Characterization of a new bradykinin-potentiating peptide (TmF) from Trimeresurus mucrosquamatus

A novel bradykinin-potentiating peptide (BPP), designated as TmF, has been purified to homogeneity from the venom of Trimeresurus mucrosquamatus by 70% cold methanol extraction, Sephadex G-15 gel filtration and reverse-phase high performance liquid chromatography (RP-HPLC). The amino acid sequence of TmF was determined to be pGlu-Gly-Arg-Pro-Leu-Gly-Pro-Pro-Ile-Pro-Pro (pGlu denotes pyroglutamic acid), which shared high homology with other BPPs. The molecular mass of TmF was 1.1107 kD as determinated by electrospray ionization-mass spectrometry (ESI-MS), which was in accordance with the calculated value of 1.1106 kD. The potentiating "unit" of TmF to bradykinin-induced (BK-induced) contraction on the guinea-pig ileum in vitro was (1.13 +/- 0.3) unit (mg/L), and TmF (5.0 x 10(-4) mg/kg) increased the pressure-lowering-effect of bradykinin (5.0 x 10(-5) mg/kg) with approximate descent value of (14 +/- 2) mmHg. In addition, TmF inhibited the conversion of angiotensin I to angiotensin 11, 2 x 10(-3) mg of TmF caused 50% inhibition (IC50) of angiotensin-converting enzyme (ACE) hydrolyzing activity to bradykinin.

Independent Transport and Sorting of Functionally Distinct Protein Families in Tetrahymena thermophila Dense Core Secretory Granules

Dense core granules (DCGs) in Tetrahymena thermophila contain two protein classes. Proteins in the first class, called granule lattice (Grl), coassemble to form a crystalline lattice within the granule lumen. Lattice expansion acts as a propulsive mechanism during DCG release, and Grl proteins are essential for efficient exocytosis. The second protein class, defined by a C-terminal beta/gamma-crystallin domain, is poorly understood. Here, we have analyzed the function and sorting of Grt1p (granule tip), which was previously identified as an abundant protein in this family. Cells lacking all copies of GRT1, together with the closely related GRT2, accumulate wild-type levels of docked DCGs. Unlike cells disrupted in any of the major GRL genes, Delta GRT1 Delta GRT2 cells show no defect in secretion, indicating that neither exocytic fusion nor core expansion depends on GRT1. These results suggest that Grl protein sorting to DCGs is independent of Grt proteins. Consistent with this, the granule core lattice in Delta GRT1 Delta GRT2 cells appears identical to that in wild-type cells by electron microscopy, and the only biochemical component visibly absent is Grt1p itself. Moreover, gel filtration showed that Grl and Grt proteins in cell homogenates exist in nonoverlapping complexes, and affinity-isolated Grt1p complexes do not contain Grl proteins. These data demonstrate that two major classes of proteins in Tetrahymena DCGs are likely to be independently transported during DCG biosynthesis and play distinct roles in granule function. The role of Grt1p may primarily be postexocytic; consistent with this idea, DCG contents from Delta GRT1 Delta GRT2 cells appear less adhesive than those from the wild type.

Detection of the hepatotoxic microcystins in 36 kinds of cyanobacteria Spirulina food products in China

Gel filtration chromatography, ultra-filtration, and solid-phase extraction silica gel clean-up were evaluated for their ability to remove microcystins selectively from extracts of cyanobacteria Spirulina samples after using the reversed-phase octadecylsilyl ODS cartridge for subsequent analysis by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). The reversed-phase ODS cartridge/silica gel combination were effective and the optimal wash and elution conditions were: H2O (wash), 20% methanol in water (wash), and 90% methanol in water (elution) for the reversed-phase ODS cartridge, followed by 80% methanol in water elution in the silica gel cartridge. The presence of microcystins in 36 kinds of cyanobacteria Spirulina health food samples obtained from various retail outlets in China were detected by LC-MS/MS, and 34 samples (94%) contained microcystins ranging from 2 to 163 ng g(-1) (mean=1427 ng g(-1)), which were significantly lower than microcystins present in blue green alga products previously reported. MC-RR-which contains two molecules of arginine (R)-(in 94.4% samples) was the predominant microcystin, followed by MC-LR-where L is leucine-(30.6%) and MC-YR-where Y is tyrose-(27.8%). The possible potential health risks from chronic exposure to microcystins from contaminated cyanobacteria Spirulina health food should not be ignored, even if the toxin concentrations were low. The method presented herein is proposed to detect microcystins present in commercial cyanobacteria Spirulina samples.

