壳寡糖和壳聚糖酶水解特性的薄层色谱分析

建立了聚合度2～10的壳寡糖的薄层层析分析(TLC)方法,在异丙醇∶水∶氨水=60∶30∶4的展开体系中,各壳寡糖可以得到很好的分离,各单体间比移值(Rf)差异明显,无拖尾现象。通过对比TLC法与高效液相色谱(HPLC)法分析壳寡糖单体的效果,得出TLC法具有快速、有效、方便的优点,可以作为定性分析壳寡糖单体纯度的方法。采用本实验室制备的壳聚糖酶对3种不同脱乙酰度的壳聚糖和DP2～6的壳寡糖进行水解,采用TLC法对水解产物进行分析。实验结果表明,随着壳聚糖脱乙酰度的降低,其酶水解产物的糖链延长。该壳聚糖酶不能水解DP2～4的壳寡糖,对DP5～6的壳寡糖水解速率缓慢。通过对该壳聚糖酶作用特点分析,表明该酶可作用于GlcN-GlcN糖苷键,而不能水解Glc-NAc-GlcNAc糖苷键。     

新型壳寡糖γ-氨基丁酸衍生物的制备及其对小麦幼苗抗旱作用的研究

利用壳寡糖氨基与γ-氨基丁酸羧基发生酰胺化反应合成新型壳寡糖-氨基丁酸衍生物,并通过傅里叶变换红外光谱(FT-IR)、核磁共振光谱(NMR)等对合成的衍生物进行结构表征。以小麦种子为材料,各样品对小麦种子进行浸种处理后干旱胁迫培养到两叶一心,测定小麦生理生化指标,评价新型衍生物的抗旱作用。结果表明,新型壳寡糖γ-氨基丁酸衍生物成功合成,并且对干旱胁迫下小麦生理生化指标有明显的影响,衍生物明显降低了小麦幼苗叶片的丙二醛含量和细胞膜透性,缓解了干旱胁迫对小麦细胞膜的损伤,增加可溶性糖含量进而增强小麦植物对土壤中水分的吸收能力,减少了干旱胁迫对生理代谢的影响。总之,成功合成了具有较强诱导小麦抗旱作用的新型壳寡糖γ-氨基丁酸衍生物,为新型植物抗逆诱导剂的开发提供了思路。     

酶法制备几丁寡糖和壳糖研究现状与进展

对用酶法制备几丁寡糖和壳寡糖研究现状与进展进行了综述。几丁质酶、壳聚糖酶、溶菌酶和 N-乙酰葡萄糖胺酶等专一性的水解酶及糖酶、蛋白酶、脂肪酶等水解酶对几丁质和壳聚糖都具有部分或完全非专一性水解作用。介绍了这些酶类的作用机理及相关微生物的种类和作用条件     

壳寡糖与三甲酰氯界面聚合制备纳滤膜及其染料分离性能

界面聚合制备方法通常用于制备纳滤膜或反渗透膜,单体一般为间苯二胺或哌嗪。但是这些水相单体具有较大的毒性、刺激性等不利因素,会对环境及人体造成危害。壳寡糖（chitosan oligosaccharide, COS）是壳聚糖经特殊的生物酶处理得到的较低分子量的海洋多糖类高分子聚合物,绿色无毒,与壳聚糖相比其溶解性大大提高。壳寡糖与壳聚糖的化学性质相似,其分子中含有氨基、羟基,适合作为水相单体通过界面聚合反应制备复合纳滤膜。本研究以壳寡糖为水相单体,均苯三甲酰氯（trimesoyl chloride, TMC）为油相单体,通过界面聚合法在聚醚砜超滤膜（PES）上成功制备COS/PES复合纳滤膜。采用场发射扫描电子显微镜（SEM）和原子力显微镜（AFM）对膜表面进行分析表征;通过错流过滤测试膜通量及染料分离性能。实验结果显示,COS/PES 2复合纳滤膜表面出现斑点状结构,具备较高的膜通量;根据Zisman的临界表面张力外推方法将液体的表面张力与cosθ作图,发现COS/PES复合纳滤膜具有较好的防生物黏着的性能;进一步采用牛血清蛋白质模拟生物质膜污染物,开展了比通量（J/Jw）下降的实验,结果表明COS/PES复合纳滤膜的比通量始终高于商业PES超滤膜,说明交联壳寡糖活性层具有一定的抗生物质膜污染的性能;染料分离实验结果显示,壳寡糖复合纳滤膜对甲基橙的截留率在0.45 ~ 0.53之间,樱草灵的截留率在0.66~0.77之间,染料分子量大于600 Da时,不同浓度（1%、2%、3%,wt）制备的壳寡糖复合膜的染料截留率均保持在90%以上,证明所制备的复合膜属于纳滤的范畴。本研究制备的COS/PES复合纳滤膜符合国内外对发展环境友好型海洋新材料的需求,在环保、医药方面极具应用潜力。     

