The antitumor effect of bromophenol derivatives <Emphasis Type="Italic">in vitro</Emphasis> and <Emphasis Type="Italic">Leathesia nana</Emphasis> extract <Emphasis Type="Italic">in vivo</Emphasis>

To investigate the antitumor effect of bromophenol derivatives in vitro and Leathesia nana extract in vivo, six bromophenol derivatives 6-(2,3-dibromo-4,5-dihydroxybenzyl)-2,3-dibromo-4,5-dihydroxy benzyl methyl ether (1), (+)-3-(2,3-dibromo-4,5-dihydroxyphenyl)-4-bromo-5,6-dihydroxy-1,3-dihydroisobenzofuran (2), 3-bromo-4-(2,3-dibromo-4,5-dihydroxybenzyl)-5-methoxymethyl-pyrocatechol (3), 2,2′,3,3′-tetrabromo-4,4′,5,5′-tetrahydroxy-diphenylmethane (4), bis(2,3-dibromo-4,5-dihydroxybenzyl) ether (5), 2,2′,3-tribromo-3′,4,4′,5-tetrahydroxy-6′-ethyloxymethyldiphenylmethane (6) were isolated from brown alga Leathesia nana, and their cytotoxicity were tested by MTT assays in human cancer cell lines A549, BGC-823, MCF-7, B16-BL6, HT-1080, A2780, Bel7402 and HCT-8. Their inhibitory activity against protein tyrosine kinase (PTK) with over-expression of c-kit was analyzed also by ELISA. The antitumor activity of ethanolic extraction of Leathesia nana (EELN) was evaluated on S₁₈₀-bearing mice. All compounds showed very potent cytotoxicity against all of the eight cancer cell lines with IC₅₀ below 10 μg/mL. In PTK inhibition study, all bromophenol derivatives showed moderate inhibitory activity and compounds 2, 5 and 6 showed significant bioactivity with the inhibition ratio of 77.5%, 80.1% and 71.4%, respectively. Pharmacological studies reveal that EELN could inhibit the growth of Sarcoma 180 tumor and increase the indices of thymus and spleen to improve the immune system remarkably in vivo. Results indicated that the bromophenol derivatives and EELN can be used as potent antitumor agents for PTK over-expression of c-kit and considered in a new therapeutic strategy for treatment of cancer. Supported by the National High Technology Research and Development Program of China (863 Program, No. 2007AA09Z410) and Knowledge Innovation Program of Chinese Academy of Sciences (No. KZCX2-YW-209)     

Chemical constituents of marine algal-derived endophytic fungus Exophiala oligosperma EN-21

Seven compounds (1–7) were identified from the cultivation of the endophytic fungus Exophiala oligosperma (EN-21) that was isolated from the inner tissue of the marine red alga Laurencia similis. Their structures were identified with spectroscopic and chemical methods as 2-phenoxynaphthalene (1), (2S, 3R, 4E, 8E)-1-O-β-D-glucopyranosyl-3-hydroxy-2-[(R)-2′-hydroxyoctadecanoyl] amino-9-methyl-4, 8-octadeca-diene (2), (22E,24R)-ergosta-7,22-dien-3β,5α,6β-triol (3), (22E, 24R)-3β, 5α, 9α-trihydroxy- ergosta-7, ...     

Steroids from green alga <Emphasis Type="Italic">Chaetomorpha basiretorsa</Emphasis> Setchell

Six steroids have been isolated from ethanolic extract of green alga Chaetomorpha basiretorsa Setchell by a combination of repeated normal phase silica gel and Sephadex LH-20 gel column chromatography as well as recrystallization. Using spectroscopic methods including MS and NMR, their structures were determined as β-1awsaritol （1）, saringosterol （2）, 24-hydroperoxy-24-vinyl- cholesterol （3）, β-stigmasterol （4）, stigmast-4-en-3α, 6β-diol （5）, 29-hydroxystigmasta-5, 24 （28）-dien-3β-ol （6）. All these compounds were obtained from this genus for the first time and they were inactive （IC50〉10μg/ml） against KB, Bel-7402, PC-3M, Ketr 3 and MCF-7 cell lines.     

