Studying the effects of secondary metabolites isolated from Cycas thouarsii R.Br. leaves on MDA-MB-231 breast cancer cells.

The various therapeutic drugs that are currently utilized for the management of cancer, especially breast cancer, are greatly challenged by the augmented resistance that is either acquired or de novo by the cancer cells owing to the long treatment periods. So, this study aimed at elucidating the possible anticancer potential of four compounds 7, 4', 7'', 4'''-tetra-O-methyl amentoflavone, hesperidin, ferulic acid, and chlorogenic acid that are isolated from Cycas thouarsii leaves n-butanol fraction for the first time. The MTT assay evaluated the cytotoxic action of four isolated compounds against MDA-MB-231 breast cancer cells and oral epithelial cells. Interestingly, ferulic acid revealed the lowest IC50 of 12.52 µg/mL against MDA-MB-231 cells and a high IC50 of 80.2 µg/mL against oral epithelial cells. Also, using an inverted microscope, the influence of ferulic acid was studied on the MDA-MB-231, which revealed the appearance of apoptosis characteristics like shrinkage of the cells and blebbing of the cell membrane. In addition, the flow cytometric analysis showed that the MDA-MB-231 cells stained with Annexin V/PI had a rise in the count of the cells in the early and late apoptosis stages. Moreover, gel electrophoresis detected DNA fragmentation in the ferulic acid-treated cells. Finally, the effect of the compound was tested at the molecular level by qRT-PCR. An upregulation of the pro-apoptotic genes (BAX and P53) and a downregulation of the anti-apoptotic gene (BCL-2) were observed. Consequently, our study demonstrated that these isolated compounds, especially ferulic acid, may be vital anticancer agents, particularly for breast cancer, through its induction of apoptosis through the P53-dependent pathway.

Phytochemical analysis, in silico study and toxicity profile of Cycas pectinata Buch.-Ham seed in mice.

Fruit of Cycas pectinata Buch.-Ham has been used as medicine by the local community in some parts of the north eastern state of India. Despite its uses for different purposes, the safety assessment study has not been conducted. Therefore, we have evaluated the acute and the sub-acute toxicity of methanolic extract of C. pectinata fruit (CPFE) in a mice model via oral route of administration. Phytochemicals analysis was carried out by liquid chromatography-mass spectroscopy (LC-MS), nuclear magnetic resonance (NMR), and Fourier-transform infrared spectroscopy (FTIR). The acute toxicity study was performed at a single dose of 1000, 3000 and 5000 mg/kg and the sub-acute toxicity study at a dose of 100, 300 and 500 mg/kg was administered daily for 28 days. The calculated Lethal dose 50 (LD50) of CPFE was found to be 4000 mg/kg. Both acute and sub-acute studies showed that 5000 mg/kg and 500 mg/kg dose was toxic to the mice. The results of acute toxicity showed CPFE could have a mild toxic effect on the kidney at a dose of 3000 and 5000 mg/kg, as some deteriorated changes in the kidney along with increase creatinine levels were observed. Acute toxicity also showed an increase in white blood cells (WBC) at a dose of 3000 mg/kg.However, sub-acute toxicity studies do not show any detrimental effects on liver, kidney and hematological parameters. Thus, it can be suggested that CPFE at a dose of 100 and 300 mg/kg would be safe for consumption. The phytochemicals analysis by LC-MS, NMR and FTIR showed the presence of 32 major chemical compounds with certain biological activity like anti-neoplastic, antioxidant, and possible modulator of steroid metabolism (cholesterol antagonist and agonist of testosterone 17ß-dehydrogenase) as predicted by PASS analysis.

Cytology-based Cancer Surgery of the Head and Neck (CyCaS-HN): a prospective, randomized, controlled clinical trial.

