A Novel Bromoindole Alkaloid from a Korean Colonial Tunicate Didemnum sp

Chemical investigation on a colonial marine tunicate, Didemnum sp. led to the isolation of a series of indole alkaloids including a new (1) and two known metabolites (2-3). Based on the spectroscopic analysis including 1D and 2D NMR along with MS spectra, the structure of 1 (16-epi-18-acetyl herdmanine D) was elucidated as a new amino acid derivative. The absolute configuration of 1 was determined by comparison of specific rotation with the known compound. The structures of compounds 2 and 3 were also identified as bromoindole containing compounds N-(6-bromo-1H-indole-3-carbonyl)-L-arginine and (6-bromo-1H-indol-3-yl) oxoacetamide, respectively, based on 1H and 13C NMR data, MS data and specific rotation value. Their pharmacological potentials as antibacterial agents and FXR antagonists were investigated, but no significant activity was found. However, the structural similarity of compound 1 to compound 4 suggested the antiinflammatory potential of compound 1.

Cryptic species and genetic structure in Didemnum granulatum Tokioka, 1954 (Tunicata: Ascidiacea) from the southern Brazilian coast / Espécies crípticas e estrutura genética em Didemnum granulatum Tokioka 1954 (Tunicata: Ascidiacea) na costa sul do Brasil

Didemnum granulatum is a colonial fouling ascidian that lives in subtidal substrates, worldwide. It exhibits two morphotypes, orange and beige. In this study, we verified if the color morphotypes and/or the spatial distribution of specimens in different islands might be associated to patterns of genetic structure of a single species, or if they represent distinct cryptic species. Specimens were collected in four islands, along the coast of the Santa Catarina state. A segment of 490 bp from the mitochondrial gene cytochrome c oxidase subunit 1 (COI) was amplified from 45 samples. Twenty-one haplotypes were identified. The total haplotype diversity (0.912) and the total nucleotide diversity (0.044) were high. The global Fst of the populations analyzed was 0.97, with most of the variation occurring between orange and beige groups (82.19%). The variation found between populations within groups was 15.37%, and 2.45% within populations. Haplotype networks and the neighbor-joining tree showed clear genetic divergence between individuals of distinct colors, and between the islands. These evidences strongly support the presence of a complex of two cryptic species for D. granulatum occupying the studied area. Both species were also highly genetically structured between islands, suggesting that the conservation process of these populations is complex.

Didemnum granulatum é uma ascídia colonial incrustante de substratos consolidados infralitorâneos, e de ampla distribuição mundial. Variação na pigmentação é comum em invertebrados marinhos, e morfotipos de D. granulatum das cores laranja e bege foram detectados na região. Neste estudo, nós verificamos se os morfotipos de cor e/ou a distribuição espacial dos espécimens nas diferentes ilhas podem estar associados aos padrões de estruturação genética de uma única espécie, ou se eles representam espécies crípticas distintas. Os espécimens foram coletados em quatro ilhas ao longo da costa do estado de Santa Catarina. Um total de 45 amostras tiveram amplificados um segmento de 490pb do gene mitocondrial citocromo oxidase subunidade I (COI). Vinte e um haplótipos foram identificados. A diversidade haplotípica total (0.912) e a diversidade nucleotídica total (0.044) foram altas. O Fst global das populações analisadas foi 0.97, e a maior parte da variação ocorreu entre os grupos laranja e bege (82.19%). A variação encontrada entre as populações dentro dos grupos foi 15.37%, e 2.45% dentro das populações. A rede de haplótipos e a árvore de Neighbor-joining mostraram nítidas divergências genéticas entre os indivíduos de cores distintas, e entre as ilhas. Tais evidências sugerem a presença de um complexo de duas espécies crípticas de D. granulatum na região. Ambas as espécies foram fortemente estruturadas geneticamente entre as ilhas, o que reforça a necessidade de que tais atributos sejam considerados em medidas de conservação e proteção do ambiente marinho, mais especificamente nestas ilhas.

