鲈鱼生长催乳素的分离纯化与N-末端序列分析

生长催乳素(SL)是生长激素/催乳素家族的一种新的脑垂体蛋白质激素,本研究首次从鲈鱼脑垂体中分离得到.鲈鱼脑垂体在碱性条件下,经葡聚糖凝胶(SephadexG-100)过滤和反相高效液相色谱(rpHPLC)分离出纯化的鲈鱼生长催乳素,(sbSL),并与大麻哈鱼生长催乳素(sSL)抗体发生特异性的Western免疫印迹交叉反应证实.SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析,发现鲈鱼生长催乳素分别是由分子量为23.2×103和28.4×103u两种形式组成(后者可能是一种糖基化形式).根据Edman降解原理,测得鲈鱼生长催乳素的N-末端16个氨基酸的序列,它与已知鱼类生长催乳素的N-末端序列比较,具有较高的同源性;由此进一步证实,分离纯化得到的鲈鱼生长催乳素是正确的.     

秋季南黄海表层海水中溶解氨基酸的分布与组成研究

用高效液相色谱法对2010年9月南黄海33个站位表层海水中总水解氨基酸(THAA)、溶解游离氨基酸(DFAA)、溶解结合氨基酸(DCAA)的分布、组成以及它们与环境因子的相关性进行了研究。结果表明:THAA平均浓度为2.08μmol/L,DFAA平均浓度为0.39μmol/L,DCAA平均浓度为1.69μmol/L。THAA浓度的分布大致呈近岸高、远岸低的特点;DFAA分布的规律性较差,在东部海域(中心约在35°N,123°E)出现高值。相关性分析显示溶解氨基酸与叶绿素a(Chl-a)、溶解无机氮(DIN)、细菌丰度、温度和盐度之间均无显著相关性。THAA中含量较高的个体氨基酸为丝氨酸、甘氨酸、天门冬氨酸、谷氨酸和丙氨酸,总和占70.59%。DFAA、DCAA中的主要成分与THAA基本相同。THAA中个体氨基酸的组成以34°N为界分为南北2个海区呈现明显的区域性特点。     

大黄鱼（Pseudosciaena crocea）一种新型抗菌肽的纯化及其生物学特性研究

以致病性溶藻弧菌为指示菌,通过酸粗提、固相萃取、Sephadex G-25柱凝胶层析和RP—HPLC等分离纯化技术,从大黄鱼胃肠组织中分离纯化抗菌肽。结果表明,大黄鱼肠道组织中存在着多种抗菌物质,并最终得到一种高纯度的新型抗菌肽AMP-W3。AMP—W3由10种氨基酸组成,其中天冬氨酸、谷氨酸和甘氨酸的含量较高。AMP...     

Seasonal trends in the abundance, composition and bioavailability of particulate and dissolved organic matter in the Chukchi/Beaufort Seas and western Canada Basin

The objectives of this study were to investigate the seasonality, abundance, sources and bioreactivity of organic matter in the water column of the western Arctic Ocean. The concentrations of particulate and dissolved amino acids and amino sugars, as well as bulk properties of particulate and dissolved organic matter (DOM), were measured in shelf, slope and basin waters collected during the spring and summer of 2002. Particulate organic matter concentrations in shelf waters increased by a factor of 10 between spring and summer. Dissolved organic carbon (DOC) and nitrogen (DON) concentrations exhibited only minor seasonal variations, whereas dissolved amino acid concentrations doubled between spring and summer, and dissolved amino sugars increased by 31% in shelf waters of the Chukchi and Beaufort Seas. Concentrations of DOC did not exhibit a significant seasonal change in surface waters of the Canada Basin, but dissolved amino acid concentrations increased by 45% between spring and summer. No significant seasonal differences were detected in the concentration or composition of DOM in waters below 100 m in depth. Concentrations of particulate and dissolved amino acids and amino sugars were strongly correlated with chlorophyll-a, indicating a plankton source of freshly produced organic matter. The amino acid and amino sugar compositions of freshly produced DOM indicated that a large portion of this material is bioavailable. While freshly produced DOM was found to be relatively bioreactive, preformed DOM in the Arctic appears to be less bioreactive but similar in degradation state to average DOM in the Atlantic and Pacific. These data demonstrate substantial summer production of POM and DOM on the Chukchi and Beaufort shelves that is available for utilization in shelf waters and export to the Canada Basin.     

