采用生物发光检测法进行Ames试验的研究

将自行构建的含有青海弧菌荧光酶基因的质粒pACYCL184引入Ames试验的4个菌株TA97、TA98、TA100、TA102中,构建成能够生物发光的工程菌,分别命名为TAL97、TAL98、TAL100、TAL102.实验表明,该4株工程菌保持原出发菌株所具有的与Ames试验相关的性状,对各类致突变剂有良好的反应,且可用平皿计数法进行致突变试验.其发光值与致突变剂浓度存在良好的量效关系,故可用发光检测取代原Ames试验的平皿计数,使检测更简单、方便、快速.图3表3参11     

己酸二乙氨基乙醇酯的致突变性研究

采用鼠伤寒沙门氏菌回复突变试验(Ames试验),用TA97,TA98,TA100和TA102菌株,加与不加S9,剂量分别设为每皿1000μg,500μg和250μg;小鼠骨髓多染红细胞微核试验,剂量设为雄性、雌性小鼠给药剂量均为250,500和1000mg*kg-1;小鼠睾丸精母细胞染色体畸变试验,剂量设为625mg*kg-1,1250mg*kg-1和2500mg*kg-1.结果表明,Ames试验中,各测试浓度的诱发回变菌落数均未超过自发回变菌落数的2倍;小鼠骨髓多染红细胞微核试验和小鼠睾丸精母细胞染色体畸变试验,各剂量组和溶剂对照组的微核率进行统计学处理,未见有显着性差异(P＞0.05),表明各项试验结果均为阴性.因此,己酸二乙氨基乙醇酯无致突变作用.     

官厅水库沉积物极性提取液化学成分分析及基因毒性效应

提取北京市官厅水库表层沉积物极性组分,定量检测了12种硝基芳香烃和5种芳香胺的含量水平;并通过鼠伤寒沙门氏菌回复突变试验(Ames试验)研究沉积物极性提取液的致突变毒性,对官厅水库沉积物的潜在基因毒性效应进行评价.结果表明,官厅水库沉积物中12种硝基芳香烃的总含量为8.4—272 ng·g~(-1),平均值为121 ng·g~(-1);5种芳香胺总含量为5.1—42.8 ng·g~(-1),平均值为4.8 ng·g~(-1).各组沉积物极性提取液对鼠伤寒沙门氏菌(Salmonella typhimurium)TA98及TA100的致突变毒性试验结果均呈阴性,但多个点位沉积物极性提取液导致的细菌突变数随提取液浓度的上升而增加,表明可能存在致突变物质.Pearson相关分析结果表明,2-硝基甲苯、3-硝基甲苯、1,3,5-三硝基苯和联苯胺的含量与TA98的突变效应具有显著相关性,推断其可能是主要的致突变物质,且其可能引起的致突变类型为移码型基因突变.     

Ames测试不确定性分析

Ames测试具有实验周期短、简便的优点,是目前普遍采用的检测化合物致突变性的初筛方法,但Ames测试仍有一定的不确定性.本文主要依据本实验室的研究结果分析了Ames测试中测试菌株、统计回复子数目的时间及2倍判定标准给测试结果带来的不确定性,对影响Ames测试结论存在假阳性及假阴性的因素做了进一步分析与讨论.图6表1参19     

氧化石墨烯遗传毒性的实验研究

研究氧化石墨烯(GO)的遗传毒性,考察其致突变作用,为GO在生物领域的安全应用提供依据。采用Ames试验、体外CHL细胞染色体畸变试验和小鼠体内染色体畸变试验,分别在细菌水平、细胞水平及整体动物水平研究GO的遗传毒性。GO各剂量组的Ames试验结果为阴性。CHL试验中,CHL细胞染色体畸变率随GO浓度的增加而升高,其中1.000 mg·mL~(-1)剂量组(+S_9)和0.500 mg·mL~(-1)剂量组(-S_9)畸变率显著升高(P 0.05)。小鼠骨髓细胞染色体畸变试验中,骨髓细胞染色体畸变率同样随GO浓度的增加而升高,1.000和0.500 mg·kg~(-1)剂量组的畸变率显著提高(P 0.05)。虽然Ames试验结果没有反映出GO的遗传毒性,但在体外及体内染色体畸变试验中,GO均表现出对哺乳动物细胞染色体有潜在的遗传毒性。     

