加热聚乙二醇法:另一种血浆分层方法

用聚乙二醇(PEG)作沉淀剂制备的血液制品证明是无毒和无热原的,并已用于制备免疫球蛋白、浓缩因子Ⅷ和人白蛋白。本文介绍用加热PEG法制备白蛋白的过程。 ACD血浆经4℃解冻,于pH6.5加39％PEG4000溶液至最终浓度为12%,离心后,沉淀再悬于12%PEG溶液,再离心,沉淀用于制备免疫球蛋白。两次离心上清合并,于pH6.5加锌酸钠0.01mol/l,搅拌下加热75℃30分钟或过滤分离沉淀。上清或滤液经石棉板过滤,调pH至4.6,加固体PEG4000至最终浓度为22%,分离白蛋白沉淀,并溶于蒸馏水(pH7.0)至蛋白浓度为8％,对4℃蒸馏水透析,除去PEG,调节蛋白浓度至5&,加锌酸钠及乙酰色氨酸钠,再经0.2μm过滤,     

免疫透射比浊法测定微量白蛋白的研究

目的 对免疫透射比浊法测定微量白蛋白（ALB）进行研究探讨,确定最佳反应条件。方法采用免疫透射比浊法,聚乙二醇（PEG）作为增敏剂,改变PEG浓度,PEG分子量,缓冲液pH值,以及缓冲液体系,对微量白蛋白进行测定。结果采用PEG6000作为增敏剂,PEG浓度为4％,PBS作为缓冲液,pH值为6．5的条件下,浊度最高,检测效果最好,检出限为0．0587μg／ml。检测线性范围为1～8μg／ml,相关系数R=0．9974。结论确定免疫透射比浊法测定微量白蛋白的最佳反应条件,对临床诊断试剂盒的研发具有指导意义。     

腹水型抗EGFR单克隆抗体纯化方法的研究

目的：研究腹水型抗EGFR单克隆抗体的纯化方法。方法：抗EGFR杂交瘤细胞诱生的腹水,经离心预处理,上清先用PEG6000沉淀,沉淀物溶解再用Protein-A柱亲和纯化,SDS-PAGE鉴定纯化效果。结果：经两步纯化,抗EGFR单抗纯度达到了98%以上,收率达到90%以上。结论：PEG6000沉淀和Protein-A柱亲和层析相结合,是纯化腹水型单抗体的一种高效方法。     

小儿七星茶泡腾片成型工艺优选

目的 研究小儿七星茶泡腾片的最佳制备工艺. 方法 采用正交实验方法,以pH、崩解时限、口感、硬度作为评定指标,对泡腾剂配比、聚乙二醇(PEG)6000、填充剂、矫味剂的用量进行优化,并对结果 进行了验证. 结果 柠檬酸与碳酸氢钠的配比为0.6:1,PEG6000用量为9.0%,甜菊糖用量1.1%. 结论 该制备工艺稳定、可行.     

猪凝血因子Ⅹ及凝血因子Ⅹa的制备工艺研究

目的从猪血中制备猪凝血因子Ⅹ(FⅩ)及凝血因子Ⅹa(FⅩa)。方法采用柠檬酸钡吸附、硫酸铵分级沉淀、DEAE 纤维素离子交换色谱等方法制备FⅩ,并优化FⅩ的激活条件,从而制备FⅩa。通过SDS PAGE进行FⅩ和FⅩa的检测。结果建立了FⅩ和FⅩa的制备工艺,收率分别为每1ml血浆34,12 μg;采用胰蛋白酶激活FⅩ,激活条件为:0 .0 2 %酶浓度、pH 8.0、37℃下作用70min。结论此工艺操作简单,收率高,适合于规模化生产     

猪凝血因子X及凝血因子Xa的制备工艺研究

目的 从猪血中制备猪凝血因子X(FX)及凝血因子Xa(FXa)。方法 采用柠檬酸钡吸附、硫酸铵分级沉淀、DEAE-纤维素离子交换色谱等方法制备FX，并优化FX的激活条件，从而制备FXa。通过SDS-PAGE进行FX和FXa的检测。结果 建立了FX和FXa的制备工艺，收率分别为每1ml血浆34，12μg；采用胰蛋白酶激活FX，激活条件为：0．02％酶浓度、pH8．0、37℃下作用70min。结论 此工艺操作简单，收率高，适合于规模化生产。     

猪血、猪脾白细胞的培养及干扰素的诱生

采用新鲜猪血、猪脾为原料,以D-Hank’s液及Hank’s液和RPMI-1640培养基培养白细胞,以新城疫鸡瘟病毒减毒株（NDV-F）为干扰素诱生剂,在 37℃、pH 7.2条件下培养18 h.制得猪脾干扰素,效价1620 u/ml,猪血干扰素,效价 1360 u/ml。     

