淡水鱼脑AChE的动力学特性及抗抑制性

用PEG2000双水相萃取、DEAE-Sephadex A-50和SephadexG-200方法分离提纯麦穗鱼(Pseudorasbora parva)、金鱼(Carassius auratus)和虹鳟鱼(Salmo gairdneri)脑乙酰胆碱酯酶(AChE),比较这3种淡水鱼脑AChE纯酶液与粗酶液的动力学特性、抗抑制性及其种类之间的差异.结果表明,这3种鱼脑AChE的性质在种类间的差异要远远小于其在粗酶状态时酶活性的差异,由此得出AChE性质在种类间的差异并非影响这3种淡水鱼脑AChE活性差异的主要因素;由AChE抗抑制试验结果可知,这3种鱼脑AChE粗酶液对经溴水氧化后的马拉硫磷和三唑硫磷的敏感性及种间的差异都远大于纯酶液.     

对有机磷农药敏感的海鱼脑AChE筛选研究

以10种海洋鱼类:鲅鱼、黄花鱼、鲈鱼、黄鱼、美国红鱼、真鲷、黑鲷、面包鱼、黑头鱼和鳗鲡为实验材料,采用半抑制浓度(IC₅₀)和双分子速率常数(K_i)为指标,通过体外抑制作用比较了这10种海鱼脑AChE对马拉硫磷和甲基对硫磷的敏感性,并分析了海鱼脑AChE作为监测海水中有机磷农药指示酶的可行性,由此筛选出适于制备海水有机磷农药酶传感器的指示酶.实验表明:10种海鱼脑AChE的基础活性在4.62～35.65μmol/(min·g)之间,面包鱼的酶活性最高,黄花鱼的酶活性最低.对马拉硫磷,由IC₅₀所得到的AChE敏感性为:鲅鱼、黄鱼、鲈鱼、真鲷>黑鲷>黑头鱼、鳗鲡>美国红鱼、黄花鱼>面包鱼;由Ki所得到的AChE敏感性为:鲅鱼>黄鱼、鲈鱼、真鲷>黑鲷、黑头鱼、鳗鲡>美国红鱼、黄花鱼>面包鱼.对甲基对硫磷,由IC50所得到的AChE敏感性为:鲅鱼>鲈鱼>黄鱼、黄花鱼、真鲷、黑鲷、黑头鱼、鳗鲡>美国红鱼>面包鱼;由Ki所得到的AChE敏感性为:鲅鱼>鲈鱼、黄鱼、黄花鱼、黑鲷、黑头鱼、鳗鲡>真鲷>美国红鱼>面包鱼.鲅鱼脑AChE对2种有机磷农药都较为敏感.影响鱼脑AChE对有机磷农药敏感性的主要因素为生物来源、农药种类和抑制方式,而与酶的基础活性无关.海鱼脑AChE活力抑制作用与水体中的有机磷农药具有良好的剂量-效应关系,在1～50mg/L浓度范围内,两者间呈现较好的线性相关性(r²≥0.93).根据AChE的敏感性及其对有机磷农药的线性响应程度,鲅鱼脑AChE适于作为监测马拉硫磷和甲基对硫磷酶传感器的指示酶.     

鲫鱼脑AChE制备及对几种有机磷农药敏感性研究

乙酰胆碱酯酶(AChE)是生物神经兴奋传导中的一种关键性酶。有机磷农药能够磷酰化AChE活性中心的丝氨酸羟基,导致神经传导阻断,因此AChE是有机磷农药残留检测的靶酶。文章选择鲫鱼脑作为酶源,对AChE提取方法进行了研究,并探讨了其对有机磷农药的敏感性。首先粗提AChE,通过正交试验确定了AChE酶活测定的最佳条件,研究了粗酶对几种有机磷农药的敏感性及检测限,之后通过层析方法纯化AChE,得到高活性的目标酶。研究表明AChE活性测定最佳条件为:200mmol/L底物、pH8.0、30℃水浴30min,这些因素对酶得活性影响顺序为:底物浓度pH温度时间,酶剂量对其影响最小;由鲫鱼脑粗提得到的AChE对4种有机磷农药的敏感性为:敌敌畏敌百虫氧乐果三唑磷辛硫磷乙酰甲胺磷;纯化后酶活达到了458.43μmo(lmin·mg),适合用于有机磷农药检测。     

