New cytochrome P450 1B1, 1C1, 2Aa, 2Y3, and 2K genes from Chinese rare minnow (Gobiocypris rarus): Molecular characterization,basal expression and response of rare minnow CYP1s and CYP2s mRNA exposed to the AHR agonist benzo[a]pyrene

Cytochrome P450 (CYP450) genes play an important role in catalyzing oxidative metabolism of toxicants. Recently, CYP1 subfamily were discovered and reported in fish, however, little is known regarding the CYP2 isoforms in fish. In the present study, the cDNA fragments of CYP 1B1 and 1C1 and CYP2Aa, 2Y3, and 2K of rare minnow were cloned and exhibited a high amino acid sequence identity compared with their zebrafish orthologs. Basal expression showed CYP1C1 and CYP 2Aa expression were observed in all eight tissues analyzed (liver, gill, intestine, kidney, spleen, brain, skin, and muscle). CYP 1A, and 1B1 expression was found in all tissues except for muscle and skin. However, CYP 2Y3 was expressed in liver, spleen, intestine and muscle whereas CYP 2 K in liver, kidney and intestine. 4 and 100 μg L⁻¹ Benzo[a]pyrene (BaP) induced patterns showed that CYP 1A, 1B1 and 1C1 expression in liver, gill, and intestine was strongly up-regulated (p < 0.05). Furthermore, CYP 2Y3 was strongly induced in liver from BaP treatments (p < 0.05). The high induction on mRNA level of CYP1s and CYP 2Y3 by BaP could be associated with catalyzing detoxification and indicated that CYP2s may also be potential biomarker to screen AHR agonist. The high responsiveness of CYP1 and 2 genes suggested Chinese rare minnow is feasible to screen and assess pollution with AHR agonist.     

Exposure of the Roots of <Emphasis Type="Italic">Populus nigra</Emphasis> L. cv. Loenen to PAHs and its Effect on growth and water balance

Poplar cuttings were cultivated for 4 weeks in a substrate, which consisted of a combination of sand and nutrient solution. The plants were treated for 24 days with BaP, Chr, Ant, Phen, P and Flt, single or in combination. The concentration of the PAHs ranged from 0.1-200 mg/kg substrate. The results of the pollution experiments can be summarized as follows: 1. The most significant deviations between the test groups and the control can be observed for transpiration, nutrient solution uptake, and root mass. 2. Although transpiration and nutrient solution uptake are significantly lower for all the treated groups than for the control group, the water content of the leaves was not affected by PAHs. 3. The biomass of the shoots and the growth in shoot length do not react as strongly to exposure to PAHs as transpiration, nutrient solution uptake and the volume of the roots. 4. The differences in leaf weight and leaf surface area are significantly less pronounced compared to the control groups. Growth inhibition is most evident with Flt. Growth and absorption of the nutrient solution dropped with just Flt 0.1 mg/kg substrate. When the substrate concentration was increased, growth and nutrient solution uptake dropped considerably and at a concentration of Flt 200, 5 of the 11 test plants died before the end of the period of exposure. Nutrient solution uptake and shoot development of the test plants decreased in the following order: BaP H approximately = Chr > Ant > Phen > Pyr > Flt.     

一株假单胞菌产脱氢酶的产酶条件优化

为提高假单胞菌（Pseudomonas sp.）降解蒽、芘过程中产生的脱氢酶量,以脱氢酶量为指标,采用Box-Behnken试验设计方法和响应面分析法（RSM）对产酶条件（如温度、盐度以及质量梯度）进行了筛选与优化.结果表明:假单胞菌降解蒽和芘模型的P均小于0.05,表明该模型差异性显著,回归效果良好.另外,假单胞菌降解蒽时的最适产酶条件为盐度3.89%、质量梯度5%、温度35.73℃,测得脱氢酶量为（140.353±6.430）μg,与预期值（141.466 μg）接近,优化结果可靠.各因素对脱氢酶量均有显著性影响,按影响程度从大到小依次为温度>质量梯度>盐度.假单胞菌降解芘时的最适产酶条件为盐度0.73%、质量梯度7%、温度34.78℃,测得脱氢酶量为（84.032±0.063）μg,与预期值（86.304 μg）接近,优化结果可靠.各因素对脱氢酶量均有显著性影响,按影响程度从大到小依次为质量梯度>盐度>温度.可见,温度是影响假单胞菌降解蒽物质的主要因素,而质量梯度则是影响芘的主要因素.研究显示,模型对假单胞菌产脱氢酶量的预测可靠,具有良好的工业应用前景.      

