Few substitutions affect the bioluminescence spectra of Phrixotrix (Coleoptera: Phengodidae) luciferases: a site-directed mutagenesis survey

Phrixotrix (railroad worm) luciferases produce bioluminescence in the green and red regions of the spectrum, depending on the location of the lanterns, and are the only luciferases naturally producing red bioluminescence. Comparison of the luciferase sequences showed a set of substitutions that could be involved in bioluminescence colour determination: (a) unique substitutions in the red luciferase replacing otherwise invariant residues; (b) conserved basic residues in the green-yellow emitting luciferases; and (c) an additional R353 residue in red-emitting luciferase (Viviani et al., 1999). To investigate whether these sites have a functional role in bioluminescence colour determination, we performed a site-directed mutagenesis. Natural substitutions in the region 220-344 and residues in the putative luciferin-binding site were also investigated. With the exception of the previously identified substitution of R215 and T226 (Viviani et al., 2002), which display dramatic red-shift effects on the spectrum of green-yellow-emitting luciferases, only a few substitutions had a moderate effect on the spectrum of the green-emitting luciferase. In contrast, no single substitution affected the spectrum of the red-emitting luciferase. The results suggest that the identity of the active site residues is not so critical for determining red bioluminescence in PxRE luciferase. Rather, the conformation assumed during the emitting step could be critical to set up proper interactions with excited oxyluciferin. Copyright ©2007 John Wiley & Sons, Ltd.

Síntese e modelagem molecular do novo derivado indolinônico como candidato a antiinflamatório COX-2 seletivo

Anti-inflammatory drugs are known to be the most widely-marketed drugs in the world, despite their serious side effects, mainly on the gastrointestinal tract. Thus, there are constant efforts to discover new prototypes with improved therapeutic activity and safety for the patient. Since the advent of the computational chemistry, the theoretical study of the physiological behavior of a new compound and hence an understanding of its supposed mechanism of action have been made a lot more accessible. Thus, molecule-receptor mathematical modeling was applied to compound I (1-(2,6-dichlorophenyl)indolin-2-one), to predict theoretically its ability to inhibit, selectively, the COX-2 isoform of prostaglandin endoperoxide synthase (PGHS-2), and the best positions to introduce chemical groups and to make molecular modifications.

Pharmacognostical evaluation of fruits of Solanum lycocarpum A. St.-Hill. (Solanaceae)

This study describes the chemical and physical-chemical profile of plant drug and ethanolic extract obtained from fruits of Solanum lycocarpum A. St.-Hill. (Solanaceae). The physical and chemical analysis involved the granulometry determination, non-compacted apparent density, loss on drying in oven and in infrared scale, pH, ash values and extractive values. The results determined the physical-chemical characteristics of the drug plant. It was also carried out the microbiological control of the plant drug. The preliminary phytochemical screening featured the presence of tannins, flavonoids and saponins in the plant drug and alkaloids and steroids in the ethanolic exctract. The solamargine and solasonine glycoalkaloids were identified through TLC and GC/ MS. The levels of total phenols and tannins were quantified in the extract (8.90% and 6,85% respectively). Such studies contribute to the chemical identification and quality control of S. lycocarpum fruits. © 2010 Phcog.net.

Estudo fitoquímico preliminar e bioensaio toxicológico frente a larvas de Artemia salina Leach. de extrato obtido de frutos de Solanum lycocarpum A. St.-Hill (Solanaceae)

The phytochemical profile of ethanolic extract of Solanum lycocarpum fruits was analyzed and preliminary toxicity tests were performed against brine shrimp larvae. The extract was subjected to preliminary phytochemical analysis to identify the main classes of secondary metabolites and tested against the larvae of A. salina to obtain the median lethal concentrations (LC50%). The phytochemical tests showed the presence of phenols, tannins, saponins, alkaloids and free steroids. The extract was fractionated with various solvents for toxicity testing against the larvae and the hydroalcoholic fraction showed considerable cytotoxicity (CL50% = 285.546 g/mL).

