力达霉素效价单位及效价测定方法的确定

目的：确定力达霉素的效价单位，并建立力达霉素效价测定方法。方法：采用DNA断裂法确定力达霉素标准品引起DNA断裂的最低浓度，并据此浓度确定力达霉素标准品的效价；采用HPLC法分析力达霉素效价与其发色团峰面积的相关性，用力达霉素发色团的峰面积表征力达霉素的效价，用HPLC法测定力达霉素的效价。结果：将首批力达霉素标准品引起pBR322DNA断裂成Ⅲ型的最低剂量规定为10^-4力达霉素单位(u)，即每1mg首批力达霉素标准品的效价为896u；力达霉素效价与其发色团峰面积呈线性关系(r=0．9995)，根据力达霉素发色团的峰面积可以确定力达霉素的效价。结论：本文采用的效价单位确定方法“溯源性”明确；采用HPLC法测定力达霉素的效价可以大大提高传统生物检定的精度。     

格尔德霉素发酵工艺的优化

目的研究格尔德霉素产生菌吸水链霉菌SIPI.A.2039的发酵工艺,以提高产生菌生物合成格尔德霉素的能力。方法以本实验室保藏的吸水链霉菌SIPI.A.2039为出发菌株,从摇床转速、摇瓶装量、碳源、50L发酵罐参数等方面进行发酵工艺的优化研究。结果采用经过优化获得的发酵培养基和培养条件,发酵周期为144h时,格尔德霉素的摇瓶发酵效价可由原来的230μg/mL提高至2500μg/mL。在50L发酵罐中采用优化的溶解氧和补料工艺能够使格尔德霉素的发酵效价达到3700μg/mL,并使发酵周期比摇瓶缩短了48h。结论本实验得到的结果比国内外报道的格尔德霉素的发酵效价提高了两倍以上,该发酵工艺已经达到了产业化的要求。     

克拉霉素的高效液相色谱法与微生物效价测定法分析比较

目的 :比较克拉霉素含量测定的HPLC法和微生物效价测定法两种方法的平行性。方法 :采用HPLC法和微生物法测定 3批样品的标示量。结果 :HPLC法测定的 3批样品的标示量为 97.98% ,99.83 % ,10 1.5 % ,微生物效价测定法为 97.3 % ,97.6% ,98.5 %。结论 :两种结果经t检验分析 ,差异无显著性 (P >0 .0 5 )。     

单链抗体与力达霉素辅基蛋白在大肠埃希菌表达的发酵工艺

目的 确定工程菌E.coli BL21(DE3)StarTM/pEFL表达抗Ⅳ型胶原酶单链抗体与力达霉素辅基蛋白构成的融合蛋白Fv-LDP的摇瓶发酵工艺.方法 比色法测定菌体浓度,干燥失重法测定细菌干重,SDS-PAGE-Spot Denso法测定融合蛋白Fv-LDP表达量,单因素或正交设计优化发酵工艺.结果 确定了融合蛋白Fv-LDP表达的最佳培养基配方和培养条件,融合蛋白Fv-LDP表达量为830μg/ml左右,比LB培养基的产量提高5.6倍.结论 优化工程菌发酵工艺大大提高了融合蛋白Fv-LDP表达水平,不仅为下一步中试研究和规模化生产奠定了基础,而且为其它生物来源的抗体和抗生素组分在基因工程大肠埃希菌的生产提供了发酵方面的实验证据.     

微生物法测定力达霉素的效价

建立力达葛素效价测定的微生物检定法，检定菌为枯草芽孢杆菌。当力达葛素的浓度为1.85～13.51μg／ml时，其抑菌圈直径与反应剂量呈线性关系，并与HPLC法进行比较，测定结果基本一致。     

金属离子对林可霉素生物合成的影响

采用优化设计方法研究金属离子在林可链霉菌(Streptomyces lincolnensis)林可霉素生物合成中的交互作用,并应用均匀设计法,以最终发酵液的生物效价为目标函数建立模型,通过软件Uniform Design Version 3.00分析得出优化配比:FeSO4.7H2O 0.45g/L,CuSO4.5H2O 1g/L,MnCl21g/L;按优化方案添加金属离子,使发酵液的最终生物效价较原配方提高20%,明显提高了林可霉素的发酵水平。     