The in vitro effects of steroids,human chorionic gonadotropin and cyanoketone on germinal vesicle breakdown of striped mullet (Mugil cephalus L.) oocytes

The in vitro effects of steroids, human chorionic gonadotropin (HCG) and cyanoketone on germinal vesicle breakdown (GVBD) of striped mullet (Mugil cephalus L.) oocytes were investigated. All concentrations of HCG (5,10,50 I.U./ml), progesterone and pregnenolone at the highest concentrations(lug/ml) were moderately effective in inducing GVBD, whereas 17β-estrodiol, cortisol, testosterone, 11β-hydroxyandrostenedione and 11-ketotestosterone did not stimulate GVBD. 17α, 20βdihydroxy-4-pregnen-3-one (17α, 20βdiOHprog) and deoxycorticosterone (DOC) were the most potent steroids in stimulating final oocyte maturation. The results indicate that C21 hydroxylated steroids are potent inducers of final maturation in mullet. Further, co-incubations with 17β-estradiol, cortisol and testosterone did not alter the maturation-inducing effects of HCG or 17α,20βdiOHprog. Cyanoketone, a blocker of 3βHSD activity, was only partially effective in blocking GVBD induced by HCG. This suggests that Δ5 (pregnenolone derived) and Δ4 steroids may be involved in final oocyte maturation in this species.     

Synthesis and protein tyrosine phosphatase 1B inhibition activities of two new synthetic bromophenols and their methoxy derivatives

3-bromo-4,5-bis(2,3-dibromo-4,5-dihydroxybenzyl)-1,2-benzenediol (1) is a natural bromophenol isolated from the red algae Rhodomela confervoides that exhibits significant inhibition against protein tyrosine phosphatase 1B (PTP1B). Based on its activity, we synthesized two new synthetic bromophenols and their methoxy derivatives from vanillin using the structure of natural bromophenol 1 as a scaffold. The structures of these bromophenols were elucidated from 1H NMR, 13C NMR, and high resolution electron ionization mass spectrometry as 2,3-dibromo-1-(2p-bromo-6p-(3q,4q-dimethoxybenzyl)- 3p,4p-dimethoxybenzyl)-4,5-dimethoxybenzene(2),2,3-dibromo-1-(2p-bromo-6p-(2q-bromo-4q,5q-dimethoxy-benzyl)-3p,4p-dimethoxybenzyl)-4,5-dimethoxybenzene(3),3,4-dibromo-5-(2p-bromo-6p-(2q-bromo-4q,5q-dihydroxybenzyl)-3p,4p-dihydroxybenzyl)pyrocatechol(4)and 3,4-dibromo-5-(2p-bromo-6p-(3q,4q-dihydroxybenzyl)-3p,4p-dihydroxybenzyl)pyrocatechol (5).PTP1B inhibition activities of these compounds were evaluated using a colorimetric assay,and compounds 3 and 4 demonstrated interesting activity against PTP1B.     

Subunit composition and chromophore content of R-phycoerythrin fromPorphyra haitanensis

R-phycoerythrin fromPorphyra haitanensis exists in two aggregation states with different molecular weights. A more highly aggregated form, RPE I, was chromatographed on Bio-Rex 70 column with urea solution (pH 3.0) as eluent, and the molecular weights of the 3 subunits (α, β, γ) obtained were determined on SDS-PAGE at 18000, 19200 and 30000, respectively. α subunit carried two phycoerythrobilin (PEB); β subunit, three PEB and one phycourobilin (PUB); γ subunit, one PEB and three PUB chromophores. The molar ratio of α, β, and γ subunits of RPE I was 6: 6: 1, and their subunit composition was confired to be (αβ)₆γ on account of the molecular weight of RPE I, 232000. A lower aggregated form, RPE II, contained α and β subunits similar to those of RPE I, but its subunit composition was the (αβ) monomer of RPE. Contribution No. 2108 from the Institute of Oceanology, Chinese Academy of Sciences     