Annual abundance of salps and doliolids (Tunicata) around Gorgona Island (Colombian Pacific), and their importance as potential food for green sea turtles / Abundancia anual de salpas y doliolidos (Tunicata) alrededor de la Isla Gorgona (Pacífico colombiano), y su importancia como alimento potencial para las tortugas marinas verdes

Gorgona National Park protects fertile waters that support large vertebrates, including green sea turtles (Chelonia mydas), and for them, gelatinous zooplankton constitute a food resource that can be found year-round in Gorgona Island´s coastal waters. This study was carried out to determine the abundance of salps and doliolids around Gorgona Island over a year, and to determine whether this is a resource that could be used reliably year-round by green turtles and other large plankton-feeding predators. The monthly abundance of salps and doliolids at eight coastal stations around Gorgona Island (Colombian Pacific) was determined between September 2005 and August 2006. Oblique tows were carried out from 50m to the surface, total zooplankton biomass was measured and the number of salps and doliolids per tow, and frequency of occurrence per station and month were determined. Superficial and bottom sea temperature, superficial and bottom salinity, and chlorophyll-a concentration were recorded at each station. There were tunicate abundance peaks in September 2005 and March 2006. The high abundances in March were probably due to a cold water intrusion into the study area, which resulted in colder saltier water and a shallower thermocline. Tunicates were probably advected to the area by currents from the southwest and aggregated due to the underwater topography. In September, the influence of continental river discharge as well as inputs from rainfall over the island could have provided increased nutrients and resulted in higher abundances. The large filter-feeding vertebrates that feed on tunicates include green sea turtle juveniles, which use coastal waters of Gorgona Island as feeding grounds, as part of their migration route in the Eastern Tropical Pacific. These turtles could be using tunicates opportunistically, as a sporadic resource that is available at certain times of the year. Rev. Biol. Trop. 62 (Suppl. 1): 149-159. Epub 2014 February 01.

Se determinó la abundancia de salpas y doliólidos en ocho estaciones costeras alrededor de Isla Gorgona (Pacífico colombiano), entre septiembre 2005 y agosto 2006. Se hicieron arrastres oblicuos desde 50m hasta la superficie; se midió la biomasa zooplanctónica total, el número de salpas y doliólidos por arrastres, y la frecuencia de ocurrencia por estación y por mes. Se registró la temperatura superficial y de fondo, la salinidad superficial y de fondo, así como la concentración de clorofila a en cada estación. Se observaron picos de abundancia de tunicados en septiembre 2005 y marzo 2006. Las altas abundancias en marzo fueron probablemente debidas a una intrusión de agua fría a la zona de estudio, la cual resultó en aguas más frías y saladas, y en una termoclina más somera. Los tunicados fueron probablemente advectados hacia el área por corrientes provenientes del suroeste, y la topografía subacuática causó una agregación de estos organismos. En septiembre, la influencia de las descargas de los ríos continentales, así como aportes por la precipitación sobre la isla pudieron haber provisto mayor cantidad de nutrientes y resultado en mayores abundancias. Los grandes vertebrados marinos filtradores que se alimentan de tunicados incluyen tortugas verdes juveniles, las cuales usan las aguas de Isla Gorgona como zona de forrajeo, como parte de su ruta de migración en el Pacífico Oriental Tropical. Estas tortugas podrían estar utilizando los tunicados oportunistamente, como un recurso esporádico que está disponible en ciertas épocas del año.

Development of Loop-Mediated Isothermal Amplification Targeting 18S Ribosomal DNA for Rapid Detection of Azumiobodo hoyamushi (Kinetoplastea)

Ascidian soft tunic syndrome (AsSTS) caused by Azumiobodo hoyamushi (A. hoyamushi) is a serious aquaculture problem that results in mass mortality of ascidians. Accordingly, the early and accurate detection of A. hoyamushi would contribute substantially to disease management and prevention of transmission. Recently, the loop-mediated isothermal amplification (LAMP) method was adopted for clinical diagnosis of a range of infectious diseases. Here, the authors describe a rapid and efficient LAMP-based method targeting the 18S rDNA gene for detection of A. hoyamushi using ascidian DNA for the diagnosis of AsSTS. A. hoyamushi LAMP assay amplified the DNA of 0.01 parasites per reaction and detected A. hoyamushi in 10 ng of ascidian DNA. To validate A. hoyamushi 18S rDNA LAMP assays, AsSTS-suspected and non-diseased ascidians were examined by microscopy, PCR, and by using the LAMP assay. When PCR was used as a gold standard, the LAMP assay showed good agreement in terms of sensitivity, positive predictive value (PPV), and negative predictive value (NPV). In the present study, a LAMP assay based on directly heat-treated samples was found to be as efficient as DNA extraction using a commercial kit for detecting A. hoyamushi. Taken together, this study shows the devised A. hoyamushi LAMP assay could be used to diagnose AsSTS in a straightforward, sensitive, and specific manner, that it could be used for forecasting, surveillance, and quarantine of AsSTS.

