Induction of apoptosis by Eleutherine bulbosa (Mill.) Urb. bulb extracted under optimised extraction condition on human retinoblastoma cancer cells (WERI-Rb-1).

ETHNOPHARMACOLOGICAL RELEVANCE: The bulb of Eleutherine bulbosa (Mill.) Urb. is an indigenous medicinal plant traditionally used among Dayak people for the management of diabetes, breast cancer, hypertension, stroke, and fertility problems in women. The bulb has been reported with a potent cytotoxic potential but with limited underlying mechanisms. AIM OF THE STUDY: This study aimed to investigate the cytotoxic properties of E. bulbosa ethanolic bulb extracted under optimised extraction condition on retinoblastoma cancer cells (WERI-Rb-1) through in vitro cell culture bioassays. The optimised extraction condition has been determined in the previous reports. MATERIALS AND METHODS: Cytotoxic assay was analysed through MTT assay. Comparison between non-optimised and optimised extraction condition from E. bulbosa ethanolic bulb extract was evaluated. Morphological assessment of apoptotic cells was conducted through acridine orange propidium iodide (AOPI) staining using fluorescence microscopy. Apoptosis assay was carried out through Annexin V-FITC and cell cycle analysis through PI staining. The effect of varying concentrations (IC25, IC50, IC75) of the optimised E. bulbosa ethanolic bulb extract was observed. The mRNA expression was also conducted to confirm the underlying mechanism. RESULTS: The optimised E. bulbosa ethanolic bulb extract markedly suppressed the proliferation of retinoblastoma cancer cells significantly with an IC50 value of 15.7 µg/mL as compared to non-optimised extract (p < 0.01). Fluorescence microscopy revealed that retinoblastoma cancer cells manifested early features of apoptosis-like membrane blebbing, chromatin condensation and formation of apoptotic bodies in a dose-dependent manner. The number of apoptotic cells were greatly observed in early and late apoptosis through Annexin V-FITC and the extract also induced cell arrestment as compared to the untreated group. The apoptosis was confirmed with the upregulation of Bax, Bad, p53, Caspase 3, Caspase 8, and Caspase 9 genes meanwhile, Bcl-2, BcL-xL, Nrf-2, and HO-1 genes were downregulated. CONCLUSION: The optimised E. bulbosa ethanolic bulb extract induced a significant cell death and cell cycle arrestment on retinoblastoma cancer cells. It could be suggested that the induction of apoptosis in retinoblastoma cancer cells may be due to the synergistic effect of the bioactive compounds extracted under optimised extraction condition. Our findings indicated that E. bulbosa bulb could be promising chemotherapeutic potential to treat retinoblastoma cancer cells.

New iridal-type triterpenoid analogues with 6/5/6-fused carbon skeleton from the rhizomes of Belamcanda chinensis.

Five new iridal-type triterpenoid derivatives with 6/5/6 tricyclic ring skeleton (1-5) were obtained from the rhizomes of Belamcanda chinensis. Their structures were determined on the basis of detailed spectroscopic data and ECD calculation. Compounds 1-5 possessed the same 6/5/6-fused carbon skeleton as Belamchinenin A, which further enriched this kind of iridals. In vitro bioassay, compounds 2 and 3 exhibited 51.95 and 54.52% inhibitory activities, respectively, against Fe2+/cysteine-induced liver microsomal lipid peroxidation at a concentration of 10 µM. A putative biogenetic pathway for compounds 1-5 was proposed.

Iridal-Type Triterpenoids Displaying Human Neutrophil Elastase Inhibition and Anti-Inflammatory Effects from Belamcanda chinensis.

The aim of this study is to explore anti-inflammatory phytochemicals from B. chinensis based on the inhibition of pro-inflammatory enzyme, human neutrophil elastase (HNE) and anti-inflammatory activities in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage. Three stereoisomers of iridal-type triterpenoids (1-3) were isolated from the roots of B. chinensis and their stereochemistries were completely identified by NOESY spectra. These compounds were confirmed as reversible noncompetitive inhibitors against HNE with IC50 values of 6.8-27.0 µM. The binding affinity experiment proved that iridal-type triterpenoids had only a single binding site to the HNE enzyme. Among them, isoiridogermanal (1) and iridobelamal A (2) displayed significant anti-inflammatory effects by suppressing the expressions of pro-inflammatory cytokines, such as iNOS, IL-1ß, and TNF-α through the NF-κB pathway in LPS-stimulated RAW264.7 cells. This is the first report that iridal-type triterpenoids are considered responsible phytochemicals for anti-inflammatory effects of B. chinensis.

