Spermatozoid life-span of two brown seaweeds, Saccharina japonica and Undaria pinnatifida, as measured by fertilization efficiency

During sexual reproduction of seaweeds, spermatozoid (sperm) discharge is triggered by chemical messengers (pheromones) released by the female gametes. The chemotactic ability of the sperm ensures fertilization success. Using unialgal male and female gametophyte material under designated standard gametogenesis testing (SGT) conditions, the potential life-span of the sperm of two seaweeds, Saccharina japonica and Undaria pinnatifida, was assessed by their ability to fertilize eggs. Results show that within 20-30 min after being discharged, sperm of both species could complete fertilization without an apparent decline in fertilization rate. Although fertilization rate 60–120 min after sperm discharge dropped significantly in both species, some sperm were viable enough to fertilize the eggs. In S. japonica, at 12°C, some sperm were able to fertilize eggs up to 12 h after discharge. In both species, egg discharge rates (EDR) in the male and female mixed positive controls were significantly higher than those of all the sperm-testing groups. Doubling the seeded male gametophytes of S. japonica in the SGT tests significantly increased the EDR, further confirming the effect of the presence of the male on the female in terms of facilitating egg discharge from oogonia.     

High genetic diversity in gametophyte clones of Undaria pinnatifida from Vladivostok, Dalian and Qingdao revealed using microsatellite analysis

Breeding practice for Undaria pinnatifida (Harvey) Suringar requires the screening of a large number of offspring from gametophyte crossings to obtain an elite variety for large-scale cultivation. To better understand the genetic relationships of different gametophyte cultures isolated from different sources, 20 microsatellite loci were screened and 53 gametophyte clone cultures analyzed for U. pinnatifida isolated from wild sporophytes in Vladivostok, Russia and from cultivated sporophytes from Dalian and Qingdao, China. One locus was abandoned because of poor amplification. At the sex-linked locus of Up-AC-2A8, 3 alleles were detected in 25 female gametophyte clones, with sizes ranging from 307 to 316 bp. At other loci, 3 to 7 alleles were detected with an average of 4.5 alleles per locus. The average number of alleles at each locus was 1.3 and 3.7 for Russian and Chinese gametophyte clones, respectively. The average gene diversity for Russian, Chinese, and for the combined total of gametophyte clones was 0.1, 0.4, and 0.5, respectively. Russian gametophyte clones had unique alleles at 7 out of the 19 loci. In cluster analysis, Russian and Chinese gametophyte clones were separated into two different groups according to genetic distance. Overall, high genetic diversity was detected in gametophyte clones isolated from the two countries. These gametophyte cultures were believed to be appropriate parental materials for conducting breeding programs in the future.     

Isolation of protoplasts from <Emphasis Type="Italic">undaria pinnatifida</Emphasis> by alginate lyase digestion

The aim of this study is to isolate protoplasts from Undaria pinnatifida. Protoplasts of the alga were isolated enzymatically by using alginate lyase, which was prepared by fermenting culture of a strain Vibrio sp. 510. Monofacterial method was applied for optimizing digestion condition. The optimum condition for protoplast preparation is enzymatic digestion at 28°C for 2h using alginate lyase at the concentration of 213.36 U (8 mL) every 0.5g fresh thalline with NaCl 50 and at the shaking speed of 150 r min⁻¹ during digestion. The protoplast yield can reach 2.62±0.09 million per 0.5 g fresh leave under the optimum condition. The enzyme activity is inhibited by Ca²⁺ and slightly enhanced by Fe²⁺ and Mn²⁺ at concentrations of 0.05, 0.08 and 0.10 mol L⁻¹.     