Ozonized Oils: A qualitative and quantitative analysis

Most of the problems of endodontic origin have a bacterial etiological agent. Thus, there is a continued interest in seeking more effective chemical substances that can replace the camphorated paramonochiorophenol or antibiotics as intracanal medicaments. Among the possible substances, ozone has some interesting biological characteristics: bactericidal action, debriding effect, angiogenesis stimulation capacity and high oxidizing power. The purpose of this study was to chemically evaluate the presence of ozone in sunflower, castor, olive and almond oil, as well as in propylene glycol and byproducts of ozonation, such as formaldehyde. These compounds were ozonized, inserted into empty and sterile vials, and analyzed by testing the reaction between ozone and indigo, for determining the presence of ozone, and subjected to the chromotropic acid test for determining the presence of formaldehyde. It was observed complete absence of ozone in all samples tested and presence of formaldehyde. The bactericidal and healing action of ozonized oils could be attributed to products formed by the ozonation of mineral oils, such as formaldehyde, not to the ozone itself.

Consideraciones al metodo de Carleton para medir cambios en la fecundidad utilizando informacion proveniente de muestras pequenas

Incluye Bibliografía

Light emission from tryptophan oxidation by hypobromous acid

The emission of ultraweak light from cells is a phenomenon associated with the oxidation of biomolecules by reactive oxygen species. The indole moiety present in tryptophan, serotonin and melatonin is frequently associated with the emission of light during the oxidation of these metabolites. This study presents results for hypobromous acid (HOBr) oxidation of tryptophan as a putative endogenous source of ultraweak light emission. We found that chemiluminescence elicited by the oxidation of tryptophan by HOBr was significantly higher than by hypochlorous acid (HOCl). This difference was related to secondary oxidation reactions, which were more intense using HOBr. The products identified during oxidation by HOCl, but depleted by using HOBr, were N-formylkynurenine, kynurenine, 1,2,3,3a,8,8a-hexahydro-3a-hydroxypyrrolo[2,3-b]-indole-2-carboxylic acid, oxindolylalanine and dioxindolylalanine. The emission of light is dependent on the free α-amino group of tryptophan, and hence, the indole of serotonin and melatonin, although efficiently oxidized, did not produce chemiluminescence. The emission of light was even greater using taurine monobromamine and dibromamine as the oxidant compared to HOBr. A mechanism based on bromine radical intermediates is suggested for the higher efficiency in light emission. Altogether, the experimental evidence described in the present study indicates that the oxidation of free tryptophan or tryptophan residues in proteins is an important source of ultraweak cellular emission of light. This light emission is increased in the presence of taurine, an amino acid present in large amounts in leukocytes, where this putative source of ultraweak light emission is even more relevant.

Phosphorus solubilizing and iaa production activities in plant growth promoting rhizobacteria from brazilian soils under sugarcane cultivation

Plant Growth Promoting Rhizobacteria (PGPR) has been used as a biofertilizer, bringing benefits to agriculture as Phosphorus Solubilizing Bacteria (PSB), indole-acetic acid (IAA) producers, and with other activites. The goal of this report was the identification of PGPR from soils under sugarcane crops by 16S rRNA sequencing, and the evaluation of the ability of phosphorus solubilizing and IAA production by biological assays. The isolates of this work were obtained from three areas of sugarcane crop from São Paulo State, Brazil. All isolates came from rhizosphere soil, and in a total of 60 isolates just 10 have showed high ability in phosphorus solubilizing. The selection of PSB may be done by phenotypic and/or genotypic characterization. Among ten isolates Enterobacter sp. (FJ890899), Entrobacter homaechei subsp. verschuerennii (FJ890998), Burkholderia sp. (FJ890895), and Labrys portucalensis (FJ890891) were able to IAA production. © 2006-2012 Asian Research Publishing Network (ARPN).

On the application of nanostructured electrodes prepared by Ti/TiO2/WO3 template : A case study of removing toxicity of indigo using visible irradiation

New assays with HepG2 cells indicate that Indigo Carmine (IC), a dye that is widely used as additive in many food and pharmaceutical industries exhibited cytotoxic effects. This work describes the development of a bicomponent nanostructured Ti/TiO2/WO3 electrode prepared by template method and investigates its efficiency in a photoelectrocatalytic method by using visible light irradiation and applied potential of 1V. After 2h of treatment there are reduction of 97% discoloration, 62% of mineralization and formation of three byproducts assigned as: 2-amine-5-sulfo-benzoic acid, 2,3-dioxo-14-indole-5-sulfonic acid, and 2-amino-α-oxo-5-sulfo-benzeneacetic acid were identified by HPLC-MS/MS. But, cytotoxicity was completely removed after 120min of treatment. © 2013 Elsevier Ltd.