Synthesis of three bromophenols from red algae as PTP1B inhibitors

Bromophenols are a set of natural products widely distributed in seaweed, most of which exhibit interesting and useful biological activities. To develop a reliable and efficient synthetic route to these natural bromophenols, three of them, 3,4-dibromo-5-(2′-bromo-3′,4′-dihydroxy-6′-methoxymethyl-benzyl)-benzene-1,2-diol (compound 9), 3,4-dibromo-5-(2′-bromo-6′-ethoxy methyl-3′,4′-dihydroxybenzyl)-benzene-1,2-diol (compound 10), and 3-bromo-4-(3′-bromo-4′,5′-dihydroxy benzyl)-5-(ethoxymethyl)-benzene-1,2-diol (compound 14), isolated from red marine algae, have been synthesized in eight steps with an overall yield of 14.4%, 14.4%, and 18.2% respectively, via a practical approach employing bromination, Wolff-Kishner-Huang reduction and a Friedel-Crafts reaction as key steps. The protein tyrosine phosphatase 1B (PTP1B) inhibitory activities of the synthetic compounds were evaluated by the colorimetric assay. The results show that these compounds are moderate PTP1B inhibitors. The synthesis of these bromophenol derivatives makes in vivo studies of their structure-activity relationships and inhibition activity against PTP1B possible.     

Comparative studies on structural feature of agar polysaccharides fromPorphyra haitanensis grown in south and north China

The structural feature of agar polysaccharides fromPorphyra haitanensis grown in south China and transplanted to the north was investigated by fractionation on DEAE—Sephadex A 50, chemical analysis, and infrared and¹³C-NMR spectroscopy. The agars composed mainly of charged molecules were eluted from DEAE—Sephadex A 50 with 1.0 mol/L NaCl solution from the southernP. haitanensis and with 0.5 mol/L NaCl from the northern one. The¹³C-NMR spectra showed that agarobiose [(1→3)-β-D-galactopyranosyl-(1→4)-3,6-anhydro-α-L-galactopyranose] and the biological precursor of agarobiose [(1→3)-β-D-galactopyra nosyl-(1→4)-6-sulfate-α-L-galactopyranose] were the major disaccharide repeating units in the charged fractions. The content of the biological precursor in the agar polysaccharides from southernP. haitanensis was higher than that in the northern one, the content of the biological precursor extracted from cold water was higher than that from hot water, and the content of 6-OMe-D-galactose in the southernP. haitanensis polysaccharides was higher than in the northern one. This distinct difference will be of significance for further study of the physiological characters ofP. haitanensis. Contribution No. 1849 from the Institute of Oceanology, Academia Sinica.     

Chemical constituents of marine medicinal mangrove plant <Emphasis Type="Italic">Sonneratia caseolaris</Emphasis>

Twenty-four compounds including eight steroids (1–8), nine triterpenoids (9–16, 24), three flavonoids (20–22), and four benzenecarboxylic derivatives (17–19, 23) were isolated and identified from stems and twigs of medicinal mangrove plant Sonneratia caseolaris. The structures of the isolated compounds were determined by extensive analysis of their spectroscopic data. Among these metabolites, compounds 1, 4–20 and 22–24 were isolated and identified for the first time from S. caseolaris. In the in vitro cytotoxic assay against SMMC-7721 human hepatoma cells, compound 21 (3′,4′,5,7-tetrahydroxyflavone) exhibited significant activity with IC₅₀ 2.8 μg/mL, while oleanolic acid (14), 3,3′-di-O-methyl ether ellagic acid (18), and 3,3′,4-O-tri-O-methyl ether ellagic acid (19) showed weak activity. None of these compounds displayed significant antibacterial activites. Supported by the National Natural Science Foundation of China (No. 30770234); Knowledge Innovation Program of Chinese Academy of Sciences (KZCX2-YW-211-04); Department of Science and Technology of Shandong Province (No.2006GG2205023)     

The chemical constituents from red alga Gymnogongrus flabelliformis Harv.