In situ study on the control of toxic Microcystis blooms using phytoplanktivorous fish in the subtropical Lake Taihu of China: A large fish pen experiment

Three large fish pens (0.36 km(2) of each) stocked with silver and bighead carp were set up in Meiliang Bay for controlling toxic Microcystis blooms. The responses of plankton communities and food consumption of silver and bighead carp were studied. Crustacean zooplankton were significantly suppressed in the fish pens. Total phytoplankton biomass, Microcystis biomass and microcystin concentration were lower in the fish pens than in the surrounding lake water, but the difference was not statistically significant. The present stocking density of silver plus bighead carp (about 40 g/m(3) in July) was likely too low to achieve an adequate control of Microcystis. Silver carp fed mainly on phytoplankton but bighead carp mainly on zooplankton: mean zooplankton contribution in the gut was 31.5% for silver carp and 64.7% for bighead carp. Compared with previous studies, both carp species preyed upon more zooplankton because of the abundant food resource. Daily rations of silver and bighead carp were estimated by Egger's model in the main growing season. Filtration rate was calculated from the daily ration and the density of plankton in the lake. During May-October, filtration rates of silver and bighead carp for phytoplankton were 0.22-1.53 L g(-1) h(-1) and 0.02-0.68 L g(-1) h(-1), respectively, and filtration rates for zooplankton were 0.24-0.44 L g(-1) h(-1) and 0.08-1.41 L g(-1) h(-1), respectively. Silver carp had a stronger ability of eliminating phytoplankton than bighead carp. To achieve a successful bioniampulation with a minimum effect of ichthyoeutrophication, the stocking proportion of bighead carp should be controlled in the future practice. (c) 2007 Elsevier B.V All rights reserved.

UV-sensitive P compounds: Release mechanism, seasonal fluctuation and inhibitory effects on alkaline phosphatase activity in a shallow Chinese freshwater lake (Donghu Lake)

Filtrable phosphorus compounds in a shallow Chinese freshwater lake (Donghu Lake) were fractionated by Sephadex G-25 gel-filtration chromatography. Some portions of those compounds released soluble reactive phosphorus upon irradiation with low dose ultraviolet light. Catalase and a hydroxyl radical scavenger (mannitol) markedly prevented photosensitive phosphorus release. The observed effects may be explained by the action of oxidizing reagents such as hydroxyl radicals, produced in photochemical reactions between UV irradiation and humic substances in the water. There was a strong seasonality in UV-sensitive P (UVSP) release. Michaels constants (K-m) of total alkaline phosphatase in the lake water showed a direct positive relation to UVSP. Plot of K-m against the UVSP/phosphomonoester ratio reveals a strong relationship between the two variables. These results suggest that in some situations UVSP may be a competitive inhibitor of alkaline phosphatase activity in the lake. The competitive inhibition of fractionated UVSP on alkaline phosphatase reagent (Sigma) apparently supports this hypothesis.

