Alkaloid and benzopyran compounds of Melicope latifolia fruit exhibit anti-hepatitis C virus activities.

BACKGROUND: New agents for developing alternative or complementary medicine to treat the hepatitis C virus (HCV) are still needed due to high rates of HCV infection globally and the current limitations of available treatments. Treatment of HCV with a combination of direct acting antivirals have been shown to be approximately 90% effective but will be limited in the future due to the emergence of drug resistance and high cost. The leaves of Melicope latifolia have previously been reported to have anti-HCV activity and are a potential source of bioactive compounds for future novel drug development. This study aimed to evaluate the efficacy of the extract of M. latifolia fruit to treat HCV and to isolate its active compounds. METHOD: M. latifolia fruit was extracted using methanol and purified using vacuum liquid chromatography (VLC) and Radial Chromatography. The anti-HCV activity was analyzed using cell culture lines Huh7it-1 and JFH1 (genotype 2a). Time-of-addition and immunoblotting studies were performed to identify the mode of action of the isolated active compounds. The structures of the active compounds were determined using nuclear magnetic resonance (NMR) spectra, UV, IR, and Mass Spectra. RESULTS: Six known compounds were isolated from M. latifolia fruit: O-methyloktadrenolon, alloevodionol, isopimpinellin, alloxanthoxyletin, methylevodionol, and N-methylflindersine. N-methylflidersine was the most active compound with IC50 value of 3.8 µg/ml while methylevodionol, isopimpinellin, and alloevodionol were less active. O-methyloktadrenolon and alloxanthoxyletin were moderately active with IC50 values of 10.9 and 21.72 µg/ml, respectively. N-methylflidersine decreased level of HCV NS3 protein expression in the cells. CONCLUSION: The alkaloid compound, N-methylflindersine which was isolated from M. latifolia possesses anti-HCV activity through post-entry inhibition and suppressed NS3 protein expression.

Biochemical and toxicological investigation of karanjin, a bio-pesticide isolated from Pongamia seed oil.

Karanjin, a furanoflavonol from Pongamia pinnata L is used in agricultural practices for its pesticidal, insecticidal and acaricidal activities. It is commercially available as a bio-pesticide targeting a wide variety of pests. The present study was intended to evaluate the biochemical interactions of karanjin with bovine serum albumin (BSA) and study its toxicological effects on mammalian and bacterial cell lines. Karanjin bound to BSA at a single site with a dissociation constant of 19.7 µM. Evaluation of BSA-karanjin interactions at three different temperatures indicated the involvement of static mode of quenching. Binding experiments in the presence of warfarin and computational docking analysis indicated that karanjin bound closer to the warfarin binding site located in the Subdomain IIA of BSA. Using Förster resonance energy transfer analysis the distance between TRP 213 of BSA and karanjin was found to be 20 Å. Collective results from synchronous fluorescence spectra analysis, differential scanning calorimetry, and circular dichroism analysis indicated that binding of karanjin induced conformational changes in the secondary structure of BSA. Karanjin exhibited low toxicity against human cervical cancer cells and normal mouse fibroblast L929 cells and modestly inhibited the growth of B. subtilis and E. coli cells. The data presented in this study provides insights for understanding the binding interactions of karanjin with BSA and its possible toxicological effects on mammalian cell lines and bacteria.

Dissolved organic matter and aluminum oxide nanoparticles synergistically cause cellular responses in freshwater microalgae.