北极放线菌BF-1发酵条件优化及代谢产物抑菌活性研究

目的优化发酵工艺以提高北极放线菌BF-1发酵液中抑菌活性物质的产量;测定BF-1次级代谢产物的体外抑菌活性。方法以发酵液抑菌活性为指标,采用单因素实验和正交实验对放线菌BF-1发酵培养基和发酵条件进行优化;琼脂稀释法测定BF-1发酵液最低抑菌浓度。结果最佳发酵培养基:淀粉5g.L-1,NH4Cl 5g.L-1,黄豆15g.L-1,MgSO40.25g.L-1,海水晶30g.L-1;最佳发酵条件:28℃,起始pH7,接种量5%;BF-1发酵液对绿脓杆菌的最低抑菌浓度(MIC)为640μg.mL-1。结论北极放线菌BF-1发酵液中次级代谢产物具有显著的体外抑菌活性,优化后BF-1发酵液的抑菌活性与优化前相比提高了约2.6倍。     

副溶血弧菌噬菌体的发酵优化及中试制备工艺探索

目的 为了以低成本的方法制备出高效价的噬菌体制剂，对副溶血弧菌噬菌体Vpas_PP24的发酵条件和关键工程化制备工艺进行探索。方法 采用单因素实验法，研究噬菌体Vpas_PP24发酵体系的不同培养基、金属离子、pH、接种量及培养时间对其效价的影响；在中试规模对优化发酵工艺进行了工程化放大；并以膜过滤结合热除菌法进行了下游工程化噬菌体分离及终端除菌工艺探索。结果 确立了副溶血弧菌噬菌体Vpas_PP24的最优培养基及最佳发酵条件。摇瓶水平最适出发培养基为2216E液体培养基；Mg2+和Ca2+可促进Vpas_PP24的增殖，且最适浓度为30 mmol·L-1；最适pH为8；VP8最适接种量为1%；最适培养时间为16 h。在摇瓶水平优化后的效价达到3.0×1010 PFU/mL，较优化前提高了14倍。随之，基于该优化条件，成功将噬菌体发酵放大到50 L中试规模，并建立上游液体深层发酵工程化生产Vpas_PP24的上游工艺，效价达到3.2×1010 PFU/mL，与此同时，建立了下游噬菌体除菌工艺，得到效价为2.5×108 PFU/mL的噬菌体Vpas_PP24批量液体制剂。结论 系统优化了噬菌体Vpas_PP24的发酵条件，并在中试规模进行了工艺放大，成功建立了中试上游噬菌体发酵工艺及下游噬菌体分离工程化工艺及末端去除宿主菌的技术手段，为噬菌体类替抗产品的工程化生产提供了可参考的范例。     

头孢菌素C两种发酵培养基的比较

本文研究了以顶头孢霉菌为试验菌株,采用200立升和2.5吨不锈钢发酵罐,作了不同培养基对顶头孢霉菌产生头孢菌素C影响的试验。 培养基用一号发酵培养基(以淀粉、糊精和玉米浆为碳、氮源)和二号发酵培养基(以植物油和玉米浆为碳、氮源),发酵中途都采用补料通氨水调节pH的工艺条件,常量法测糖;甲醛法测氨基氮;用定量发酵液以每分钟3000转离心五分钟,计算发酵过程中菌丝量的百分数(V/V％);用紫外分光光度法、菸酰胺法、高压液相色谱仪测定发酵液中的总效价、头孢菌素C及去乙酰头孢菌素C的效价。 为选择较佳的发酵工艺条件,首先在200立升罐中做了三个试验内容,结果:以发酵中途不补料设其产量为100％,则发酵中途补     

RP-HPLC法检测发酵液中利普司他汀的效价

目的建立奥利司他中间体利普司他汀发酵效价的检测方法。方法采用RP-HPLC法。色谱柱为Ultimate ODS硅胶柱(4.6 mm×250 mm,5μm),流动相为乙腈-水(体积比4∶1),柱温为35℃,进样量为10μL,流速为1.1 mL·min-1,检测波长为210 nm。结果发酵70、100和130 h时测得发酵液中利普司他汀效价分别为2.518、4.323、6.981 g·L-1。结论该方法可有效地检测发酵液中利普司他汀效价。     

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.     

Pharmacological treatment of COVID-19: an opinion paper

The precocity and efficacy of the vaccines developed so far against COVID-19 has been the most significant and saving advance against the pandemic. The development of vaccines has not prevented, during the whole period of the pandemic, the constant search for therapeutic medicines, both among existing drugs with different indications and in the development of new drugs. The Scientific Committee of the COVID-19 of the Illustrious College of Physicians of Madrid wanted to offer an early, simplified and critical approach to these new drugs, to new developments in immunotherapy and to what has been learned from the immune response modulators already known and which have proven effective against the virus, in order to help understand the current situation.     

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.     

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.     

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.