at 237 nm, column oven temperature was 25 degrees C. Results showed that the contents of swertiamarin and swertisin were closely related the different species and producing areas. The content range of swertiamarin in L. rotatum from different habitats was 1.73% - 2.72%, 0.43% - 0.96% for the swertisin content; the content of swertiamarin in G. barbata from Alxa Left Banner was 0.38%, and the content of swertiamarin and swertisin in G. barbata from the others habitats and G. Acuta from different habitats were all detected qualitatively. The contents of swertiamarin and swertisin among these medicinal plants showed a significant difference due to the different species and producing areas. As a consequence, these medicinal plants should not be put together for clinical applications.

[Chemical Constituents in Petroleum Ether Extract of Mongolian Medicine Halenia corniculata].

OBJECTIVE: To study the chemical constituents of Mongolian medicine Halenia corniculata. METHODS: Positive phase and reversed phase silica gel, as well as Sephadex LH-20 methods were used to separate and purify. The structure of the isolated constituents was identified according to the NMR spectroscopy data and the literature data. RESULTS: Nine compounds were isolated from 95% ethanol extracts of petroleum ether part of Halenia corniculata and identified as: 1-hydroxy-2,3,4,6-tetramethoxyxanthone (1), 1-hydroxy-2,3, 5-trimethoxyxanthone (2) 1-hydroxy-3,7-dimethoxyxanthone (3), 1-hydroxy-3,5,6,7,8-pentamethoxyxanthone (4), 1-hydroxy-2,3,4, 7-tetramethoxyxanthone (5), 1-hydroxy-3,5-dimethoxyxanthone (6),1-hydroxy-2,3,4,5,7-pentamethoxyxanthone (7), palmitic acid (8) and ß-sitosterol (9). CONCLUSION: Compounds 3, 4 and 8 are isolated from this genus for the first time, Compound 1 is isolated from this plant for the first time.

Hypoglycaemic effects of methanolic extract of Canscora decussata (Schult) whole-plant in normal and alloxan-induced diabetic rabbits.

In present study hypoglycaemic effects of the crude powdered C. decussata and its methanolic extract (ME) in alloxan diabetic rabbits were evaluated. The hypoglycaemic effect was measured by blood glucose, insulin level, HbA1c and his to pathology of pancreas. Glucose lowering effect of the ME was studied in diabetic rabbits. The effects of extract on blood glucose, body weight, food in take, fluid intake, OGTT were also evaluated. The results showed that 0.5,1 and 2g/kg of the powder significantly decreased blood glucose levels in normal rabbits and diabetic rabbits at the intervals checked. Oral intake of pioglitazone also reduced the levels in these rabbits. Synergistic hypoglycaemic effect of 600mg/kg of ME with different doses of insulin (2 & 3unit/kg, s/c) further reduced blood glucose levels of treated alloxan-diabetic rabbits. The oral glucose tolerance test revealed lowered area under curve values in ME treated rabbits. Treatment with ME (400 and 600 mg/kg) for 30 days showed highly significant decrease in blood glucose level by augmenting insulin secretion, HbA1cand significant increase in body weight, serum insulin levels in treated diabetic rabbits. Histopathology study showed regeneration of ß-cells. These studies have, therefore, supported the traditional use of this herb in diabetic patients.

The first stereoselective total synthesis of neosemburin and isoneosemburin.

The Claisen rearrangement of sugar derived allyl vinyl ether, Wittig olefination and intramolecular acetalization reactions were used as key steps in the first stereoselective total synthesis of neosemburin and isoneosemburin through the formation of a 3-C-branched sugar precursor as a key intermediate.

Hepatoprotective saikosaponin homologs from Comastoma pedunculatum.

Eight new triterpenoid saponins, the saikosaponin homologs comastomasaponins A-H (1-8), as well as a known triterpenoid (9) and eight known saponins (10-17) were isolated from the aerial portions of Comastoma pedunculatum. The structures of these compounds were elucidated spectroscopically, and their hepatoprotective activity and cytotoxic activity were evaluated against five human tumor cell lines in vitro. Compounds 1, 5-12, 14, 15, and 17 exhibited potent hepatoprotective activity, and compound 11 displayed cytotoxic activity against HCT-8, Bel-7402, BGC-825, A549, and A2780 human tumor cell lines.