乙型脑炎减毒活疫苗病毒滴度单瓶平行检测与多瓶混样检测的对比分析

目的:比较乙型脑炎减毒活疫苗病毒滴度单瓶平行检测与多瓶混样检测之间的差异性,为进一步完善中国药典中相关技术要求奠定基础。方法:1人份和5人份乙型脑炎减毒活疫苗各抽取20批,同一批次疫苗样品随机取样6瓶,3瓶用于病毒基础滴度检测,3瓶用于病毒热稳定性滴度检测。检测方式分别为3瓶单瓶平行检测与3瓶混样检测：单瓶平行检测是从...     

乙型脑炎减毒活疫苗单一病毒收获液在2～8℃条件下储存的稳定性研究

目的 考察在现行工艺规定的2～8℃储存条件下,乙型脑炎减毒活疫苗(乙脑活疫苗)单一病毒收获液病毒滴度的稳定性.方法 取常规生产中收获的高、中、低滴度的单一病毒收获液各6批,按正常生产程序和工艺条件储存于2～8℃,自第5天起每隔1d无菌取样,进行病毒滴定,采用极差控制图、线性回归、单因素方差分析等方法对收获液的病毒滴度结果进行统计分析.结果 常规生产中高、中、低滴度段的单一病毒收获液在2～8℃储存21 d,各单一病毒收获液的病毒滴度均呈下降趋势,并且不同滴度段病毒收获液间的日均下降滴度差异无统计学意义(F=1.67,P=0.222),总体日均下降滴度为0.036 lg蚀斑形成单位/ml.结论 掌握了单一病毒收获液在工艺规定储存条件下的滴度下降趋势及日均下降水平,为乙脑活疫苗半成品的点配制及其病毒滴度的一致性控制提供了数据支持.     

细胞工厂在乙型脑炎减毒活疫苗生产中的应用

目的  采用40层细胞工厂（CF40）替代15 L转瓶工艺制备乙型脑炎（乙脑）减毒活疫苗。方法  分别采用CF40和15 L转瓶工艺制备乙脑减毒活疫苗,通过显微镜观察和细胞计数来考察细胞质量。病毒收获时观察细胞病变情况,并对2种工艺制备的原液、半成品以及成品进行病毒滴度检测对比,成品37 ℃放置7 d考察成品热稳定性。结果  两种工艺相比,细胞工厂内单位面积内细胞数量差异存在统计学意义（P＜0.05》）,生长一致性和单只地鼠制备的细胞数量均优于15 L转瓶;CF40和15 L转瓶制备的单一病毒收获液平均滴度分别为7.59 lgPFU/ml和7.56 lgPFU/ml,没有显著差异(P＞0.05),但CF40单一病毒收获液收率一致性更好;两种工艺制备的原液、半成品、成品滴度以及7 d后37 ℃热稳定性均符合企业注册标准规定。结论  使用CF40制备乙脑减毒活疫苗是可行的,并且在产品收率和一致性方面优于15 L转瓶工艺。     

乙型脑炎减毒活疫苗病毒滴定用BHK21细胞密度和代次的确认

 目的  确定乙型脑炎（乙脑）减毒活疫苗病毒滴定使用的BHK₂₁细胞的密度和代次范围。方法  抽取20批不同代次BHK21细胞在6孔细胞板内培养成单层,进行细胞计数,计算细胞密度,滴定同批次乙脑病毒滴度国家参考品。将BHK₂₁细胞从P95传代至P105,观察细胞形态及生长情况,与已用于检定的BHK21细胞（＜P95）对比。用P95~P105 BHK₂₁细胞重复6次滴定同批参考品,对滴度结果进行双因素方差分析,并与21细胞的病毒滴度检测结果进行趋势分析比较。结果  20批6孔板单层BHK21细胞密度均值1.55×10⁶  个/ml,差异较大（95%置信区间：7.89×10⁴～3.02×10⁶  个/ml）。病毒滴度结果均在规定范围内（6.48~7.12 lg蚀斑形成单位/ml,相对标准偏差<5%）,单层细胞密度对滴度结果无显著影响。P95~P105 BHK21细胞的生长情况及细胞形态与F=0.13,P=0.721 4）、不同细胞代次（F=1.17,P=0.337 6）及两者的交互作用（F=0.30,P=0.978 0）对检测结果的影响无统计学意义,试验结果与较低代次细胞（21细胞密度对滴度检测结果无显著影响。     

乙型脑炎减毒活疫苗稳定性评价

目的 对乙型脑炎减毒活疫苗(乙脑疫苗)的稳定性进行评价.方法 选取2种规格(1、5次人用剂量)的成品乙脑疫苗进行(25±2)、(37±2)℃条件下保存的加速稳定性试验,以及2～8℃条件下的长期稳定性考察,检定关键质量指标,并对长期稳定性数据进行回归分析.结果 4批1次人用剂量、2批5次人用剂量疫苗于(25±2)℃、相对湿度(60±5)％存放3个月,(37±2)℃、相对湿度(75±5)％存放14d,其外观、病毒滴度、水分、pH值均符合质量标准.9批1次人用剂量、7批5次人用剂量疫苗在2～8℃存放24个月的检定结果均符合质量标准.回归分析显示,在2～8℃存放时间每增加1个月,1、5次人用剂量疫苗的病毒滴度分别平均降低0.020和0.017 lg蚀斑形成单位/ml,水分分别平均增加0.055％和0.047％.根据回归方程因变量均值95％置信区间计算出的乙脑疫苗有效期远长于现行注册标准规定的18个月.结论 被检乙脑疫苗质量稳定性良好,产品安全可靠.     