Eleutherine bulbosa (Mill.) Urb. Bulb: Review of the Pharmacological Activities and Its Prospects for Application.

Natural product is an excellent candidate for alternative medicine for disease management. The bulb of E. bulbosa is one of the notable Iridaceae family with a variety therapeutic potential that is widely cultivated in Southeast Asia. The bulb has been used traditionally among the Dayak community as a folk medicine to treat several diseases like diabetes, breast cancer, nasal congestion, and fertility problems. The bulb is exceptionally rich in phytochemicals like phenolic and flavonoid derivatives, naphthalene, anthraquinone, and naphthoquinone. The electronic database was searched using various keywords, i.e., E. bulbosa, E. americana, E. palmifolia, E. platifolia, and others due to the interchangeably used scientific names of different countries. Scientific investigations revealed that various pharmacological activities were recorded from the bulb of E. bulbosa including anti-cancer, anti-diabetic, anti-bacterial, anti-fungi, anti-viral, anti-inflammatory, dermatological problems, anti-oxidant, and anti-fertility. The potential application of the bulb in the food industry and in animal nutrition was also discussed to demonstrate its great versatility. This is a compact study and is the first study to review the extensive pharmacological activities of the E. bulbosa bulb and its potential applications. The development of innovative food and pharma products from the bulb of E. bulbosa is of great interest.

Isolation and identification of belamcandaoids A-N from Belamcanda chinensis seeds and their inhibition on extracellular matrix in TGF-ß1 induced kidney proximal tubular cells.

Belamcandaoids A-N (1-14), fourteen new triterpenoids were isolated from the seeds of Belamcanda chinensis. Their structures including absolute configurations were assigned by using spectroscopic, computational, and crystallographic methods. All the compounds except 1 and 2 are 3,4-seco-triterpenoids belonging to fernane type. Biological evaluation results indicated that 3 and 13 could reduce fibronectin and collagen I expression respectively in TGF-ß1 induced kidney proximal tubular cells.

Activity and Toxicity of Eleutherine palmifolia (L.) Merr. Extract on BALB/c Mice Colitis-Associated Colon Cancer Model.

OBJECTIVE: Eleutherine palmifolia (L.) Merr. extract (EPE) containing isoliquiritigenin and oxyresveratrol is believed to be an anticancer agent. This study evaluates colon histopathology, TNF-α, TGF-ß, and hepatotoxicity on BALB/c mice colitis-associated colon cancer (CAC) model treated with EPE. METHODS: In vivo study was performed on BALB/c mice CAC model induced by 10 mg/kgBW AOM on the first day followed by administration that each cycle consisted of 5% DSS in water for seven days and regular water for seven days. The indicators of the formation of CAC were observed by a fecal occult blood test (FOBT) and serum amyloid α (SAA) test. The treatment was conducted once a week started from the seventh week up to the twentieth week with six treatment groups: I was administrated by regular water only (negative control), II was administrated by AOM and DSS only (positive control), III was administrated by doxorubicin,  IV-VI were treated by EPE (0.25 mg/kg BW, 0.50 mg/kg BW, and 1.00 mg/kg BW) respectively. The colon and liver's histopathology was observed using hematoxylin-eosin (HE) staining, TNF-α with immunohistochemistry (IHC), and level measurement of TGF-ß colon with ELISA reader. The data were used one-way ANOVA followed by post hoc as statistical analysis. RESULTS: The administration of EPE increased the expression of TNF-α, the total of goblet cells of the colon, and decreased the level of TGF-ß. Administration of EPE 0.50 mg/20g BW decreased a liver histopathological score but induced a histopathological alteration of the liver at a dose of 1.00 mg/20g BW. CONCLUSION: This study indicate that EPE could be recommended as a colon anticancer through increase the goblet cells, induce apoptosis through increase TNF-α, and decrease TGF-ß.