BaP和Cd单一复合对BaP蚯蚓亚细胞分配的影响

选用钙离子通道阻断剂氯化镧（LaCl₃）和巯基蛋白阻断剂N-乙马来酰亚胺（NEM）对赤子爱胜蚓进行预暴露,然后构建BaP单一或Cd-BaP复合污染,研究不同阻断剂对BaP在蚯蚓不同亚细胞组分（Fraction C：细胞溶质组分;Fraction D：固体颗粒组分;Fraction E：细胞碎片组分）中分配积累特征的影响.结果表明,无论单一或复合污染,BaP主要分布于细胞碎片组分中（占55.42%~69.96%）,其次为固体颗粒组分（占27.91%~32.90%）,在细胞溶质组分中的浓度最低（占2.13%~11.67%）.单一BaP污染下,两种阻断剂对BaP的作用相近,即LaCl₃和NEM的加入均能不同程度地促进3个亚细胞组分中BaP的积累.而在Cd和BaP复合污染下,两种阻断剂对BaP的分布积累的影响略有不同,LaCl₃能够促进复合污染下BaP在3个亚细胞组分中的积累,而NEM促进了复合污染下BaP在固体颗粒组分和细胞碎片组分中的积累,但抑制了BaP在细胞溶质组分中的积累,浓度从原来的0.99mg/kg降低至0.59mg/kg.因此,钙离子通道和巯基蛋白可能参与BaP在蚯蚓亚细胞的分配积累,相比单一污染,Cd的复合污会进一步改变BaP的积累分配特征.     

A study of 2,4,6-trinitrotoluene inhibition of benzo[a]pyrene uptake and activation in a microbial mutagenicity assay

A number of in vitro and in vivo studies have determined that binary and complex mixtures may interact to produce a toxicity that could not be predicted based on the individual chemicals. The present study was conducted with a binary mixture of model compounds to investigate possible interactions affecting their mutagenicity. The compounds included Benzo[a]pyrene (BAP), a polycyclic aromatic hydrocarbon that is an indirect-acting mutagen of great environmental concern, and 2,4,6-Trinitrotoluene (TNT), a nitro-aromatic compound that is a direct-acting mutagen frequently found as a soil contaminant at munitions sites. This study indicated that a binary mixture of BAP and TNT failed to induce the positive mutagenic response in Salmonella typhimurium strain TA98 characteristic of either compound alone. Spectrofluorometric analysis of BAP, and kinetic analyses of ³HBAP uptake in the presence or absence of TNT using TA98 cells that were treated or untreated with activated rat liver microsomes were performed. In cells preloaded with BAP, cellular BAP fluorescence was rapidly suppressed in the presence of TNT. Mass spectroscopy of BAP and TNT mixtures revealed a number of products, believed to be the result of complexation and nitration, that may account for the antagonistic action of TNT on BAP-induced mutagenicity in TA98 cells. Further, kinetic studies indicated that TNT inhibited the incorporation of BAP into cells.     

Effect of expandable clays and cometabolism on PAH biodegradability

Pyrene and phenanthrene degradation was examined in both single and binary slurry systems for three different natural soils. It was found that the amount of total expandable clays (smectite and vermiculite) was in a good agreement with the achieved rate and extent of biodegradation. For instance, the intrinsic phenanthrene biodegradation rate was 626 microg/L/day for the soil with the largest expandable clay and 3203 microg/L/day for the soil with the least. Similarly, the smallest total pyrene biodegradation (65%) was found for the soil rich in expandable clays, compared to an 82% pyrene reduction in the soil that had the lowest amount. Mass transfer limitation after compound sorption to the clays was more pronounced for the more hydrophobic pyrene. In the presence of phenanthrene, total pyrene biodegradation increased by 2 to 7% due to cometabolism, while the total phenanthrene biodegradation was only enhanced by 0.5 to 5% in the binary system. This research demonstrated that expandable clays might govern the substrate availability to microorganisms and microbial accessibility to substrates. Therefore, the contribution of organic matter and expandable clays to sorption, desorption and biodegradation should be taken equally into account in order to better understand complex bioremediation issues.     