低分子量氨基糖及其衍生物对角膜细胞生长影响的研究

体外培养兔角膜上皮细胞和基质细胞,采用显微观察和MTT相结合的方法分析低分子量氨基糖(包括壳寡糖(chitosan)、羧甲基壳寡糖(Carboxymethyl chitosan oligosaccharide)、羧甲基甲壳寡糖(Carboxymethyl chitin oligosaccharide)、N-乙酰氨基葡萄糖(N-acetyl-glucosamine))对细胞生长的影响。结果在0~1000μg/mL浓度范围内4种氨基糖对角膜上皮细胞和基质细胞均没有细胞毒性,且均能促进角膜上皮细胞的生长,以壳寡糖和羧甲基甲壳寡糖效果最佳,二者与对照组相比均具有显著性差异(P<0.05)。壳寡糖、羧甲基甲壳寡糖和N-乙酰氨基葡萄糖能明显促进角膜基质细胞的生长,与对照组相比具有显著性差异(P<0.05),其中壳寡糖的促进作用最明显。提示低分子量氨基糖可适用于角膜细胞促生长的培养,为壳聚糖衍生物材料用于眼科研究提供一定的实验依据。     

丁二酰化壳寡糖稀土配合物的抗氧化活性测定

合成了丁二酰化壳寡糖稀土(La,Nd,Eu)配合物,并考察其稀土配合物对羟自由基和超氧阴离子自由基的清除活性及其还原能力。实验表明:对超氧阴离子自由基的清除能力以及还原能力依次为:稀土配合物丁二酰化壳寡糖壳寡糖。在低浓度下对羟基自由基清除能力顺序为:丁二酰化壳寡糖稀土配合物壳寡糖,高浓度下丁二酰化壳寡糖和壳寡糖对羟基自由基清除能力急剧下降,而稀土配合物的清除能力稳定。这可能是由于稀土配合物的稀土离子含量及稀土离子性质、分子电荷密度以及对自由基清除机理不同所致。     

酶促反应制备壳寡糖及壳寡糖分析

用壳聚糖酶降解法对壳聚糖进行降解制备壳低聚糖，研究了温度、pH、底物浓度、脱乙酰度、反应时间对酶促反应的影响。对酶解产物进行HPLC分析，用Bio—Gel P-4柱对酶解产物进行初步分离。结果表明，壳聚糖酶降解壳聚糖的最适温度为45℃，pH为6．0，最适底物质量分数为6％，脱乙酰度越高酶解反应所得产物的数均相对分子质量越小，24h后酶解反应达到平衡，酶解产物壳寡糖水溶性极好。P-4柱分离得到聚合度20以上，10～20，10以下3种壳寡糖。     

薄层色谱法分析壳寡糖

建立壳寡糖的薄层色谱 (TL C)分析条件。通过对各种展开剂的实验 ,确定壳寡糖的 TL C最佳的展开剂是乙酸乙酯 :乙醇 :水 :氨水 =5 :5 :4 :0 .3。壳寡糖溶液上行展距为 8cm,采用浸渍法显色。本文方法分离效果好 ,简便 ,重现性好。     

烟酰化壳寡糖季铵盐的制备及抑菌活性研究

海洋生物体内蕴含着丰富的天然活性物质,许多都具有抑菌活性,据此研制新型海洋生物源抑菌药物,是海洋生物产业中的一个新领域,也是新型抑菌药物研发的重要途径。壳聚糖是来自海洋中虾蟹等甲壳类动物外壳的甲壳素的脱乙酰产物,壳寡糖是壳聚糖的降解产物,具有比壳聚糖更好的水溶性和生物活性,在食品和生物医药等领域具有更广阔的应用前景。为了提高壳寡糖的抑菌活性,本研究以壳寡糖和烟酸为原料、N, N-羰基二咪唑为催化剂合成了壳寡糖烟酸接枝衍生物,并与不同链长的溴代烷烃亲核反应得到其阳离子化衍生物。壳寡糖及其衍生物的化学结构通过红外光谱(FTIR)和核磁共振波谱(NMR)进行分析表征,壳寡糖及其衍生物对2种海洋致病细菌和3种植物致病真菌的体外抑菌能力分别采用肉汤稀释法和菌丝生长速率法进行研究和评估。结果表明,与壳寡糖相比,合成的终产物对致病菌的抑制能力得到了极大增强,其中含有己基的烟酰化壳寡糖季铵盐表现出最强的抑制海洋致病细菌活性(MIC=0.125mg/mL、MBC=0.25mg/mL),并且该化合物表现出较强的抗真菌活性,在1.0 mg/mL时,对灰葡萄孢菌和围小丛壳的抑制率分别为65.7%和51.5%。...     

