Unexpected fine-scale population structure in a broadcast-spawning Antarctic marine mollusc

Several recent empirical studies have challenged the prevailing dogma that broadcast-spawning species exhibit little or no population genetic structure by documenting genetic discontinuities associated with large-scale oceanographic features. However, relatively few studies have explored patterns of genetic differentiation over fine spatial scales. Consequently, we used a hierarchical sampling design to investigate the basis of a weak but significant genetic difference previously reported between Antarctic limpets (Nacella concinna) sampled from Adelaide and Galindez Islands near the base of the Antarctic Peninsula. Three sites within Ryder Bay, Adelaide Island (Rothera Point, Leonie and Anchorage Islands) were each sub-sampled three times, yielding a total of 405 samples that were genotyped at 155 informative Amplified Fragment Length Polymorphisms (AFLPs). Contrary to our initial expectations, limpets from Anchorage Island were found to be subtly, but significantly distinct from those sampled from the other sites. This suggests that local processes may play an important role in generating fine-scale population structure even in species with excellent dispersal capabilities, and highlights the importance of sampling at multiple spatial scales in population genetic surveys.     

Structure and biological evaluation of novel cytotoxic sterol glycosides from the marine red alga Peyssonnelia sp

Bioactivity-guided fractionation of the extract from a Fijian red alga Peyssonnelia sp. led to the isolation of two novel sterol glycosides 19-O-β-d-glucopyranosyl-19-hydroxy-cholest-4-en-3-one (1) and 19-O-β-d-N-acetyl-2-aminoglucopyranosyl-19-hydroxy-cholest-4-en-3-one (2), and two known alkaloids indole-3-carboxaldehyde (3) and 3-(hydroxyacetyl)indole (4). Their structures were characterized by 1D and 2D NMR and mass spectral analysis. The sterol glycosides inhibited cancer cell growth with mean IC₅₀ values (for 11 human cancer cell lines) of 1.63 and 1.41 μM for 1 and 2, respectively. The most sensitive cancer cell lines were MDA-MB-468 (breast) and A549 (lung), with IC₅₀’s in of 0.71–0.97 μM for 1 and 2. Modification of the sterol glycoside structures revealed that the α,β-unsaturated ketone at C-3 and oxygenation at C-19 of 1 and 2 are crucial for anticancer activity, whereas the glucosidic group was not essential but contributed to enhanced activity against the most sensitive cell lines.     

Isolation and structure of axinastatin 5 from a republic of comoros marine sponge

The new cancer cell growth (human and murine) inhibitory (GI₅₀ 0.3 to 3.3 μg/ml) cyclo-octapeptide axinastatin 5 (2) was isolated in 3.8 x 10⁻⁷% yield from the Western Indian Ocean marine sponge Axinella cf. carteri. Structural elucidation was achieved by high field (400 and 500 MHz) 2 D-NMR techniques and the amino acid sequence was confirmed by results of NMR and high resolution mass spectral MS/MS interpretations. Other peptide cancer cell growth inhibitory constituents of this orange sponge were found to be the cyclic hepta- and octapeptides axinastatin 1 and hymenistatin 1 (1).     

Isolation and structure elucidation of new cytotoxic steroids from the gorgonian Leptogorgia sarmentosa

The gorgonian Leptogorgia sarmentosa contains three new steroids, (20S)-20-hydroxycholestane-3,16-dione (1), (16S, 20S)-16,20-dihydroxycholestan-3-one (2), and (20S)-20-hydroxycholest-1-ene-3,16-dione (3) together with a known related compound (4). Their structures were defined by spectroscopic analysis. The new steroids exhibited significant cytotoxicity against four tumor cell lines (ED50 = 1 microg/ml).     

Isolation and structure elucidation of cytotoxic polyacetylenes and polyenes from Echinacea pallida

Bioassay-guided fractionation of n-hexane extracts of Echinacea pallida (Asteraceae) roots led to the isolation and structure elucidation of two polyacetylenes (1, 3) and three polyenes (2, 4, 5). Two are known hydroxylated compounds, namely 8-hydroxy-pentadeca-(9E)-ene-11,13-diyn-2-one (1) and 8-hydroxy-pentadeca-(9E,13Z)-dien-11-yn-2-one (2). Two dicarbonylic constituents, namely pentadeca-(9E)-ene-11,13-diyne-2,8-dione (3) and pentadeca-(9E,13Z)-dien-11-yne-2,8-dione (4), were isolated and characterized for the first time. Furthermore, the structure elucidation of pentadeca-(8Z,13Z)-dien-11-yn-2-one (5) is described. The structure of the compounds isolated was determined on the basis of UV, IR, NMR (including 1D and 2D NMR experiments, such as 1H-1H gCOSY, gHSQC-DEPT, gHMBC, gNOESY) and MS spectroscopic data. The cytotoxic activity of the isolated constituents against MIA PaCa-2 human pancreatic adenocarcinoma cells was evaluated in the concentration range 1-100 microg/ml. Results show that the hydroxylated compounds (1, 2) have low cytotoxicity, while the more hydrophobic polyacetylenes (3) and polyenes (4, 5) displayed moderate activity.     