二甲基苯蒽和苯并(a)芘对小鼠肝脏金属硫蛋白及氧化损伤的诱导作用

观察两种致癌性多环芳烃化合物二甲基苯蒽（DMBA）和苯并（a)芘（BaP）对小鼠肝脏脂质过氧化产物和金属硫蛋白含量的影响。选用雄性C57BL／OLA129小鼠，观察DMBA和BaP各50mg/kg一次腹腔注射染毒后24、48、72和144h后的动物肝脏重量、金属硫蛋白（MT）含量和脂质过氧化程度的变化。MT测定采用镉－血红蛋白亲和分析法；脂质过氧化指标为硫代巴比妥酸测定丙二醛法。结果表明，DMBA和BaP均能引起小鼠肝脏重量增加、MT含量增加和MDA含量增加。随着DMBA和BaP染毒后的时间延长小鼠肝脏的重量不断增加，呈现时间效应关系。小鼠肝脏MT的含量在染毒后24和48h后明显增加1倍。肝脏MDA的含量增加在染毒后24h后达到高峰。本研究观察到DMBA和Bap可以MT增加。MT的增加与MDA的增加呈现一定平行的关。提示MT诱导可能与多环芳烃化合物所致的氧化损伤有关。     

载带苯并[a]芘的纳米碳／二氧化硅颗粒引起大鼠氧化应激作用的研究

通过人工制备载带B[a]P的纳米碳(C)和纳米二氧化硅(SiO2)颗粒,采用气管滴注染毒方式,以7.5mg·kg-1(以体重计)的染毒剂量急性染毒大鼠,观察染毒24小时后载带B[a]P的纳米C/SiO2颗粒对机体产生氧化应激损伤的联合毒性效应.结果表明,在急性染毒后大鼠外周血中反映机体脂质过氧化损伤程度指标的丙二醛(MDA)含量表现为染毒组较对照组显著增加(p<0.05),表明纳米颗粒诱发机体发生了氧化应激反应.在急性染毒后各组大鼠肺泡灌洗液中谷胱苷肽过氧化物酶(GSH-PX)和超氧化物岐化酶(T-SOD)活力与对照组相比显著增加(p<0.05)(载带B[a]P的纳米SiO2组除外);载带B[a]P的纳米SiO2组肺泡灌洗液中GSH-PX活力与对照组相比无显著差异,而与单纯纳米SiO2组和B[a]P组比较显著降低,推测与抗氧化酶的一过性增高有关;载带B[a]P的纳米C组肺泡灌洗液T-SOD活力与其单纯纳米C组和单纯B[a]P组比较显著增加(p<0.05),由此表明载带B[a]P的复合纳米C/SiO2颗粒在致机体氧化损伤效应方面二者存在一定的协同作用.     

苯并(a)芘对大弹涂鱼肝脏和卵巢还原型谷胱甘肽 含量影响的比较研究

分析了大弹涂肌肉、腮、脏脏、内脏（不含肝脏）以及卵巢等不同器官的还原型谷胱甘肽含量，并以不这原型谷胱甘肽含量较高的肝脏和 卵巢为研究对象，在实验生态条件下，将大弹涂鱼暴露于0，0.05，0.2和0.5mg/L等不同浓度的苯并（a）芘3d，比较这两个器官中还原型谷胱甘肽含量的变化。结果显示，苯并（a）芘肋迫下，大弹 涂鱼肝脏和卵巢还原型谷胱甘肽含量均显著升高（P≤0.05)；表明肝脏和卵巢都是代谢苯并（a）芘的主要器官，还原型谷胱甘肽含量的显著升高表明机体对苯并（a）芘胁迫的适应性反应。     

苯并(a)芘和苯并(k)荧蒽混合物对栉孔扇贝毒理学指标的影响

检测了暴露于不同浓度多环芳烃中栉孔扇贝(Chlamys farreri)消化盲囊和鳃丝的几项毒理学指标变化.苯并(a)芘和苯并(k)荧蒽混合物的浓度梯度设置为0.5、1.0、10.0、50.0μg·L-1.结果表明,低浓度PAHs对消化盲囊EROD活力无显著影响,对GST有一定的诱导作用,而高浓度PAHs对消化盲囊EROD有明显的诱导作用,对GST先诱导后抑制;在PAHs作用下消化盲囊和鳃丝的3种抗氧化酶(SOD、CAT和GPx)活力呈现一定的峰值变化,且在高浓度PAHs下均被显著抑制,同时鳃丝的酶活力较消化盲囊抑制显著;消化盲囊和鳃丝的LPO水平在PAHs处理下随时间不断上升,并表现出明显的剂量和时间效应.暴露于PAHs中栉孔扇贝的EROD、GST和抗氧化酶活力的变化反映了机体的解毒代谢过程和能力,而LPO水平则直接反映了机体的氧化损伤程度,而且各毒理学指标在解毒过程中相互关联,具有很强的规律性.