Production of the Blood Pressure Lowing Peptides from Brown Alga （ Undaria pinnatifida ）

Brown alga （ Undaria pinnatifida） was treated with alginate lyase and hydrolyzed using 17 kinds of proteases and the inhibitory activity of the hydrolysates for the angiotensin-I-converting enzyme （ACE） was measured. Four hydrolysates with potent ACE-inhibitory activity were administered singly and orally to spontaneously hypertensive rats （SHRs）. The systolic blood pressure of SHRs decreases significantly after single oral administration of the brown alga hydrolysates by pro- tease S ＇ Amano＇ （from Bacillus stearothermophilus） at the concentration of 10 （mg protein） （kg body weight）^ - 1. In the 17 weeks of feeding experiment, 7-week-old SHRs were fed standard diet supplemented with the brown alga hydrolysates for 10 weeks. In SHRs fed 1.0 and 0.1% brown alga hydrolysates, elevating of systolic bloodpressure was significantly suppressed for 7 weeks. To elucidate the active components, the brown alga hydrolysates were fractionated by 1-butanol extraction and HPLC on a reverse-phase column. Seven kinds of ACE-inhibitory peptides were isolated and identified by amino acid composition analysis, sequence analysis, and LC-MS with the results Val-Tyr, Ile-Tyr, Ala-Trp, Phe-Tyr, Val-Trp, Ile-Trp, and Leu-Trp. Each peptide was determined to have an antihypertensive effect after a single oral administration in SHRs. The brown alga hydrolysates were also confirmed to decrease the blood pressure in humans.     

Production of the Blood Pressure Lowing Peptides from Brown Alga ( Undaria pinnatifida )

1 IntroductionIncreased blood pressure appears to be one of theprimary risk factors of circulatory organ diseases suchas encepharo-apoplexy, encepharo-infarction and cardi-ac infraction. Angiotensin-I-converting enzyme (ACE)plays an important role in the rennin-angiotensin sys-tem by regulating blood pressure. Antihypertensivedrugs such as captopril and enalapril are potent ACEinhibitors (Ondetti et al., 1977). Recently, severalinhibitory peptides derived from food proteins havebeen isolat…     

Structural characterization of sulfated galactofucan from Undaria pinnatifida and its effect on type 2 diabetic mice

Journal of Oceanology and Limnology - A sulfated galactofucan designated as UpG was obtained from the brown algae Undaria pinnatifida by calcium chloride extraction. Chemical analyses showed that...     

Optimization of culturing condition and medium composition for the production of alginate lyase by a marine <Emphasis Type="Italic">Vibrio</Emphasis> sp. YKW-34

Carbohydrases secreted by marine Vibrio sp. YKW-34 with strong Laminaria cell wall degrading ability were screened, and among them alginate lyase was found to be dominant. The effects of medium composition and culturing condition on the production of alginate lyase by marine Vibrio sp. YKW-34 in flask were investigated in this study. In the culture medium of marine broth, no alginate lyase was produced. The activity of the alginate lyase, after being induced, reached 5 UmL^-1. The best inoculum volume and inoculum age were 10% and 12h, respectively. The optimal temperature for alginate lyase production was 25℃. The fermentation medium was composed of 0.5% of Laminaria powder and 0.2% of KNO3 with an initial acidity of pH 8.0. Alginate could induce alginate lyase production but not as efficiently as Laminaria powder did. The addition of fucoidan, cellulose and glucose had negative effect on the alginate lyase production. Other kinds of nitrogen sources, such as yeast extract, beef extract and peptone, had positive effect on the growth of the microorganism and negative effect on alginate lyase production. In addition, the time course of alginate lyase production under the optimized condition was described. The optimal harvest time was 48 h.     