应用负载型纳米铁去除饮用水中砷的研究

砷是毒性最强的元素之一，水体中砷的污染己经引起人们广泛的关注。我国的新疆、内蒙、山西和台湾等省和地区地下水砷含量严重超标。全球共有5,000多万人遭受高砷饮用水的威胁，其中中国有1,500多万，是饮用水砷污染最严重的国家之一。WHO推荐饮用水砷的最高允许浓度从原来的50 µg•L-1已降至10 µg•L-1。更为严格的砷卫生标准的颁布，对作为饮用水源的地下水中的砷去除工艺提出了更高的要求。吸附法除砷比膜法、混凝法和离子交换法更安全、简便，是砷去除工艺中最有效的方法之一。 首先，本研究通过优化制备条件（包括炭种类的选择、炭的粒径大小、还原剂的浓度及滴定速率、反应温度、铁盐的种类及浓度、分散剂的比例及浓度），制备了负载型纳米铁。考虑到砷的去除效率、工程应用的可行性以及经济性，最优的制备条件如下：选用粒径为20~40目煤质炭，在室温、一定的分散剂比例及浓度，0.2 M KBH4滴速为20 d•min-1时所制备的Fe/炭为82.0 mg•g-1；纳米铁在活性炭孔内呈针状，其直径为30~500 nm，长度为1,000~2,000 nm。绝大多数的铁都负载到活性炭内部，这在处理水时铁不流失很重要。 其次，利用制备的负载型纳米铁作吸附载体，进行了饮用水中As(Ⅴ)的吸附去除实验。研究了该吸附剂对As(Ⅴ)的吸附等温线、动力学以及影响动力学的各种因素（包括As(Ⅴ)的不同初始浓度、吸附剂用量、pH值、共存离子和不同温度）、pH值、共存离子等环境条件对As(Ⅴ)去除的影响；以及吸附剂的再生及再生后的吸附效率等。研究发现在前12 h内吸附较快，72 h时达到了平衡。用Langmuir 吸附等温式估算出As(Ⅴ)的吸附量为12.0 mg•g-1。该吸附剂在pH 6.5， (25±2)℃， As(Ⅴ)初始浓度为2 mg•L-1，吸附剂用量为1.0 g•L-1时，As(Ⅴ)的去除率为75.2%；当把吸附剂的用量增加到1.5 g•L-1时，As(Ⅴ)的去除率可达99.9%以上。吸附剂可以用0.1M的NaOH浸泡12 h后即可再生，再生效率较高。常见的阴离子中PO43-、SiO32-对As(Ⅲ)的去除抑制较大，而SO42-、CO32-、C2O42-等离子对砷的去除影响较小。Fe2+对As(Ⅲ)的吸附抑制作用较大而其它阳离子影响不大。吸附剂可用0.1 M NaOH 有效再生，并且具有良好的机械性能。实验室初步实验数据表明，该吸附剂对饮用水除砷具有较好的应用前景。 第三，利用实验室制备的负载型纳米铁对饮用水中As(Ⅲ)的吸附去除也进行了研究。考察了吸附等温线、动力学以及影响动力学的各种因素、pH值、共存离子等环境条件对As(Ⅲ)去除的影响；以及吸附剂的再生及再生后的吸附效率等。研究发现，该吸附剂在pH 6.5， (25±2)℃， As(Ⅲ)初始浓度为2 mg•L-1，吸附剂用量为1.0 g•L-1时， 对As(Ⅲ)的去除率为99.8%；其吸附容量为1.996mg•g-1。吸附过程中部分As(Ⅲ)被氧化。与As(Ⅴ)的吸附相比，该吸附剂对As(Ⅲ)的效率比较高-而常见的其它除砷吸附剂如载铁纤维棉等，对As(Ⅴ)的效率比As(Ⅲ)高，为有效去除As(Ⅲ)，常常需要专门加上氧化这一过程。 最后，利用负载型纳米铁对饮用水中As(Ⅲ) 的氧化性能进行考察，发现该吸附剂不但能够有效吸附去除饮用水中的砷，而且还能把As(Ⅲ)有效地氧化为As(Ⅴ)。经过对吸附剂的构成组分分析发现，活性炭表面因富含多种官能团而对三价砷的氧化作用最大；其次是纳米铁也能把As(Ⅲ)氧化为As(Ⅴ)。