PURPOSE: Liquid-based cytology (LBC) is routinely used in gynecology but is rarely applied in head and neck oncology though many suspicious lesions are easily accessible. While several studies have evaluated the potential use of LBC for early detection and molecular characterization of head and neck squamous cell carcinomas (HNSCCs), no study investigated its potential role in surgical management and therapy planning so far. METHODS: Twenty-five patients with cT1-2 squamous cell carcinomas of the oral cavity and oropharynx were prospectively enrolled in this study and were randomized to two treatment arms: in the control arm, a diagnostic panendoscopy with incisional biopsy was followed by a second operation with transoral tumor resection ± neck dissection and tracheostomy. In the intervention arm, patients underwent LBC diagnostics and in case of a positive result received one single operation with panendoscopy and incisional biopsy for confirmation of LBC result by rapid section histology followed by transoral tumor resection ± neck dissection and tracheostomy in the same session. RESULTS: Time between clinical diagnosis and definitive surgical treatment was significantly shorter in the intervention group compared with the control group (p < 0.0001). Additionally, time of hospitalization (p < 0.0001) and cumulative operation time (p = 0.062) were shorter in the intervention group. No significant differences in overall, progression-free, and disease-specific survival were observed. CONCLUSION: Cytology-based cancer surgery is a promising therapeutic strategy that can potentially be considered for a well-defined group of early-stage HNSCC patients and help to avoid repetitive general anesthesia, shorten the diagnosis-to-treatment interval and spare operation as well as hospitalization time.

Population genetics of cycas necrotic stunt virus and the development of multiplex RT-PCR diagnostics.

Cycas necrotic stunt virus (CNSV) has an extensive host range and is detected in an accelerated pace around the globe in several agricultural crops. One of the plant species affected is peony (Paeonia lactiflora Pall.). The virus is asymptomatic in most peony cultivars, but there have been reports of symptoms in others. It is thus important to study CNSV and its population structure to gain insights into its evolution and epidemiology. The outputs of this study, in addition to the in-depth analysis of the virus population structure, include the development of a multiplex RT-PCR detection protocol that can amplify all published CNSV isolate sequences; allowing for accurate, reliable detection of the virus and safeguarding its susceptible, clonally-propagated hosts.

Polyphenol fingerprinting and hypoglycemic attributes of optimized Cycas circinalis leaf extracts.

BACKGROUND: Cycas circinalis leaves are used to treat diabetes mellitus in local medicinal systems without any scientifically proved information on their medicinal potential and phytochemicals. In this study, the total phenolic contents, total flavonoid contents, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging and inhibitory effects on α-glucosidase and α-amylase were determined for optimized hydroethanolic leaf extracts. Secondary metabolites were identified using ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS/MS). In vivo studies on diabetic albino mice were also carried out to evaluate the impact of the most active extract on their blood glucose levels. RESULTS: The 60% ethanolic extract showed the highest extract yield (209.70 ± 0.20 g kg-1 ) and total phenolic (154.24 ± 3.28 mg gallic acid equivalent) and flavonoid (78.52 ± 1.65 mg rutin equivalent per gram dried extract) contents and exhibited the maximum DPPH scavenging activity (IC50 = 59.68 ± 2.82 µg mL-1 ). The IC50 values for inhibition of α-glucosidase (58.42 ± 2.22 µg mL-1 ) and α-amylase (74.11 ± 1.70 µg mL-1 ) were also significant for the 60% ethanolic extract. The untargeted UHPLC-QTOF-MS/MS-based metabolite profiling confirmed the presence of iridoid glucoside, gibberellin A4, O-ß-d-glucosyl-4-hydroxy-cinnamate, 3-methoxy-2-phyenyl-4H-furo[2,3-h]chromen-4-one, kaempferol, withaferin A, amentoflavone, quercitin-3-O-(6″-malonyl glucoside), ellagic acid, and gallic acid. Plant extract at a dose of 500 mg kg-1 body weight reduced the blood glucose level by a considerable extent and also improved the lipid profile of diabetic mice after a 28-day trial. CONCLUSION: The findings revealed the medicinal potential of C. circinalis leaves to treat diabetes mellitus and provided the nutraceutical leads for functional food development. © 2020 Society of Chemical Industry.

The synergistic effects of cinnamon essential oil and nano TiO2 on antimicrobial and functional properties of sago starch films.