This study investigated the impact of dissolved organic matters (DOM) on the ecological toxicity of aluminum oxide nanoparticles (Al2O3NPs) at a relatively low exposure concentration (1 mg L-1). The unicellular green alga Scenedesmus obliquus was exposed to Al2O3NP suspensions in the presence of DOM (fulvic acid) at various concentrations (1, 10, and 40 mg L-1). The results show that the presence of DOM elevated the growth inhibition toxicity of Al2O3NPs towards S. obliquus in a dose-dependent manner. Moreover, the combination of DOM at 40 mg L-1 and Al2O3NPs resulted in a synergistic effect. The relative contribution of Al-ions released from Al2O3NPs to toxicity was lower than 5%, indicating that the presence of the particles instead of the dissolved ions in the suspensions was the major toxicity sources, regardless of the presence of DOM. Furthermore, DOM at 10 and 40 mg L-1 and Al2O3NPs synergistically induced the upregulation of intercellular reactive oxygen species levels and superoxide dismutase activities. Analysis of the plasma malondialdehyde concentrations and the observation of superficial structures of S. obliquus indicated that the mixtures of DOM and Al2O3NPs showed no significant effect on membrane lipid peroxidation damage. In addition, the presence of both DOM and Al2O3NPs contributed to an enhancement in both the mitochondrial membrane potential and the cell membrane permeability (CMP) in S. obliquus. In particular, Al2O3NPs in the presence of 10 and 40 mg L-1 DOM caused a greater increase in CMP compared to Al2O3NPs and DOM alone treatments. In conclusion, these findings suggest that DOM at high concentrations and Al2O3NPs synergistically interrupted cell membrane functions and triggered subsequent growth inhibition toxicity.

Reproductive toxicity of brazilein in ICR mice.

Brazilein is an active small molecular compound extracted from Caesalpinia sappan L. with favorable pharmacological properties on immune system, cardiovascular system, and nervous system. C. sappan has been used as a traditional medicine in China for hundreds of years for various diseases. However, the general reproductive toxicity of brazilein is still unknown. The purpose of the present study was to thoroughly evaluate the general reproductive toxicity of brazilein in ICR mice to support the future drug development and modernization of this potent traditional Chinese medicine. The results showed that, although no apparent toxicity on the reproducibility of the male was observed, brazilein might cause considerable risks to the fetuses and females as indicated by the ratios of dead fetuses and reabsorptions. In conclusion, our results from the present study provided some useful insights about the safety profile of brazilein, suggesting that brazilein should be used with caution in pregnant women.

Antitrypanosomal isoflavan quinones from Abrus precatorius.

Human African trypanosomiasis is a neglected tropical disease in sub Saharan Africa that is fatal if left untreated. In a search for new natural products with antitrypanosomal activity, we recently identified abruquinones B and I from Abrus precatorius as potent in vitro trypanocidal compounds with high selectivity indices. To obtain sufficient compound for in vivo efficacy tests in mice, a second batch of plant material was re-collected and extracted. However, the chemical profiles of the two batches differed, and additional abruquinones were isolated and identified by HR-ESI-MS, and 1D and 2D NMR ((1)H, (13)C, COSY, HMBC, HSQC, and NOESY) spectroscopy. Abruquinones J (1), K (2), and L (3) were new, while abruquinones A (4) and D (5) were known from the first batch of plant material. The absolute configuration of compounds 1 to 3 was determined by comparison of electronic circular dichroism (ECD) spectra with calculated ECD data. Compounds 2 to 5 showed high in vitro activity against T. b. rhodesiense (IC50 of 0.01, 0.02, 0.02 and 0.01 µM, respectively), and remarkable SIs of 508, 374, 1379, and 668, respectively.

Antioxidant enzyme activities and lipid peroxidation in earthworm Eisenia fetida exposed to 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethyl-cyclopenta-γ-2-benzopyran.

Polycyclic musks have been indicated to cause lethal and sublethal effects on exposed biota. However, knowledge about the effect of polycyclic musks on the antioxidant defense system in earthworms is vague. In this work, the activities of antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and malondialdehyde (MDA) exposed to 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethyl-cyclopenta-γ-2-benzopyran (HHCB) were systematically investigated. The investigation shows that their activities are closely related to the exposed dose and time of HHCB. For SOD and CAT, the activities increased monotonically with increased exposed dose of HHCB, which indicates a dose-dependent change pattern. POD exhibited its peak activity in 0.0157 µg cm(-2) HHCB treatment and decreased at higher concentrations. These two changing patterns were complementary, which reveals the cooperation of enzymes in response to oxidative stress. MDA content in earthworms was basically unaffected with a 1-day exposure and significantly increased after 2-day and 3-day exposures, correlating with changes in the activities of SOD and CAT when the concentration of HHCB was high. It was also found that the sensitivity of Eisenia fetida to HHCB increased over time. These results may support the theoretical hypothesis that oxidative stress is an important component for the response of earthworms to the toxicity of HHCB in environment. Among the studied enzymes, SOD and CAT appeared to be the most responsive biomarkers of oxidative stress caused by HHCB. © 2010 Wiley Periodicals, Inc. Environ Toxicol, 2012.