Fucoxanthin Isolated from Undaria pinnatifida Can Interact with Escherichia coli and lactobacilli in the Intestine and Inhibit the Growth of Pathogenic Bacteria

Fucoxanthin is a xanthophyll-type carotenoid that provides many benefits to human health. However, the mechanism by which fucoxanthin interacts with microbes and inhibits pathogenic bacteria is unknown. In this study, we investigated the effects of fucoxanthin isolated from the edible seaweed Undaria pinnatifida on pathogenic bacteria Escherichia coli and lactobacilli both in vitro and in vivo. Fucoxanthin strongly inhibited the growth of Gram-positive pathogenic bacteria but was less effective against Gram-negative bacteria. Fucoxanthin extracted from the crude mixture had a recovery rate of 93.38% and a purity of 82.70%, which were higher than those of fucoxanthin extracted using a previous method. Fucoxanthin also promoted the growth of intestinal microbes in mice. Fucoxanthinol, a metabolite of fucoxanthin, was generated in the culture media. Fucoxanthin can be deacetylated into fucoxanthinol not only by conventional digestive enzymes in the digestive tract, but also by E. coli and lactobacilli in the intestine. These results indicate that fucoxanthin interacts with and influences E. coli and lactobacilli in the intestine. Therefore, fucoxanthin isolated from Undaria pinnatifida possibly can be applied in human health maintenance.     

Preparation,Characterization and Pharmacokinetics of Fluorescence Labeled Propylene Glycol Alginate Sodium Sulfate

A rapid and sensitive fluorescence labeling method was developed and validated for the microanalysis of a sulfated polysaccharide drug,namely propylene glycol alginate sodium sulfate （PSS）,in rat plasma.Fluorescein isothiocyanate （FITC） was selected to label PSS,and 1,6-diaminohexane was used to link PSS and FITC in order to prepare FITC-labeled PSS （F-PSS） through a reductive amination reaction.F-PSS was identified by UV-Vis,FT-IR and ^1H-NMR spectrum.The cell stability and cytotoxicity of F-PSS were tested in Madin-Darby canine kidney （MDCK） cells.The results indicated that the labeling efficiency of F-PSS was 0.522%±0.0248% and the absolute bioavailability was 8.39%.F-PSS was stable in MDCK cells without obvious cytotoxicity.The method was sensitive and reliable; it showed a good linearity,precision,recovery and stability.The FITC labeling method can be applied to investigating the absorption and metabolism of PSS and other polysaccharides in biological samples.     

Construction of Porphyra yezoensis Pure Line from Protoplasts and Its 18S rDNA Sequence Determination

1　Introduction　PorphyraisthemainobjectforalgafarmingandplaysaveryimportantroleinChinesemarineindus tries.Recently ,therehasbeenagreatlossinPor phyracultivationduetothedegenerationoftheculti var,sothereisanincreasingdemandforgoodPor phyracultivars .Theprerequisiteofthetraditionalbreed ingandbioengineeringresearchofPorphyraistheconstructionofpurelines .Traditionally ,theclassifi cationofPorphyrawasaccordingtotheirmorphologi calcharacteristics .However ,mostmorphologicalfea turesofPorphyraar…     

Isolation of plasmid from the blue-green algaSpirulina platensis

CCC plasmid was isolated from an economically important blue-green alga —Spirulina platensis (1.7×10⁶ dalton from the S₆ strain and 1.2×10⁶ dalton from the F₃ strain) using a rapid method based on ultrasonic disruption of algal cells and alkaline removal of chromosomal DNA. The difference in the molecular weight of the CCC DNAs from the two strains differing in form suggests that plasmid may be related with the differentiation of algal form. This modified method, which does not use any lysozyme, is a quick and effective method of plasmid isolation, especially for filamentous blue-green algae. Contribution No. 79 of the Experimental Marine Biology Laboratory and 2153 of the Institute of Oceanology, Academia Sinica. This research was supported in part by The President of the Chinese Academy of Sciences Foundation.     