Purine and carboline alkaloids from ascidian Symplegma oceania / 中国中药杂志

Five purine and carboline alkaloids were isolated from the methanol extract of the ascidian Symplegma oceania. Classic chromatographies including preparative HPLC were used for isolation and purification of the compounds. The structures were established as 6-methoxy-7-methyl-8-oxoguanine (1), 2-methylimino-3-methyl-6-methylamino- 9H-purine (2), 1,2,3,4-tetrahydro-betacarboline (3), 1,2,3,4-tetrahydro-1-methyl-beta-carboline (4) and 1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (5) by comparison the spectroscopic data (MS, 1H, 13C-NMR) with those reported in the literatures. Compounds 2-5 were reported from the the genus Symplegma for the first time. The purine and carboline were the major alkaloid types of S. oceania.

Apendicularias (Urochordata) y quetognatos (Chaetognatha) del Parque Nacional Isla del Coco, Costa Rica / Appendicularians (Urochordata) and chaetognaths (Chaetognatha) of Isla del Coco National Park, Costa Rica

Appendicularians and chaetognaths are important members of the zooplancton but little studied in Costa Rica and not at all at Isla del Coco Nacional Park, eastern tropical Pacific. Epipelagic (0-50m) zooplankton samples from the oceanic area off Isla del Coco, were collected in November 2001, and over patch reefs in 2009. We analized the species composition of appendicularians and chaetognaths present in 31 samples. Nine species of appendicularians, and eight species of chaetognaths were identified. Oikopleura longicauda was the most abundant in the surveyed area (217 specimens, 65.56% of total appendicularian numbers), followed by Oikopleura rufescens with 35 (10.57%). All the appendicularian species found in this study are common in warm ocean waters, but six species herein recorded for the first time from waters off Isla del Coco, and two species are new records for Costa Rica. Among the chaetognaths, Flaccisagitta enflata was the commonest species (85 specimens, 50.6% of total numbers) recorded in this study, followed by Serratosagitta pacifica with 38 organisms (20.62%). Zonosagitta bedoti was represented by a single specimen. These species have been recorded from tropical waters of the Eastern Pacific, but six species are new records for Isla del Coco. Data on the distribution and morphology of the species recorded are presented together with keys for the identification of both taxa in the surveyed area.

Las apendicularias y quetognatos son miembros importantes del zooplancton pero poco estudiados en Costa Rica y no del todo en el Parque Nacional Isla del Coco, Pacífico tropical este. Se recolectaron muestras epipelágicas de zooplancton de la zona oceánica (0-50m) de la Isla del Coco, en noviembre de 2001 y sobre parches arrecifales en 2009. Se analizó la composición de especies de apendicularias y quetognatos presentes en 31 muestras. Nueve especies de apendicularias, y ocho especies de quetognatos fueron identificadas. Oikopleura longicauda fue la más abundante en el área estudiada (217organismos, representando el 65.56% del número total de apendicularias), seguida por Oikopleura rufescens, con 35 (10.57%). Todas las especies de apendicularias encontradas en este estudio son comunes en las aguas cálidas de los océanos, seis especies se registran aquí por primera vez en aguas de la Isla del Coco y dos son nuevos registros para Costa Rica. Entre los quetognatos, Flaccisagitta enflata fue la especie más común (85 ejemplares, representando el 50.6% del número total) registrada en este estudio, seguida por Serratosagitta pacifica con 38 organismos (20.62%). Zonosagitta bedoti estuvó representada por un solo espécimen. Estas especies también se han registrado en las aguas tropicales del Pacífico oriental, pero seis especies son nuevos registros para Isla del Coco. Los datos sobre la distribución y morfología de las especies registradas se presentan junto con las claves para la identificación de los taxones en el área estudiada.

Identification of active components in Styela clava by HPLC-eSI-Tof/MS and the study on their HPLC specific chromatograms / 药学学报

A new method based on high performance liquid chromatography-electrospray ionization time of flight-mass spectrometry (HPLC-ESI-TOF/MS) was developed for the rapid identification of active compounds in Styela clava and the development of its specific chromatograms. Samples were extracted by ultrasonic-assisted extraction, and the extraction conditions were optimized. The developed HPLC-ESI-TOF/MS method was used to identify the components in Styela clava extract, and a specific chromatogram based on HPLC analysis was established. Ten compounds in Styela clava extract have been primary identified by HPLC-ESI-TOF/MS on-line detection combined with literature review. The result of similarity evaluation for specific chromatograms indicated that the quality of different Styela clava samples was not entirely consistent. This method has the advantages of simple operation, rapid measurement and it is a powerful tool for identification of active components in Styela clava and its quality control.