In vitro assessment of the cytotoxic, apoptotic, and mutagenic potentials of Isatin

Isatin (1H-indole-2,3-dione) is a chemical found in various medicinal plant species and responsible for a broad spectrum of pharmacological and biological properties that may be beneficial to human health, as an anticonvulsant, antibacterial, antifungal, antiviral, and anticancer agent. The aim of the present study was to determine in vitro the cytotoxic, mutagenic, and apoptotic effects of isatin on CHO-K1 and HeLa cells using the MTT viability assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide), micronucleus (MN) test, apoptosis index, and nuclear division index (NDI). The 5 isatin concentrations evaluated in the mutagenicity and apoptosis tests were 0.5, 1, 5, 10, and 50 μM, selected through a preliminary MTT assay. Positive (doxorubicin, DXR) and negative (phosphate buffered saline, PBS) control groups were also included in the analysis. Isatin did not exert a mutagenic effect on CHO-K1 after 3 and 24 h of treatment or on HeLa cells after 24 h. However, 10 and 50 μM concentrations inhibited cell proliferation and promoted apoptosis in both CHO-K1 and HeLa cells. Data indicate that the cytotoxic, apoptotic, and antiproliferative effects of isatin were concentration independent and cell line independent. The authors thank Profa Dra Eiko Nakagawa Itano for the use the spectrophotometer and the Conselho Nacional para o Desenvolvimento Científico e Tecnológico for master's scholarships to P. M. Cândido-Bacani and grants to T. R. Calvo, W. Vilegas, E. A. Varanda and I. M. S. Cólus. The Conselho Nacional para o Desenvolvimento Científico e Tecnológico provided funding for this study. © 2013 Taylor & Francis Group, LLC.

7-Hydroxycassine: A new 2,6-dialkylpiperidin-3-ol alkaloid and other constituents isolated from flowers and fruits of senna spectabilis (Fabaceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effects of liver S9 enzymes on somalargine and solasodine cytotoxicity and mass spectrometric fragmentation

The steroidal glycoalkaloid solamargine and its parent aglycone solasodine, isolated from Solanum palinacanthum, were studied in vitro for cytotoxicity and biotransformation by the hepatic S9 fraction as the metabolic activating system. The MTT uptake assay was used to determine viability after 24 h in RAW 264.7 mouse macrophage-like and SiHa cells exposed to various concentrations of the alkaloids in the presence and absence of the hepatic S9 microsomal fraction. The dose-response curves were established for solamargine and solasodine in the presence and absence of external metabolizing system. From these data, the cytotoxic index (CI50) was calculated with mean values of 7.2 and 13.6 μg/mL for Raw cells and 8.6 and 26.0 μg/mL for SiHa cells, respectively. Mass spectrometry was performed to compare the fragmentation patterns of the alkaloids to predict metabolism by the S9 fraction. The mass spectra demonstrated a distinct fragmentation patterns for solamargine and solasodine after the addition of the S9 fraction. In the present study, we demonstrate that the cytotoxic effect of solamargine and solasodine and their metabolites prepared in vitro by biotransformation with the S9 fraction are comparable. These findings suggest that the metabolic activation system S9 fraction may fail to suppress the cytotoxicity of these alkaloids. © 2013 Springer-Verlag Berlin Heidelberg.

Atividade antimicrobiana in vitro de extrato de Punica granatum L. sobre staphylococcus aureus isolado em leite bovino

Bovine mastitis is considered important disease causing major economic losses in dairy herds. Antimicrobial drug can promote resistance, chemical residues in food and environmental contamination. The purpose of this study was to evaluate the antibacterial activity in vitro of pomegranate extract on Staphylococcus aureus isolated from bovine milk, evaluate its antioxidant activity, and quantify levels of total phenols and fravonoides of the different extracts used. Aqueous extracts were used in nature and dry, from the peel of the fruit (EAC) and leaves (EAF). Additionally, it was evaluated the antioxidant activity (AA%), total phenols and flavonoids. Milk samples were inoculated, incubated, and the colonies were characterized as Staphylococcus aureus were adjusted to 1.0x106 UFC/mL in the 6 standard of the MacFarland scale. The sensitivity of the microbial isolates were determined in quintuplicate, in disk diffusion test. The minimum inhibitory concentration was determined by visible inhibition zone greater than 15mm. The results were evaluated by ANOVA, Tukey 5%, using the program SISVAR 5.3 - DEX/UFLA. Results implied that the aqueous extract from the bark of dried fruit was capable of inhibiting bacterial growth at concentrations of 3%. The other treatments only showed this activity, from the concentrations of 15%, 20% and 30% for dry EAF, in nature EAC and in nature EAF, respectively. Regarding the action antioxidant of dry EAC, was not correlated with total phenols and flavonoids. Probably other alkaloids substances present in the extract studied, may have been responsible for this activity. It is concluded that extracts of Punica granatum L., especially those obtained by the shell of the fruit, showed inhibitory activity against S. aureus, indicating your use potential for the control of bovine mastitis.