This study describes a sago starch-based film by incorporation of cinnamon essential oil (CEO) and nano titanium dioxide (TiO2-N). Different concentrations (i.e., 0%, 1%, 3%, and 5%, w/w) of TiO2-N and CEO (i.e., 0%, 1%, 2%, and 3%, v/w) were incorporated into sago starch film, and the physicochemical, barrier, mechanical, and antimicrobial properties of the bionanocomposite films were estimated. Incorporation of CEO into the sago starch matrix increased oxygen and water vapor permeability of starch films while increasing TiO2-N concentration decreased barrier properties. Moisture content also decreased from 12.96% to 8.04%, solubility in water decreased from 25% to 13.7%, and the mechanical properties of sago starch films improved. Sago starch bionanocomposite films showed excellent antimicrobial activity against Escherichia coli, Salmonella typhimurium, and Staphylococcus aureus. Results also showed that incorporation of TiO2-N and CEO had synergistic effects on functional properties of sago starch films. In summary, sago starch films incorporated with both TiO2-N and CEO shows potential application for active packaging in food industries such as fresh pistachio packaging.

Metabolite profiling and in-vitro colon cancer protective activity of Cycas revoluta cone extract.

The methanolic extract of Cycas revoluta cone (MECR) was analyzed by GC-MS and UHPLC for metabolite profiling and was evaluated for anti-colon cancer property by using in vitro assays like Cell Viability Assay, Colony Formation Assay, ROS Determination, Flowcytometry, DAPI staining assay, Tunel assay. GC-MS and HPLC analysis confirmed the presence of different phytochemicals in the extract of Cycas revoluta cone. In-vitro studies showed MECR extract showed significant anti-colon cancer activity by reducing proliferation and inducing apoptosis in colon cancer cell (HCT-8) line, but no such activity was seen in normal colon cell (CCD-18Co) line. The investigation confirms that MECR may be a promising candidate in colon cancer protection.

Extract of Cycas revoluta Thunb. enhances the inhibitory effect of 5-fluorouracil on gastric cancer cells through the AKT-mTOR pathway.

BACKGROUND: Gastric cancer is one of the most common and deadly malignancies worldwide. Despite recent medical progress, the 5-year survival rate of gastric cancer is still unsatisfactory. 5-fluorouracil (5-Fu) is one of the first-line antineoplastic treatments for gastric cancer, as it can effectively induce cancer cell apoptosis. However, the effect of 5-Fu is limited due to drug resistance of the malignant tumor. Previous studies have reported that Sotetsuflavone from Cycas revoluta Thunb. can markedly suppress lung cancer cell proliferation by apoptosis, though its effect on gastric cancer remains unknown. AIM: To investigate the inhibitory effect of Cycas revoluta Thunb. and to determine whether it can overcome gastric cancer cell drug resistance to 5-Fu. METHODS: Cell viability was examined to determine whether the natural extract of Cycas revoluta Thunb. induced gastric cancer cell death. The half-maximal effective concentration and the half-maximal lethal concentration were calculatede. Wound-healing and transwell assays were performed to examine gastric cancer cell motility. Clonogenic assays were performed to investigate the synergistic effects of Cycas revoluta Thunb. with 5-Fu, and apoptotic bodies were detected by Hoechst staining. Western blotting was performed to examine the expression of related proteins and to investigate the molecular mechanism of Cycas revoluta Thunb.-induced cancer cell apoptosis. The expressions of proteins, including mammalian target of rapamycin (mTOR) and p-AKT, were detected in different combinations of treatments for 48 h, then analyzed by ECL detection. RESULTS: Gastric cancer cells were more sensitive to the natural extract of Cycas revoluta Thunb. compared to normal gastric epithelial cells, and the extract effectively inhibited gastric cancer cell migration and invasion. The extract improved the anti-cancer effect of 5-Fu by enhancing the chemosensitization of gastric cancer cells. Extract plus 5-Fu further reduced the expression of the drug-resistance-related proteins p-AKT and mTOR after 48 h compared to 5-Fu alone. Compared to 5-Fu treatment alone, mTOR and p-AKT expression was significantly reduced by about 50% and 75%, respectively. We also found that the natural extract of Cycas revoluta Thunb. further increased 5-Fu-induced gastric cancer cell apoptosis. Expression of apoptosis-related protein X-linked inhibitor of apoptosis protein and apoptosis inducing factor were significantly reduced and increased, respectively, in the 5-Fu-resistant gastric cancer line SGC-7901/R treated with extract plus 5-Fu, while the expression of survivin did not change. CONCLUSION: The natural extract of Cycas revoluta Thunb. effectively inhibited gastric cancer cell growth and enhanced the anti-cancer effect of 5-Fu through the AKT-mTOR pathway.