Eleucanainones A and B: Two Dimeric Structures from the Bulbs of Eleutherine americana with Anti-MRSA Activity.

Two naphthoquinone-derived heterodimers with unprecedented carbon skeletons, eleucanainones A (1) and B (2), were isolated from the bulbs of Eleutherine americana. Their structures were elucidated by comprehensive spectroscopic methods. The structures of 1 and 2 were determined to be the first examples of dibenzofuran- and naphthalenone-containing naphthoquinone dimers. Compound 1 exhibited significant anti-MRSA activity in vitro with minimum inhibitory concentration (MIC) values of 0.78 µg/mL by downregulation of basal expression of agrA, cidA, icaA and sarA in methicillin-resistant S. aureus (MRSA).

Effect in white shrimp Litopenaeus vannamei of Eleutherine bulbosa (Mill.) Urb. Powder on immune genes expression and resistance against Vibrio parahaemolyticus infection.

This study investigated the influence of Eleutherine bulbosa (Mill.) Urb. on the immune responses, bacterial population in the intestines, and resistance of white shrimp, Litopenaeus vannamei, against infection with Vibrio parahaemolyticus. Shrimp were fed with three dosages of powder, at 6.25 g kg-1 (P6.25), 12.5 g kg-1 (P12.5), and 25 g kg-1 (P25). One dosage of the crude extract was provided, 1.25 g kg-1 (E1.25), and the controls without administration of E. bulbosa consisted of a positive control (PC) and a negative control (NC). Feed supplementation was carried out for 30 days; then shrimp from all treatments were challenged by intramuscular injection with V. parahaemolyticus (106 cfu/mL), except for the NC. The results showed that supplementation with the powder and extract of E. bulbosa for 30 days resulted in significantly higher (P < 0.05) immune responses (total hemocyte count (THC), phenoloxidase activity (PO), respiratory bursts (RBs)), gene expression (prophenoloxidase (proPO), lipopolysaccharide- and ß-1,3-glucan-binding protein (LGBP)), and total bacterial count (TBC) compared to PC/NC. In post challenge testing, there were significantly higher levels for THC, PO, RBs, proPO, LGBP, and PE (peroxinetin), and the treatments were able to suppress V. parahaemolyticus in the intestines, hepatopancreas, and muscles and to reduce damage to the muscles and hepatopancreas. The survival rate with P12.5 was significantly higher compared to the other treatments. It was concluded that the shrimp receiving supplementation with the powder and extract of E. bulbosa had increased immunity and resistance against V. parahaemolyticus infection, with the best dosage being the P12.5 treatment.

Montbresides A-D: antibacterial p-coumaroyl esters of a new sucrose-based tetrasaccharide from Crocosmia × crocosmiiflora (montbretia) flowers.

Crocosmia × crocosmiiflora (montbretia) flowers yielded four esters (montbresides A-D) of a new sucrose-based tetrasaccharide, 3-O-ß-d-glucopyranosyl-4´-O-α-d-rhamnopyranosyl-sucrose [ß-d-Glc-(1 → 3)-α-d-Glc-(1↔2)-ß-d-Fru-(4 ← 1)-α-d-Rha]. All four possess O-p-coumaroyl residues on C-3 of fructose and C-4 of α-glucose, plus O-acetyl residues on C-2 and C-3 of rhamnose and C-6 of fructose. Montbresides A and B are additionally O-acetylated on C-1 of fructose. The p-coumaroyls are trans- in montbresides A and C and cis- in B and D. Elemental compositions were determined from MS data, and structures from 1D and 2D NMR spectra. Monosaccharide residues were identified from selective 1D TOCSY spectra and TLC, and acylation sites from 2D HMBC spectra. Enantiomers were distinguished by enzymic digestion. Montbretia flower extracts were cytotoxic against six human cancerous cell-lines, but purified montbresides lacked cytotoxicity. Each montbreside displayed antibacterial activity against Staphylococcus aureus (minimal inhibitory concentration ~6 µg/ml). Montbretia is a potential source of new cytotoxins and antibacterial agents.

Evaluation of Lactobacillus plantarum encapsulated with Eleutherine americana oligosaccharide extract as food additive in yoghurt.