Physicochemical characterization and toxicity of decursin and their derivatives from Angelica gigas.

Angelica gigas NAKAI is used to treat dysmenorrhea, amenorrhea, menopause, abdominal pain, injuries, migraine, and arthritis. The present study provided a physicochemical and toxicological characterization of compounds in A. gigas NAKAI (decursin, decursinol angelate, diketone decursin, ether decursin, epoxide decursin and oxim decursin). Diketone decursin (173.16 µg/mL) and epoxide decursin (122.12 µg/mL) exhibited >100 µg/mL kinetic solubility after applying nephelometry, suggesting a highly soluble compound. The Student's t-test revealed significant differences in the pKa ranges of the compounds by automatic titration from capillary electrophoresis (p<0.05). Diketone decursin, epoxide decursin and oxim decursin might be formulated into an oral dosage form (log P: 0-3) by an automatic titration analysis. A parallel artificial membrane permeability assay demonstrated permeability coefficients of <10 x 10⁻6 cm/s for all of the compounds, suggesting poor permeability. Ether decursin exhibited a toxic effect after being applied to mouse (NIH 3T3, EC50: 57.9 µM) and human (HT-29, EC50: 36.1 µM; Hep-G2, EC50: 4.92 µM) cells. Additionally, epoxide and oxim decursin were toxic through acute oral toxicity (four and three deaths of Institute of Cancer Research (ICR) mice) and mutation toxicity testing by applying Salmonella typhimurium cells with and without S9. Although diketone decursin exhibited less permeability, it is potentially valuable pharmacological compound that should be investigated.

Barakol-induced apoptosis in P19 cells through generation of reactive oxygen species and activation of caspase-9.

ETHNOPHARMACOLOGICAL RELEVANCE: Barakol, an anxiolytic agent isolated from Senna siamea leaves which has been traditionally used for producing natural sleep, has been described as toxic to patients. AIM OF THE STUDY: The aim of current study was to investigate the molecular mechanism of barakol-induced toxicity in mouse embryonal carcinoma P19 cell model. MATERIALS AND METHODS: XTT assay was used to determine cell viability in P19 cells treated with barakol. Apoptotic cells were detected by Hoechst 33342 staining. Intracellular reactive oxygen species (ROS) generation was analyzed by flow cytometry using a fluorescent dye, DCFH-DA. Detection of apoptotic protein expression in P19 cells was performed by Western blot analysis. Caspase-9 activity was measured using a fluorescent immunosorbent enzyme assay kit. RESULTS: Treatment with barakol decreased cell viability in a concentration- and time-dependent manner with an IC(50) value of 1.5mM in 24-h treated cells. A Hoechst 33342 assay revealed that barakol cytotoxicity was due to a significant increase in the number of apoptotic cells. Different scavengers to characterize ROS were utilized and revealed that hydroxyl radicals played a major role in ROS-induced apoptosis in barakol-treated cells. Western blot analysis demonstrated that barakol-induced apoptosis was mediated by the increase in expression ratio of Bax/Bcl-2. Furthermore, increase in caspase-9 activity after exposure to barakol for 24h was also observed. Pretreatment of cells with N-acetyl-l-cysteine (NAC) attenuated intracellular ROS generation, the Bax/Bcl-2 protein expression, and apoptosis. CONCLUSIONS: The mechanism of barakol-mediated toxicity in P19 cells is mainly associated with the ROS generation, followed by the imbalance of the Bax/Bcl-2 ratio, and caspase-9 activation leading to apoptotic cell death. Pretreatment of cells with NAC could antagonize the toxicity produced by barakol.

Phytotoxic activities of (2R)-6-hydroxytremetone.