5种海鱼脑AChE对2种有机磷农药的敏感性比较

以鲈鱼、黄鱼、美国红鱼、真鲷和黑鲷为实验材料,采用半抑制浓度(IC_(50))和双分子速率常数(K_i)为指标,通过体外抑制作用比较了5种海鱼脑乙酰胆碱酯酶[AChE(EC 3.1.1.7)]对马拉硫磷和甲基对硫磷的敏感性。结果表明:(1)5种海鱼脑AChE基础活性为10.18～22.64[μmol/(min·g)],黄鱼的酶活性最高,鲈鱼的酶活性最低。(2)对马拉硫磷,由IC_(50)所得到的AChE敏感性为:黄鱼、鲈鱼、真鲷>黑鲷>美国红鱼;由K_i所得到的AChE敏感性为:鲈鱼、黄鱼、真鲷>黑鲷>美国红鱼。对甲基对硫磷,由IC_(50)所得到的AChE敏感性为:鲈鱼>黄鱼、真鲷、黑鲷>美国红鱼;由K_i所得到的AChE敏感性为:鲈鱼、黄鱼、黑鲷>真鲷>美国红鱼。鲈鱼和黄鱼对2种有机磷农药都较为敏感。(3)鱼脑AChE活力抑制作用与水体中的有机磷农药具有良好的剂量—效应关系。在1～100mg/L浓度范围内,两者间呈现较好的线性相关性,其相关系数基本上都在0.98以上。根据AChE的敏感性及其对有机磷农药的线性响应程度,黄鱼和鲈鱼脑AChE分别适于作为监测马拉硫磷和甲基对硫磷的指示酶。     

麻黄碱在斑马鱼体内的器官特异性蓄积及毒代动力学

麻黄碱（ephedrine,EPH）是一种生物碱,用于减轻感冒、过敏性鼻炎、鼻炎及鼻窦炎引起的鼻充血症状,控制支气管哮喘等,同时还是制造毒品冰毒的原料.EPH已经在地表水中广泛检出,并可能对水生生物甚至生态系统产生不利影响.但目前,EPH在水生生物中的摄取、器官分配和毒代动力学过程还没有受到关注.本研究中将斑马鱼（Danio rerio）在半静态系统中暴露于EPH,研究斑马鱼的不同器官对EPH的吸收、蓄积以及EPH在斑马鱼体内的毒物代动力学.暴露浓度分别为0.01、1.00和100.00 μg·L^-1,暴露14 d后,在斑马鱼脑、肝、肠、卵巢及肌肉中均检测到EPH,高浓度组斑马鱼脑中EPH最高达到84.97 ng·g^-1;EPH的平均含量的相对大小遵循以下顺序：脑 > 卵巢 > 肝 >肠 > 肌肉.斑马鱼器官中的麻黄碱的摄取速率常数（K_u）为0.23~570.31 L·（kg·d）^-1,消除速率常数（K_e）为1.22~6.11 d^-1,半衰期为0.12~0.57 d.观察到的生物富集系数（BCF_o）和动力学来源的生物富集系数（BCF_k）范围分别为0.24~337.33 L·kg^-1和0.13~316.43 L·kg^-1,处于相同的水平.     

两性复配修饰黏土对苏丹草修复Cu2+污染紫色土的影响

为了探索添加吸附材料对植物修复Cu²⁺污染土壤的影响,选取不同十二烷基二甲基甜菜碱 （BS-12）和十四烷基磺酸钠 （STS）复配修饰比例（50%BS-12+25%STS、50%BS-12+50%STS和50%BS-12+100%STS）的膨润土作为吸附材料,将其分别以1%的质量比加入到紫色土（PS）中,形成PS（对照）、PBS₂₅（50%BS-12+25%STS修饰膨润土混合紫色土）、PBS₅₀和PBS₁₀₀四种混合土样。采用盆栽实验对比苏丹草在0（CK）、50、100、200和500 mg/L Cu²⁺污染混合土样中的发芽率、株高、地上部和根系生物量,并分析混合土样和苏丹草对Cu²⁺污染的修复效果。结果显示,随着Cu²⁺处理浓度的增加,苏丹草在PS上的种子发芽率逐渐降低,而在各PBS土样上的发芽率基本保持先增加后稳定的趋势;不同Cu²⁺污染混合土样上的苏丹草株高均随生长时间的增加而增加。苏丹草地上部、根系干重和鲜重在PBS₂₅、PBS₅₀土样上均随Cu²⁺处理浓度的增大而降低,而在PS和PBS₁₀₀土样上均随Cu²⁺处理浓度的增加先增加后降低,峰值分别出现在200和100 mg/L的Cu²⁺处理。Cu²⁺污染处理下苏丹草各生理指标之间均保持中度以上正相关关系,且相关系数表现为PBS₁₀₀最高、PBS₂₅和PBS₅₀次之、PS最低的趋势。各混合土样对Cu²⁺的吸附能力表现为PBS₂₅ > PBS₅₀ > PBS₁₀₀ > PS,苏丹草对Cu²⁺的富集含量表现为PS > PBS₁₀₀ > PBS₅₀ > PBS₂₅的趋势,苏丹草联合两性复配黏土修复Cu²⁺污染紫色土的修复率达99%以上。     