Eight compounds were isolated from red alga Gymnogongrus flabelliformis Harv. In normal phase silica gel, Sephadex LH-20 gel column chromatography, reverse phase HPLC, and recrystallization. Based on MS and ID NMR spectroscopic data, their structures were determined as： stigmast-4-en-3-one （Ⅰ）, cholest-4-en-3-one （Ⅱ）, cholesterol （m）, uracil （Ⅳ）, uridine （V）, adenosine （Ⅵ）, succinic acid （Ⅶ）, and 5-hydroxy-4-methyl-5-pentyl-2,5-dihydro-furan-2-on （Ⅷ）. All of them were obtained from this species for the first time. Cytotoxicity of these compounds was screened using standard MTT method, but all the compounds were inactive （IC50 〉 10 μg/ml）.     

Structural studies on agar fractions extracted sequentially from chinese red sea weeds:Gracilaria sjeostedtii,G. textorii andG. salicornia using13C-NMR and IR spectroscopy

The polysaccharides of three species of ChineseGracilaria (Rhodophyta) were extracted by sequential solvent extraction and the chemical structures of the fractions obtained were investigated by using¹³C-NMR and IR spectroscopy. The¹³C-NMR spectra showed that the main extract, cold water fraction, fromG. sjeostedtii consisted of repeating disaccharides of agarobiose and (1→3)-β-D-galactopyranosy 1-(1→4)-α-L-galactopyranose 6-sulfate, the latter being a precursor to agarobiose and converted to agarobiose after alkali modification. The component of the 60% ethanol fraction fromG. textorii was composed mainly of agarose 6-sulfate, 6-O-methyl-agarose and some amount of agarose, whse structures remained unchanged after alkali treatment. The autoclave extract fromG. salicornia revealed a complex structure composed of 6-O-methyl-agarose, 2-O-methyl-agarose, branched 4-O-methyl-L-galactose and agarose 4-sulfate. All the substituents in these agaroses were stable against alkali treatment. The alkali extracts fromG. sjeostedtii andG. textorii showed the distinct spectra of amylopectin-like floridean starch, while that fromG. salicornia showed a mixture of agaroses and floridean starch. From the IR spectra the relative contents of total sulfate, 3,6-anhydro-galactose, 4-sulfate and 6-sulfate in some fractions of polysaccharides of the algae were qualitatively estimated by the ratios of absorption bands of 1250/2920, 930/2920, 850/2920 and 820/2920, respectively. Contribution No. 1489 from Institute of Oceanology, Academia Sinica     

Effects of Mg2+, NaCl, citric acid, and other factors on synthesis and accumulation of β-carotene inDunaliella salina

The effect of Mg²⁺, NaCl and citric acid on the accumulation of β-carotene inDunaliella Salina was studied. The experimental results showed that 10.5 mmol/L Mg²⁺, 5 mol/L NaCl, 3 μmol/L citric acid, and CO₂ are favorable forDunaliella Salina cell growth and β-carotene accumulation. After 144 h culture under the above conditions, theDunaliella Salina biomass increased by 7.18 times; β-carotene reached 9.61%. Project 89023990 supported by the Shandong Natural Science Fund.     