This study aimed to determine the survival and antibacterial activity of Lactobacillus plantarum TISTR1465 encapsulated with Eleutherine americana oligosaccharide extract. Capsules were stored at 4 °C for 0, 2, and 4 weeks. The encapsulated cells were evaluated for their survival after sequential exposure to simulated gastric and intestinal juices, then evaluated in terms of their antibacterial activity. Survival of the encapsulated cells was higher than that of free cells at weeks 2 and 4. Highest levels of viable cells were observed with encapsulation in E. americana oligosaccharide extract. No surviving free cells were found in week 4. Yoghurt prepared with encapsulated cells showed less acidification than with free cells. Antibacterial activity of L. plantarum TISTR1465 before pH neutralisation against Clostridium perfringens ATCC13124, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Salmonella Typhimurium ATCC13311 was higher than after pH neutralisation. Encapsulation by extrusion enhanced antibacterial activity of the cells against enteropathogenic bacteria. The antibacterial activity of encapsulated cells against Gram-positive bacteria was higher than that against Gram-negative bacteria. Results indicates that L. plantarum TISTR1465 encapsulated with E. americana oligosaccharide extract showed potential for application as a functional food additive.

Four New Compounds Isolated from the Aerial Part of Belamcanda chinensis (L.) and Their Effect on Vascular Smooth Muscle Cell (VSMC) Proliferation.

Bio-guided fractionation of the 70% ethanol extract of Belamcanda chinensis (L.) DC. revealed four new compounds, including 6″-O-acetylembinin (5), 3″-O-acetylembinin (6), irigenin 3'-O-ß-glucopyranoside (8), and 2'-acetyl-1,3-O-diferuloylsucrose (9), along with five known compounds (1-4, 7). Their chemical structures were determined using extensive NMR data, mass spectroscopy, and comparison with published literature. Among the isolates, compounds 1 and 4-7 achieved good regulation of the growth and proliferation of vascular smooth muscle cells.

Chemotherapeutic options for the treatment of human trichomoniasis.

Trichomonas vaginalis is the causative agent of the most common non-viral sexually transmitted disease worldwide. The infection may be associated with severe complications, including infertility, preterm labour, cancer and an increased risk of human immunodeficiency virus (HIV) transmission. Treatment remains almost exclusively based on 5-nitroimidazoles, but resistance is on the rise. This article provides an overview of clinically evaluated systemic and topical treatment options for human trichomoniasis and summarises the current state of knowledge on various herbal, semisynthetic and synthetic compounds evaluated for their anti-Trichomonas efficacy in vitro.

New naphtoquinones derivatives from the edible bulbs of Eleutherine americana and their protective effect on the injury of human umbilical vein endothelial cells.

Seven new naphthoquinone derivatives, named eleutherins A-G (1-7), were isolated from the edible bulbs of Eleutherine americana (Hong-Cong), which belongs to the Iridaceae family. Their structures were elucidated on the basis of spectroscopic analysis including high-resolution-electron spray ionization-mass spectrometry (HR-ESI-MS) and 1D and 2D nuclear magnetic resonance (NMR) techniques. The absolute configurations of compounds 1-4 were determined by experimental and calculated data. Additionally, a hypothetical biosynthetic pathway of 1-7 was postulated. All the isolates were evaluated for their protective effect against the injury of human umbilical vein endothelial cells (HUVECs) in vitro. Eleutherins A and B (1-2) showed a potential protective effect on microvessels.

[A new naphthalene derivative from bulbs of Eleutherine americana].

A new naphthalene derivative and three known compounds were isolated from the petroleum ether extract of the bulbs of Eleutherine americana by using various chromatographic techniques, such as column chromatography over silica gel and semi-preparative HPLC. Their structures were elucidated by spectroscopic date (MS, UV, IR, NMR), which were identified as eleutherol B (1), 4,8-dihydroxy-3-methoxy-1-methylanthraquinone-2-carboxylic acid methyl ester (2), 8-hydroxy-3,4-dimethoxy-1-methylanthraquinone-2-carboxylic acid methyl ester (3), and isoeleutherine (4). Compound 1 is a new compound. The diastolic blood vessels activity of compound 1 and 2 were potent, reaching 82.5% and 85.3% at the concentration of 10 µmol·L⁻¹, which were basically the same as that of the positive drug tanshinone ⅡA.