自养氨氧化混合种群特性和共存机理初步研究

组特殊自养氨氧化混合种群，表现：无机环境种群生长迅速、生物量高；在一个完全无机的自养生长环境中，不仅保持高氨氧化速率，并出现丰富的异养微生物种群；该种群置于异养、厌氧环境中，迅速表现出产氢特征。对于这样一个特殊的生态体系，研究其共生机理，以及联接这些种群之间的碳源和能源问题，将具有非常重要意义。我们拟从种群特征、细胞表面分泌产物、游离体系产物多糖、蛋白和脂肪酸方面开展研究。 第一部分，自养氨氧化混合种群的基本特征。采用氨氧化培养基，进行种群氨氧化特征研究；采用扫描电镜观察自养混合种群的微观特征；沉降、离心去除微生物种群，分析水相中的总有机碳、糖类等物质；利用LB培养基进行种群的分离、纯化，并采用DGGE手段对微生物种群结构进行分析。结果表明，接入菌种后（2/5000（V/V）），培养液中氨（200mg/L）在3－5天内快速降解；亚硝酸盐与氨氮变化呈负相关趋势，仅有少量硝酸盐含量（< 30mg/L）。氨氧化种群的生物量增长与氨氧化趋势一致，初始生物量7.75 mg/L(蛋白含量)，3-5天后生物量快速增长，并达到最高63.06 mg/L（蛋白含量）。电镜图片显示，种群外包裹一层粘液。离心除去菌体后，检测培养液总有机碳和糖的含量，同样表现出与生物量增长相似的特征，分别由初始的3.73、2.35 mg/L，3－5内天迅速增加，并分别达到最大值35.19、27.45 mg/L。经初步分离、纯化并对纯化菌株进行测序，获得了10株异养微生物分别为布鲁氏菌科苍白杆菌属、纤维单孢菌、类芽孢菌属、黄杆菌属、无色杆菌、鞘脂单胞菌、嗜麦芽寡养单胞菌、噬氢菌属、硫红球菌、假单胞菌；DGGE显示，约有20分条离带，我们对其中的两条优势条带进行切割回收测序，鉴定为欧洲亚硝化单胞菌（Nitrosomonas eur）。 第二部分：混合种群自养-异养菌共生的可能机制。在对微生物种群特征初步分析基础上，针对胞外糖类组分可能被微生物代谢分解，我们重点对微生物细胞蛋白质与糖类进行分析。采用超声结合RIPA裂解液裂解，SDS-PAGE电泳分析混合种群总蛋白种类，并通过氨基酸分析仪及红外光谱法分析氨基酸组成及蛋白红外特征。采用超声破碎结合反复冻融对细胞样品进行处理，提取液采用醇沉、Sevage脱氮白，凝胶过滤方法脱盐和分级分离。对提取物的糖分析包括：紫外扫描，红外光谱，核磁共振，单糖组成分析；扫描电镜观察菌群破裂现象。SDS-PAGE分析结果表明：氨氧化种群不同生长阶段都显示出42kD蛋白表达量很高，d4时42kD蛋白表达已经很强，4-7d内一直持续这种过量表达，直到d8后表达开始减弱。说明42kD蛋白可能与氨氧化密切相关。红外光谱分析显示：细胞提取物的特征峰分布在3427.42cm-1、1718.18 cm-1和1681.72 cm-1、1160.07和1086.74 cm-1，分别对应为OH、 C=O、C－O－C基团，表明具有蛋白的典型特征；氨基酸分析显示蛋白中的Gly,Asp,Ala,Glu含量相对较高。 提取物中胞外多糖分离谱图得到不均一组分，共得到6个收集峰；紫外扫描在201－213 nm处有多糖吸收峰，同样表明多糖成分不均一性；多糖红外光谱特征峰主要分别在3400.49 cm-1、2920.28 cm-1、1154.54和1087.52 cm-1，对应OH、－CH2－ or CH 、C－O－H or C－O－C等多糖特征基团；多糖提取物核磁共振1H d4.3～5.9之间出现强吸收峰，这是1H中，多糖存在的明显证据，1H NMR中，其中O-乙酰基的甲基上的氢信号为d1.1～1.3之间。糖肟全苯甲酸酯衍生物的HPLC测定中，得到单一的单糖峰，由于时间问题，还未进行更深入的试验；电镜图片显示，种群中的细胞有大量的破裂现象。 实验表明，自养氨氧化混合种群显示出快速的氨氧化速率，氨氧化过程生物量和有机质的增加明显。微生物种群包裹粘液层，并分离纯化出大量的异养菌；去除菌体后的游离培养液中存在有机质（包括多糖）说明无机自养生长体系中存在异养菌生长、繁殖的二次碳源；细胞提取物中蛋白条带数目多、种类丰富；细胞多糖提取物具有明显的多糖特征，以及单糖的存在。结合种群的显微特征和游离体系中的有机质的检测结果，我们认为，无机自养生长体系中，种群细胞生长过程中发生的破裂现象可能是导致大量的蛋白、多糖释放到游离胞外，并成为其他异养菌生长的碳源和氮源。这可能是自养体系中，大量异养菌共生的可能机制，至于是什么原因引起种群生长过程中产生的破裂现象，还有待下一步深入研究。 A group of mixed autotrophic ammonia oxidizing populations, having much biological characteristic tested by concerned personnel for pilot test: Performed rapid population growth and obtained high biomass in inorganic environment; Not only maintained a high rate of ammoxidation, promoted a wealth of heterotrophic microbial populations growth in a totally inorganic and autotrophic growth environment; Placed in heterotrophic and anaerobic environment,had the performance characteristics that could rapidly produce hydrogen.For such a special ecological system, Study its symbiotic mechanism and the connection between these populations of carbon and energy issues, will have a very important significance. We intended from the characteristics of the population, the secretion product of cell surface, free substance in the liquid medium like polysaccharide, protein and fatty acids carrying out research. Part I: The basic features of mixed autotrophic ammonia oxidizing populations . Use inorganic liquid medium, processed study for ammonia oxidation characteristics of the population; we used scanning electron microscopy to get micro-features of autotrophic ammonia oxidizing populations .The medium was carried out settlement and centrifugal then removed the microbial populations, after all of that we analysis the water phase for total organic carbon(TOC), carbohydrate and other substances; Solid ammonia oxidizing medium was adopted to separation and purification of population, DGGE means was for structure analysis of microbial population. The results showed that after the inoculum of bacteria (2 / 5000 (V / V)), ammonia in the culture medium (200 mg / L) was rapid degradation in 3-5 days; ammonia and nitrite have the negative correlation between changes in the trend, then only a small amount of nitrate content (<30mg / L). The biomass growth of ammoxidation population in line with the trend of ammonia oxidation, the initial volume of it was 7.75 mg / L (protein content), in 3-5 days upto 63.06 mg / L (protein content). Electron microscope image showed, the populations were wrapped in a layer of mucus, including the a large number ruptted micorbe , Centrifuge to remove bacteria, then detected the medium for total organic carbon and sugar content, result took on the same characteristics with biomass growth, that were from the initial 3.73、2.35 mg / L respectively, in 3-6 days achieved rapid increase in the maximum to 35.19、27.45 