THE DYNAMIC EFFECTS OF A FINITE DEPTH AMBIENT FLUID ON BOTTOM EDDY

Scaling analysis shows that if o(ε~2, β_1ε, γε) ~o (δ), frontal geostrophic dynamics governs the behav-ior of an isolated bottom eddy in a finite depth ambient fluid; and that the ambient flow induced bybottom eddy migration satisfies quasi-geostrophic dynamics. This two layer model includes the impor-tant processes of advection of bottom eddy due to ambient flow, baroclinic instability, and form dragintroduced by Rossby waves. The numerical results show that the three processes enhance theinstability and alter the migration speed of the bottom eddy, and that the form drag induces asignificant meridional drift of the eddy.     

Structural studies on the oligomers from the polysaccharide ofGracilaria textorii (Rhodophyta) using β-agarase and13C-NMR spectroscopy

The 60% ethanolic extract fromGracilaria textorii (Rhodophyta) was degraded with β-agarase, and certain charged (sulfated) and neutral oligosaccharides were separated by using DEAE Sephadex A25 and Bio- gel P6, P2 chromatographic techniques. Some of the charged oligomers were verified to be neoagarotetraose-6³-sulfate (DP2), neoagarohexaose-6³, 6⁵-disulfate (DP3) and neoagarooctaose-6³, 6⁵, 6⁷-trisulfate (DP4) by using¹³C- and¹H-NMR spectroscopy. One neutral oligomer was assumed to be a mixture of methylated neoagarotetraoses (DP2) by¹H-NMR spectroscopy. These oligomers assigned by their chemical shifts may be used as the model compounds for the structural investigation of the agar-type sulfated polysaccharides using the β-agarase degradation method. Contribution No. 1749 from the Institute of Oceanology, Academia Sinica.     

Cloning and Characterization of an Abalone （Haliotis discus hannai ） Actin Gene

1　Introduction　Actinisanimportantcontractileproteinineukary oticcells .Itisoneofthetwomajorcomponentsin volvedinthecontractionofmusclecells .Innon mus clecells ,itisthemajorpartofcytoskeletoninvolvedinmanyprocessessuchascellmotility ,endocytosis ,exocyt…     

Effects of heavy metals （Pb^2＋ and Cd^2＋） on the ultrastructure, growth and pigment contents of the unicellular cyanobacterium Synechocystis sp. PCC 6803

The unicellular cyanobacterium Synechocystis sp. PCC 6803, a model organism known for its unique combination of highly desirable molecular genetic, physiological and morphological characteristics, was employed in the present study. The species was cultured in BG11 liquid medium contained various initial concentrations of Pb²⁺ and Cd²⁺ (0, 0.5, 1, 2, 4, 6 and 8 mg/L). The experiment was conducted for six days and the metal induced alterations in the ultrastructure, growth and pigment contents were assessed. Alterations in the ultrastructure of the Synechocystis sp. PCC 6803 cells became evident with the increased (>4 mg/L Pb²⁺) metal concentration. The photosynthetic apparatus (thylakoid membranes) were found to be the worst affected. Deteriorated or completely destroyed thylakoid membranes have made large empty spaces in the cell interior. In addition, at the highest concentration (8 mg/L Pb²⁺), the polyphosphate granules became more prominent both in size and number. Despite the initial slight stimulations (0.2, 3.8 and 6.5% respectively at 0.5, 1 and 2 mg/L Pb²⁺), both metals inhibited the growth in a dose-dependent manner as incubation progressed. Pigment contents (chlorophyll α, β carotene and phycocyanin) were also decreased with increasing metal concentration. Cells exposed to 6 mg/L Pb²⁺, resulted in 36.56, 37.39 and 29.34% reductions of chlorophyll α, β carotene and phycocyanin respectively over the control. Corresponding reductions for the same Cd²⁺concentrations were 57.83, 48.94 and 56.90%. Lethal concentration (96 h LC₅₀) values (3.47 mg/L Cd²⁺ and 12.11 mg/L Pb²⁺) indicated that Synechocystis sp. PCC 6803 is more vulnerable to Cd²⁺ than Pb²⁺. Supported by the Chinese Scholarship Council     