Dayak onions ( Eleutherine bulbosa (Mill.) Urb. ) bulbs extracts reduces blood pressure of hypoestrogen model rats by controlling lipids profile / El extracto de bulbos de cebollas dayak ( Eleutherine bulbosa (Mill. ) Urb. ) reduce la presión sanguínea en un modelo hipoestrogénico en ratas controlando el perfil lipídico

To determine the effects of 70 % ethanolic extract of Eleutherine bulbosa (Mill.) L. bulbs contained naphtoquinone in blood pressure and lipids profile level of ovariectomized rats. Thin layer chromatography was performed to analyze the content of the Eleutherine bulbosa (Mill.) bulbs extracts using Chloroform : Methanol (7:1) as eluent. This study used white female Sprague-Dawley rats which were divided into 6 groups: SHAM and OVX, both given 0.5 % carboxymethyl cellulose; the positive control group that was given tamoxifen (0.36 mg/200 g B.W.); the three various of doses of extract namely dose 1, 2, and 3 that were given 8, 12, and 18 mg/200 g BW respectively on day 29 until 50. All groups were ovariectomized on day 1, except sham group. Measurement of blood pressure were performed on the day before ovariectomy using non-invasive tool CODA®; and on the second, third, and fourth weeks post-ovariectomy. Three weeks after treatment of hypoestrogen rats by the dose 18 mg/200 g B.W, Eleutherine bulbosa extracts could decrease the systolic blood pressure level to 28.06 %, the diastolic to 30.47 %. Lipid profile of Dose 3 also showed recovery of Triglyceride, LDL and also Total Cholesterol. Eleutherine bulbosa extracts could improve blood pressure of ovariectomized rats by controlling lipids profile level.

Para determinar los efectos del 70 % de extracto etanólico de Eleutherine bulbosa (Mill.) L. en bulbos que contenían naftoquinona, se analizó la presión arterial de ratas ovariectomizadas. Se realizó un estudio de cromatografía de capa fina para analizar el contenido de los extractos de bulbos de Eleutherine bulbosa (Mill.) usando cloroformo:metanol (7:1) como diluyente. En este estudio se utilizaron ratas blancas Sprague-Dawley hembras, que se dividieron en 6 grupos: SHAM y OVX, ambos grupos fueron administrados con carboximetilcelulosa al 0,5 %; grupo control positivo, recibió tamoxifeno (0,36 mg / 200 g de B.W.); y tres grupos restantes, que recibieron tres dosis diferentes de extracto, es decir, dosis 1, 2 y 3 a las que se les administró 8, 12 y 18 mg / 200 g de por peso, respectivamente el día 29 hasta el día 50. Todos los grupos fueron sometidos a ovariectomía en el día 1, excepto el grupo simulado. La medición de la presión arterial se realizó el día anterior a la ovariectomía con la herramienta no invasiva CODA®, y, posteriormente, en la segunda, tercera y cuarta semanas de realizada la ovariectomía. Tres semanas después del tratamiento aplicado a las ratas hipoestrógenas, con la dosis de 18 mg / 200 g por peso, los extractos de Eleutherine bulbosa podrían disminuir el nivel de presión arterial sistólica al 28,06 %, y la diastólica al 30,47 %. La dosis 3 del perfil lipídico mostró mejoría en los niveles de Triglicéridos, LDL y Colesterol Total. Los extractos de Eleutherine bulbosa podrían mejorar la presión sanguínea de ratas ovariectomizadas controlando el nivel del perfil lipídico.

Probing the Antiallergic and Anti-inflammatory Activity of Biflavonoids and Dihydroflavonols from Dietes bicolor.