Food and Meal Preference of Workers in the Chonnam Yeosu Industrial Area / 대한지역사회영양학회지

This study was conducted to investigate food and meal preference of workers (435 male and 212 female) in the Chonnam Yeosu industrial area. The results of the survey were analyzed by principal components analysis. The results were obtained as follows: 19.3% of the subjects were twenties, 28.3% were thirties, 28.8% were forties and 23.7% were fifties. Females liked rice rolled in dried laver, rice cakes, janchi-noodle and breads, on the other hand males liked thick beef soup, loach soup, an eel stew and soju. The young liked instant noodles, fried chicken, sweet and sour pork, pork cutlet, pizza, hamburger, ham, sausage and fruit beverage, on the other hand the old liked a fish pot stew, loach soup, eel stew, fish boiled in soy with spices, panfried fish, sea slug, ascidian, bunder, green laver and boiled burdock-lotus root in soy. Females who are young liked hamburger and sweet and sour pork whereas males who are young liked instant noodles, pork cutlet, ham, sausage and fruit beverage. Aged females liked fish boiled in soy with spices, panfried fish, bunder, green laver, boiled burdock-lotus root in soy whereas aged males liked a fish pot stew, loach soup, eel stew, sea mussel and oysters. Boiled rice was located within the middle irrespective of age and sex, but noodles, wheat flour meal, fast foods and fruits were situated at the young female side. Aged males liked soup and pot stew. Young males liked meats and eggs whereas fish and shellfish and kimch were located at the aged people side. Aged females liked sea weeds and most people disliked vegetables but females liked some vegetables irrespective of age. Processed foods, salted foods, and alcohol were generally disliked foods by subjects but males liked those foods. Soybean curd was liked more males than females, and teas, except coffee, was liked by males.

New record of Thalia cicar van Soest 1973 (Urochordata: Thaliacea) in the Equatorial Atlantic / Novo registro de Thalia cicar van Soest 1973 (Urochordata: Thaliacea) no Atlântico Equatorial

Thalia cicar van Soest 1973 (Urochordata, Thaliacea) is considered a tropical-subtropical species, registered in the Atlantic Ocean at latitudes between 7-34º S and 6-32º N. This work enlarges the occurrence of this species for Equatorial Atlantic waters. The specimens were found in a shelf break area of the Brazilian northeastern (07º 50'-07º 70' S and 34º 23' W) during the expedition JOPS-II (Joint Oceanographic Projects II) in March 1995; and, in São Pedro e São Paulo arquipelago (0º 55' N and 29º 20' W) in May and June, 2005. These two areas are characterized by the presence of local upwelling that induces the ascent of bottom waters rich in nutrients, generating areas of larger productivity than the typically oligotrophic Equatorial Atlantic waters. The new occurrence of Thalia cicar reported in this work can be related to these more productive waters of Equatorial Atlantic. The species that is most commonly found in the Atlantic Ocean is T. democratica, and the lack of past records of T. cicar might have been caused by the taxonomic difficulties determining of the solitary and aggregate zooids of these two species. This work suggests the potential use of the ratios among tunic lengths as an additional character to differentiate T. cicar and T. democratica oozooids.

Thalia cicar van Soest 1973 (Urochordata, Thaliacea) é considerada uma espécie tropical-subtropical, registrada no oceano Atlântico desde 7 a 34º S e 6 a 32ºN. Este trabalho amplia a ocorrência desta espécie para águas do Atlântico equatorial. Os espécimes foram encontrados na plataforma continental do Nordeste brasileiro (07º 50'-07º 70' S e 34º 23' W) durante a expedição JOPS-II (Joint Oceanographic Projects II) em março de 1995 e no arquipélago de São Pedro e São Paulo (0º 55' N e 29º 20' W) em maio e junho de 2005. Essas duas áreas se caracterizam pela presença de ressurgências locais que induzem a ascensão de nutrientes, gerando locais de alta produtividade, nas águas tipicamente oligotróficas do Atlântico equatorial. O registro de Thalia cicar nesses dois locais pode estar relacionado com essas águas de maior produtividade no Nordeste brasileiro. A espécie de Thalia comumente encontrada no oceano Atlântico é T. democratica e a ausência de registros passados de T. cicar pode decorrer das dificuldades taxonômicas em diferenciar os zoóides solitários e agregados dessas duas espécies. Este trabalho sugere a utilização da proporção entre os comprimentos das projeções da túnica como um caráter adicional para diferenciar os oozoóides destas duas espécies.