O2、NO-3、NO-2为电子受体的生物除磷比较

通过平行运行3个分别以O₂、NO^-₃、NO^-₂为电子受体的SBR除磷系统,探讨了反硝化除磷区别于好氧除磷的工艺特征及其内在微生物竞争机制.NO^-₂不会对经驯化后反硝化聚磷菌（DPAO）的缺氧吸磷产生直接抑制作用,但其作为反硝化除磷电子受体的效能远低于NO^-₃;具备利用NO^-₃而缺乏NO^-₂反硝化酶系的DPAO（DPAO₅）流失及聚糖菌（GAO）增殖是根本原因.而NO^-₃是一种高效电子受体,其反硝化除磷效能与以O₂为受体的好氧除磷系统相当,两者在除磷计量学和功能菌群构成上十分接近.作为聚磷菌（PAO）的竞争者,GAO在3个研究系统中均大量存在.基于对不同电子受体的利用能力,PAO包含P_O、P_N、P_n、P_Nn、P_ON、P_ONn等6个种类.P_ON和P_ONn等兼性PAO是生物除磷的主体,其在污泥中比例越高,系统的除磷负荷也越大,即SBR_o＞SBR_n5＞SBR_n3.     

基于固定化AChE的流动注射型酶传感器研究

以固定化鲅鱼脑组织乙酰胆碱酯酶[acetylcholinesterase,AChE(EC 3.1.1.7)]为识别元件,以pH电极为换能器,构建流动注射型AChE酶传感器.该传感器在以磷酸盐缓冲液为载液的条件下具有良好的重现性(RSD=1.427%,n=10)和敏感性,可实现对有机磷化合物的在线监测;对甲基对硫磷具有线性响应的浓度范围为4.29×10-10^-4.29×10^-8mol·L^-1,最低检测限为1.3×10^-     

新型饮用水除氟材料Bio-F的除氟特性和比较研究

对3种传统除氟剂活性氧化铝、骨炭和改性沸石与自制的新型生物除氟剂Bio-F的除氟性能及影响因素（材料粒径、pH值、吸附时间、水样含氟浓度、其它离子、再生能力等）进行了比较,并模拟动态实验评估了这4种除氟材料对实际高氟地下水处理的效果.结果表明,Bio-F生物除氟剂对F^-的吸附过程符合Lagergren一级吸附动力学特征（R²=0.958 0）,吸附速率较快,且该过程属于吸热反应; Bio-F吸附F^-符合Langmuir吸附等温模型（R²=0.999 2）,吸附容量高,静态吸附容量可达4.088 3 mg·g^-1,分别约是活性氧化铝和改性沸石的1.8和 5.8倍.4种除氟材料吸附容量与氟浓度正相关,与吸附剂粒径负相关.高浓度的CO^2-₃、HCO^-₃明显抑制Bio-F的除氟（p<0.05）,但高浓度的Ca²⁺、NO^-₃、HPO^2-₄有利于Bio-F的除氟（p<0.001）.Bio-F除氟最佳停留时间3～4 min,远远低于沸石20 min和活性氧化铝11 min.在pH 4.0～9.0范围内Bio-F可保持90％以上吸附F^-的能力.再生性能稳定,10次再生后吸附容量变化不超过15%.Bio-F综合性能优于其它3种传统除氟剂,在我国广大农村地区推广有显著优越性.     