Sotetsuflavone inhibits proliferation and induces apoptosis of A549 cells through ROS-mediated mitochondrial-dependent pathway.

BACKGROUND: Sotetsuflavone is isolated from Cycas revoluta Thunb., which has biological activity against tumors. However, the anti-proliferative effects of sotetsuflavone on A549 cells and its mechanism are not fully elucidated. METHODS: This study investigated the mechanisms of growth inhibition, cell cycle arrest and apoptosis in non-small cell lung cancer A549 cells induced by sotetsuflavone and evaluated whether sotetsuflavone can be safely utilized by humans as therapeutic agent. RESULTS: We found that sotetsuflavone had significant antiproliferative activity against A549 cells. At the same time, the reactive oxygen species (ROS) content increased while the mitochondrial membrane potential and the ratio of Bcl-2/Bax decreased. Cleaved caspase-3, cleaved caspase-9, cytochrome C and Bax expression increased, and Cyclin D1, CDK4, cleaved caspase-8 and Bcl-2 expression decreased. Interestingly, we demonstrated that sotetsuflavone could effectively inhibit the G0/G1 cycle progression, and then induce the endogenous apoptosis pathway. Our results show that sotetsuflavone could inhibit the growth of A549 cells by up-regulating intracellular ROS levels and causing the mitochondrial membrane potential to collapse, inducing G0/G1 phase arrest and endogenous apoptosis. CONCLUSIONS: In short, we confirm that sotetsuflavone had an inhibitory effect on A549 cells and discovered that it causes apoptosis of A549 lung cancer cells. Sotetsuflavone may be used as a novel candidate for anti-tumor therapy in patients with lung cancer.

Recovery of Palm Oil and Valuable Material from Oil Palm Empty Fruit Bunch by Sub-critical Water.

Oil palm empty fruit bunch (EFB) is one of the solid wastes produced in huge volume by palm oil mill. Whilst it still contains valuable oil, approximately 22.6 million tons is generated annually and treated as solid waste. In this work, sub-critical water (sub-cw) was used to extract oil, sugar and tar from spikelet of EFB. The spikelet was treated with sub-cw between 180-280°C and a reaction time of 2 and 5 minutes. The highest yield of oil was 0.075 g-oil/g-dry EFB, obtained at 240°C and reaction time of 5 minutes. Astonishingly, oil that was extracted through this method was 84.5% of that obtained through Soxhlet method using hexane. Yield of oil extracted was strongly affected by the reaction temperature and time. Higher reaction temperature induces the dielectric constant of water towards the non-polar properties of solvent; thus increases the oil extraction capability. Meanwhile, the highest yield of sugar was 0.20 g-sugar/g-dry EFB obtained at 220°C. At this temperature, the ion product of water is high enough to enable maximum sub-critical water hydrolysis reaction. This study showed that oil and other valuable material can be recovered using water at sub-critical condition, and most attractive without the use of harmful organic solvent.

Hepatoprotective activity of hepatoplus on isonaizid and rifampicin induced hepatotoxicity in rats.

Present study deals with the hepatoprotective activity of polyherbal formulation Hepatoplus (HP) as an oral supplement to the INH and RIF induced hepatitis in experimental rats. Rats treated with INH and RIF show abnormal liver function with significant increase in serum transaminases, bilirubin and clotting time (CT) and significant decrease in total protein and Albumin, which is brings to near normal levels by HP and LIV 52 treatments. Rats treated with INH and RIF suffer from oxidative stress in the hepatocytes, due to the decrease in Glutathione (GSH), Glutathione peroxidase (GPX), Catalase (CAT), Super oxide dismutase (SOD) and significant increase in Lipid Per oxidation (LPO). HP decreases the oxidative stress and protects the liver cells membrane from LPO. 85% of DNA damage (comet tail) seen with RIF and INH treatment is reduced to 34.1% on HP application. A decrease of hepatocytes mitochondrial dehydrogenase activity is observed in INH and RIF treatment is restored by HP supplementation. Hepatic apoptotic and CYP2E1 gene expressions were also studied, BAX, p53, Caspase 3 and CYP2E1 were significantly up regulated and Bcl2 was down- regulated in INH and RIF treated rats. Concomitant application of HP prevents the modulation of these gene expressions. It is concluded that high dose of HP (100mg/kg) supplemented along with INH and RIF effectively prevents the toxicity induced by INH and RIF, as effective as 100mg/kg of LIV52.