Distribution of a pelagic tunicate, Salpa fusiformis in warm surface current of the eastern Korean waters and its impingement on cooling water intakes of Uljin nuclear power plant.

Impingement of a large amount of gelatinous plankton, Salpa fusiformis on the seawater intake system-screens in a nuclear power plant at Uljin was firstly recorded on 18th June 2003. Whole amount of the clogged animals was estimated were presumptively at 295 tons and the shortage of cooling seawater supply by the animal clogging caused 38% of decrease in generation capability of the power plant. Zooplankton collection with a multiple towing net during the day and at night from 5 to 6 June 2003 included various gelatinous zooplanktons known to be warm water species such as salps and siphonophores. Comparatively larger species, Salpa fusiformis occupied 25.4% in individual density among the gelatinous plankton and showed surface distribution in the depth shallower than thermocline, performing little diel vertical migration. Temperature, salinity and satellite data also showed warm surface current predominated over the southern coastal region near the power plant in June. The results suggested that warm surface current occasionally extended into the neritic region may transfer S. fusiformis, to the waters off the power plant. The environmental factors and their relation to ecobiology of the large quantity of salpa population that are being sucked into the intake channel of the power plant are discussed.

A Clinical Study on the Efficacy of Cosmetics Containing the Ascidian tunic in Reducing Wrinkles / 대한피부과학회지

BACKGROUND: In the last few decades, there has been a substantial increase in the population of aged people. Aging skin is a common concern for many people these days. OBJECTIVE: The purpose of this study was to evaluate the efficacy of cosmetics containing Ascidian tunic in reducing wrinkles. METHODS: Cream containing the Ascidian tunic was applied to right lateral epicanthal areas of 30 subjects for 8 weeks. As control group, a placebo which does not contain the Ascidian tunic was applied to the left lateral epicanthal areas of the subjects. Skin barrier function was evaluated and skin replicas were molded on both epicanthal areas every 4 weeks. The changes of the skin wrinkles were analyzed with skin visiometer. RESULTS: In comparison of the changes between groups treated with placebo and Ascidian tunic, there was statistically significant differences in hydration and sebum secretion. But there were no statistically significant differences in the result of R-value by visiometer. CONCLUSION: The treatment of Ascidian tunic extract seems to be partially effective in improving wrinkles, but further study is required.

Antibiotic, cytotoxic and enzyme inhibitory activity of crude extracts from Brazilian marine invertebrates

Herein we present the results of a screening with 349 crude extracts of Brazilian marine sponges, ascidians, bryozoans and octocorals, against 16 strains of susceptible and antibiotic-resistant bacteria, one yeast (Candida albicans), Mycobacterium tuberculosis H37Rv, three cancer cell lines MCF-7 (breast), B16 (murine melanoma ) and HCT8 (colon), and Leishmania tarentolae adenine phosphoribosyl transferase (L-APRT) enzyme. Less than 15 percent of marine sponge crude extracts displayed antibacterial activity, both against susceptible and antibiotic-resistant bacteria. Up to 40 percent of marine sponge crude extracts displayed antimycobacterial activity against M. tuberculosis H37Rv. Cytotoxicity was observed for 18 percent of marine sponge crude extracts. Finally, less than 3 percent of sponge extracts inhibited L-APRT. Less than 10 percent of ascidian crude extracts displayed antibacterial activity. More than 25 percent of ascidian crude extracts were active against M. tuberculosis and the three cancer cell lines. Only two crude extracts from the ascidian Polysyncraton sp. collected in different seasons (1995 and 1997) displayed activity against L-APRT. Less than 2 percent of bryozoan and octocoral crude extracts presented antibacterial activity, but a high percentage of crude extracts from bryozoan and octororal displayed cytotoxic (11 percent and 30 percent, respectively) and antimycobacterial (60 percent) activities. The extract of only one species of bryozoan, Bugula sp., presented inhibitory activity against L-APRT. Overall, the crude extracts of marine invertebrates herein investigated presented a high level of cytotoxic and antimycobacterial activities, a lower level of antibacterial activity and only a small number of crude extracts inhibited L-APRT. Taxonomic analysis of some of the more potently active crude extracts showed the occurrence of biological activity in taxa that have been previously chemically investigated. These...