饲料中添加枯草芽孢杆菌和壳寡糖对大黄鱼幼鱼血清免疫指标的影响

本实验采用2×3双因子实验设计,分别在基础饲料中添加0(对照)、0.5×107和1.0×107/g枯草芽孢杆菌,在每个枯草芽孢杆菌添加水平分别添加0(对照)、0.3％和0.6％的壳寡糖,配制出9种等氮等能的实验饲料,在海水浮式网箱中进行为期70 d的实验,探讨饲料中枯草芽孢杆菌和壳寡糖的添加对大黄鱼(Pseudosciaena crocea)血清免疫指标的影响.实验结果表明,在每个枯草芽孢杆菌水平下,饲料中添加0.3％和0.6％的壳寡糖显著提高了大黄鱼幼鱼血清溶菌酶的活性(P＜0.05),而对大黄鱼血清替代途径补体活力,超氧化物歧化酶(SOD)活力及过氧化氧酶( CAT)活力没有显著性影响.在每个壳寡糖水平,饲料中枯草芽孢杆菌的添加对大黄鱼血清免疫指标的影响不显著.饲料中枯草芽孢杆菌和壳寡糖对大黄鱼幼鱼血清免疫指标的影响不存在交互作用.总的来说,饲料中添加一定量的壳寡糖(0.3％～0.6％)在一定程度上影响了大黄鱼幼鱼的非特异性免疫反应.     

广西钦州内湾贝类养殖海区三氮的含量和百分组成

本文根据1998年10月和1999年5月的调查资料，报道了钦州内湾帆类养殖海区三态氮的含量和百分组成。结果表明：NO3－N是本水域无机氮的主要存在形式；三态氮之间的相互转化以春季进行得比较充分，95．70％均已转化为NO3－N，基本达到了热力学平衡状态，NO3－N与NH4－N和NO2－N之间均具有显著的正相关关系；秋季，只有79．87％的无机氮转化为NO3－N，没有达到热力学平衡状态，但NO3－N与NH4－N却具有高度显著的负相关关系，与NO2－N的相关性较差。     

胶州湾及周边海域大气和海水中N_2O和CH_4的分布及海气交换通量

分别于2006年8月,12月和2007年4月,10月采集胶州湾及周边海域大气和海水样品,对氧化亚氮(N2O)和甲烷(CH4)浓度进行了测定,并设置1个连续站进行24 h连续观测.结果表明:大气中N2O春、夏、秋、冬的平均浓度(体积分数)分别为(3.17±0.03)×10-7,(3.24±0.15)×10-7,(3.19±0.02)×10-7和(3.08±0.25)×10-7;大气中CH4春、夏、秋、冬的平均浓度(体积分数)分别为(1.89±0.04)×10-6,(1.79±0.04)×10-6,(2.09±0.21)×10-6和(2.01±0.09)×10-6.胶州湾表、底层海水中N2O和CH4的浓度和饱和度表现出明显的季节变化,其中N2O浓度和饱和度冬季最高,春、秋季次之,夏季最低;CH4浓度和饱和度夏季最高,冬季最低.利用Liss and Merlivat(1986)公式和Wanninkhof(1992)公式估算出胶州湾海域N2O的年平均海-气交换通量分别为(11.16±14.15)和(22.42±27.56)μmol m-2·d-1;CH4分别为(7.75±6.19)和(17.76±14.84)μmol m-2·d-1.胶州湾大部分海域表层海水中N2O和CH4呈过饱和状态,是大气中N2O和CH4的净源.     