水源水雄激素受体干扰效应及在水处理工艺中的变化

我国很多饮用水源水已检测出雌激素受体干扰效应,但是其他效应如雄激素受体干扰效应却罕见报道。为了弥补这方面资料的缺乏,应用雄激素受体双杂交酵母结合S9代谢方法评价了我国6大流域23个水源地水源水以及3个供水厂不同工艺出水样品的雄激素受体干扰效应。结果显示,所有水源水样品浓缩液均没有检测出雄激素受体直接或间接诱导效应,但都观察到了抑制效应,抑制效应以抑制剂氟他胺当量浓度进行表征:直接抑制效应在0.67~3.68μg·L-1之间,间接抑制效应在0.52~3.02μg·L-1之间;长江三角洲和淮河流域水源地的水源水雄激素抑制效应明显高于其他流域;经S9代谢后,有16个水源地的氟他胺当量比代谢前降低。3个饮用水厂处理工艺能够使源水的雄激素受体抑制效应降低19.1%~70.5%。研究表明,我国水源水中普遍含有雄激素受体干扰效应物质,目前水厂常规处理工艺对雄激素受体干扰效应有一定的去除效果,但不能完全去除。雄激素受体双杂交酵母测试能够快速、综合地对水体中的雄激素受体干扰效应进行评价,是饮用水安全评价的重要补充手段。     

废弃农药厂污染场地土壤浸出液的急性毒性和遗传毒性筛查

污染土地再开发是城市土地可持续利用的关键。为进行废弃农药厂的生态风险评价,采集常州市某废弃农药厂污染场地6个不同区域的土壤样品(编号为S1、S2、S3、S4、S5和S6),选取发光细菌、大型溞和蚕豆根尖细胞为试验生物,运用成组生物毒性试验对6个土壤样品的浸出液进行生物毒性检测。结果表明:6个土壤样品的浸出液都具有急性毒性和遗传毒性;发光菌和大型溞急性毒性试验表明,S3和S5的土壤浸出液毒性最高,S6的土壤浸出液毒性最低;蚕豆根尖微核遗传毒性试验表明S3和S5的土壤浸出液毒性最高,S4和S6的土壤浸出液毒性最低。蚕豆根尖微核遗传毒性试验结果与发光菌和大型溞急性毒性效应基本相同,表明成组生物毒性测试法可以用于污染土壤浸出液的毒性评价。污染物质和毒性检测结果的相关性分析结果表明,土壤中所含污染物种类和浓度已经对土壤毒性效应产生影响,结合化学分析与生物毒性检测可为污染场地进行综合评价与风险评估提供依据。     

2-硝基芴的致突变性受共存多环芳烃的影响

本文以Ames试验的TA_(98)菌株研究了2-硝基芴(2-Nitroflurene)在其它多环芳烃共存时,其致突变性的变化.结果显示:它与其它硝基多环芳烃共存时.直接致突变性增强;而与其它无直接致突变性的多环芳烃共存时,其直接致突变性减弱,且多环芳烃的环数越多、减弱作用越强,这可能是因为硝基多环芳烃具有亲电子性,DNA的碱基有亲核性,易于结合,造成DNA的损伤;而多环芳烃一般具有供电子性,阻碍了硝基多环芳烃与DNA碱基的结合.     

部分离子液体及其混合物对发光菌的毒性作用

离子液体(ILs)因其环境安全和良好的非挥发性而得以广泛应用,尽管其理化性质与工程数据一直在不断扩充,但其可用的毒性及生态毒性数据很少.以青海弧菌Q67为指示生物,应用微板发光毒性测试方法,测定了C6H11BF4N(2S1)、C8H15ClN2(S2)、C8H15BF4N2(S3)、C9H14BF4N(S4)、C9H17BF4N2(S5)、C9H17BrN2(S6)、C11H13BF4N2(S7)、C11H13ClN2(S8)、C12H23BrN2(S9)、C14H27BF4N(2S10)、C14H27ClN(2S11)和C16H31ClN(2S12)等12种ILs对发光菌的发光抑制毒性.结果表明,4种ILs(S9、S10、S11、S12)具有高抑制毒性(pEC50>4.5),而另外8种毒性相对较小(pEC50<3.5).为研究混合ILs的联合毒性,根据单个ILs的剂量-效应关系,构建了两组混合物,即由S9、S10、S11和S12构成的高毒性组(简称H组)以及由S2、S3、S4、S5、S6和S8构成的低毒性组(简称L组)混合物.应用非线性模拟技术与剂量加和(DA)及独立作用(IA)模型对混合物毒性数据进行拟合与预测分析,结果表明,以等效应浓度比法设计的混合物,无论是对于H组的4个混合物还是L组的4个混合物,其联合毒性大小均可用DA模型准确预测.对于均匀试验设计浓度比法设计的混合物,H组的6个混合物的毒性可用DA模型有效预测,而L组的6个混合物由于剂量-效应曲线在低浓度区翘起,其混合物毒性用DA或用IA模型预测均有一定误差.     

