基于枯草芽孢杆菌的CellSense生物传感器的重金属联合毒性分析

代森联检测方法的研究现状引言代森联检测方法是一种用于检测代森联（Dioxin-like PCBs）的方法，代森联是一类具有毒性的有机污染物，对人类健康和环境造成潜在威胁。

本文将全面、详细、完整且深入地探讨代森联检测方法的研究现状。

代森联的背景代森联的定义1.代森联是一类多氯联苯类化合物。

2.代森联具有类似戴奥辛的毒性作用。

代森联的来源1.工业生产过程中的废弃物。

2.燃烧过程中产生的废气和废渣。

3.环境中的自然分解产物。

代森联检测方法的分类生物学方法1.生物传感器–基于酶的生物传感器–基于细胞的生物传感器–基于抗体的生物传感器2.生物指示器–鱼类生物指示器–贝类生物指示器–植物生物指示器化学方法1.色谱法–气相色谱法–液相色谱法–超高效液相色谱法2.质谱法–气质联用法–液质联用法–高分辨质谱法物理方法1.红外光谱法2.紫外-可见光谱法3.核磁共振法代森联检测方法的研究进展生物学方法的研究进展1.基于酶的生物传感器–研究A–研究B–研究C2.基于细胞的生物传感器–研究A–研究B–研究C3.基于抗体的生物传感器–研究A–研究B–研究C化学方法的研究进展1.气相色谱法–研究A–研究B–研究C2.液相色谱法–研究A–研究B–研究C3.超高效液相色谱法–研究A–研究B–研究C物理方法的研究进展1.红外光谱法–研究A–研究B–研究C2.紫外-可见光谱法–研究A–研究B–研究C3.核磁共振法–研究A–研究B–研究C代森联检测方法的优缺点比较生物学方法的优缺点1.优点–高选择性–低检测限–环境友好2.缺点–操作复杂–易受干扰化学方法的优缺点1.优点–快速–灵敏度高–可定量分析2.缺点–耗时耗能–需要复杂的仪器设备物理方法的优缺点1.优点–非破坏性分析–无需样品前处理2.缺点–检测限较高–可选择性较差结论代森联检测方法的研究现状包括生物学方法、化学方法和物理方法。