乙型脑炎减毒活疫苗的国外临床研究

 乙型脑炎减毒活疫苗（乙脑活疫苗）是中国自主研发的疫苗,也是中国第一个通过世界卫生组织预认证的疫苗。此文介绍了乙脑活疫苗在国外进行的临床研究,包括该疫苗在亚洲不同地区人群中的免疫原性、免疫效力和安全性研究,从而为该疫苗在中国以外的乙型脑炎流行区的推广和使用提供科学依据。     

乙型脑炎病毒及疫苗研究进展

乙型脑炎病毒是黄病毒科中致病性最强的病毒之一,目前对该病毒的分子生物学研究进展较快.现阶段国内外主要使用三种类型疫苗,即鼠脑纯化灭活疫苗、原代地鼠肾细胞灭活疫苗和原代地鼠肾细胞减毒活疫苗.灭活疫苗和减毒活疫苗各有利弊.基因工程疫苗(包括载体重组活疫苗、嵌合减毒活疫苗、病毒样颗粒、亚单位疫苗及DNA疫苗)成为乙型脑炎疫苗发展的方向.     

乙型脑炎减毒活疫苗主种子批猴体神经毒力试验

 目的   为了解乙型脑炎（乙脑）减毒活疫苗主种子批弱毒株的神经毒力减弱程度,对其进行猴体神经毒力试验,以观察其安全性。 方法  对恒河猴脑内和脊髓内同时注射乙脑病毒强毒株SA14或减毒活疫苗主种子批弱毒株SA14-14-2,然后进行为期21 d的临床体征观察,并进行组织病理学检查。 结果   强毒株组动物接种后的临床体征为典型的神经毒性反应,动物发病并死亡。组织病理学检查显示,中枢神经系统严重受损,并有神经细胞坏死。弱毒株组动物仅出现短时发热,组织病理学检查可见轻微炎症反应,未见神经细胞坏死现象。 结论   乙脑减毒活疫苗主种子批的神经毒力高度减弱, 对恒河猴是安全的。     

乙型脑炎减毒活疫苗批签发管理与质量评价

目的比较乙型脑炎减毒活疫苗实施批签发前后的质量变化,评价疫苗的质量控制。方法通过对市场抽检疫苗样品和批签发送检的疫苗样品质控关键项目测定结果及制造和检定记录摘要审核,对疫苗的质量和改进趋势进行分析、归纳和总结。结果3个企业生产的乙脑活疫苗在1990～2004年期间,尤其是生产初期,疫苗病毒滴度≥5.7lgpfu.mL-1的批次合格率为58.6%～100%;疫苗热稳定性试验合格批次占15%～100%;其中A企业2001～2004年,疫苗热稳试验100%合格。从2005年实施批签发后,3个企业生产的乙脑活疫苗病毒滴度均为100%合格。2006和2008年国家市场监督抽验的疫苗病毒滴度结果合格率达97.4%。结论乙脑活疫苗实施批签发后的病毒滴度合格率达100%,较实施前有明显提高。国家市场监督抽验的疫苗测定结果显示,乙脑活疫苗的质量稳定,病毒滴度合格率为97.4%。通过疫苗批签发可以准确、具体的了解企业生产和质量管理情况,督促企业规范生产、完善工艺、加强质量管理。     

乙型脑炎减毒活疫苗SA14-14-2在BHK21细胞上传代后的基因序列研究

目的：研究乙型脑炎减毒活疫苗SA14-14-2在BHK₂₁细胞上传代后的病毒基因序列,以深入控制乙型脑炎减毒活疫苗的安全性。方法：将乙型脑炎减毒活疫苗SA14-14-2在BHK₂₁细胞上连续传代,选取第二代病毒（P2）、第五代病毒（P5）、第十五代病毒（P15）提取RNA,反转录为cDNA后,进行各代病毒全基因序列分析,分析与乙型脑炎病毒毒力密切相关的关键位点基因是否发生改变。结论：乙型脑炎减毒活疫苗SA14-14-2在BHK₂₁细胞上传5代后,病毒E蛋白上关键基因第279位（E279）氨基酸由甲硫氨酸（M）突变为赖氨酸（K）。     

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.     

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.     

Alzheimer’s disease – current trends in basic and clinical research. Highlights from the 16th ECNP

Advances in the molecular biological knowledge of neuronal nicotinic acetylcholine receptors (nAChRs) have led to a growing interest by the pharmaceutical industry in the development of novel compounds that selectively modulate nAChR function. The ability of (-)-nicotine, an activator of nAChRs, to enhance attentional aspects of cognition in animals and humans, to exert neuroprotective and anxiolytic-like effects, and presumably to mediate the negative correlation between smoking and Alzheimer's (and Parkinson's) Disease, has focused interest on the potential therapeutic utility of modulators of nAChR function for treatment of some of the deficits associated with these progressive, neurodegenerative conditions. Numerous compounds are known which activate nAChRs and which might serve as lead compounds toward the development of such agents. The pharmacologic diversity of neuronal nAChR subtypes suggests the possibility of developing selective compounds which would have more favourable side-effect profiles than existing agents. This broader class of agents, collectively called cholinergic channel modulators (ChCMs), is anticipated to encompass compounds which would have more favourable side-effect profiles than existing agents, which generally exhibit low selectivity. This selectivity may be achieved by preferentially activating some subtypes of nAChRs (i.e., Cholinergic Channel Activators, ChCAs) or inhibiting the function of other subtypes (Cholinergic Channel Inhibitors, ChCIs). An overview of the biology of nAChRs and the rationale for the use of ChCMs for the treatment of dementia related to neurodegenerative diseases are presented, followed by a discussion of lead compounds and compounds under consideration for clinical evaluation.     

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.