N,N-二烷基酰胺类物质的膜状态及其在水面溢油集油剂中的应用研究

合成了 5种 N,N-二烷基酰胺型表面活性剂 ,并对其所成膜的状态进行了测定。表明当N,N-二烷基酰胺类物质单独成膜时获得的集油膜为多层膜 ,而当选用合适的溶剂与其配制成溶液以后 ,所成的集油膜则为转变膜或液态凝聚膜。在 5种 N,N-二烷基酰胺类物质中 ,酰基碳链越长、氮烷基碳链越短则其集油能力越强。应用 N,N-二烷基酰胺类物质配制了 4个配方 ,均具备良好的集油能力 ,并且对原油的集油能力大于对柴油的集油能力     

营养盐因子对龙须菜和菊花江蓠氮磷吸收速率的影响

本文在实验室条件下,研究了营养盐因子对龙须菜和菊花江蓠N、P吸收速率的 影响,分别对不同氮和磷的浓度、氮磷比及不同化合态氮对这两种藻类氮、磷吸收速 率的影响进行了实验与分析．结果表明:在适宜范围内,两种海藻对氮、磷的吸收速率 随介质中氮、磷浓度的增加而增加;不同氮磷对两种海藻N吸收速率的影响不存在 显著差异,但对磷吸收速率的影响非常显著,最高对磷吸收速率在介质的氮磷比为 10:1时;海藻对NH4-N的吸收速率随着NO3-N／NH4-N比的降低而增加,对PO43-P的 吸收速率与对NH4-N的吸收相似,而对NO3-N的吸收速率与对NH4-N的吸收相反．     

Nitrogen and phosphorus in New Zealand streams and rivers: Control and impact of eutrophication and the influence of land management

Abstract

Given sufficient light and heat, the growth of aquatic macrophytes and algae associated with eutrophication is generally controlled by the concentration, form and ratio between nitrogen (N) and phosphorus (P). Data from 1100 freshwater sites monitored for the last 10 years by New Zealand's regional councils and unitary authorities were assessed for streams and rivers with mean nitrate/ nitrite‐N (NNN), dissolved reactive P (DRP), total N (TN) and total P (TP) concentrations in excess of New Zealand guidelines, and to generate a data set of N:P ratios to predict potential periphyton response according to the concentration of the limiting nutrient. The frequency of sites exceeding the guidelines varied from 0 to 100% depending on the parameter and region, but South Island regions were generally more compliant. The dissolved inorganic N (DIN) to dissolved reactive P (DRP) ratio was used to group data into three nutrient limitation classes: <7:1 (N‐limited), between 7:1 and 15:1 (co‐limited), and>15:l (P‐limited), by mass. P‐limitation was the most frequent scenario in New Zealand streams (overall, 76% of sites were P‐limited, 12% N‐limited, and 12% co‐limited). The mean concentration of the limiting nutrient for each site was combined with empirical relationships to predict periphyton densities (the average of N‐and P‐limited growth was used for sites with co‐limitation). This assessment predicted that 22 sites were likely to exceed the periphyton guideline for protecting benthic biodiversity (50 mg chlorophyll a m^?2), but this assessment is likely to be highly changeable in response to climatic conditions and present and future land use. As an example, we modelled N and P losses from an average sheep and a dairy farm in Southland (South Island, New Zealand) in 1958, 1988, 2008 and 2028. We predicted that with time, as farm systems have and continue to intensify, N losses increase at a greater rate than P losses. Since the pathway for N to reach fresh waters may be more tortuous and take longer than P to reach a stream or river, focusing mitigation on P losses may have a quicker effect on potential algal growth. In addition, with time, it is expected that P‐limitation in New Zealand's rivers and streams will be more widespread as N‐losses are unabated. Hence, although strategies to decrease N losses should be practised, mitigating P losses is also central to preventing eutrophication.     

Egg production and early development of the subarctic copepods Neocalanus cristatus,N. plumchrus and N. flemingeri

Reproduction and early development of the large subarctic copepods Neocalanus cristatus, N. plumchrus and N. flemingeri were investigated in the laboratory. All three species produced eggs at intervals of 5.0 to 14.7 days, depending on species and temperature. The females of N. flemingeri released the fewest clutches of the three species, but with the largest clutch size. Clutch size of N. cristatus was smallest with longest intervals between clutches. Mean±1 S.E. of total fecundities were 386±116 eggs for N. cristatus, 840±214 eggs for N. plumchrus, and 924±346 eggs for N. flemingeri. The egg laying period of a female at 2°C was estimated to be 91 days for N. cristatus, 60 days for N. plumchrus and 47 days for N. flemingeri. The color and outer characteristics of eggs and nauplii of these species were quite different. C : N ratios of the eggs were 9.3–10.5, which were slightly higher than that of females or CV. Egg hatching times for each species were 4.6–5.7 days at 2°C and decreased with increasing temperature at a Q₁₀ of 2.8–3.0. N. cristatus nauplii developed to copepodid stage I (CI) without feeding, and the developmental time in days from hatching to CI was expressed as a Bělehrádek equation, D_CI=17068×(T+14.7)^−2.05. Reproduction strategies of the three species of Neocalanus are discussed with reference to their life history strategies.     