Crystal structures and inhibitor binding properties of plant class V chitinases: the cycad enzyme exhibits unique structural and functional features.

A class V (glycoside hydrolase family 18) chitinase from the cycad Cycas revoluta (CrChiA) is a plant chitinase that has been reported to possess efficient transglycosylation (TG) activity. We solved the crystal structure of CrChiA, and compared it with those of class V chitinases from Nicotiana tabacum (NtChiV) and Arabidopsis thaliana (AtChiC), which do not efficiently catalyze the TG reaction. All three chitinases had a similar (α/ß)8 barrel fold with an (α + ß) insertion domain. In the acceptor binding site (+1, +2 and +3) of CrChiA, the Trp168 side chain was found to stack face-to-face with the +3 sugar. However, this interaction was not found in the identical regions of NtChiV and AtChiC. In the DxDxE motif, which is essential for catalysis, the carboxyl group of the middle Asp (Asp117) was always oriented toward the catalytic acid Glu119 in CrChiA, whereas the corresponding Asp in NtChiV and AtChiC was oriented toward the first Asp. These structural features of CrChiA appear to be responsible for the efficient TG activity. When binding of the inhibitor allosamidin was evaluated using isothermal titration calorimetry, the changes in binding free energy of the three chitinases were found to be similar to each other, i.e. between -9.5 and -9.8 kcal mol(-1) . However, solvation and conformational entropy changes in CrChiA were markedly different from those in NtChiV and AtChiC, but similar to those of chitinase A from Serratia marcescens (SmChiA), which also exhibits significant TG activity. These results provide insight into the molecular mechanism underlying the TG reaction and the molecular evolution from bacterial chitinases to plant class V chitinases.

Nuclear ribosomal ITS functional paralogs resolve the phylogenetic relationships of a late-Miocene radiation cycad Cycas (Cycadaceae).

Cycas is the most widespread and diverse genus among the ancient cycads, but the extant species could be the product of late Miocene rapid radiations. Taxonomic treatments to date for this genus are quite controversial, which makes it difficult to elucidate its evolutionary history. We cloned 161 genomic ITS sequences from 31 species representing all sections of Cycas. The divergent ITS paralogs were examined within each species and identified as putative pseudogenes, recombinants and functional paralogs. Functional paralogs were used to reconstruct phylogenetic relationships with pseudogene sequences as molecular outgroups, since an unambiguous ITS sequence alignment with their closest relatives, the Zamiaceae, is unachievable. A fully resolved and highly supported tree topology was obtained at the section level, with two major clades including six minor clades. The results fully supported the classification scheme proposed by Hill (2004) at the section level, with the minor clades representing his six sections. The two major clades could be recognised as two subgenera. The obtained pattern of phylogenetic relationships, combined with the different seed dispersal capabilities and paleogeography, allowed us to propose a late Miocene rapid radiation of Cycas that might have been promoted by vicariant events associated with the complex topography and orogeny of South China and adjacent regions. In contrast, transoceanic dispersals might have played an important role in the rapid diversification of sect. Cycas, whose members have evolved a spongy layer in their seeds aiding water dispersals.

Two new dihydroamentoflavone glycosides from Cycas revoluta.