Nitensidine A, a guanidine alkaloid from Pterogyne nitens, induces osteoclastic cell death

Nitensidine A is a guanidine alkaloid isolated from Pterogyne nitens, a common plant in South America. To gain insight into the biological activity of P. nitens-produced compounds, we examined herein their biological effects on osteoclasts, multinucleated giant cells that regulate bone metabolism by resorbing bone. Among four guanidine alkaloids (i.e., galegine, nitensidine A, pterogynidine, and pterogynine), nitensidine A and pterogynine exhibited anti-osteoclastic effects at 10 μM by reducing the number of osteoclasts on the culture plate whereas galegine and pterogynidine did not. The anti-osteoclastic activities of nitensidine A and pterogynine were exerted in a concentration-dependent manner, whereas nitensidine A exhibited an approximate threefold stronger effect than pterogynine (IC50 values: nitensidine A, 0.93 ± 0.024 μM; pterogynine, 2.7 ± 0.40 μM). In the present study, the anti-osteoclastic effects of two synthetic nitensidine A derivatives (nitensidine AT and AU) were also examined to gain insight into the structural features of nitensidine A that exert an anti-osteoclastic effect. The anti-osteoclastic effect of nitensidine A was greatly reduced by substituting the imino nitrogen atom in nitensidine A with sulfur or oxygen. According to the differences in chemical structures and anti-osteoclastic effects of the four guanidine alkaloids and the two synthetic nitensidine A derivatives, it is suggested that the number, binding site, and polymerization degree of isoprenyl moiety in the guanidine alkaloids and the imino nitrogen atom cooperatively contribute to their anti-osteoclastic effects. © 2013 Springer Science+Business Media Dordrecht.

Pulmonary and hepatic lesions caused by the dehydropyrrolizidine alkaloid-producing plants Crotalaria juncea and Crotalaria retusa in donkeys

The effects and susceptibility of donkeys to Crotalaria juncea and Crotalaria retusa poisoning were determined at high and low doses. Seeds of C. juncea containing 0.074% of dehydropyrrolizidine alkaloids (DHPAs) (isohemijunceines 0.05%, trichodesmine 0.016%, and junceine 0.008%) were administered to three donkeys at 0.3, 0.6 and 1g/kg body weight (g/kg) daily for 365 days. No clinical signs were observed and, on liver and lung biopsies, the only lesion was a mild liver megalocytosis in the donkeys ingesting 0.6 and 1g/kg/day. Two other donkeys that received daily doses of 3 and 5g seed/kg showed initial respiratory signs 70 and 40 days after the start of the administration, respectively. The donkeys were euthanized following severe respiratory signs and the main lung lesions were proliferation of Clara cells and interstitial fibrosis. Three donkeys ingested seeds of C. retusa containing 5.99% of monocrotaline at daily doses of 0.025, 0.05 and 0.1g/kg for 365 days. No clinical signs were observed and, on liver and lung biopsies, the only lesion was moderate liver megalocytosis in each of the three donkeys. One donkey that received a single dose of 5g/kg of C. retusa seeds and another that received 1g/kg daily for 7 days both showed severe clinical signs and died with diffuse centrilobular liver necrosis. No lung lesions were observed. Another donkey that received a single dose of 2.5g/kg of C. retusa seeds showed no clinical signs. The hepatic and pneumotoxic effects observed are consistent with an etiology involving DHPAs. Furthermore, the occurrence of lung or liver lesions correlates with the type of DHPAs contained in the seeds. Similarly as has been reported for horses, the data herein suggest that in donkeys some DHPAs are metabolized in the liver causing liver disease, whereas others are metabolized in the lung by Clara cells causing lung disease. © 2013 Elsevier Ltd.