Benzofurans are bioactive compounds isolated from the Asteraceae family. Benzofuran derivatives have been extensively studied because of their toxic effects on humans and animals. The phytotoxic activity of the benzofuran derivative (2R)-6-hydroxytremetone was studied on germination, seedling development, and cytotoxic and genotoxic effects on monocotyledoneous (onion and wheat) and dicotyledoneous (lettuce and tomato) species. Results of these assays demonstrated that (2R)-6-hydroxytremetone is a potent germination inhibitor of onion, lettuce, and tomato seeds. Germination reductions of approximately 80% were measured when seeds were exposed to 100 mg l(-1) of the compound, and showed considerably effects on the posterior development of the sprouts, including rootlets and hypocotyl elongations. Moreover, this benzofuran derivative also significantly reduced the root length and mitotic division of Allium cepa bulbs, although DNA damages were not observed. Our findings suggest that a mechanism of mitosis inhibition may play a role in the phytotoxic effects of plants producing these compounds.

Acute toxicity, biochemical and gene expression responses of the earthworm Eisenia fetida exposed to polycyclic musks.

AHTN (Tonalide) and HHCB (galaxolide) are recognized as ubiquitous contaminants in soil and have potential adverse impacts on soil organisms. The aim of this study is to investigate the effects of AHTN and HHCB on the earthworm (Eisenia fetida) as an important soil animal with attention to the acute toxicity, biochemical and transcriptional changes of representative antioxidant enzymatic (SOD, CAT) and stress-response gene (Hsp70). The 48 h-LC(50) value was 20.76 µg cm(-2) for AHTN and 11.87 µg cm(-2) for HHCB respectively in the acute lethal studies. The time-dependent elevation in the level of malondialdehyde (MDA) suggests that reactive oxygen species (ROS)-induced cellular oxidative injury of E. fetida might be one of the main toxic effects of AHTN and HHCB. SOD and CAT were both up-regulated at low exposure dose (0.6 µg cm(-2) AHTN and 0.3 µg cm(-2) HHCB) during 48 h testing period, which protected earthworms from oxidative stresses. However, the down-regulation of SOD and CAT after 48 h exposure to high dose contaminants might be caused by the extreme oxidative stress levels (maximum up-regulation 1.70-fold and 1.40-fold for MDA levels at 6.0 µg cm(-2) AHTN and 3.0 µg cm(-2) HHCB compared to the controls, respectively). The Hsp70 gene expression did not show variation during 48 h, except that it had a significant down-regulation (P<0.05) after 48 h of exposure to high doses of contaminants. These results showed that the dermal contact of AHTN and HHCB could result in pronounced biochemical and physiological responses to earthworms, and the transcriptional level changes in antioxidant genes could be potential molecular biomarkers for the stress of the pollutants.

Genotoxicity evaluation of three benzopyrans from Hypericum polyanthemum.

In the present study we used the micronuclei test and the comet assay in mice to investigate the genotoxic and mutagenic effects of three benzopyrans--6-isobutyryl-5,7-dimethoxy-2,2-dimethyl-benzopyran (HP1); 7-hydroxy-6-isobutyryl-5-methoxy-2,2-dimethyl-benzopyran (HP2); and 5-hydroxy-6-isobutyryl-7-methoxy-2,2-dimethyl-benzopyran (HP3)--isolated from Hypericum polyanthemum. No significant difference in polychromatic erythrocyte (PCE) percentage between the vehicle group and groups treated with HP1, HP2, or HP3 was detected, indicating no toxicity to the bone marrow of the animals. Only HP1 increased the frequency of micronucleated cells (MNPCEs) in bone marrow sampled after 24 h in comparison with the vehicle group, suggesting a weak mutagenic effect. The damage index and damage frequency did not show a significant increase after treatment with HP1, HP2, or HP3 in comparison with the vehicle group. The antitumor activity previously reported in vitro for these benzopyrans, the lack of acute toxicity, the MN induction only for HP1, and the relatively low DNA damage make all compounds good candidates for in vivo studies on antitumor action.

Synthesis of novel substituted 7-(benzylideneamino)-4-methyl-2H-chromen-2-one derivatives as anti-inflammatory and analgesic agents.