No presente estudo apresentamos resultados da triagem biológica realizada com 349 extratos obtidos de esponjas marinhas, ascídias, briozoários e octocorais do Brasil, em testes contra 16 linhagens de bactérias comuns e resistentes à antibióticos, uma levedura (Candida albicans), Mycobacterium tuberculosis H37Rv, três linhagens de células tumorais MCF-7 (mama), B16 (melanoma murínico) e HCT8 (cólon), e de inibição da enzima adenina fosforribosil transferase de Leishmania tarentolae (L-APRT). Menos de 15 por cento dos extratos de esponja marinhas apresentaram atividade antibacteriana, contra linhagens resistentes ou não a antibióticos. Quase 40 por cento dos extratos de esponjas marinhas apresentaram atividade antimicobacteriana contra Mycobacterium tuberculosis H37Rv. Foi observada citotoxicidade para 18 por cento dos extratos de esponjas marinhas. Finalmente, menos de 3 por cento dos extratos de esponjas apresentaram atividade inibitória da enzima L-APRT. Menos de 10 por cento dos extratos de ascídias apresentaram atividade antibacteriana. Mais de 25 por cento dos extratos de ascídias apresentaram atividade contra M. tuberculosis e as três linhagens de células tumorais. Somente dois extratos obtidos da ascídia Polysyncraton sp. coletada em duas diferentes épocas (1995 e 1997) apresentaram atividade contra L-APRT. Menos de 2 por cento dos extratos de briozoários e octocorais apresentaram atividade antibacteriana, mas uma alta percentagem de extratos destes animais apresentaram atividades citotóxica (11 por cento e 30 por cento, respectivamente) e antimicobacteriana (60 por cento). O extrato de somente uma espécie de briozoário, Bugula sp., apresentou atividade inibitória da enzima L-APRT. A análise taxonômica de algumas espécies de invertebrados que forneceram alguns dos extratos mais ativos, indicou a ocorrência de atividade biológica em espécies pertencentes a grupos taxonômicos que já foram anteriormente investigados do ponto de vista...

Alimentación de la ascidia Ecteinascidia turbinata (Ascidiacea: Perophoridae) en dos áreas de manglar de Cuba

Diet of the ascidian Ecteinascidia turbinata (Ascidiacea: Perophoridae) in two mangrove areas of Cuba. Stomach contents of 88 zooids of Ecteinascidia turbinata Herdman 1880 and the qualitative and quantitative composition of phytoplankton in the water column were studied in Santa Fe (North Coast of Havana) and Punta del Este (SW of Cuba). We identified 59 microalgal species and four tintinnids in the stomachs. Cell size was 75-165 µm in length and 2-105 µm in width. There were not significant differences in microalgal diversity in the water column in the two locations. In both locations, the diatoms had the largest number of species and individuals in stomachs and water. In Santa Fe, dinoflagellate biomass was larger in water and stomach contents, while in Punta del Este the contribution of each group to the stomach content was similar to that of the water column. This species filters in a constant and irregular way during the day, independently of food availability. Rev. Biol. Trop. 55 (2): 499-507. Epub 2007 June, 29.

Se estudiaron los contenidos estomacales de 88 zooides de Ecteinascidia turbinata Herdman 1880 y la composición cualitativa y cuantitativa del fitoplancton en la columna de agua en Santa Fe (Litoral Norte de la Habana) y Punta del Este (SW de Cuba). En el contenido estomacal de la ascidia se identificaron 59 especies de microalgas y cuatro tintínidos. El tamaño de las células varió entre 75 y 165 µm de largo y de 2 a 105 µm de ancho. No se encontraron diferencias significativas en la diversidad de especies de microalgas en la columna de agua de las dos localidades. Las diatomeas tuvieron la mayor cantidad de especies y el mayor número de individuos, tanto en los estómagos como en la columna de agua en ambas localidades. La biomasa de dinoflagelados en Santa Fe fue mayor en agua y estómagos. En Punta del Este el aporte de cada grupo al contenido estomacal es similar al encontrado en la columna de agua. Esta especie filtra de forma constante e irregular durante las 24 horas del día, independientemente de la disponibilidad de alimento.