Composition and bacterial utilization of free amino acids in tropical mangrove sediments

The composition and bacterial utilization of dissolved free amino acids (DFAA) in tropical mangrove sediments was examined. Amino acid concentrations (300–900 ng total DFAA ml⁻¹) and composition were similar to that of other organic-rich, anaerobic sediments with lowest and highest concentrations in the low and mid intertidal zones, respectively. The non-protein amino acid, β-glutamic acid, rarely reported in previous studies, was found as a major component of the interstitial DFAA pool. Intracellular amino acids from some cultured strains of sulphate-reducing bacteria (e.g. Desulfobacter app) showed the presence of β-glutamic acid as a major cellular constituent suggesting that these bacteria may be a source of this amino acid in mangrove pore waters. In high intertidal sediments, bacterial growth rates (μ) correlated significantly with total DFAA concentrations with depth.Amino acid concentrations and composition differed significantly between sediments and overlying tidal waters. Flux chamber experiments showed negligible amino acid flux out of the sediments in untreated chambers, but rates of amino acid flux ranged from 27 to 69 mgN m⁻² day⁻¹ (= 81–207 mg C.m⁻² day⁻¹) in chambers where poisons were applied to the sediment surface. Such fluxes could account for between 9–38% and 5–19% of the nitrogen and carbon required to support the levels of bacterial productivity measured in surface (0–1 cm) sediments. These experiments suggest that bacterial populations in surface sediments are capable of utilizing all of the amino acid flux to the sediment-water interface in tropical mangroves.     

扇贝碱性磷酸酶中Zn的作用

以华贵栉孔扇贝Chlamys nobilis(Reeve)为材料,获得聚丙烯酰胺凝胶电泳为单一蛋白区带的酶制剂。用凝胶过滤法测得该酶的分子量为159000,由20种氨基酸组成,共1118个氨基酸残基。每个酶蛋白含有4个Zn原子。将该酶的酶制剂制成脱Zn酶后,与相同浓度过度性两价金属离子作用,其活力回升以Zn最强,Co次之。若向脱Zn酶蛋白分别加入一定量Zn原子或Co原子,根据实验结果初步推测:每个酶蛋白的4个Zn原子中有2个Zn原子先与该酶结合,与酶的催化作用有密切关系;另外2个Zn原子与酶结合,则起维持酶结构的作用,与哺乳动物和Eschericha coli来源的碱性磷酸酶研究结果一致。此外,本实验采用可见吸收光谱和圆二色谱(CD)来确证Co能很好地取代该酶蛋白内的Zn,与Simpon报道相比较,得知当Co原子加入脱Zn酶时,先加2个Co~(2 )离子结合到该酶的结构部位,而后加2个Co~(2 )离子才结合到催化部位。     

汞、铅对非洲鲫鱼肝线粒体功能影响的初步研究

为了研究和探讨重金属毒物对海洋生物的致毒机理及其对生物体内各种生化过程的影响,继镉、铜对非洲鲫鱼肝线粒体功能影响的研究之后,本实验继续以非洲鲫鱼(Tilapia mossambica)为实验材料,采用氧电极法测定了氯化汞、硝酸铅对鱼肝线粒体(离体)呼吸控制率和氧化磷酸化效率的影响。     

东海陆架典型断面颗粒态氨基酸的分布及控制因素分析

采用高效液相色谱法,通过现场调查对东海典型PN断面(文中为C断面)的颗粒态氨基酸(Particulate Amino Acids,PAA)进行了分析,并结合叶绿素a(Chla)、颗粒有机碳(POC)、颗粒有机氮(PON)及颗粒态氨基酸的构型特征(D和L型)等参数探讨了该区颗粒有机氮的来源和降解情况。结果表明,在长江口最大浑浊带,受咸淡水混合和生物现场生产双重作用影响,POC、PON以及PAA的总浓度均达到了极大值,其中,受再悬浮作用影响,底层水体中的有机物呈现高度降解的状态;近岸水华区域的颗粒态氨基酸则更多来源于现场生产,而且POC/Chla质量比值与降解因子DI值的负相关特征表明冲淡水向海洋输送的过程中,现场生产力对颗粒有机碳的贡献比重逐渐增大,悬浮颗粒物也变得越来越新鲜。值得关注的是,一些D型氨基酸[如D型天冬氨酸(D-Asp),D型丙氨酸(D-Ala)]与细菌生物量之间存在良好的正相关性,暗示颗粒态氨基酸在受到物理水团和生物现场生产作用控制的同时,还受控于微微型浮游生物以及异养细菌。     