Phytochemical investigation of the ethyl acetate extract of Cycas revoluta Thunb. leaflets afforded five compounds including two new dihydroamentoflavone glucosides, (2S)-I-(2,3)-dihydro-I-7-O-ß-D-glucopyranosylamentoflavone (1) and (2S)-I-(2,3)-dihydro-I-7,II-7-di-O-ß-D-glucopyranosylamentoflavone (2), in addition to the known compounds prunin (3), vitexin-2″-rhamnoside (4) and protocatechuic acid (5). Compounds (3) and (4) being reported for the first time in this plant. The structures of these compounds were established by the detailed analysis of their spectroscopic data, mainly 1D NMR, 2D NMR, CD and HR-MSD-TOF. The ethyl acetate extract showed weak cytotoxicity against HepG2 (IC50 = 207.6 µg/mL) and RAW 264.2 cells (IC50 = 160.8 µg/mL). Compound 4 showed significant activity towards Leishmania donavani (IC50 = 13.8 µM, IC90 = 34.6 µM). The isolated compounds showed weak antimicrobial activity (IC50>10 µg/mL).

Estrategia estatal de conservación de las cícadas (Zamiaceae): una propuesta para el estado de Hidalgo, México / State strategy for Cycad (Zamiaceae) conservation: A proposal for the State of Hidalgo, Mexico

Mexico has the second largest cycad diversity in the world, and the Sierra Madre Oriental (SMO) is one of the richest biogeographic regions for these plants. Despite there is a general Cycad National Program in the country, there are no state-level cycad conservation strategies or programs. Thus the aim of this study was to propose a cycad conservation strategy for the state of Hidalgo, which is located in the Southern part of the SMO. For this, a cycad species inventory was made in the state, for which three methods were used: review of published literature; consultation in the main Mexican herbaria to verify botanical specimens; and exhaustive field research to compare findings with previously reported species and to recognize new records at the county and state level. The proposed research work strategy combined the following elements: prioritize the county and local areas with greatest cycad species richness; prioritize the species least resistant to environmental change and/or having restricted geographic distribution; and to consider the main uses of these plants by local residents. The results showed that Hidalgo has three genera and eight species of cycads: Ceratozamia fuscoviridis, C. latifolia, C. mexicana, C. sabatoi, Dioon edule, Zamia fischeri, Z. loddigesii and Z. vazquezii, all of which are considered by the International Union for Conservation of Nature (IUCN). This study added two new species records for Hidalgo and 21 at the county level. The species are distributed in 26 counties, of which Chapulhuacán and Pisaflores are notable for their high species richness. Hidalgo has the fourth-greatest cycad species richness among Mexican states, although its area accounts for only 1.07% of the country. The state’s diversity is greater than in other states with larger area, and even than in some other entire countries in Mesoamerica. The presented state cycad conservation strategy proposes that a total of some 11 325ha to be conserved in nine zones, including different vegetation types, distributed in seven counties. The strategy involves a mixed scheme that incorporates conservation in Protected Natural Areas (ANP), Small Farmer Reserves (Reservas Campesinas) and Environmental Management Units (UMA in Spanish). This proposal will be useful for government agencies to take into account in the process of designating land use for the Cloud Forest Biological Corridor (CBBMM in Spanish), a ANP in creation. The state of Hidalgo urgently needs a detailed analysis of trends in changes in vegetation cover and land use, and demographic studies of the cycads. It is recommended that the implementation phase of this state strategy be carried out jointly with local communities, academia, and state and federal agencies responsible for biodiversity conservation.

México es el segundo país con mayor diversidad de cícadas en el mundo. En él, la Sierra Madre Oriental (SMO) es una de las regiones biogeográficas con mayor riqueza de estas plantas. El objetivo de este estudio fue plantear la estrategia de conservación de cícadas para el estado de Hidalgo, ubicado en la parte sur de la SMO; para esto se revisó la literatura científica, se consultaron los principales herbarios de México y se hicieron recorridos de campo. Los resultados muestran que Hidalgo alberga tres géneros y ocho especies: Ceratozamia fuscoviridis, C. latifolia, C. mexicana, C. sabatoi, Dioon edule, Zamia fischeri, Z. loddigesii, Z. vazquezii, posicionándose en el cuarto lugar nacional en riqueza de cícadas. Las especies se distribuyen en 26 municipios. Se encontró que la diversidad de cícadas es mayor a la de otras entidades con mayor superficie e incluso a la de algunos países de Centroamérica. La estrategia estatal de conservación de cícadas aquí desarrollada propone conservar cerca de 11 325ha, distribuidas en siete municipios, bajo un esquema mixto de conservación. Esta propuesta será útil para que las instancias gubernamentales establezcan nuevas ANP’s en la región biogeográfica de la SMO.