生物学方法中的生物传感器和生物指示器在代森联检测中具有广泛应用。

电化学传感器在水质重金属检测中的应用研究一、电化学传感器的原理电化学传感器是利用电化学原理进行测量的一种传感器。

其基本原理是基于电极在溶液中的电化学反应。

电化学传感器通常由工作电极、参比电极和计时电极组成。

当被测物质与电极表面发生化学反应时，会产生电流或电压变化，通过测量这些电流或电压的变化，就可以实现对被测物质的检测。

根据测量的信号类型，电化学传感器可以分为安培计（测量电流）传感器和库仑计（测量电压）传感器。

1. 重金属离子的检测电化学传感器主要应用于重金属离子的检测，如铅、汞、镉、铬等。

这些重金属离子通常以阳离子的形式存在于水中，通过与电极表面发生化学反应，可以产生特定的电流或电压信号。

研究表明，通过合理设计电化学传感器的电极材料和表面修饰，可以实现对不同重金属离子的高灵敏度、高选择性的检测。

利用微纳米技术和生物技术，还可以提高电化学传感器的检测性能，实现对微量重金属离子的快速检测。

除了重金属离子外，一些重金属有机物也是水质中常见的污染物之一，如苯基汞、有机锡化合物等。

这些重金属有机物对生态系统和人体健康同样具有潜在的危害。

电化学传感器可以通过选择性的化学反应，实现对重金属有机物的检测。

利用催化电极和生物传感器技术，可以实现对重金属有机物的高灵敏度和高选择性的检测。

水体中往往存在多种重金属污染物，传统的检测方法往往需要多次取样和分析，耗时耗力。

电化学传感器具有快速、在线监测的优势，可以实现对多种重金属污染物的同时检测。

通过数据处理和模式识别技术，还可以实现对不同重金属污染物的准确识别和定量分析。

三、电化学传感器在水质重金属检测中的挑战与展望虽然电化学传感器在水质重金属检测中具有诸多优势，但同时也面临着一些挑战。

对电极材料的选择和表面修饰的研究，需要不断深入，以实现对不同重金属污染物的高灵敏度和高选择性的检测。

电化学传感器的样品预处理和环境干扰抑制也是需要重点关注的问题。

电化学传感器的实时监测和在线分析技术需要进一步完善，以满足水质重金属监测的实际需求。

微生物对土壤中重金属污染物的影响研究重金属污染是当今环境问题中的一个重要方面。

许多废水和废气中含有大量的重金属，它们会进入土壤并影响生物的生长和发展。

然而，微生物在土壤中具有重要的生物地球化学作用，可以对土壤中的重金属进行转化和去除，从而减轻土壤污染的程度。

本文将探讨微生物对土壤中重金属污染物的影响，并介绍其作用机制和应用前景。

一、微生物对重金属的转化作用微生物可将土壤中的重金属离子转化成可溶性有机络合物或不溶性沉淀物，从而减少其毒性和迁移性。

一些微生物具有还原、氧化、沉淀和吸附等特性，可以转化土壤中的重金属形态。

举例来说，硫酸还原菌可以将重金属离子还原成金属沉淀物，硫醇基功能化微生物可以通过产生硫醇将重金属离子络合成沉淀物。

这些微生物的作用有助于将重金属离子固定在土壤中，减少其对生物体的毒性影响。

二、微生物对重金属的去除作用微生物可通过吸附、螯合、沉淀和矿化等途径将重金属离子从土壤中去除。

一些细菌和真菌可以通过草酸、胞外多糖和胞内蛋白质等物质与重金属离子螯合，从而减少其毒性。

此外，微生物还可通过沉淀作用使重金属离子形成不溶性沉淀物，进而进行去除。

一些微生物还具有矿化功能，可以将重金属转化为无毒的无机形态，从而完全去除其对环境的污染。

三、微生物的应用前景由于微生物在土壤中处理重金属污染中具有独特的优势，因此其应用前景广泛。

一方面，微生物修复可以在原地进行，不需要对土壤进行大规模开挖和运输，因此具有较低的成本和环境风险。

另一方面，微生物修复对土壤生态环境的破坏相对较小，能够保持土壤的水、肥结构，并且不会产生二次污染。

此外，微生物修复适用于不同类型的土壤和不同程度的污染，具有较高的适应性和灵活性。

然而，微生物修复技术在实际应用中还存在一些问题和挑战。

首先，不同微生物对不同重金属的转化和去除效果存在差异，因此需要针对具体的重金属污染物选择适宜的微生物种类。

其次，微生物修复过程需要一定的时间和环境条件，无法实现即时修复。

重金属测试报告1. 引言重金属污染是当今环境问题的重要组成部分之一。

重金属对人体健康和环境造成的潜在危害已经引起了广泛关注。

为了确认某一环境样品中是否存在重金属，本文对样品进行重金属测试，并提供了详细的测试报告。

2. 测试目的本次测试的主要目的是确定样品中是否含有以下常见的重金属元素：1.铅（Pb）2.汞（Hg）3.镉（Cd）4.铬（Cr）5.铜（Cu）6.锌（Zn）重金属的存在可能会对生态系统和人体健康产生潜在危害。

因此，通过测试，我们可以了解样品中重金属元素的含量，并评估其对环境和人体的影响。

3. 测试方法本测试采取了以下步骤：1.样品采集：从目标区域采集样品，并尽量表示该区域的典型特征。

2.样品预处理：将样品进行必要的处理，如研磨，过滤等。

3.仪器分析：使用X射线荧光光谱仪（XRF）或火焰原子吸收光谱仪（FAAS）等仪器对样品进行测试。

4.数据分析：根据测试结果，计算样品中各重金属元素的含量，并与相关标准进行比较。

4. 测试结果根据我们的测试结果显示，样品中的重金属含量如下：•铅（Pb）： 10.2ppm•汞（Hg）： 0.05ppm•镉（Cd）： 2.3ppm•铬（Cr）： 1.8ppm•铜（Cu）： 50.7ppm•锌（Zn）： 80.1ppm5. 结果分析根据相关标准，我们可以对测试结果进行分析和评估。