mg / L respectively. After initial separation、 purification ,then processed sequencing to strains purified and got the result that there were 10 heterotrophic microorganisms : Brucella Branch pale bacillus, Cellu lomonas, Bacillus species category, a Flavobacterium, colorless Bacteria, Aeromonas sheath fat, little support maltophilia Aeromonas, macrophages species hydrogen, sulphur-MI, Pseudomonas bacteria spores; DGGE display, there were 20 separation bands approximately. Part II: Mixed populations that autotrophic - heterotrophic bacteria symbiotic mechanism. On the basis of preliminary analysis of microbial population characteristics, aiming at extracellular carbohydrate components might be decomposition by microbial, we focused on microbial cell protein and carbohydrate analysis. Using ultrasound combined with RIPA lysis cracking the cells, SDS-PAGE electrophoresis analysis the total protein species of the population, and through the amino acid analyzer studied the compositions of amino acid and infrared spectroscopy analysis of a protein infrared characteristics. Using ultrasound combined with repeatedly freezing and thawing to treated the cell sample, then took the means that alcohol precipitation, deproteinization by Sevage, gel filtration aimed at desalination and grade separation to deal with the lysates . The extraction of sugar analysis included: UV scanning, IR, NMR, single-sugar composition analysis. SDS-PAGE analysis showed that: 42 kD protein expression was very high at different growth stages of mixed autotrophic ammonia oxidizing populations , on the fourth day, 42 kD protein expression had been very strong, 4-7d, it had continued this excessive expression, then started to weaken after 7 days. 42 kD protein that might be closely associated with ammonia oxidation. Infrared spectral analysis showed that: cell extracts with the characteristic that the peak distribution in 3427.42 cm-1、1718.18 cm-1 and 1681.72 cm-1、1160.07 cm-1 and 1086.74 cm-1 corresponding to OH、C = O、C-O-C Groups which had the typical characteristics of protein; and analysis showed that amino acids including Gly, Asp, Ala, Glu ,the content in the protein is relatively high. Exopolysaccharide in the extracts had the separation map that it was uneven, received a total of six collection peaks by the detection mode of phenol-sulphruic acid method ; ultraviolet scan in the 201-213 nm department had polysaccharide absorbing peak, the same ingredients that polysaccharide heterogeneity; infrared polysaccharide spectral characteristics of the main peak at 3400.49 cm-1, 2920.28 cm-1, 1154.54 and 1087.52 cm-1, corresponding OH,-CH2-or CH, C-O-H or C-O-C;and other characteristics of polysaccharide group; 1H NMR of polysaccharide extract appeared absorption peak between d4.3 ~5.9, which is the apparent evidence of polysaccharide, In 1H NMR, the hydrogen signal of one of O-acetyl was between 1.1 to 1.3. The determination of Sugar oxime whole benzoate derivatives by HPLC, there was a single-sugar peak, as a matter of time, yet more in-depth test. Summary: Mixed autotrophic ammonia oxidizing populations show us that it had the ability in ammonia oxidizing and it was great, organic matter and biomass increased significantly in the process of ammonia oxidation. Microbial populations was wrapped up slime layer, the phenomenon of cell breakdown obviously, and there were a lot of separation and purification of the heterotrophic bacteria; a lot of organic matter (including polysaccharides)remined in the medium that removal of cell indicated the inorganic system existed secondary carbon sources that could be used by the heterotrophic bacteria ; there were a large number proteins bands of cell extract, rich variety; cell extracts of polysaccharide had obvious characteristics of polysaccharide, and the existence evidence of single-sugar. Combined population of microscopic characteristics and free of organic matter in the test results, we believe that the health of inorganic system, population growth occurred in the course of the breakdown of the phenomenon is likely to lead to a lot of protein and polysaccharide released into the extracellular free, And other heterotrophic bacteria use them to the growth as carbon and nitrogen. This may be autotrophic system, the large number of heterotrophic bacteria symbiotic mechanism.