水库型饮用水水源保护区划分方法与实证研究

饮用水水源保护区划分研究对水源保护及管理具有重要意义。通过解读国内外水库型饮用水水源保护区划分的标准和方法,分析常用的5种保护区划分方法的适用条件和技术要求;根据不同水库水文条件、水源水质、地形环境及周边污染源情况,确定48种不同的划分预设场景,总结出20种划分方法体系。以广东省67个水库型水源保护区划分方法作为实例举证,证明划分方法体系具有科学性和可操作性,为合理划分水库型饮用水水源保护区提供依据。     

Analyses of PAHs (polycyclic aromatic hydrocarbons) and mutagenicity in raw and chlorinated water

Both raw water and chlorinated drinking water samples were collected from and the Liu‐Du water treatment plant in northern Taiwan from October 1990 to April 1992. The polycyclic aromatic hydrocarbons (PAHs) and mutagenicity in these water samples were analyzed by GC/MS and Ames test. The Mutagenicity/DMSO (Dimethyl Sulfoxide) ratio in S. typhimurium TA98 and TA100 with or without S9 mixture increased, even higher than 2, following the sequence of unit process. It was observed that the mutagenicity with TA98 (S9+) was highly related to most of PAHs in the raw water; while the mutagenicity with TA98 (S9+) was only correlated with DbA and BghiPr in the treated water. It could be expected that the mutagenicity level was controlled by other predominant components after the raw water was treated, for example, the chlorination process.     

Aliphatic and aromatic halocarbons as potential mutagens in drinking water

Chlorinated phenols are either products of industrial chemical processes or the result of chlorination of drinking water. Often, the formation of chlorinated phenols is based upon naturally occurring phenol. The following chlorinated phenols have been selected for testing in the Ames‐test for their mutagenic activity: 3‐chlorophenol, 4‐chlorophenol, 2,3,6‐, 2,4,5‐, 2,4,6‐trichlorophenol, 4‐chloro‐2‐methylphenol and 4‐chloro‐3‐methylchlorophenol. The tester strains TA97, TA98, TA100 and TA104 were employed. All tested compounds produced mutagenic activity at least in one tester strain. The highest numbers of revertants were detected for 2,3,6‐ and 2,4,6‐trichlorophenol. But in contrast to the other substances, these two induced only frameshift mutations in presence of a metabolizing system. The evidence of their presence in drinking water and of their mutagenic activity makes them to a potential health hazard.     

Humic Acids from Endemic Arsenicosis Areas in Inner Mongolia and from the Blackfoot-Disease Areas in Taiwan: A Comparative Study

The physical and chemical properties of humic acids (HA) extracted from drinking waters from the endemic arsenicosis areas in Inner Mongolia and from the Blackfoot disease (BD) areas in Taiwan are studied by using AAS (atomic absorption spectrophotometry), ICP-MS (inductively coupled plasma emission-mass spectrometry), IR (Infrared spectroscopy), FR (fluorescence spectrometry) and TLS (total luminescence spectroscopy) in order to shed light on the pathogenesis of BD with the concern as to whether the disease may eventually occur in arsenicosis-affected areas in the Mainland of China. Ames test and lipid peroxidation experiments were also conducted on these HA samples. It is found that water samples from the two regions are high in arsenic (As) with strong fluorescence and apparent positive correlations between As content, fluorescence, pH and total dissolved solids (TDS). The water samples are similar in fluorescence spectra but differ somewhat in IR and TLS between the two regions. The difference may be a reflection of the difference in radicals and structure of the HA owing to different hydrogeological conditions, and may also be related to the difference in their biological effects, i.e., HA from Inner Mongolia have a stronger ability to cause lipid peroxidation while HA from Taiwan exhibits a more prominent effect of mutation with respect to TA98(±S9).     

Evaluation of the antimutagenic and antiradical potentials of extracts from the tubers of (Tunisian) Cyperus rotundus

The mutagenic potential of aqueous, total oligomers flavonoids (TOF), ethyl acetate and methanol extracts from tubers of Cyperus rotundus L. was assessed using Ames Salmonella tester strains TA98 and TA100, and SOS chromotest strain Escherichia coli PQ37 with and without metabolic activation (S9). None of the different extracts produced a mutagenic effect. Likewise, the antimutagenicity of the same extracts was tested using the “Ames test” and the “SOS chromotest”. Our results showed that C. rotundus extracts possess antimutagenic effects against S. typhimurium TA98 and TA100 strains towards the mutagen aflatoxin B1 (AFB1), similar to E. coli PQ37 strain against AFB1 and Nifuroxazide mutagens using the SOS chromotest assay. A free radical scavenging test was used in order to explore the antioxidant capacity of the extracts obtained from the tubers of C. rotundus. TOF, ethyl acetate and methanol extracts showed an important free radical scavenging activity towards the 1,1-diphenyl-2-picrylhydrazine (DPPH) free radical. These extracts showed an IC50 value of respectively 5, 20 and 65?µg?mL^?1. The beneficial effects of TOF, ethyl acetate and methanol extracts of C. rotundus were assessed by antioxidant and antimutagenic activities.     