The free amino acid composition of seawater obtained without desalting and preconcentration

A conventional amino acid analyzer was modified to allow detection of dissolved free amino acids in a direct injection of 0.5 to 2.0 ml of seawater. The amino acids were monitored fluorimetrically by reaction with o-phthaldialdehyde after column chromatography. The accuracies of two commonly employed desalting methods, cation exchange resin and copper Chelex, were determined by comparison with the direct injection method. After correcting the results of the desalting methods for the procedural blank and recovery rate for each individual amino acid, the amino acid compositions on a mole percent basis as determined by these methods resembled approximately the true composition determined by direct injection. However, on an absolute basis the cation resin method overestimates and the copper Chelex method underestimates the total amount of dissolved free amino acids. The reasons for these discrepancies are discussed. Using the direct injection method, the spectra of dissolved free amino acids in surface seawater samples from different locations were determined. As opposed to the relatively uniform patterns reported in most past investigations, the amino acid spectra found were highly variable. In a number of samples, the acidic amino acids dominated the spectra. Valine was also high in many samples. In a depth profile the deep samples were dominated by arginine. Application of the method to real-time shipboard analyses of dissolved free amino acids is discussed.     

REPRODUCTIVE MECHANISM OF RUDITAPES PHILIPPINARUM ISOLATION OF SHEDDING HORMONE AND DETERMINATION OF BIOLOGICAL ACTIVITY

An active substance, shedding hormone capable of inducing parent clams to spawn and sper-miate, has preliminarily been isolated from the ovaries of Ruditapes philippinarum by means of ammonia sulfate precipitate of 25% concentration and Sephadex G-25 column chromatography in present study. The biological activity of the shedding hormone is above 95%. It is composed of sorts of amino acids by determination with an amino acid automatic analyzer. The experimental results indicate that the active substance is a polypeptide.     

海水中氨基酸对Cu（Ⅱ）在r—MnOOH上吸附的影响

实验测定了海水中氨基酸存在下Ｃｕ（Ⅱ）在ｒ－ＭｎＯＯＨ上吸附的等温线、Ｃｕ（Ⅱ）％－－ｐＨ曲线和Ｃｕ（Ⅱ）％－氨基酸深度曲线。结果表明，较低浓度氨基酸对Ｃｕ（Ⅱ）在ｒ－ＭｎＯＯＨ上吸附没有影响，而较高浓度氨基酸却起抑制作用，这主要是由于海水中Ｃｕ（Ⅱ）－氨基酸络合反应与ｒ－ＭｎＯＯＨ竞争Ｃｕ（Ⅱ）所致。此外，海水中较高浓度氨基酸只使Ｃｕ（Ⅱ）在ｒ－ＭｎＯＯＨ上吸附的表面分级离子交换稳定常数Ｋｉ（ｉ     

浙江十种经济海藻的氨基酸含量分析

采用以异硫氰酸苯脂（ＰＩＴＣ）为柱前衍生化的ＰＩＣＯ－ＴＡＧ反相高效液相色谱法对浙江沿岸１０种经济海藻的氨基酸含量进行测定,得出以上海藻的氨基酸色谱图。发现１０种海藻的氨基酸组成基本相同。氨基酸总含量达１０．５７％～３３．５１％,其中必需氨基酸占总量的２８．２８％～４５．２２％。该测定结果可以为海藻经济开发提供参考。     

Development of lipids during a spring plankton bloom in the northern North Sea: II. Dissolved lipids and fatty acids

During the spring plankton bloom (1976) in the northern North Sea the fatty acid and lipid distribution of the chloroform extractable fraction of seawater was analyzed by gas chromatography and thin-layer chromatography. The fatty acids in the seawater made up about 3% of the total dissolved organic matter. Maximum concentrations during the first phytoplankton bloom were in the range of 5 μmol C dm^?3. Palmitic acid (30.4%) and oleic acid (21.4%) as well as myristic, stearic and palmitoleic acids were the main fatty acids. The concentrations of fatty acids were higher below the thermocline in the deeper layers of the water column. The oleic acid showed large fluctuations, especially below the thermocline and seemed to be bound in a polar fraction. Different lipid classes containing fatty acids were determined by thin-layer chromatography. An estimation showed that the free fatty acid fraction made up the main portion (47.9%), followed by the triacylglycerol fraction (22.8%) and a more polar fraction with 25.1%. The triacylglycerol fraction appears to be more stable than the other fraction in the water column.     