用高浓度对苯二甲酸溶液产电的微生物燃料电池

以高浓度对苯二甲酸（TA）溶液为底物,研究微生物燃料电池的产电效果.以厌氧活性污泥作为接种体,经过210 h驯化,开路电压达到0.54 V,证明了TA可以作为微生物燃料电池的底物进行产电.深入研究了不同pH值和底物浓度对产电的影响,实验结果表明,当体系pH为8.0时,负载两端（R=1 000 Ω）电压最大,底物浓度越高,负载两端电压越大,并逐渐趋近于一个最大值,通过Monod方程回归得到该微生物燃料电池体系输出电压的最大值U_max为0.5 V,K_s值为785.2 mg/L.当底物浓度（以COD计）为4 000 mg/L时,最大输出功率密度为96.3 mW/m²,库仑效率为2.66%,COD去除率为80.3%.     

Measurement of cholinesterases in amniotic fluid using a naphthyl acetate as substrate: A possible initial test for prenatal diagnosis of open neural tube defects

A quantitative method for cholinesterases in amniotic fluid using the non-specific substrate α naphthyl acetate and the cholinesterase-specific inhibitor, eserine, is described. This assay was used to test 671 samples of amniotic fluid. The diagnoses for fetal ONTDs, based on the levels of AChE + ChE, were compared with those made for the same samples by the AFP method. Correct diagnoses were made by both methods with amniotic fluid from 35 women carrying fetuses with ONTDs and 631 carrying normal fetuses. There were five false-positive test results for normal fetuses by both methods when the cut-off points were 5 standard deviations above the mean for AFP and above the upper limit of the normal range (7. 5 milliunits) for cholinesterase (AChE + ChE). None of the false-positive samples from either method had the acetylcholinesterase band of activity characteristic of ONTDs after gel electrophoresis. In addition to the above 671 samples, 37 pregnancies with serious fetal abnormalities other than ONTDs were tested. Two were identified by both the AFP and AChE + ChE methods, two more by AFP assay and one other by the AChE + ChE assay.     

家蝇AChE检测毒死蜱残留的最佳条件研究

以纯化的家蝇(Museadomestica)乙酰胆碱酯酶(AChE)为试验材料,研究了缓冲液pH值、底物(ATCh)浓度、温度三个因素对酶活性的影响。结果表明:在pH值为8.5￣9.0,温度为30℃,底物浓度为2.5mmol/L的条件下,该酶具有最大活性。在此基础上,测定了不同pH值的缓冲反应体系和不同抑制时间条件下毒死蜱(Chiorpyifos)对AChE活性的抑制,发现pH值为8.0时AChE对毒死蜱最为敏感,而且对毒死蜱具有良好的线性响应关系(R2=0.9973),半抑制浓度仅为39.1μg/(L抑制时间为20min),检测下限为4.4μg/L。随着抑制时间的延长,毒死蜱对AChE的抑制程度逐渐增强,检测限可进一步降低。可见家蝇AChE是指示环境中毒死蜱污染程度的良好材料。     

Application of environmental DNA analysis to inform invasive fish eradication operations

Environmental DNA (eDNA) detection of non-native species has considerable potential to inform management decisions, including identifying the need for population control and/or eradication. An invasive species of European concern is the Asian cyprinid fish, topmouth gudgeon (Pseudorasbora parva). Here, eDNA analyses were applied at a commercial angling venue in southern England to inform operations aiming to eradicate P. parva, which had only ever been observed in one of the venue’s seven unconnected angling ponds. Eradication of P. parva was initially attempted by repeated depletion of the population using fish traps (crayfish traps fitted with 5 mm mesh netting) and the introduction of native predators over a 4-year period. The very low number of P. parva captured following these eradication efforts suggested a possible population crash. Conventional PCR analysis of water samples using species-specific primers was applied to all seven ponds to confirm that P. parva was present in only one pond, that the eradication attempt had indeed failed and that the species’ distribution in the pond appeared to be restricted to three bankside locations. The continued presence of P. parva at these locations was confirmed by subsequent trapping. Water samples from an adjacent, unconnected stream were also analysed using the eDNA methodology, but no DNA of P. parva was detected. The results suggest that further management action to eradicate P. parva be focused on the pond shown to contain the isolated P. parva population and thereby eliminate the risk of further dispersal. This study is the first to apply eDNA analysis to assess the efficacy of an eradication attempt and to provide evidence that the species was unlikely to be present in the other ponds, thus reducing the resources needed to control the species.     