Genotoxic potential of organic extracts from municipal reclaimed water in Tianjin,China

Despite increasing pressure to make more efficient use of water resources, the widespread use of reclaimed water still remains a contentious issue, primarily because of risks to human health arising from water pollution by organic compounds. Therefore, safety evaluation of reclaimed water is an urgent need. The aim of this study was to determine the genotoxic potential of reclaimed water during low and high water periods. Reverse phase C-18 solid-phase extraction (RP-C18SPE) was used for the extraction of organic compounds from water samples. The tests, namely, Ames, micronucleus and chromosome aberrations were used to determine the damage caused by water samples on genetic material. Ames tests showed that both influent and effluent reclaimed water, except influent after metabolism by S9 in high water period, induced mutation of TA98 and TA100 strains of Salmonella typhimurium in a concentration-response manner. There were no significant differences in micronucleus and chromosome aberration tests. Even after treatment, reclaimed water in Tianjin still showed mutational effects and new strategies for reduction of genotoxins need to be considered.     

Modification of mutagenic activity of benzo[a]pyrene by trace elements in the Ames assay

Benzo[a]pyrene (BaP) is one of the most ubiquitous environmental contaminants and exerts potent carcinogenicity and mutagenicity. In the presence of rat liver S9 mix, four trace elements, germanium (Ge), iron (Fe), zinc (Zn), and molybdenium (Mo), were included in the Ames Salmonella microsome/mutagenic assay to test their inhibitory effects on BaP-induced mutations. All trace elements were found to exert an inhibitory effect in tester TA98 and TA100 strains; Fe was the most potent among the elements tested, whereas Zn was weakly inhibitory to prevent mutations. The present findings demonstrate that the antimutagenic potential of trace elements, notably Fe, may be attributed to its modulatory effect on the bioactivation and detoxification processes of BaP.     

Assessment of mutagenic potency of source water treated by ozone and adsorption processes

This research utilized the Ames test to determine the mutagenicity of water treated by advanced processes, including ozonation and granular activated carbon (GAC). Raw water samples for this research included those obtained from the Pan Hsin waterworks as well as samples containing humic acids. Treated samples were collected from the pilot‐scale advanced treatment plant. The Ames test was used to measure the mutagenicity of the water after each treatment process. For the Pan Hsin raw water samples treated with ozone or GAC, it was indicated that, regardless of whether samples were preozonated or not, they all showed a mutagenic potency less than 2 once the S9 enzyme was added. This level of mutagenicity is insignificant. The prepared humic acid samples, on the other hand, demonstrated a significant reduction in mutagenicity after the pre‐ozonation process, indicating that preozonation can lower the degree of mutagenicity. Furthermore, the mutagenicity of the prepared humic acid samples gradually decreased after the advanced treatment process. However, when chlorine was added later to these samples, the mutagenicity increased again. This research shows that the use of O3/GAC processes to treat water can successfully lower mutagenicity, indicating a great potential for applications in the treatment of drinking water.     

Mutagenicity of diesel engine exhaust in the Ames / Salmonella assay using a direct exposure method

The aim of this study was to investigate the potential mutagenic activity of diesel engine exhaust in the Ames/Salmonella assay using a direct aerosol exposure system. So, TA 98 and TA 100 strains, with or without added S9 mix, were exposed to diesel emissions after varying degrees of filtration. Variants of these two strains, deficient in nitroreductase (TA 98NR and TA 100NR) or over-expressing O-Acetyl Transferase (YG 1024 and YG 1029), were also exposed to total (unfiltered) diesel exhaust to highlight the putative mutagenicity of any nitro-PAHs present in these emissions. Mutagenic activity of the diesel exhaust was demonstrated on Salmonella typhimurium, strains TA 100 and variants TA 100 NR and YG1029. The use of a particle filter did not modify the genotoxicity of the diesel emissions, indicating a major contribution of the gas phase to the mutagenicity of these diesel emissions. The prominent role of the particulate-associated nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) claimed by some authors working on diesel exhaust organic extracts was not confirmed by our results with native diesel exhaust exposure. Our results show that the gas phase is potentially more mutagenic than the particles alone.