Response of embryos of the american oyster, Crassostrea virginica, to heavy metal mixtures

The response of embryos of the American oyster, Crassostrea virginica, to various combinations of two or three metals (copper, mercury and zinc as the nitrates and chlorides) was studied in the laboratory at the optimal temperature and salinity regime for their development. The experimental design was a 3 × 3 × 3 factorial experiment using copper concentrations of 0,8 and 16 ppb; zinc concentrations of 0, 100 and 200 ppb and mercury concentrations of 0, 6 and 10 ppb. Highly significant toxic synergism was observed in the copper-zinc mixtures and three-metal mixtures of both salts, particularly at high concentrations. Although the chloride salt appeared to be slightly more toxic than the nitrate salt, the overall difference was not significant.     

The Importance of Posidonia oceanica and Cymodocea nodosa as Contributors of Free Amino Acids in Water and Sediment of Seagrass Beds

Abstract. Dissolved free amino acids (DFAA) were measured in sediment porewater and overlying sea water in seagrass beds of Posidonia oceanica and Cymodocea nodosa at a water depth of 8–10 m in Corsica, France. In the sea water, DFAA concentrations from 0.5 to 1.9μM were found. Lowest concentrations occurred in the afternoon in surface water samples, while higher concentrations were measured in morning surface samples and in all bottom samples. The most abundant amino acids were serine, glycine, and alanine. In the sediment, the upper 4cm contained the highest concentrations of DFAA (177nmol cm^-3 or 367 μM), while lower concentrations (<50nmol cm^-3 or 104 μM) prevailed deeper in the sediment. Glycine, glutamic acid, alanine, and β-alanine were the major components of the DFAA. The photosynthetic activity of the seagrasses influenced sediment porewater pools of DFAA. When a Posidonia bed was covered with a black polyethylene shield, the interstitial amino acids were reduced at a rate of 16nmol cm^-2 h^-1 (0-10cm depth). Conversely, extended daylight (floodlight) increased the DFAA pool in sediment depths below 4 cm. Free amino acids were released to the overlying sea water from both seagrass sediment and individual shoots, the rates being highest in the dark. The present investigation indicates that a substantial portion of the N-cycling of the seagrass sediment includes free amino acids.     

Growth Dynamics in Posidonia oceanica (L.) Delile

Abstract. Amounts of photosynthate in the rhizomes, in photosynthetically inactive basal parts of the leaves and different old leaf blades were examined. Winter leaf growth was supported by mobilisation of starch in the rhizome. This winter growth enables Posidonia oceanica to utilize the increased energy influx in early spring via the substantial leaf area already developed and to approach highest productivity in spring. During summer and autumn considerable concentrations of soluble carbohydrates were found in the leaves and rhizomes. Starch was stored in the rhizomes in concentrations of up to 6.8 % of dry weight. Levels of nitrogen and free amino acids were correlated with growth rates. The percentage of total nitrogen present as free amino acid-nitrogen decreased from November (35 %) to summer (less than 1 %), by which time leaf growth had stopped. In contrast to this, amounts of organic anions were low in winter and spring and reached their maximum in summer.
Within a shoot, sites of leaf growth were characterized by high amounts of total nitrogen and free amino acids (innermost leaves), while carbon Fixation was highest in the leaves #2–#4. Particularly high concentrations of soluble carbohydrates were found midway along leaf blades.
It is evident that this unusual growth rhythm of Posidonia oceanica was only possible because of the ability to store considerable amounts of carbon and nitrogen in the rhizomes.
The results of this investigation demonstrate on the one hand that the chemical composition of the plant is strongly correlated with growth and production, and on the other hand that it is dependent on environmental factors, such as energy influx and temperature, which change with season.     

中国对虾(Fenneropenaeus chinensis)血淋巴中凝血蛋白的分离纯化、鉴定及 cDNA 片段克隆、 组织表达谱分析

采用层析技术、N末端氨基酸序列分析及同源基因克隆等技术对中国对虾中国对虾血淋巴中的凝血蛋白(CP)进行了研究。结果表明,纯化的CP约为380kDa,其亚基的分子量约为190kDa,说明该蛋白是由两个相同的亚基组成的同源二聚体;中国对虾CP的N末端氨基酸序列与凡纳滨对虾、保罗美对虾具有100%的相似性,与其它对虾的也具有很高的相似性;获得了CP基因518bp的cDNA片段,分析表明该基因序列与其它虾类CP基因序列的具有很高的同源性;CP基因的组织表达谱分析发现心脏和表皮是该基因主要的合成表达部位。     