Distribución y abundancia de la ascidia Ecteinascidia turbinata (Ascidiacea: Perophoridae) en Cuba / Distribution and abundance of the ascidian Ecteinascidia turbinata (Ascidiacea: Perophoridae) in Cuba

Permanently submerged mangrove roots (Rhizophora mangle) are the main habitat of the ascidian Ecteinascidia turbinata in Cuba. It was occasionally found on black coral (Antiphates caribeana) between 22 and 38 meters deep. This species exhibits a wide distribution in all the mangrove keys surrounding the Island of Cuba but does not occur in riparian or fringing mangroves. Populations of this species are abundant in Cuba: in 75% of the 58 localities sampled the species was present and in 57% more than 50% of the roots held at least one colony. The highest colony densities were found in the northern coast of Pinar del Rio province with values near one colony per lineal meter of mangrove root. We found the highest density (1.46 col/m) and greatest biomass at Jutías Key, with values between 25 and 660 g/m. The average of wet biomass in the studied mangroves was 73.63 g/m.

In vivo antithrombotic properties of a heparin from the oocyte test cells of the sea squirt Styela plicata(Chordata-Tunicata)

In the ascidian Styela plicata, the oocytes are surrounded by two types of accessory cells named follicle cells and test cells. A heparin-like substance with an anticoagulant activity equivalent to 10 percent of mammalian heparin and about 5 percent as potent as the mammalian counterpart for the inhibition of thrombin by antithrombin was isolated from the oocyte test cells. In the present study, we compared the antithrombotic and hemorrhagic effects of sea squirt oocyte test cell heparin with those of porcine heparin in rat models of venous thrombosis and blood loss. Intravenous administration of the oocyte test cell heparin to Wistar rats (both sexes, weighing ~300 g, N = 4 in each group) at a dose of 5.0 mg/kg body weight, which produced a 1.8-fold increase in plasma activated partial thromboplastin time, inhibited thrombosis by 45 ± 13.5 percent (mean ± SD) without any bleeding effect. The same dose of porcine heparin inhibited thrombosis by 100 ± 1.4 percent, but produced a blood loss three times greater than that of the saline-treated control. However, 10-fold reduction of the dose of porcine heparin to 0.5 mg/kg body weight, which produced a 5-fold increase in plasma-activated partial thromboplastin time, inhibited thrombosis by 70 ± 13 percent without any bleeding effect. The antithrombotic properties of a new heparin isolated from test cells of the sea squirt S. plicata, reported here for the first time, indicate that, although sea squirt oocyte test cell heparin was a poor anticoagulant compared to porcine heparin, it had a significant antithrombotic effect without causing bleeding.

Experimental study of peripheral nerve regeneration by using non-tubular natural cellulose membrane nerve conduit

Styela clava, called non-native tunicate or sea squirt, is habitat which include bays and harbors in Korea and several sites in the sea faced world. We fabricate cellulose membrane nerve conduit (CMNC) from this native sea squirt skin, and evaluate the capacity of promoting peripheral nerve regeneration in the rat sciatic nerve defect model. After processing the pure cellulose membrane from the sea squirt skin as we already published before, CMNC was designed as a non-tubular sheet with 14 mm length and 4 mm width. Total eleven male Spraque-Dawley rats (12 weeks, weighing 250 to 300g) were divided into sham group (n=2), silicone tube grafted control group (n=3) and experimental group (n=6). Each CMNC grafted nerve was evaluated after 4, 8 and 12 weeks in the experimental group, and after 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were all examined using image analyzer and electromicroscopic methods in the all groups. The regenerated axon and nerve sheath were found only in the inner surface of the CMNC after 4 weeks and became more thicker after 8 and 12 weeks. In the TEM study, CMNC grafted group showed more abundant organized myelinated nerve fibers with thickened extracellular matrix than silicone conduit grafted group after 12 weeks. The sciatic function index (SFI) and ankle stance angle (ASA) in the functional evaluation were -47.2+/-3.9, 35.5.+/-4.9.in CMNC grafted group (n=2) and -80.4+/-7.4, 29.2.+/-5.3.in silicone conduit grafted group (n=3), respectively. And the myelinated axon was 41.59% in CMNC group and 9.51% in silicone conduit group to the sham group. The development of a bioactive CMNC to replace autogenous nerve grafts offers a potential and available approach to improved peripheral nerve regeneration. As we already published before, small peptide fragment derived from the basement membrane matrix proteins of squirt skin, which is a kind of anchoring protein composed of glycocalyx, induced the effective axonal regeneration with rapid growth of Schwann cells beneath the inner surface of CMNC. So the possibilities of clinical application as a peripheral nerve regeneration will be able to be suggested.