A series of novel Schiff' base-containing a 7-amino-4-methylcoumarin moiety have been synthesized III a-l, characterized by spectroscopic data and studied for their anti-inflammatory and analgesic activity. The results of the anti-inflammatory and analgesic activity evaluation of 7-(substituted benzylideneamino)-4-methyl-2H-chromen-2-one derivatives III a-l proved to be comparable or more potent with respect to the reference drugs. In particular, compounds 7-(4-chlorobenzylideneamino)-4-methyl-2H-chromen-2-one III f, 7-(2,4-dichlorobenzylideneamino)-4-methyl-2H-chromen-2-one III g and 7-(4-bromobenzylideneamino)-4-methyl-2H-chromen-2-one III h exhibited potent anti-inflammatory and analgesic activity.

Synthesis and pharmacological activities of some condensed 4-chloro-2,2-dialkyl chromene-3-carbaldehyde derivatives.

Some new hydrazono 5a,b, thiosemicarbazono 6a-c, and oximo chromenes 7a-c were prepared via the reaction of the corresponding beta-chlorocarbaldehyde 3 with hydrazine, aromatic hydrazine, thiosemicarbazide and hydroxylamine hydrochloride, respectively. In addition, ether derivatives 8a-h were prepared from the corresponding aldoximes 7a-c. The new products were tested for anti-inflammatory and ulcerogenic score activities compared to indomethacin.

Barakol contents in fresh and cooked Senna siamea leaves.

Senna siamea (Lam.) Irwin and Barneby is a medicinal plant popularly used in Thailand. Young leaves and/or young flowers of this plant have been consumed by Thai people as a Khi Lek curry for a long time. The fresh young leaves and flowers are boiled with water 2-3 times to get rid of the bitterness and the boiled mush is used for curry cooking. Barakol, a major constituent of Senna siamea leaves was analyzed for its content in the fresh young leaves, the boiled leaves and the boiled filtrates by a high-performance thin-layer chromatographic method. Fresh young leaves of S. siamea contained 0.4035% w/w barakol. The amount of barakol in the first and second boiled filtrates were 0.2052 and 0.1079% fresh weight, while the first and second boiled leaves samples were 0.1408 and 0.0414% fresh weight, respectively. The results show the process of preparation of Khi Lek curry by boiling S. siamea young leaves twice with water reduced barakol content up to 90% and the content of barakol in boiled leaves used for curry has much less tendency to cause liver toxicity. This may explain the reason why Thai Khi Lek curry has not caused hepatotoxicity, unlike S. siamea leaves consumed as a powdered capsule.

Cytotoxic flavonoids and new chromenes from Ficus formosana f. formosana.

Two new chromenes, ficuformodiol A and ficuformodiol B, and nine known compounds including one chromene, spatheliachromen, six flavonoids, obovatin, carpachromene ( 5), apigenin ( 6), norartocarpetin ( 7), steppogenin, 6-prenylpinocembrin, one benzopyran, chromenylacrylic acid, and one isocoumarin, ( R)-(-)-mellein, were obtained from the cytotoxic chloroform-soluble fraction of the stems of Ficus formosana f. formosana (Moraceae). Their structures were determined by means of spectroscopic analyses. Compounds 5, 6 and 7 exhibited significant cytotoxicity against HepG2, PLC/PRF/5 and Raji cancer cell lines in vitro.

High prevalence of potentially hepatotoxic herbal supplement use in patients with fulminant hepatic failure.

HYPOTHESIS: The use of potentially hepatotoxic herbal and dietary supplements is highly prevalent in the fulminant hepatic failure (FHF) patient population at our institution, and this subgroup of patients has a worse prognosis. DESIGN: Retrospective case series. Settings An adult tertiary care university hospital and a Veterans Affairs hospital in Oregon. PATIENTS: All patients referred to the liver transplantation service for FHF from January 2001 through October 2002 (N = 20). We defined FHF as onset of encephalopathy within 8 weeks of onset of jaundice in the absence of preexisting liver disease. All patients underwent investigation for potential causes of liver injury. Potentially hepatotoxic supplements were defined as those with previously published reports of hepatic injury related to their use. RESULTS: Ten patients (50%) were recent or active users of potentially hepatotoxic supplements or herbs; 10 had no history of supplement use. In the supplement group, 7 patients (35%) had no other identified cause for hepatic failure. Six patients in the supplement group and 2 patients in the nonsupplement group underwent orthotopic liver transplantation. Five patients in each group died. There were no significant differences in transplantation rate (P =.07) or survival (P>.99) between groups. Supplement use alone accounted for the most cases of FHF during this period, exceeding acetaminophen toxicity and viral hepatitis. CONCLUSIONS: Herbal and dietary supplements were potential hepatotoxins in a high proportion of patients with FHF at our institution. Enhanced public awareness of the potential hepatotoxicity of these commonly used agents and increased regulatory oversight of their use is strongly urged.