巨噬细胞来源的外泌体不同提取方法的比较

目的根据聚乙二醇(polyethylene glycol,PEG)沉淀原理,结合超高速离心改进一种简便、快速的细胞外泌体分离方法。方法采用超高速离心法、PEG6000沉淀结合超高速离心和EXO Quick试剂盒,分别提取巨噬细胞外泌体。通过透射电镜、NanoSight ZS及Nanoparticle tracker(NTA)分析外泌体的形态和粒径;蛋白印迹法鉴定细胞外泌体标志物Alix、TSG101的表达情况。结果 PEG6000沉淀结合超高速离心法获得的外泌体,其粒径分布和差速离心法相当,小于商品化的EXO Quick试剂盒。而外泌体表面蛋白Alix、TSG101在3种方法提取的沉淀物中都有表达。透射电镜结果显示,3种方法均获得典型的盘状囊泡。结论实验改进的细胞外泌体提取方法操作简洁、效率高,并且成本较低。     

妇阴康洁栓剂制备工艺研究

目的 研究妇阴康洁栓剂的制备工艺。方法采用满因子分析设计法，以栓剂体外溶化时间为指标，选定处方中聚乙二醇（PEG）4000／PEG 6000的比例和栓剂制备时水浴加热的温度作为考察因素。结果 处方间PEG 4000／PEG 6000比例对栓剂体外溶化时间有显著影响，水浴温度影响较小，处方中PEG 4000／PEG 6000比例最佳为7：1。结论 该制备工艺合理、可行。     

正交设计法优选骨肽注射液纯化参数的研究

目的:探讨优选骨肽注射液提取的工艺条件。方法:采用正交设计法,骨肽注射液多肽含量及操作可行性为考察指标,对骨肽注射液提取过程中乙醇沉淀浓度、酸沉淀(pH 2.0～4.0)、碱沉淀(pH 6.8～7.2)3个因素进行筛选。结果:对于选定的三因素三水平,骨肽注射液最佳纯化参数组合为乙醇沉淀浓度70%、酸沉淀pH 4.0,碱沉淀pH 8.0,结果具有较显著的统计学意义。结论:根据骨肽注射液最佳提取条件对猪骨进行骨多肽提取,提取效率高,成本低。     

Efficacy of gut lavage,hemodialysis, and hemoperfusion in the therapy of paraquat or diquat intoxication

Clinical and in vitro investigations were carried out to test the efficacy of gut lavage, hemodialysis, and hemoperfusion in the treatment of poisoning with paraquat or diquat. In a patient suffering from diquat intoxication 130 times more diquat was removed by gut lavage 30 h after ingestion than was removed by complete aspiration of the gastric contents.Determination of in vitro clearances for paraquat and diquat by hemodialysis showed that, at serum concentrations of 1–2 ppm, such as are frequently encountered in poisoning in man, toxicologically relevant quantities of herbicide cannot be removed from the body. At a concentration of 20 ppm, on the other hand, hemodialysis proved to be effective, the clearance being 70 ml/min at a blood flow rate of 100 ml/min. The efficacy of hemoperfusion with coated activated charcoal was on the whole better. Especially at concentrations around 1–2 ppm, the clearance values for hemoperfusion were some 5–7 times higher than those for hemodialysis.In a patient suffering from paraquat poisoning, both hemodialysis as well as hemoperfusion were carried out. The in vitro results could be confirmed: At serum concentrations of paraquat less than 1 ppm no clearance could be obtained by hemodialysis while by hemoperfusion with activated charcoal quite high clearance values were measured and the serum level dropped down to zero.

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Zusammenfassung Klinische Untersuchungen und Laboratoriumsversuche wurden durchgeführt, um die Wirksamkeit von Darmspülung, Hämodialyse und Hämoperfusion bei Paraquat- und Deiquat-Vergiftungen zu prüfen.Bei einem Patienten wurde 30 Std nach Deiquat-Aufnahme durch Darmspülung 130mal mehr Deiquat entfernt als durch vollständige Aspiration des Mageninhaltes. In vitro-Versuche ergaben, daß bei Blutserumkonzentrationen von 1–2 ppm, die bei Vergiftungen oft gemessen werden, durch Hämodialyse keine toxikologisch relevanten Paraquat- oder Deiquat-Mengen entfernt werden können. Dagegen erwies sich die Hämodialyse bei 20 ppm und einer Blutumlaufgeschwindigkeit von 100 ml/min mit einer Clearance von 70 ml/min als wirksam. Die Hämoperfusion mit beschicheter Aktivkohle war in diesen Versuchen aber eindeutig überlegen, denn insbesondere bei Konzentrationen um 1–2 ppm waren die Clearance-Werte 5–7mal höher als bei der Hämodialyse.Die in vitro-Ergebnisse wurden bei einem Patienten mit einer Paraquat-Vergiftung bestätigt: Bei Konzentrationen unter 1 ppm war die Hämodialyse wirkungslos, während durch Hämoperfusion relativ hohe Clearance-Werte erreicht wurden, so daß der Serumspiegel rasch unter die Nachweisgrenze abfiel.