Nitrogen and phosphorus species in the coastal and shelf waters of Southeastern North Carolina, Mid-Atlantic U.S. coast

The diversity of small-scale wetlands, high salinity tidal creeks, salt marshes, estuaries, and a wide and shallow shelf with the Gulf Stream close to the break makes the coastal zone of south-eastern North Carolina (U.S.) a natural laboratory for the study of the cycling of nitrogen (N) and phosphorus (P) in coastal and shelf waters. We assessed the summer concentrations, forms, and ratios for each N (total dissolved N, nitrate + nitrite, ammonium and dissolved organic N) and P (total dissolved P, o-phosphate and dissolved organic P) pool as these nutrients travel from tidal creeks, salt marshes and two large estuaries to Long and Onslow Bays. Additionally, we measured ancillary physical (temperature, salinity and turbidity) and chemical (dissolved oxygen, chlorophyll a and pH) water properties. Highest concentrations of all individual N and P compounds were found in the upper parts of each tributary and were attributed to loads from agricultural and urban sources to the coastal watersheds, continuing downstream to receiving estuaries. In all areas, dissolved organic N and P species were predominant constituents of the total dissolved N and P pools (64–97% and 56–93%, respectively). The lower parts of estuaries and surface shelf waters were characterized by oceanic surface values, indicating removal of N and P downstream in all tributaries. The different watershed and hydrological characteristics also determined the different speciation of N and P pools in each estuary. Despite a high level of anthropogenic pressure on the uppermost coastal waters, there is self-regulation in this coastal ecosystem with respect to human perturbations; i.e. significant amounts of the N and P load are retained within estuarine and nearshore waters without reaching the shelf.     

Spatial and temporal variations in free polyamine distributions in Uranouchi Inlet, Japan

Polyamines are an essential component of living organisms that influence normal cell growth and development. Despite the possible importance of polyamines for marine microbial ecology, concentrations of polyamines in seawater are not well known.Using HPLC analysis with a cation exchange column and OPA reagent, we determined polyamine concentrations in seawater without carrying out desalting, concentration or derivatization steps prior to chromatography.In seawater collected from Uranouchi Inlet, putrescine and spermidine were the predominant polyamines, ranging from undetectable (N.D.) to 4.4 nM and N.D. to 2.8 nM, respectively. Though other polyamines were detected, the frequency of detection was much lower than that of putrescine and spermidine; concentrations ranged from N.D. to 0.9 nM for cadaverine, and N.D. to 3.2 nM for norspermidine. Norspermine and spermine were detected in only a few samples; their concentrations varied from N.D. to 3.2 nM and N.D. to 0.5 nM, respectively. These concentrations were higher in the summer.     

三种织纹螺性畸变的发展过程与雄性结构效应

从2000年3月到2002年10月对中国沿海海产腹足类性畸变的大范围调查过程中．在广东省汕头港、湛江港和厦门港分别采集到西格织纹螺(Nassarius siquijorensis)、胆形织纹螺(Nassarius thersites)和节织纹螺(Nassarius hepaticus)等3个种，解剖观察发现3种织纹螺的雌性个体都发生了性畸变现象。根据织纹螺性畸变的发展过程，对目前的性畸变发展划分体系作了新的修改和补充。通过对比性畸变个体与正常雄性个体的特征．探讨了雄性个体结构特征在性畸变过程中的效应。     

渤黄海营养盐结构及其潜在限制作用的时空分布

根据2006-2007年4个季节的现场调查资料,分析探讨了渤海和黄海营养盐结构分布变化特征及其对浮游植物生长的潜在的限制状况.结果表明,渤黄海水体 Si/N/P 比值均偏离 Redfield 比值,季节变化明显;春夏冬季 N/P和 Si/P比值由近岸向远岸海域递减,高值区主要分布在黄河口、鸭绿江口及苏北近岸,秋季上层水体N/P和Si/P比值的分布趋势有所不同,高值区主要分布在南黄海的中部海域.受陆源输入的影响,近岸特别是河口区 N/P和 Si/P比值均较高,温跃层的生消变化和生物活动调控着黄海中部海域营养盐结构的变化.渤黄海浮游植物生长主要受P的潜在限制,部分季节受N、Si的潜在限制;营养盐限制状况存在着明显的时空变化及不同营养盐的同时或交替限制的现象.