Antibacterial and cytotoxic activity of prenylated bicyclic acylphloroglucinol derivatives from Hypericum amblycalyx.

Two new bicyclic acylphloroglucinol derivatives, hypercalyxone A (1-[5,7-dihydroxy-2-methyl-3-(3-methyl-but-2-enyl)-2-(4-methyl-pent-3-enyl)-chroman-8-yl]-2-methyl-propan-1-one, 1) and B (1-[5,7-dihydroxy-2-methyl-3-(3-methyl-but-2-enyl)-2-(4-methyl-pent-3-enyl)-chroman-8-yl]-2-methyl-butan-1-one, 2), have been isolated from the petroleum ether extract of the aerial parts of Hypericum amblycalyx, together with two further compounds (1-[5,7-dihydroxy-2-methyl-2-(4-methyl-pent-3-enyl)-chroman-8-yl]-2-methyl-propan-1-one, 3 and 1-[5,7-dihydroxy-2-methyl-2-(4-methyl-pent-3-enyl)-chroman-8-yl]-2-methyl-butan-1-one, 4), which have been described only as semi-synthetic products. In addition, the known triterpene lup-20(29)-en-3-one was obtained. Structure elucidation was based on 1D and 2D NMR studies, as well as on data derived from mass spectrometry. The four acylphloroglucinol derivatives were evaluated for their cytotoxic and antibacterial activity. All compounds showed moderate cytotoxic activity against KB and Jurkat T cancer cells. Especially compounds 3 and 4 exhibited a strong antibacterial activity against different Gram-positive strains.

Repinotan Bayer.

Bayer is developing the aminomethylchroman derivative, repinotan, a 5-HT1A agonist, as a potential treatment for ischemic stroke and traumatic brain injury [216172]. It has completed phase III trials in Canada and the US [342562], [344747]. In June 2001, NDA filing was expected by 2004 [411649]. As of March 2002, it was undergoing phase III trials for both indications, and Bayer expected to launch the product in 2006 in Europe and the US [443846]. Repinotan was being developed initially as an i.v. formulation for the treatment of ischemic stroke and brain injury and as an oral formulation for the treatment of depression; however, development has been discontinued for the latter indication [317452]. In March 2002, Lehman Brothers predicted a launch date of 2006, estimating peak sales of US $1000 million and a 20 to 30% probability of reaching market [450456], while in April 2002, Bank Vontobel also predicted a launch in 2006 with potential sales of Euro450 million in its fifth year of sales [450454]. Bayer predicts peak sales of Euro450 million [397137].

The potential anti-migraine compound SB-220453 does not contract human isolated blood vessels or myocardium; a comparison with sumatriptan.

The mechanistically novel benzopyran derivative SB-220453, which is undergoing clinical evaluation in migraine, exhibits a high affinity for a selective, but not yet characterized, binding site in the human brain. It inhibits nitric oxide release and cerebral vasodilatation following cortical spreading depression as well as carotid vasodilatation induced by trigeminal nerve stimulation in the cat. The aim of our study was to investigate the contractile properties of SB-220453 on a number of human isolated blood vessels (coronary artery, saphenous vein and middle meningeal artery) as well as atrial and ventricular cardiac trabeculae. While sumatriptan induced marked contractions in three blood vessels investigated, SB-220453 was devoid of any effect. In atrial and ventricular cardiac trabeculae, neither SB-220453 nor sumatriptan displayed a positive or negative inotropic effect. Since SB-220453 did not contract the middle meningeal artery, we conclude that potential anti-migraine effects are not mediated via a direct cerebral vasoconstriction. The lack of activity of SB-220453 in coronary artery, saphenous vein and cardiac trabeculae demonstrates that the compound is unlikely to cause adverse cardiac side-effects.