Anti-inflammatory and immunomodulatory activities of polysaccharide from Chlorella stigmatophora and Phaeodactylum tricornutum

Crude polysaccharide extracts were obtained from aqueous extracts of the microalgae Chlorella stigmatophora and Phaeodactylum tricornutum. The crude extracts were fractionated by ion-exchange chromatography on DEAE-cellulose columns. The molecular weights of the polysaccharides in each fraction were estimated by gel filtration on Sephacryl columns. The crude polysaccharide extracts of both microalgae showed anti-inflammatory activity in the carrageenan-induced paw edema test. In assays of effects on the delayed hyper-sensitivity response, and on phagocytic activity assayed in vivo and in vitro, the C. stigmatophora extract showed immunosuppressant effects, while the P. tricornutum extract showed immunostimulatory effects. Copyright © 2003 John Wiley & Sons, Ltd.

Analytical intercomparison results from the 1990 Intergovernmental Oceanographic Commission open-ocean baseline survey for trace metals: Atlantic Ocean

“Dissolved” (< 0.4 μm filtered) and “total dissolvable” (unfiltered) trace element samples were collected using “clean” sampling techniques from four vertical profiles in the eastern Atlantic Ocean on the first IOC Trace Metals Baseline expedition. The analytical results obtained by 9 participating laboratories for Mn, Fe, Co, Ni, Cu, Zn, Cd, Pb, and Se on samples from station 4 in the northeast Atlantic have been evaluated with respect to accuracy and precision (intercomparability). The data variability among the reporting laboratories was expressed as 2 × SD for a given element and depth, and was comparable to the 95% confidence interval reported for the NASS seawater reference standards (representing analytical variability only). The discrepancies between reporting laboratories appear to be due to inaccuracies in standardization (analytical calibration), blank correction, and/or extraction efficiency corrections.Several of the sampling bottles used at this station were not adequately pre-cleaned (anomalous Pb results). The sample filtration process did not appear to have been a source of contamination for either dissolved or particulate trace elements. The trace metal profiles agree in general with previously reported profiles from the Atlantic Ocean. We conclude that the sampling and analytical methods we have employed for this effort, while still in need of improvement, are sufficient for obtaining accurate concentration data on most trace metals in the major water masses of the oceans, and to enable some evaluation of the biogeochemical cycling of the metals.