以下是我们对每种重金属含量的评估：1.铅（Pb）：样品中的铅含量为10.2ppm。

根据环保局标准，铅的接受水平为5ppm，该样品超过了标准限值。

2.汞（Hg）：样品中的汞含量为0.05ppm。

根据环保局标准，汞的接受水平为0.03ppm，该样品超过了标准限值。

3.镉（Cd）：样品中的镉含量为2.3ppm。

根据环保局标准，镉的接受水平为0.5ppm，该样品超过了标准限值。

4.铬（Cr）：样品中的铬含量为1.8ppm。

根据环保局标准，铬的接受水平为1ppm，该样品稍微超过了标准限值。

5.铜（Cu）：样品中的铜含量为50.7ppm。

Toxicology237(2007)126–133Toxic effects of di(2-ethylhexyl)phthalate on mortality,growth, reproduction and stress-related gene expression inthe soil nematode Caenorhabditis elegansJi-Yeon Roh,In-Ho Jung,Jai-Young Lee,Jinhee Choi∗Faculty of Environmental Engineering,College of Urban Science,University of Seoul,90Jeonnong-dong,Dongdaemun-gu,Seoul130-743,Republic of KoreaReceived16March2007;received in revised form19April2007;accepted2May2007Available online18May2007AbstractIn this study,di(2-ethylhexyl)phthalate(DEHP)toxicities to Caenorhabditis elegans were investigated using multiple toxic endpoints,such as mortality,growth,reproduction and stress-related gene expression,focusing on the identification of chemical-induced gene expression as a sensitive biomarker for DEHP monitoring.The possible use of C.elegans as a sentinel organism in the monitoring of soil ecosystem health was also tested by conducting the experiment on the exposure of nematode tofield soil. Twenty-four-hour median lethal concentration(LC50)data suggest that DEHP has a relatively high potential of acute toxicity to C. elegans.Decreases in body length and egg number per worm observed after24h of DEHP exposure may induce long-term alteration in the growth and reproduction of the nematode population.Based on the result from the C.elegans genome array and indicated in the literatures,stress proteins,metallothionein,vitellogenin,xenobiotic metabolism enzymes,apoptosis-related proteins,and antioxidant enzyme genes were selected as stress-related genes and their expression in C.elegans by DEHP exposure was analyzed semi-quantitatively.Expression of heat shock protein(hsp)-16.1and hsp-16.2genes was decreased by DEHP exposure.Expression of cytochrome P450(cyp)35a2and glutathione-S-transferease(gst)-4,phase I and phase II of xenobiotic metabolism enzymes, was increased by DEHP exposure in a concentration-dependent manner.An increase in stress-related gene expressions occurred concomitantly with the deterioration on the physiological level,which suggests an increase in expression of those genes may not be considered as a homeostatic response but as a toxicity that might have physiological consequences.The experiment with the soil from the landfill site suggests that the potential of the C.elegans biomarker identified in laboratory conditions should be calibrated and validated for its use in situ.©2007Elsevier Ireland Ltd.All rights reserved.Keywords:Caenorhabditis elegans;Di(2-ethylhexyl)phthalate;Stress-related gene expression;Ecotoxicity monitoring1.IntroductionDi(2-ethylhexyl)phthalate(DEHP)is widely used in flexible polyvinyl chloride(PVC)as a plasticizer to∗Corresponding author.Tel.:+82222105622;fax:+82222442245.E-mail address:jinhchoi@uos.ac.kr(J.Choi).soften plastic materials.Owing to its utility and cost effectiveness,DEHP has been used in a broad range of applications,such as wires and cables,floor tiles,garden hoses,containers,footwear and clothing(De Jonge et al., 2002;Chao and Cheng,2007).Due to the widespread use of DEHP,it is ubiquitous in the environment and many environmental species are thus exposed to vari-ous levels of DEHP in their natural habitat.At present,0300-483X/$–see front matter©2007Elsevier Ireland Ltd.All rights reserved. doi:10.1016/j.tox.2007.05.008J.-Y.Roh et al./Toxicology237(2007)126–133127hazard or risk assessments of DEHP conducted by inter-national authorities are available(WHO,1992;EPA, 1999;EU,2001;ATDSR,2002),some of which include an assessment of the ecological risk of DEHP to aquatic life.However,few studies have been performed on the ecotoxicity of soil organism.Caenorhabditis elegans,a free-living nematode that lives mainly in the liquid phase of soils,is thefirst multicellular organism to have its genome completely sequenced.The genome showed an unexpectedly high level of conservation with the vertebrate genome,which makes C.elegans an ideal system for biological studies, such as those in genetics,molecular biology and devel-opment biology(Brenner,1974;Bettinger et al.,2004; Leacock and Reinke,2006;Schafer,2006;Schroeder, 2006).C.elegans is also a good animal model for the study of ecotoxicology.Due to its abundance in soil ecosystems,its convenient handling in the labora-tory,and its sensitivity to different kinds of stresses, C.elegans is frequently used in ecotoxicological stud-ies utilizing various exposure media,including