Toxic effects of certain heavy metals (Hg,Cd, Pb,As and Se) on the intertidal crab scylla serrata

The toxic effects of certain heavy metals (Hg, Cd, Se, As and Pb) on Scylla serrata, the common edible Indian marine crab, are evaluated. Mercuric chloride, phenyl mercuric acetate (PMA), cadmium chloride, selenium dioxide, arsenic trioxide and lead nitrate were tested individually in laboratory static bioassays using artificial seawater to determine the 24 to 96 h LC₅₀ values. As indicated by 96 h LC₅₀ values PMA was found to be the most toxic and lead nitrate the least toxic. CdCl₂ showed a 6·6 fold increase in toxicity with increase in the time of exposure from 24 to 96 h. Marked histopathological changes were noticed in the hepatopancreas and gill of Scylla serrata exposed to acute 96 h LC₅₀ values of these metal compounds. 30 days' long-term exposure to sublethal concentrations of mercury and cadmium brought about degenerative changes in the hepatopancreas and gill of exposed animals. Extensive cellular proliferation (hyperplasia) of secondary lamellae accompanied by cyst formation and necrotic regions were often seen in the gills of cadmium treated crabs. Interestingly enough, the free amino acid pool from muscle and hepatopancreas of treated animals, either after acute or long-term exposure, did not show any reduction in number or quantity     

Chemical and GC-MS characterization of marine dissolved lipids

The chloroform-extractable ‘lipid’ fraction of dissolved organic matter in seawater was analyzed by gravimetry, liquid chromatography, gas chromatography (GC), and gas chromatography—mass spectrometry (GC—MS). Gravimetric concentrations of dissolved lipids in the Gulf of Mexico were in the range of 60–160 μg 1^?1 in near-surface waters and 61–116 μg 1^?1 in near bottom waters and accounted for ～4% of the dissolved organic carbon. Over a 12-h sampling period and a 5-d sampling period extensive variability in dissolved lipid quantity and quality were observed. The major percentage of extractable weight was collected in the polar liquid chromatographic fraction (55–95%). Gas chromatographic concentrations of the aliphatic fractions were in the range of 0.014-0.187 μg 1^?1. Concentrations derived from gas chromatography were consistently lower than gravimetrically-derived concentrations. A number of compounds were tentatively identified by a combination of GC, GC—MS, and authentic standards. The major components of the analyzable dissolved lipids were n-alkanes (C₁₆C₃₂), pristane, phytane, methyl, ethyl and propyl esters of fatty acids. Minor components included olefins and cycloalkanes, aromatics, short-chained acids, and possibly a lactone and an alcohol. All concentrations and compounds were indicative of a fairly pristine environment. The n-alkane distribution appears to be the result of marine and terrestrial inputs superimposed on a chronic low-level background of oil pollution. It is suggested that the fatty acid esters and other fragment molecules are the résult of the degradation of humic substances. A number of potential indicators of source were isolated.     

金乌贼(Sepia esculenta)蛋白抗氧化肽的酶解制备及活性评价

采用木瓜蛋白酶水解金乌贼(Sepia esculenta)蛋白, 利用超滤、凝胶色谱和反相高效液相色谱从金乌贼蛋白水解物中制备抗氧化肽, 采用氨基酸序列分析仪和质谱(ESI-MS)确定抗氧化肽结构, 采用自由基清除实验和脂质过氧化抑制实验对多肽抗氧化能力进行评价。结果表明, 金乌贼蛋白经木瓜蛋白酶水解和分离纯化得到1个抗氧化肽(AEH-P3), 经氨基酸序列分析和质谱(ESI-MS)确定其结构为Ala-Pro-Pro-Glu-Asn-Gly-Met-Ala-Gln-Met (APPENGMAQM), 分子量为1045.22Da。体外抗氧化实验结果表明: AEH-P3对DPPH自由基(EC50 4.01mg/mL)、羟自由基(EC50 4.66mg/mL)、ABTS自由基(EC50 3.44mg/mL)和超氧阴离子自由基(EC50 6.03mg/mL)具有良好的清除作用, Ala-Pro-Pro-Glu-Asn-Gly-Met-Ala-Gln-Met (APPENGMAQM)亦显示出了良好的脂质过氧化抑制作用, 可以用于抗氧化相关的功能食品、药物或者食品添加剂。