Extraction, purification and analysis of conotoxin of Conus textile captured from Persian Gulf and the investigation of analgesic effects of conotoxins in an animal model

Identification of venomous species of Persian Gulf cone snails and characterization of venom composition and their features is so important from the point of medical importance. Marine cone snails from the genus Conus are estimated to consist of up to 700 species. The venom of cone snails has yielded a rich source of novel neuroactive peptides or conotoxins. The present study was aimed to study the analgesic effect of Persian Gulf Conus textile and its comparison with morphine in mouse model. The specimens of Conus textile were collected of Larak Island from depth of 7 m. The collected samples were transferred to laboratory alive and were stored at -700 c. he veno s ducts were separated and ho ogenized with deionized water he ixture centrifuged at rp for inutes upernatant was considered as extracted veno and stored at - C after lyophylization. The protein profile of venom determined by using SDS-PAGE and HPLC used to investigate the extracted venom and to evaluate the analgesic activity, formalin test was carried out. SDS-PAGE indicated several bands ranged between 6 and 250 kDa. Chromatogram of the venom demonstrated more than 44 large and small fractions. The amount of 10 ng of Conus crude venom and analgesic peptide showed the best anti-pain activity in formalin test. No death observed up to 100 mg/kg, which is 250,000 times higher than the effective dose.Venom characterization of Persian Gulf Conus textile may be of medical importance and potential for new pharmaceutical drugs as well.

The study of the ergosan and vibromax effects on growth performances, survival rate and increase of the resistance to some of the environmental stresses in post larvae stage of the Indian white shrimp

In this study, which has been done in Hormoz larve Hatchery at Kohestak in Minab at 1385, the efficiency of Ergosen and Vibromax vaccine and the effect of them on growth factors such as total length, Carapase, dry weight and the number of upper mordents of rostrum and survival of the stages of larvae and post larvae of Indian white shrimp was studied. Thus in order to comparison the effects of Vibromax and Ergosen, each of them separately, in one treatment, and in another simultaneously with one control treatment was used. Vaccination against larvae shrimps was done through Artemia. This study used four treatments with three replicates in a completely randomized design and comparison of means was done through Duncan test. Breeding larvae and post larvae of Indian white shrimp from zoa I stage to PL 15 was done in 20 litter plastic buckets. Present results indicated that the highest amount of growth and survival factors in larvae stage (from zoa to PL1), and also in stages of PL5 and PL15, in the treatment of Ergoson effect + vaccine and it was with a little difference from that treatment of Ergoson effect which was in high significance difference in regard to control treatment at α<0.01 level and treatment of vaccine effect and control treatment at α<0.01 level often have no significant difference. This research used environmental stress tests to study the quality of post larvae under experiment. Studying in this field showed that feeding vaccine to larvae of Indian shrimps which was done through Artemia nauplii enrichment ,and ergosen , in treatment of ergosen vaccine lead to more resistance of post larvaes against salinity stress tests and formalin .This case was observed in every three stages ,so that in stress formalin test 100 parts per million and also 10 and 20 salinity parts in thousands the most survival was observed in treatment of Ergosan effect+vaccine and after that in treatment of Ergoson effect and with a little difference in treatment of vaccine effect. Of course this case, in treatment of Ergoson effect + vaccine due to the synergistic properties vaccine with Ergoson was more than to other treatments, while every three treatments, in most stages had significant difference toward control treatment at α<0.01 level and the control treatment because of not having Ergoson and nauplii artemia with vaccine, having the least survival rate in this stages.

Refolding of omega conotoxin peptide derived from marine snail, Conus magus, to evaluate its analgesic effects