Isolation and structure of a novel peptide inhibitor of HIV-1 integrase from marine polychaetes

A homogeneous peptide with a mass of 683 Da which inhibits HIV-1 integrase with IC₅₀ 3 × 10^?5 M was separated from aqueous extracts of a marine worm Eunicidae sp. by multistage chromatography purification. The Asp-Leu-Hse-His-Ala-Gln structure was proposed for this peptide according to amino acid analysis, automated amino acid Edman sequences, and TLC with witness homoserine and MS/MS fragmentation. The proposed structure is the first example of a natural peptide containing an amino acid homoserine residue.     

Ophirasterol, a new C31 sterol from the marine sponge Topsentia ophiraphidites

Analysis of the sterol fraction obtained from the Colombian Caribbean sponge Topsentia ophiraphidites revealed that this sponge is a rich source of C30 and C31 sterols. Among them, a new C31 sterol, named ophirasterol, was isolated, and its structure was established as (22E,24R)-24-(1-buten-2-yl)cholesta-5,22-dien-3beta-ol (1) by spectral means and comparison with synthetic C-24 epimers with known configuration. Other isolated C30 and C31 sterols were the known 24-ethyl-24-methyl-22-dehydrocholesterol (2), 24-isopropyl-22-dehydrocholesterol (3), 24-isopropylcholesterol (4), 24-ethyl-24-methylcholesterol (5), 24-isopropenyl-25-methyl-22-dehydrocholesterol (6) and 24-isopropenyl-25-methylcholesterol (7), and 24-isopropenyl-22-dehydrocholesterol (8).     

Isolation of functional chloroplasts from the sacoglossan mollusc Elysia viridis Montague

A novel technique has been developed in which functional chloroplasts have been isolated and purified from the symbiotic sacoglossan mollusc Elysia viridis , Montague. This entailed the progressive filtration and gel-filtration of a hyaluronidase-incubated homogenate. Isopycnic centrifugation of the resultant crude chloroplast suspension reduced the cytoplasmic/mitochondrial and microbody contamination by approx. 70 and 90% respectively. Purified chloroplast suspensions were capable of linear rates of ¹⁴CO₂ fixation for at least 25 min at 50 μmol m² s⁻¹ (PPFD), even though approx. 54% of assimilate was being released into the bathing medium. Values for CO₂ fixation and assimilate release were similar to those obtained for the intact mollusc.     

Haemocyanin mRNA from arthropod and mollusc origin. Evidence for a multi-unit structure in mollusc haemocyanin mRNA.

Hepatocytes were preincubated with 10mM-glucagon and 100 microM-corticosterone to increase phosphatidate phosphohydrolase activity. Addition of 10 nM-glucagon or 100 microM-8-bromo cyclic GMP to a second incubation mixture that contained cycloheximide increased the half-life of the phosphohydrolase activity. Dexamethasone (100 nM) had no significant effect, but insulin (500 pM) or spermine (1 mM) decreased the half-life. None of these compounds altered the general rate of degradation of proteins labelled with [3H]leucine. There appears to be a specific control of the half-life of phosphatidate phosphohydrolase activity, which could contribute to its long-term regulation in the liver.     

Interactions of temperature and pH on the regulatory properties of pyruvate kinase from organs of a marine mollusc