     

In vitro studies on sterically stabilized liposomes (SL) as enzyme carriers in organophosphorus (OP) antagonism

This study describes a new approach for organophosphorous (OP) antidotal treatment by encapsulating an OP hydrolyzing enzyme, OPA anhydrolase (OPAA), within sterically stabilized liposomes. The recombinant OPAA enzyme was derived from Alteromonas strain JD6. It has broad substrate specificity to a wide range of OP compounds: DFP and the nerve agents, soman and sarin. Liposomes encapsulating OPAA (SL)* were made by mechanical dispersion method. Hydrolysis of DFP by (SL)* was measured by following an increase of fluoride ion concentration using a fluoride ion selective electrode. OPAA entrapped in the carrier liposomes rapidly hydrolyze DFP, with the rate of DFP hydrolysis directly proportional to the amount of (SL)* added to the solution. Liposomal carriers containing no enzyme did not hydrolyze DFP. The reaction was linear and the rate of hydrolysis was first order in the substrate. This enzyme carrier system serves as a biodegradable protective environment for the recombinant OP-metabolizing enzyme, OPAA, resulting in prolongation of enzymatic concentration in the body. These studies suggest that the protection of OP intoxication can be strikingly enhanced by adding OPAA encapsulated within (SL)* to pralidoxime and atropine.     

Synthesis,Cytotoxicity and Antileishmanial Activity of Some N-(2-(indol-3-yl)ethyl)-7-chloroquinolin-4-amines

We report herein the condensation of 4,7-dichloroquinoline (1) with tryptamine (2) and D-tryptophan methyl ester (3) . Hydrolysis of the methyl ester adduct (5) yielded the free acid (6) . The compounds were evaluated in vitro for activity against four different species of Leishmania promastigote forms and for cytotoxic activity against Kb and Vero cells. Compound (5) showed good activity against the Leishmania species tested, while all three compounds displayed moderate activity in both Kb and Vero cells.     

Steroidal sapogenin contents in some domestic plants

In order to find out the values of the steroid resources for the future use. the compositions and contents of steroidal sapogenins from 13 domestic plants have been investigated. As a result,Dioscorea nipponica, D. quinqueloba andSmilax china were found to have large amount of diosgenin. And pennogenin inTrillium kamtschaticum andParis verticillata, yuccagenin inAllium fistulosum, hecogenin inAgave americana and neochlorogenin inSolanum nigum were appeared to be major steroidal sapogenins.     

Aquatic Insects and Trace Metals: Bioavailability,Bioaccumulation, and Toxicity

Abstract

The uptake of metals from food and water sources by insects is thought to be additive. For a given metal, the proportions taken up from water and food will depend both on the bioavailable concentration of the metal associated with each source and the mechanism and rate by which the metal enters the insect. Attempts to correlate insect trace metal concentrations with the trophic level of insects should be made with a knowledge of the feeding relationships of the individual taxa concerned. Pathways for the uptake of essential metals, such as copper and zinc, exist at the cellular level, and other nonessential metals, such as cadmium, also appear to enter via these routes. Within cells, trace metals can be bound to proteins or stored in granules. The internal distribution of metals among body tissues is very heterogeneous, and distribution patterns tend to be both metal and taxon specific. Trace metals associated with insects can be both bound on the surface of their chitinous exoskeleton and incorporated into body tissues. The quantities of trace meals accumulated by an individual reflect the net balance between the rate of metal influx from both dissolved and particulate sources and the rate of metal efflux from the organism. The toxicity of metals has been demonstrated at all levels of biological organization: cell, tissue, individual, population, and community. Much of the literature pertaining to the toxic effects of metals on aquatic insects is based on laboratory observations and, as such, it is difficult to extrapolate the data to insects in nature. The few experimental studies in nature suggest that trace metal contaminants can affect both the distribution and the abundance of aquatic insects. Insects have a largely unexploited potential as biomonitors of metal contamination in nature. A better understanding of the physico-chemical and biological mechanisms mediating trace metal bioavailability and exchange will facilitate the development of general predictive models relating trace metal concentrations in insects to those in their environment. Such models will facilitate the use of insects as contaminant biomonitors.