Oxidative refolding is one of the key challenges hampering the development of peptide based compounds as therapeutics. The correct refolding for three disulfide peptide like w-Conotoxi n MVIIA is difficult and crucial for biological activity. This work advanced knowledge of chemical and biological for improve oxidative refolding of synthetic w-Conotoxi n MVIIA in base of Conus magus venom. The present study aimed to set up an appropriate and effective protocols for refolding of disulfide-rich w-Conotoxin MVIIA. In this study, the crude peptide was protected with Acm group, according to the right amino acid sequences (Synthesized by Australian Company). The crude peptide was purified by H PLC. To prepare the peptide to refolding, innovative deprotection applied molar ratio (AMR) method was performed based on mercury. Accuracy of deprotection was approved by reverse phase chromatography. The deprotected target peptide (omega-conotoxin) was determined by SDS-PAGE. Then the Oxidative refolding of target peptide was performed in six protocol based on Guanidinium chloride and oxidized and reduced Glutathione. Analgesic effect of refolded peptide was surveyed with formalin test in mice Balb/c. Non neurotoxic effects of target peptides were survey with ICV injection in mice model (C57/BL6). The innovative deprotection protocol performed based on the best ratio of mercury/2-mercaptoethanol adjusted to 1mg/10p1 in 90 minute. The results showed the yield and purity of omega-conotoxin MVIIA as 93 and 95%, respectively. Refolding of 40 mg omega Conotoxin with GSSG and GSH on ratio of 10:1 and 20 mM ammonium acetate showed the best analgesic effect compared with the other methods. The result showed 95.5% yield and 98% purity of omega-conotoxin MVIIA in this refolding method. Related refolding method reduced 85% pain in experimented mice using 7 ng of the peptide. That was 71.5 fold stronger than morphine and 2 times than standard Prialt®. And it was not neurotoxic in mice. In this study, refolding method for omega-conotoxin MVIIA was optimized in the fourth factor including: reducing the time, amount and number of reagent and increase the efficiency. We introduced new method for deprotection of omega-conotoxin MVIIA. Effective, economic and applied refolding and deprotecti on method was performed in this research may al so be applied to similar omega conotoxin peptides.

Test well exploration in the Myakka River Basin area, Florida

In recent years, difficulties encountered in obtaining ground-water supplies with acceptable chemical characteristics in the Myakka River basin area led to the implementation of a test drilling program. Under this program, well drilling and data collection were executed in such a manner that all water-producing zones of the local aquifers, together with the quality and quantity of the water available, were effectively identified. A step-drilling method was utilized which allowed the collection of formation cuttings, water samples, and water-level data, from isolated zones in the well as drilling proceeded. The step drilling procedure is described. The driller's logs, geophysical logs, and chemical quality of water tables are presented.(Document has 66 pages.)

A Method for siting and prioritizing the removal of derelict vessels in Florida Coastal Waters: test applications in the Florida Keys

Increased boating activities and new waterfront developments have contributed an estimated 3,000 dismantled, abandoned, junked, wrecked, derelict vessels to Florida coastal waters. This report outlines a method of siting and prioritizing derelict vessel removal using the Florida Keys as a test area. The data base was information on 240 vessels, obtained from Florida Marine Patrol files. Vessel location was plotted on 1:250,000 regional and 1:5,000 and 1:12,000 site maps. Type of vessel, length, hull material, engine, fuel tanks, overall condition, afloat and submerged characteristics, and accessibility, were used to derive parametric site indices of removal priority and removal difficulty. Results indicate 59 top priority cases which should be the focus of immediate clean up efforts in the Florida Keys. Half of these cases are rated low to moderate in removal difficulty; the remainder are difficult to remove. Removal difficulty is a surrogate for removal cost: low difficulty -low cost, high difficulty - high cost. The rating scheme offers coastal planners options of focusing removal operations either on (1) specific areas with clusters of high priority derelict vessels or on (2) selected targeted derelicts at various, specific locations. (PDF has 59 pages.)

Test von Fanggeräten zur Untersuchung von Grundfischfaunen in Windparkgebieten der Ostsee

With respect to future environmental impact assessment different bottom trawls are presently investigated for their suitability for sampling the fish fauna in potential windpark areas asses. For this purpose the international Baltic Sea Trawl, the Box trawl, two different small beam trawls and small eel fykes were compared on a research cutter in June 2002. The best catchability with the highest abundance of demersal fish species and widest length distribution was obtained with the Box trawl. During another cruise in December 2002 the Box trawl was again compared with the Windpark trawl. As a result the Box trawl performed best, with regard to the overall catchability. However the Windpark Trawl is easier to handle under the conditions onboard small fishing cutters.