The effects of low temperature assay (5 °C) on the properties of the aerobic (low phosphate) vs. anoxic (high phosphate) forms of pyruvate kinase (PK) from foot muscle and gill of the whelk Busycon canaliculatum (L.) were assessed at two pH values, pH 7.00 and 7.25, and compared to control conditions of 20 °C and pH 7.00 (all assayed in imidazole buffer). When pH was held constant at 7.00, the decrease in assay temperature to 5 °C had large effects on the measured kinetic parameters of all PK forms, as compared to 20 °C and pH 7.00. However, when assay pH was allowed to rise, from 7.00 to 7.25, with the temperature decrease to 5 °C there were fewer alterations of kinetic parameters and quantitatively smaller changes to enzyme properties. It appears, then, that when pH rises with decreasing temperature following alphastat predictions, kinetic properties of PK are largely conserved. Low temperature, at either pH value, had several significant effects on PK properties. For example, low temperature raised the S_0.5 for phosphoenolpyruvate of PK-anoxic from gill by 3–6 fold and decreased the I₅₀ Mg · ATP for PK-anoxic from foot by the same amount. Arrhenius plots of PK activity for the gill PK forms showed a distinct break at 10 °C; > 10 °C Q₁₀ was 2.5 whereas < 10 °C Q₁₀ was 8.4. Temperature-dependent changes in all cases affected enzyme properties in a manner that would restrict enzyme function at low temperature.     

Enkephalins increase dopamine levels in the CNS of a marine mollusc.

Intracardiac administration of methionine enkephalin and leucine enkephalin increased dopamine but not serotonin levels in the CNS of the marine mollusc $\text{Mytilus}$$\text{edulis}$. Naloxone blocked the effects of the enkephalins. These responses displayed a time dependent desensitization to methionine enkephalin. The study suggests the presence of an opiate receptor mechanism in this invertebrate species.     

Isolation and structure of novel peptide inhibitor of HIV-1 integrase from marine polychaetes

A homogeneous peptide with m683 Da which inhibits HIV-1 integrase with IC50 3 x 10(-5) M was separated from aqueous extracts of marine worm Eunicidae sp. by multi-stage chromatography purification. The structure Asp-Leu-Hse-His-Ala-G1n was proposed for this peptide according to the amino acid analysis, automated amino acid Edman sequencing, TLC with the witness and homoserine MS/MS fragmentation. The proposed structure is the first example of natural peptide containing amino acid homoserine residue.     

Lagunamide C, a cytotoxic cyclodepsipeptide from the marine cyanobacterium Lyngbya majuscula

Lagunamide C (1) is a cytotoxic cyclodepsipeptide isolated from the marine cyanobacterium, Lyngbya majuscula, from the western lagoon of Pulau Hantu Besar, Singapore. The complete structural characterization of the molecule was achieved by extensive NMR spectroscopic analysis as well as chemical manipulations. Several methods, including the advanced Marfey’s method, a modified method based on derivatization with Mosher’s reagents and analysis using LC–MS, and the use of ³J_H–H coupling constant values, were utilized for the determination of its absolute configuration. Compound 1 is related to the aurilide-class of molecules and it differs mainly in the macrocyclic structure by having a 27 membered ring system due to additional methylene carbon in the polyketide moiety. Lagunamide C displayed potent cytotoxic activity against a panel of cancer cell lines, such as P388, A549, PC3, HCT8, and SK-OV3 cell lines, with IC₅₀ values ranging from 2.1 nM to 24.4 nM. Compound 1 also displayed significant antimalarial activity with IC₅₀ value of 0.29 μM when tested against Plasmodium falciparum. In addition, lagunamide C exhibited weak anti-swarming activity when tested at 100 ppm against the Gram-negative bacterial strain, Pseudomonas aeruginosa PA01.     

N-Methylsansalvamide, a cytotoxic cyclic depsipeptide from a marine fungus of the genus fusarium

N-Methylsansalvamide (1), a new cyclic depsipeptide, was isolated from extracts of a cultured marine fungus, strain CNL-619, identified as a member of the genus Fusarium. N-Methylsansalvamide exhibits weak in vitro cytotoxicity in the NCI human tumor cell line screen (GI50 8.3 microM). The structure of 1 was determined by combined spectral and chemical methods.     

Enzymatic syntheses and selective hydrolysis of O-beta-D-galactopyranosides using a marine mollusc beta-galactosidase

The use of crude extract of the hepatopancreas of Aplysia fasciata, a large mollusc belonging to the order Anaspidea containing a beta-galactosidase activity, was reported for the synthesis of different galactosides. Good yields with polar acceptors and the uncommon beta-1-3 selectivity in the transgalactosylation reactions with most of the acceptors were observed. A beta-1-2 selectivity in the hydrolytic conditions was also observed and discussed.     

Isolation of a natural sterol and an aliphatic acid from Vernonia cinerea

From the roots of Vernonia cinerea a new natural sterol and a new aliphatic acid characterized as stigmast-5,17(20)-dien-3β-ol and 26-methylheptacosanoic acid, respectively, have been isolated together with stigmasterol and sitosterol.