Oil palm empty fruit bunch as alternative substrate for acetone-butanol-ethanol production by Clostridium butyricum EB6.

Acetone-butanol-ethanol (ABE) production from renewable resources has been widely reported. In this study, Clostridium butyricum EB6 was employed for ABE fermentation using fermentable sugar derived from treated oil palm empty fruit bunch (OPEFB). A higher amount of ABE (2.61 g/l) was produced in a fermentation using treated OPEFB as the substrate when compared to a glucose based medium that produced 0.24 g/l at pH 5.5. ABE production was increased to 3.47 g/l with a yield of 0.24 g/g at pH 6.0. The fermentation using limited nitrogen concentration of 3 g/l improved the ABE yield by 64%. The study showed that OPEFB has the potential to be applied for renewable ABE production by C. butyricum EB6.

Identification and characterization of a bactericidal and proapoptotic peptide from Cycas revoluta seeds with DNA binding properties.

Nowadays, novel pharmacies have been screened from plants. Among them are the peptides, which show multiple biotechnological activities. In this report, a small peptide (Ala-Trp-Lys-Leu-Phe-Asp-Asp-Gly-Val) with a molecular mass of 1,050 Da was purified from Cycas revoluta seeds by using reversed-phase liquid chromatography. This peptide shows clear deleterious effects against human epidermoid cancer (Hep2) and colon carcinoma cells (HCT15). It caused inhibition of cancer cell proliferation and further disruption of nucleosome structures, inducing apoptosis by direct DNA binding. A remarkable antibacterial activity was also observed in this same peptide. Nevertheless, no significant lysis of normal RBC cells was observed in the presence of peptide. Additionally, an acetylation at the N-termini portion is able to reduce both activities. Bioinformatics tools were also utilized for construction of a three-dimensional model showing a single amphipathic helix. Since in vitro binding studies show that the target of this peptide seems to be DNA, theoretical docking studies were also performed to better understand the interaction between peptide and nucleic acids and also to shed some light on the acetyl group role. Firstly, binding studies showed that affinity contacts basically occur due to electrostatic attraction. The complex peptide-ssDNA was clearly oriented by residues Ala(1), Lys(3), and Asp(6), which form several hydrogen bonds that are able to stabilize the complex. When acetyl was added, hydrogen bonds are broken, reducing the peptide affinity. In summary, it seems that information here provided could be used to design a novel derivative of this peptide which a clear therapeutic potential.

Pathobiochemical effect of acylated steryl-ß-glucoside on aggregation and cytotoxicity of α-synuclein.

Cycad seed consumption by the native islanders of Guam is frequently associated with high rates of amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS/PDC); furthermore, accompanying pathological examination often exhibits α-synuclein inclusions in the neurons of the affected brain. Acylated steryl-ß-glucoside (ASG) contained in cycad seeds is considered as causative environmental risk factor. We aimed to investigate whether ASG influences aggregation and cell toxicity of α-synuclein. To understand whether ASG is a causative factor in the development of ALS/PDC, soybean-derived ASG was tested for its effect on in vitro aggregation of α-synuclein using Thioflavin-T. ASG was also tested to determine whether it modulates α-synuclein cytotoxicity in yeast cells. In addition, we determined whether an interaction between ASG and α-synuclein occurs in the plasma membrane or cytoplasm using three factors: GM1 ganglioside, small unilamellar vesicles, and ATP. In the present study, we found that ASG-mediated acceleration of α-synuclein aggregation is influenced by the presence of ATP, but not by the presence of GM1. ASG accelerated the α-synuclein aggregation in the cytoplasm. ASG also enhanced α-synuclein-induced cytotoxicity in yeast cells. This study demonstrated that ASG directly enhances aggregation and cytotoxicity of α-synuclein, which are often observed in patients with ALS/PDC. These results, using assays that replicate cytoplasmic conditions, are consistent with the molecular mechanism that cytotoxicity is caused by intracellular α-synuclein fibril formation in neuronal cells.