Der Fischei-Test: Ein Toxizitätstest für ökotoxikologische Untersuchungen

Zebrafish (Danio rerio) embryos have been used to quantify the teratogenic potential of environmental samples and harmful substances respectively. The short spawning interval renders this species a good test organism in toxicological research. Due to the transparency of the eggs several lethal and non-lethal endpoints can be detected in parallel after 48 h of embryonic development. Zebrafishembryos have been shown to be sensitive to a number of environmental relevant contaminants, as well as to ex-tracts from polluted sediments

Determination of the activity of nitrifying bacteria in surface waters by a modified BOD-test. [Translation from: Zentbl.Bakt.Parasitkde 156(1) 414-421, 1972. ]

A modified method of the Biochemical oxygen demand (BOD)test was applied in order to find out the seasonal changing activity of the nitrifying bacteria in surface waters. Samples were taken from the River Elbe near Teufelsbrueck.

Ecological effects of trial pumpings in the Upper Lambourn: Stage 1 group pumping test Upper Lambourn group, September - November 1975

This report to the Thames Water Authority and Central Water Planning Unit is on research carried out in conjunction with the Stage 1 Group Pumping Test of five boreholes in the upper Lambourn Group for a period of three months in September, October and November 1975. The aim of the study was to assess the ecological effects of the pumpin g of five bore-holes in the upper Lambourn. That is, to determine how the seasonal sequence of ecological events in the river differed from what would hav e occurred had no pumping taken place. Since this 'experiment' has no control it is not possible to make a direct assessment. Nevertheless, by careful monitoring of ecological events before, during and after the pumping it is possible to document changes in th e river and by reference to the data already available for the Rive r Lambourn, normal seasonal changes in the flora and fauna can be separated from changes which may be attributable to the pumping and subsequent events.

Progress report on fish counting on the Rivers Itchen and Test

This progress report summarises work on NSHEB Mark 10 fish counters which are installed at Woodmill on the River Itchen and Nurseling Mill and Connegar Bridge on the River Test. Counters are evaluated and salmon behaviour regarding the counters examined. The report includes a a list of equipment needed for the efficient running of the project in the future.

A Test of Computer-assisted Matching Using the North Pacific Humpback Whale, Megaptera novaeangliae, Tail Flukes Photograph Collection

Testing was conducted of a computer-assisted system for matching humpback whale tail flukes photographs. Trials with a 12,000-photographs database found no differences in match success between matching by computer and matching by comparing smaller catalogs ranging in size from 200 to 400 photographs. Tests with a 24,000-photographs database showed that, on average, the first match was found after examining about 130 photographs whether the photograph quality was excellent, good, or poor. Match success did not appear to be strongly related to whether the tail flukes had especially distinctive markings or pigment patterns (recognition quality). An advantage of computer-assisted matching is the ability to compare new photographs to the entire North Pacific collection, where no bias is introduced based on expectation of resightings within or between specific areas, or based on expectation of behavioral role (e.g. matching “known” females to “known” females).

DNA extraction from formalin-fixed Franciscana tissues

The present paper reports the extraction of DNA from formalin-fixed Pontoporia blainvillei tissues. Following the Vachot and Monerot (1996) protocol, fragmented DNA (300-700bp) was extracted from more than 95% of liver and muscle samples. DNA yield in liver samples was significantly higher than in muscle samples (4.574 ± 1.169mg DNA/mg versus 0.808 ± 0.297mg DNA/mg). Similar results were obtained from nine other species of cetaceans and five species of pinnipeds. It is of special interest to have a method that allows the utilisation of museum specimens not originally preserved for genetic studies, which may include rarely available, declining or extinct species. SPANISH: El presente trabajo reporta la extracción de ADN a partir de tejidos formolizados de Pontoporia blainvillei. Siguiendo el protocolo de Vachot y Monerot (1996) se pudo extraer ADN degradado (300-700pb) en más del 95% de las muestras de hígado y músculo analizadas. El rendimiento en ADN fue significativamente mayor en muestras de hígado que en muestras de músculo (4.574 ± 1.169mg DNA/mg tejido húmedo versus 0.808 ± 0.297mg DNA/mg tejido húmedo). Resultados similares se obtuvieron en otras nueve especies de Cetáceos y cinco de Pinnípedos. Resulta de gran interés contar con un método que permita la utilización de especímenes depositados en museos y que no hayan sido originalmente colectados para estudios genéticos, incluyendo especies de difícil obtención, en franca declinación o extintas.