Isolation of novel psychrophilic bacteria from Arctic sea ice

The phylogenetic diversity of cuhurable psychrophilic bacteria associated with sea ice from the high latitude regions of Canadian Basin and Chukchi Sea, Arctic, was investigated. A total of 34 psychropilic strains were isolated using three methods of （ Ⅰ ） dilution plating （ at 4 ℃ ）, （ Ⅱ ） bath culturing （ at - 1 ℃ ） and dilution plating, and （ Ⅲ） cold shock （ -20 ℃ for 24 h）, bath culturing and dilution plating under aerobic conditions. Sea-ice samples were exposed to -20 ℃ for 24 h that might reduce the number of common microorganisms and encourage outgrowth of psyehrophilic strains. This process might be able to be introduced to isolation psychrophilic bacteria from other environmental samples in future study. 16S rDNA nearly full-length sequence analysis revealed that psychrophilic strains felled in two phylo- genetic divisions, γ-proteobacteria （in the genera ColweUia,Marinobaeter、Shewanella,Glaciecola,Marinomortas and Pseudoalteromortas ） and Cytophaga-Flexibacter-Baeteroides （ Flavobacterlum and Psyehroflexus）. Fifteen of bacterial isolates quite likely represented novel species （16S rDNA sequence similarity below 98% ）. One of strains （BSi20002） from Canadian Basin showed 100% sequence similarity to that of Marinobacter sp. ANT8277 isolated from the Antarctic Weddell sea ice, suggesting bacteria may have a bipolar distribution at the species level.     

Isolation and pharmacological activities of bromophenols from Rhodomela confervoides

Four bromophenols were isolated from the extract of marine red alga, Rhodomela confervoides by column chromatography and HPLC methods. By means of spectroscopic methods including IR, MS, 1D, and 2D-NMR techniques, their structures were elucidated as (1) 3-bromo-4,5-dihydroxy benzoic acid methyl ester; (2) his (2,3-dibromo-4,5-dihydroxybenzyl) ether; (3) 3,4-dibromo-5-(methoxymethyl)-1,2-benzenediol and (4)3-bromo-4,5-dihydroxy-benzaldehyde. Compound 1 was first isolated from the algae in nature, and 1, 4 were found to have selective cytotoxic activities against KB, Bel 7402 and A549 cells, 2 showed powerful antibacterial activities against Staphylococcus aurens and Pseudomonas aeruginosa.     

Isolation and characterization of phosphate-solubilizing bacteria from seagrass rhizosphere soil

Phosphate-solubilizing bacterial strains(6 Nos.) were isolated from the rhizosphere soils of two seagrasses(Halophila ovalis(R.Br.) Hook and Halodule pinifolia(Miki) Hartog) in the Vellar estuary.Experimental studies found that the strain PSSG6 was effective in phosphate solubilization with Phosphate Solubilization efficiency index E = 375 ± 8.54,followed by the strain PSSG5 with Phosphate Solubilization efficiency index E = 275 ± 27.3.Of the 6 strains isolated,the strains PSSG4 and PSSG5 be-longed to the genus Bacillus,and PSSG1,PSSG2 and PSSG3 were identified as Citrobacter sp.,Shigella sp.,and Klebsiella sp.,respectively,by conventional method,and PSSG6 was identified as Bacillus circulans using conventional and molecular methods.     

Isolation and characterization of a fucoidan-degrading bacterium from Laminaria japonica

Fucoidan, a polysaccharide containing abundant fucose and sulfate ester group, was prepared from Laminaria japonica. In order to obtain fucoidan-degrading enzyme, bacteria capable of degrading fucoidan were screened from kelp. A bacterial strain named RC2-3 was obtained, which degraded fucoidan by the maximum extent of 54% ± 1.3%, the highest among all bacterial isolates. High-performance size exclusion chromatography(HPSEC) showed that the molecular weight of fucoidan was gradually reduced by RC2-3 with culturing time, suggesting the production of fucoidan-degrading enzyme by RC2-3. Phylogenetic analysis of partial 16S ribosomal RNA gene(16S rDNA) sequence showed that RC2-3 belonged to the family Flavobacteriaceae. However, it showed different physiological and biochemical characteristics from the known Flavobacteriaceae members producing fucoidan-degrading enzyme, thus RC2-3 was proposed to be a new member of this family.     