Dietes bicolor (Iridaceae) is an ornamental plant used by African local healers to treat diarrhea and dysentery. A new dihydroflavonol, (2R,3R)-3,5,7-trihydroxy-8-methoxyflavanone (1); two known dihydroflavonols, trans-3-hydroxy-5-methoxy-6,7-methylenedioxyflavanone (2) and trans-3-hydroxy-5,7-dimethoxyflavanone (3); the known isoflavone orobol 7,3'-di-O-methyl ether (4); the known biflavones lanaroflavone (5), robustaflavone (6), and amentoflavone (7); and ß-sitosterol (8) were isolated from the CH2Cl2 fraction of D. bicolor leaves. The extract showed potent activity in antiallergic and anti-inflammatory assays. The structures of the isolates were identified by spectroscopic and spectrometric methods. Compounds 6 and 7 (400 µM) exhibited antiallergic activity by inhibiting antigen-induced ß-hexosaminidase release at 45.7% and 46.3%, respectively. Moreover, 6 and 7 exerted anti-inflammatory activity as demonstrated by the inhibition of superoxide anion generation with an IC50 value of 1.0 µM as well as the inhibition of elastase release with IC50 values of 0.45 and 0.75 µM, respectively. The anti-inflammatory activity was further explained by the virtual docking of the isolated compounds to the binding sites in the human neutrophil elastase (HNE) crystal structure using Discovery Studio 2.5. It was concluded that the biflavonoids bind directly to HNE and inhibit its enzymatic activity based on the CDOCKER algorithm. The data provided evidence for the potential use of D. bicolor against certain diseases related to allergy and inflammation.

The impact of mangiferin from Belamcanda chinensis on experimental colitis in rats.

BACKGROUND: Inflammatory bowel disease (IBD) [including Crohn's disease (CD) and ulcerative colitis (UC)] constitutes an important clinical problem. The pathogenesis of IBD remains unclear. It is believed that immune dysfunction, inflammatory mediators and oxidative damage play crucial roles in development of IBD. The condition is clinically associated with symptoms ranging from mild to severe during relapses, depending on the affected segment of the gastrointestinal tract. Bloody diarrhea with mucus, abdominal pain, weight loss and anemia are initial symptoms of both CD and UC. Differences between diseases become more evident in time, along with the development of intestinal and extraintestinal complications. Mangiferin (1,3,6,7-tetrahydroxyxanthone-C-2-ß-D-glucoside), a natural polyphenol in plants, exerts antioxidant and anti-inflammatory effects making it an interesting option for the treatment of inflammatory pathologies associated with oxidative stress in humans, such as IBD. PURPOSE: The aim of the current study was to elucidate the impact of mangiferin on colon tissues in 2,4,6-trinitrobenzensulfonic acid (TNBS)-induced colitis in rats. METHODS: Mangiferin was obtained from Belamcanda chinensis rhizomes by a multistage process. Groups of rats were pre-treated with 10, 30 or 100 mg/kg of mangiferin, or with distilled water administered intragastrically for 16 days. An ethanol solution of TNBS or saline was given rectally on the day 15 of the experiment. The experiment was terminated on the day 17. The colon was removed, cleaned, weighed and examined macro- and microscopically. Determination of tumor necrosis factor α (TNF-α), interleukin 17 (IL-17), malondialdehyde (MDA) levels and superoxide dismutase (SOD) activity were performed spectrophotometrically in homogenates of colon tissues. RESULTS: Rats in the TNBS group developed symptoms of colitis, including: body weight loss, colon mass index increase and damage of intestinal tissues with concomitant increase in TNF-α, IL-17, MDA levels and decreased SOD activity. In non-TNBS-treated rats mangiferin did not cause any changes of studied parameters. Pre-treatment with mangiferin exerted a protective effect, reducing the intensity of damage caused by TNBS. Mangiferin at the doses of 30 and 100 mg/kg reduced the macro- and microscopic damage score and the MDA level in colon tissues. Only at the dose of 100 mg/kg, mangiferin decreased TNF-α and IL-17 concentrations, and SOD activity in colon tissues. CONCLUSION: Mangiferin attenuates inflammatory changes of colon tissues in experimental, TNBS-induced colitis in rats. Protective effect exerted by mangiferin depends primarily on its anti-inflammatory activity and secondarily on its antioxidant properties.

Gladiolus dalenii lyophilisate reverses scopolamine-induced amnesia and reduces oxidative stress in rat brain.