Sleep alterations in an environmental neurotoxin-induced model of parkinsonism.

Parkinson's disease (PD) is classically defined as a motor disorder resulting from decreased dopamine production in the basal ganglia circuit. In an attempt to better diagnose and treat PD before the onset of severe motor dysfunction, recent attention has focused on the early, non-motor symptoms, which include but are not limited to sleep disorders such as excessive daytime sleepiness (EDS) and REM behavioral disorder (RBD). However, few animal models have been able to replicate both the motor and non-motor symptoms of PD. Here, we present a progressive rat model of parkinsonism that displays disturbances in sleep/wake patterns. Epidemiological studies elucidated a link between the Guamanian variant of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC) and the consumption of flour made from the washed seeds of the plant Cycas micronesica (cycad). Our study examined the effects of prolonged cycad consumption on sleep/wake activity in male, Sprague-Dawley rats. Cycad-fed rats exhibited an increase in length and/or number of bouts of rapid eye movement (REM) sleep and Non-REM (NREM) sleep at the expense of wakefulness during the active period when compared to control rats. This hypersomnolent behavior suggests an inability to maintain arousal. In addition, cycad-fed rats had significantly fewer orexin cells in the hypothalamus. Our results reveal a novel rodent model of parkinsonism that includes an EDS-like syndrome that may be associated with a dysregulation of orexin neurons. Further characterization of this early, non-motor symptom, may provide potential therapeutic interventions in the treatment of PD.

Efecto de D-004, un extracto lipídico extraído de las frutas de la palma real (Roystonea regia) sobre las contracciones provocadas por la Fenilefrina sobre la próstata aislada de rata / Effect of D-004, a lipid extract from royal palm (Roystonea regia) fruits, on phenylephrine-induced contractions of isolated rat prostate

D-004, a lipid extract of Roystonea regia fruits, inhibits norepinephrine-elicited contraction in rat vas deferens. Nevertheless, the effect of D-004 on a1-adrenergic contractile responses in isolated prostate preparations had not been studied. Therefore, this study investigated the effects of D-004 on phenylephrine-induced contractions in isolated rat prostate strips. D-004 (125-500 mg/mL) was added to prostate preparations in which contractions were induced with phenylephrine (10-6-10-4 M) or with ClK (50 mM). D-004 (250-500 mg/mL) significantly (p < 0,01).and dose-dependently inhibited the phenylephrine-induced contractions through a non-competitive mechanism, since it reduced (12-60 percent) the maximal contraction to phenyle phrine with respect to control preparations. D-004 at 500 mg/mL inhibited similarly both ClK and phenylephrine-contractile responses. Tamsulosin (0.01 mg/mL) abolished (100 percent inhibition) the phenylephrine, but unaffected the ClK-induced contractions.

D-004, un extracto lipídico de las frutas de Roystonea regia, inhibe la contracción provocada por la norepinefrina en el vas deferens de las ratas. No obstante, el efecto de D-004 sobre las respuestas contráctiles adrenérgicas a1 en las preparaciones aisladas de la próstata no ha sido estudiado. Por lo tanto, este estudio investigó los efectos de D-004 sobre las contracciones provocadas por la fenilefrina en las extracciones aisladas de la próstata de la rata. D-004 (125-500 mg/mL) fue adicionado a las preparaciones a partir de la próstata en la que las contracciones fueron provocadas por la fenilefrina (10-6-10-4 M) o por CIK (50 mM). D-004 (250-500 mg/mL) en forma significativa y la dosis en forma dependiente inhibieron las contracciones provocadas por la fenilefrina mediante un mecanismo no competetitivo, reduciendo así (12-60 por ciento) la contracción máxima por la fenilefrina con respecto a las preparaciones controles. D-004 a 500 µg/mL inhibieron de la misma forma tanto CIK como las respuestas contráctiles-fenilefrina. Tamsulosin (0,01 mg/mL) eliminó la acción de la fenilefrina (100 por ciento de inhibición), pero no afectó las contracciones provocadas por CIK.