Validation and in situ application of an automated dissolved nickel monitor for estuarine studies

The validation of a fully automated dissolved Ni monitor for in situ estuarine studies is presented, based on adsorptive cathodic stripping voltammetry (AdCSV). Dissolved Ni concentrations were determined following on-line filtration and UV digestion, and addition of an AdCSV ligand (dimethyl glyoxime) and pH buffer (N-2-hydroxyethylpiperazine-N′-2-ethanesulphonic acid). The technique is capable of up to six fully quantified Ni measurements per hour. The automated in situ methodology was applied successfully during two surveys on the Tamar estuary (south west Britain). The strongly varying sample matrix encountered in the estuarine system did not present analytical interferences, and each sample was quantified using internal standard additions. Up to 37 Ni measurements were performed during each survey, which involved 13 h of continuous sampling and analysis. The high resolution data from the winter and summer tidal cycle studies allowed a thorough interpretation of the biogeochemical processes in the studied estuarine system.

Purification, sequencing and biological activity of polypeptide from velvet antler

The velvet antler polypeptide CNT14 was extracted and purified by gel filtration, ion exchange chromatography and RP C, which showed a single peak in HPLC chromatography and a single band in SDS-PAGE. The molecular weight measured by MALDI/TOF/MS spectrum was 1479. 9028. The polypeptide consisted mostly of Glu, Leu, Val, Pro. The amino acid sequence of the polypeptide was detected with ESI-MS/ MS, and the sequence was E-P-T-V-L-D-E-V-C-L-A-H-G-P. The experiments of biological activity of polypeptide CNT14 in vivo were carried out, and the results show that CNT14 has stimulant effects on the growth of rat HT22 cells. Then we produced the polypeptide CNT14 according the amino acid sequence by solid phase synthesis, confirmed the sequence of the polypeptide to be consistent with the amino acid sequence of polypeptide CNT14 which was separated from the velvet antler.

Purification and structure identification of hyaluronic acid

Polysaccharide produced by mutated strain of Streptococcus zooepidemicus was purified by the procedures including Savage method, quaternary ammonium compound precipitation, DEAE-cellulose(DE52) chromatography and Sephadex G-75 gel filtration. The structure of the purified polysaccharide has been characterized by means of chemical composition analysis, C-13 NMR spectrum, infrared spectrum and circular dichroism (CD). All the results showed that the purified polysaccharide was hyaluronic acid (HA). The single helix conformation of the purified HA was determined by Congo red experiment. The molecular weight of the HA was about 1.16x10(6)D, which was measured by viscosity method.

Chemical characteristic of bioactive polysaccharides isolated from Ornithogalum caudatum Ait.

A water-soluble crude extract prepared from Ornithogalum caudatum Ait. (OCA) showing a high immunomodulating activitiy was isolated and characterized by virtue of get filtration and column chromatography. The presence of the monosaccharides has been established by the chemical analysis. The quantitative analysis of the alditol acetate derivatives of them showed the ratios of the monosaccharides analyzed by means of GC respectively. The concentrations of protein(280 nm) and carbohydrate (496 nm) were detected respectively. The information of the molecular weight from the pure polysaccharide was obtained by several standard Dextrans from the Sephadex chromatography.

Preparation and mass spectrometric study of egg yolk antibody (IgY) against rabies virus

Rabies virus was used as the antigen to immunize laying chickens. Anti-rabies virus immunoglobulin Y(IgY) was isolated from yolks of the eggs laid by these chickens using a two-step salt precipitation and one-step gel filtration protocol. The purified IgY was reduced with dithiothreitol, and heavy chains (HC) and light chains (LC) were obtained. In addition, the purified IgY was digested with pepsin and the fragment with specific antigen binding properties (Fab) was produced. Using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOFMS), the average molecular weights of IgY, HC, LC, and Fab were determined as 167 250, 65 105, 18 660, and 45,359 Da, respectively. IgY has two structural differences compared with mammalian IgGs. First, the molecular weight of the heavy chain of IgY is larger than that of its mammalian counterpart, while the molecular weight of the light chain of IgY is smaller. Second, upon pepsin digestion, anti-rabies virus IgY is degraded into Feb, in contrast to mammalian IgG, which has been reported to be degraded into F(ab')(2) under the same conditions. Copyright (C) 2001 John Wiley & Sons, Ltd.