Development of a bioactive cellulose membrane from sea squirt skin for bone regeneration - a preliminary research

OBJECTIVES: To develop a bioactive membrane for guided bone regeneration (GBR), the biocompatibility and bone regenerating capacity of the cellulose membrane obtained from the Ascidians squirt skin were evaluated. MATARIALS AND METHODS: After processing the pure cellulose membrane from the squirt skin, the morphological study, amino acid analysis and the immunoreactivity of the cellulose membrane were tested. Total eighteen male Spraque-Dawley rats (12 weeks, weighing 250 to 300g) were divided into two control (n=8) and another two experimental groups (n=10). In the first experimental group (n=5), the cellulose membrane was applicated to the 8.0 mm sized calvarial bone defect and the same sized defect was left without cellulose membrane in the first control group (n=4). In the another experimental group (n=5), the cellulose membrane was applicated to the same sized calvarial bone defect after femoral bone graft and the same sized defect with bone graft was left without cellulose membrane in the another control group (n=4). Each group was sacrificed after 6 weeks, the histological study with HandE and Masson trichrome stain was done, and immunohistochemical stainings of angiogenin and VEGF were also carried out. RESULTS: The squirt skin cellulose showed the bio-inductive effect on the bone and mesenchymal tissues in the periosteum of rat calvarial bone. This phenomenon was found only in the inner surface of the cellulose membrane after 6 weeks contrast to the outer surface. Bone defect covered with the bioactive cellulose membrane showed significantly greater bone formation compared with control groups. Mesenchymal cells beneath the inner surface of the bioactive cellulose membrane were positive to the angiogenin and VEGF antibodies. CONCLUSION: We suppose that there still remains extremely little amount of peptide fragment derived from the basement membrane matrix proteins of squirt skin, which is a kind of anchoring protein composed of glycocalyx. This composition could prevent the adverse immunological hypersensitivity and also induce bioactive properties of cellulose membrane. These properties induced the effective angiogenesis with rapid osteogenesis beneath the inner surface of cellulose membrane, and so the possibilities of clinical application in dental field as a GBR material will be able to be suggested.

Structure and anticoagulant properties of sulfated glycosaminoglycans from primitive Chordates

Dermatan sulfates and heparin, similar to the mammalian glycosaminoglycans, but with differences in the degree and position of sulfation were previously isolated from the body of the ascidian Styela plicata and Ascidia nigra. These differences produce profound effects on their anticoagulant properties. S. plicata dermatan sulfate composed by 2-O-sulfatedalpha-L-iduronic acid and 4-O-sulfated N-acetyl-beta-D-galactosamine residues is a potent anticoagulant due to a high heparin cofactor II activity. Surprisingly, it has a lower potency to prevent thrombus formation on an experimental model and a lower bleeding effect in rats than the mammalian dermatan sulfate. In contrast, A. nigra dermatan sulfate, also enriched in 2-O-sulfated alpha-L-iduronic acid, but in this case sulfated at O-6 of the N-acetyl-beta-D-galactosamine units, has no in vitro or in vivo anticoagulant activity, does not prevent thrombus formation but shows a bleeding effect similar to the mammalian glycosaminoglycan. Ascidian heparin, composed by 2-O-sulfated alpha-L-iduronic acid, N- and 6-O-sulfated glucosamine (75 percent) and alpha-L-iduronic acid, N- and 6-O-sulfated glucosamine (25 percent) disaccharide units has an anticoagulant activity 10 times lower than the mammalian heparin, is about 20 times less potent in the inhibition of thrombin by antithrombin, but has the same heparin cofactor II activity as mammalian heparin

Minimum effective release rate of antifoulants.1. Effects of zosteric acid at Ford Island, Hawaii (USA).

Minimum effective release rate (MERR) of zosteric acid (ZA) was studied at Ford Island. At laboratory condition, spore attachment in Enteromorpha intestinalis was inhibited significantly at 28.2 microg cm2 of ZA. More than 95% reduction in spore attachment was achieved in Ulva fasciata at 2,000 microM of ZA through membrane diffusion system. At 40 cm depth level larval settlement was high with maximum number of Hydroides elegans settlement. The overall larval settlement was increased with increasing days of exposure. The MERR of 10 microg cm(-2) d(-1) ZA inhibited 50% of the larval settlement. However, the inhibition rate was not proportionate to the antifouling ZA concentration. Using MERR membrane diffusion system an effective antifouling strategy was suggested for Ford Island.