Learning and memory are the most important executive functions performed by the human brain, the loss of which is a prominent feature in dementia. Gladiolus dalenii is traditionally used to treat a number of illnesses such as epilepsy and schizophrenia in Cameroon. This study aims to investigate the anti-amnesia effect of Gladiolus dalenii in scopolamine-induced amnesia in rats and its possible antioxidant properties in this model. Morris water maze, novel object location and recognition tasks were used to assess spatial and working memory. Male rats were treated for 12 days with saline, G. dalenii or Tacrine. Experimental animals were co-treated with scopolamine once daily from day 9 to 12. Acetylcholinesterase activity was measured in the prefrontal cortex and hippocampus. Malondialdehyde and glutathione levels were measured in the hippocampus. G. dalenii reversed memory impairment induced by scopolamine in the Morris water maze, novel object location and recognition tasks. It decreased acetylcholinesterase activity in the hippocampus and prefrontal cortex. It also decreased the level of malondialdehyde and increased the level of glutathione in the hippocampus. The results of this study show that G. dalenii ameliorates the cognitive impairment induced by scopolamine, through inhibition of oxidative stress and enhancement of cholinergic neurotransmission. It can therefore be useful for treatment of conditions associated with memory dysfunction as seen in dementia.

Ionic-liquid-based ultrasound-assisted extraction of isoflavones from Belamcanda chinensis and subsequent screening and isolation of potential α-glucosidase inhibitors by ultrafiltration and semipreparative high-performance liquid chromatography.

The separation of a compound of interest from its structurally similar homologues to produce high-purity natural products is a challenging problem. This work proposes a novel method for the separation of iristectorigenin A from its structurally similar homologues by ionic-liquid-based ultrasound-assisted extraction and the subsequent screening and isolation of potential α-glucosidase inhibitors via ultrafiltration and semipreparative high-performance liquid chromatography. Ionic-liquid-based ultrasound-assisted extraction was successfully applied to the extraction of tectorigenin, iristectorigenin A, irigenin, and irisflorentin from Belamcanda chinensis. The optimum conditions for the efficient extraction of isoflavones were determined as 1.0 M 1-ethyl-3-methylimidazolium tetrafluoroborate with extraction time of 30 min and a solvent to solid ratio of 30 mL/g. Ultrafiltration with liquid chromatography and mass spectrometry was applied to screen and identify α-glucosidase inhibitors from B. chinensis, followed by the application of semipreparative high-performance liquid chromatography to separate and isolate the active constituents. Four major compounds including tectorigenin, iristectorigenin A, irigenin, and irisflorentin were screened and identified as α-glucosidase inhibitors, and then the four active compounds abovementioned were subsequently isolated by semipreparative high-performance liquid chromatography (99.89, 88.97, 99.79, and 99.97% purity, respectively). The results demonstrate that ionic liquid extraction can be successfully applied to the extraction of isoflavones from B. chinensis.

Ultrafiltration-LC-MS combined with semi-preparative HPLC for the simultaneous screening and isolation of lactate dehydrogenase inhibitors from Belamcanda chinensis.

Stroke represents the fourth leading cause of death in the USA and the second leading cause of death worldwide. Lactate dehydrogenase inhibitors are widely used in the treatment of ischemic stroke and natural products are considered a promising source of novel lactate dehydrogenase inhibitors. In this study, we used PC12 cells to determine the protective effect of extracts from the herb Belamcanda chinensis following toxic challenge. Using ultrafiltration high-performance liquid chromatography coupled with photo-diode array detection and electrospray ionization mass spectrometry, we screened and identified isoflavonoids from Belamcanda chinensis extracts. Semi-preparative high-performance liquid chromatography was then applied to separate and isolate the active constituents. Using these methods, we identified six major compounds in Belamcanda chinensis as lactate dehydrogenase inhibitors: tectoridin, iristectorin A, iridin, tectorigenin, irigenin, and irisflorentin, which were then isolated to >92% purity. This is the first report that Belamcanda chinensis extracts contain potent lactate dehydrogenase inhibitors. Our results demonstrate that the systematic isolation of bioactive components from Belamcanda chinensis guided by ultrafiltration high-performance liquid chromatography coupled with photo-diode array detection and electrospray ionization mass spectrometry represents a feasible and efficient technique that could be extended for the identification and isolation of other enzyme inhibitors.