Isolation and pharmacological activities of bromophenols from Rhodomela confervoides

Four bromophenols were isolated from the extract of marine red alga,Rhodomela confervoides by column chromatography and HPLC methods. By means of spectroscopic methods inclding IR, MS, 1D, and 2D-NMR techniques, their structures were elucidated as (1) 3-bromo-4,5-dihydroxy benzoic acid methyl ester; (2) bis (2,3-dibromo-4,5-dihydroxybenzyl) ether; (3) 3,4-dibromo-5-(methoxymethyl)-1,2-benzenediol and (4) 3-bromo-4,5-dihydroxy-benzaldehyde. Compound 1 was first isolated from the algae in nature, and 1, 4 were found to have selective cytotoxic activities against KB, Bel 7402 and A549 cells, 2 showed powerful antibacterial activities againstStaphylococcus aurens andPseudomonas aeruginosa. Supported by National “863” Program (No. 2004AA625030, 2001AA620503), Quingdao marine sciences project (No. 04-2-NN-26) and Key Knowledge Innovative Project of the Chinese Academy of Sciences (KZCX3-SW-215).     

徐闻8种珊瑚共附生真菌的分离及初步鉴定

从徐闻珊瑚保护区的8种珊瑚中分离共附生真菌,以获得珊瑚共附生可培养真菌的多样性信息。选用5种培养基,采用平板涂布法培养分离共附生真菌,并进行形态鉴定。8种珊瑚共分离121株真菌,鉴定了其中97株真菌,分属于14个属,青霉属和枝孢霉为优势种属,其次是曲霉属和木霉属。二异角孔珊瑚上分离出22株,数量最多;盔型珊瑚上分离菌株数量最少仅8株。除优势种属外,不同珊瑚样品上真菌种类分布有较大差异,炭角菌属和葡萄穗霉菌属均来源于盔形珊瑚,镰刀菌属在盔形珊瑚和角孔珊瑚上较为常见,盔形珊瑚样品上还分离出2株散囊菌属,1株节菱孢属,因此盔形珊瑚的种群最为丰富;不同培养基分离真菌的能力不同,淀粉培养基分离的菌株最多占所有分离菌株的30.6%,CDA培养基最少,但种类特异。     

Isolation and characterization of venom from nematocysts of jellyfish Rhopilema esculentum Kishinouye

The present work is the first report of the biochemical characterization of the venom from nematocysts of the jellyfish Rhopilema esculentum Kishinouye. The nematocysts were isolated by autolysis and centrifugation and separated by flow cytometry. Four types of nematocysts were identified: mastigophores, euryteles, and atrichous and holotrichous isorhiza. SDS-PAGE and amino acid analyses demonstrated that most of the proteins in the nematocyst extract were between 10 kDa and 40 kDa, and that glutamic acid was the main amino acid. A hemolytic activity assay showed that the activity of the nematocyst venom (RNV) was strongest in Tris-HCl buffer (50 mmol/L, pH 7.8, 5% glycerol, 0.5 mmol/L EDTA, 0.1 mol/L NaCl). The hemolytic activity was related to protein concentration and the HU50 against chicken erythrocytes was 0.91 μg/mL.     

Isolation and Characterization of Collagen from Squid(Ommastrephes bartrami) Skin

Collagen of squid(Ommastrephes bartrami) skin was examined in the present study. Histology showed that collagen fiber in the skin was partially cross-linked with muscle fiber. Acid-solubilized collagen(ASC) and pepsin-solubilized collagen(PSC) were extracted from the skin and characterized. The results of amino acid composition and electrophoretic patterns revealed that ASC and PSC were both type Ⅰ collagen,containing α1 and α2 chains. FTIR(fourier transform infrared spectroscopy) investigations confirmed t...