The Antitumor Components from Marine-derived Bacterium Streptoverticillium luteoverticillatum 11014 Ⅱ

Eight known compounds were isolated from a marine-derived bacterium Streptoverticillium luteoverticillatum 11014 using bioassay-guided fractionations. Their structures were identified by spectral analysis as bis （4-hydroxybenzyl） ether （1）, p-hydroxyphenylethyl alcohol （2）, N-（4-hydroxyphenethyl） acetamide （3）, indole-3 carboxylic acid methyl ester （4）, dibenzo[b,e] [1,4]dioxine （5）, thymine （6）, cytosine deoxyribonucleoside （7） and 2, 3-butanediol （8）. These compounds were evaluated for their cytotoxic activities against K562 cell line with the SRB method for the first time. Compounds 2 and 4 showed cytotoxcities with IC50 values of 101.1 and 165.3 μolL^-1, respectively. All compounds were isolated from S. luteoverticillatum 11014 for the first time.     

Bioaccumulation of Pb<Superscript>2+</Superscript> and its effects on growth,morphology and pigment contents of <Emphasis Type="Italic">Spirulina</Emphasis> (<Emphasis Type="Italic">Arthrospira</Emphasis>) <Emphasis Type="Italic">platensis</Emphasis>

A laboratory experiment was conducted to assess the bioaccumulation of Pb²⁺ and its effects on growth, morphology and pigment contents of Spirulina (Arthrospira) platensis. The specimen cultured in Zarrouk liquid medium was treated with various initial metal concentrations (0, 5, 10, 30, 50 and 100 μg mL⁻¹). The growth of S. platensis was adversely affected by Pb²⁺ at high concentrations (30, 50 and 100 μg mL⁻¹). However, at low concentrations (5 μg mL⁻¹), Pb²⁺ could stimulate its growth slightly. The pigment contents (chlorophyll α and β carotene) were decreased in a dose-dependent manner. The highest reductions (67% and 53% respectively in chlorophyll α and β carotene) were observed in 100 μg mL⁻¹ treatment group. The LC₅₀ (96 h) of Pb²⁺ was measured as 75.34 μg mL⁻¹. Apart from a few cases of filament breakages at elevated concentrations (50 and 100 μg mL⁻¹), morphological abnormalities are not specific. Metal bioaccumulation increased with Pb²⁺ concentrations, but decreased with exposure time. The maximum accumulated amount was 188 mg g⁻¹ dry weight. The bioconcentration factor (BCF) reached to a peak at day 2, followed by a gradual reduction for all the exposure concentrations. S. platensis is able to tolerate considerably high Pb²⁺ concentrations. Consequently it can be used as a potential species to remove heavy metal from contaminated waters.     

A comparative analysis of four kinds of polysaccharides purified from Furcellaria lumbricalis

Four kinds of polysaccharides (FA, FB, FC and FD) were extracted and purified from a red alga Furcellaria lumbricalis. Their monosaccharide composition, average molecular mass and structural feature were determined by gas chromatography (GC), high performance gel permeation chromatography (HPGPC) and Fourier transform infrared spectroscopy (FTIR), respectively. The results showed that FA and FB were composed of galactose (Gal), 3,6-anhydro-galactose (AnG) and 4-sulfated-galactose (G4S), which were structurally similar to β-carrageenan and κ-carrageenan. FC was composed of sulfated galactose and small amount of AnG, indicating that it was a hybrid-sulfated galactan. FD was composed of α-1,4-glucose and belonged to floridean starch. The respective yields of FA, FB, FC and FD were 4.84%, 33.74%, 8.76% and 13.5%, and the corresponding were 537 kD, 432 kD, 513 kD and 51 kD respectively. Owning to its high content (>60%) of polysaccharide, F. lumbricalis will be a good source of marine sulfated polysaccharide and shows its potential utilization in food and medicine.