热污染对金鲫鱼组织内四种酶活性的影响

在实验室模拟条件下,研究了热污染对金鲫鱼(Carasslusauratus)肝脏、肾脏与血清中的谷丙转氨酶(GPT)、谷草转氨酶(GOT)和过氧化氢酶(CAT)活性及脑中乙酰胆碱酯酶(AChE)活性的影响.结果表明,金鲫鱼肝脏和血清中的GOT、肝脏和肾脏中的CAT以及脑中的AChE活性均可以用作其组织细胞因热污染受胁迫及损伤的生物标志物,金鲫鱼可作为热污染环境下的指示生物.同时分析热污染时各种酶活性的变化表明,酶活性的改变是鱼体对环境增温产生的一种应激机制,用以减小危害;若超过其耐受范围,这一机制则被破坏,而使机体受损.     

Predictive value of amniotic acetylcholinesterase analysis in the diagnosis of fetal abnormality in 3700 pregnancies

The value of acetylcholinesterase (AChE) analysis as an adjunctive test to amniotic alpha fetoprotein (amAFP) for the diagnosis of fetal abnormality has been investigated in a series of 3785 amniotic fluid samples. Quantitative analysis of AChE performed retrospectively on a selected group of 541 amniotic fluid samples failed to discriminate between normal and open neural tube defect pregnancies. Qualitative analysis of AChE by polyacrylamide gel (PAG) electrophoresis in the same series of 541 fluids correctly identified 251 of the 255 pregnancies with open neural tube defect and 29 of the 31 pregnancies with false positive amAFP results. The failure of the test to diagnose 4 cases of open neural tube defect was probably attributable to the age and condition of the stored AF samples. Routine diagnostic testing of AChE isoenzymes in a further 3244 AF samples successfully identified all 170 cases of open neural tube defect and 20 cases with other fetal defects. Thirteen fluids gave false positive AChE results (0.4 per cent) compared to 59 of the series in which there were false positive amAFP results (1.8 per cent). Six of the 13 false positive AChE cases had AChE bands of low intensity which would not be regarded as diagnostic of fetal abnormality, and in five the AChE band may have been the result of significant blood contamination. False positive AChE results contributed to the decision to abort three apparently normal fetuses, but a normal AChE result undoubtedly helped to save a number of pregnancies with false positive amAFP results. Our experience suggests that repeating the amniocentesis may help in resolving the rare diagnostic difficulty of a positive AChE result with or without an elevated amAFP in the absence of ultrasound evidence of fetal abnormality, particularly where there is blood contamination of the amniotic fluid sample.     

纳米二氧化钛对金鱼脑组织的氧化应激损伤

为探讨光催化型nano-TiO₂(纳米二氧化钛)对金鱼(Carassius auratus L.)脑组织的氧化应激损伤效应,以5 nm的锐钛型钛白粉为试验材料,分别在ρ(nano-TiO₂)为0、16、32、64、128 mg/L下对金鱼进行处理,测定金鱼脑组织MDA(丙二醛)、PC(羰基化蛋白)和H₂O₂含量,SOD(超氧化物歧化酶)和GPx(谷胱甘肽过氧化物酶)活性以及Cu/Zn-SOD蛋白表达量. 结果表明:ρ(nano-TiO₂)从0 mg/L升至128 mg/L,金鱼脑组织Cu/Zn-SOD活性从1.70×10-3 kat/kg(以鲜质量计,下同)降至0.35×10-3 kat/kg,再回升至0.46×10-3 kat/kg; Mn-SOD活性呈降—升—降趋势,MDA含量从0.85 nmol/g升至2.83 nmol/g,之后降至2.54 nmol/g;PC含量一直上升;H₂O₂含量从4.32 μmol/g升至9.28 μmol/g,之后降至8.38 μmol/g. 随着ρ(nano-TiO₂)的加大,Cu/Zn-SOD蛋白表达量逐渐减少. 研究表明,nano-TiO₂对金鱼脑组织具有很强的氧化损伤作用,MDA与PC含量升高与Cu/Zn-SOD和GPx活性下降相关,H₂O₂含量升高与GPx活性下降等因素有关.      

敌百虫对鲤鱼组织乙酰胆碱酯酶和腺三磷酶活力的影响

在系列浓度的敌百虫农药暴露中,鲤鱼脑乙酰胆碱酯酶(Acetyl-cholinesterase,AChE)和鳃?肾腺三磷酶(Adenosinetriphosphatase,ATPase)的敏感性依次为脑AChE>肾ATPase>鳃ATPase?引起死亡的脑AChE抑制率在急性暴露中可大于95%,而鳃?肾ATPase抑制率达50%~60%时,鱼通常难以存活?实验结果表明,AChE抑制并非是有机磷农药中毒中引起死亡的唯一原因,非胆碱能毒性作用也具有重要的作用?