soil and water(Peredney and Williams,2000;Willams et al., 2000;Boyd and Williams,2003a).The range of C.ele-gans studies in ecotoxicology focuses on organism-level endpoints,such as mortality,behavior,growth or repro-duction.However,using these classical test endpoints, it is difficult tofind significant effects.Therefore,more specific and sensitive systems than classical ecotoxico-logical tests are needed.In this study,to identify a suitable tool to develop a screening system for ecotoxicity monitoring,DEHP toxicities to C.elegans were investigated using multiple toxic endpoints,such as mortality,growth,reproduc-tion and stress-related gene expression,focusing on the identification of chemical-induced gene expression as a sensitive biomarker for DEHP toxicity.C.elegans whole genome microarray was conducted for screen-ing the differentially expressed gene list by DEHP exposure.Tested stress-related genes were selected according to the results of microarray data and liter-ature.Alteration on stress-related gene expression by DEHP exposure was examined in a semi-quantitative manner.The response of greenfluorescent protein (gfp)transgenic nematode,incorporated full-length heat shock protein(hsp)-16.2and hsp-16.48genes to DEHP exposure was also examined to test a possibility of transgenic worm as a biosensor of environmen-tal monitoring.Additionally,in situ application of C. elegans toxicity indicator was investigated on the nema-todes exposed to soil from landfill sites,using the same endpoints applied for the exposure of laboratory condition.2.Materials and methodsanismsThe wild-type C.elegans Bristol strain N2was used in this study.C.elegans were maintained on nematode growth medium(NGM)plates seeded with Escherichia coli strain OP50,at20◦C,using the standard method previously described by Brenner(1974).Young adults(3days old)from an age-synchronized culture were used in all the experiments.Worms were incubated at20◦C for24h without a food source,and were then subjected to the analysis.2.2.Sample preparationFour types of endpoints(mortality,growth,reproduction, and stress-related gene expression)were assessed for exposure to DEHP.Pure analytical-grade DEHP(Sigma–Aldrich Chem-ical,St.Louis,MO,USA)was used in the experiment and it was dissolved in dimethyl sulfoxide(DMSO,Sigma–Aldrich Chemical).Nematodes were exposed to DEHP prepared in a K-medium(0.032M KCl,0.051M NaCl)(Williams and Dusenbery,1990).Three replicates for each concentration and a control were conducted for all the test types.The DEHP concentrations in a K-medium were nominal values.Soil toxicity testing with C.elegans was performed as described in the American Society for Testing and Materials (ASTM,2001).Briefly,for each sample,2.33g of soil as loaded in to a35mm×10mm petridish.The moisture was adjusted to35%(dry weight)using K-medium,worms were added and were incubated at20◦C for24h without a food source.The worms were then recovered with centrifugation/flotation using Ludox(Sigma–Aldrich).Three replicates were conducted for all the test types.2.3.Lethal toxicity testsEach test consisted of four concentrations and a control,in which10±1of young C.elegans adults were transferred onto 24-well tissue culture plates containing1ml of the test solution for each of thefive wells.The worms were exposed for24h at20◦C.After the24h,the numbers of live and dead worms were determined through visual inspection and by probing the worms with a platinum wire under a dissecting microscope.2.4.Measurement of growth and reproductionFollowing the24h incubation with exposure to sub-lethal concentrations of DEHP,growth and reproduction were assessed.Growth was assessed by measuring the length of the worms that had been killed by the heat through microscopy, with a scaled lens in each sample.The average length of the unexposed control worm was in the range of1.0–1.2mm. Reproduction was assessed by counting the eggs of each worm through the microscopic inspection of the transparent C.ele-gans body in each sample.Although this procedure differs from128J.-Y.Roh et al./Toxicology237(2007)126–133more commonly used reproduction tests of offspring counting from an age-synchronized single worm,this simple detection method seems appropriate for the rapid screening of the repro-duction effect(Roh et al.,2006).The average number of eggs per worm in the unexposed controls was in the range of10–25. One hundred worms were examined per treatment for growth and reproduction experiments.2.5.RNA extractionFollowing the24h incubation with exposure to sub-lethal concentrations of DEHP,nematodes were harvested for the preparation of RNA.Total RNA was prepared by phenol–chloroform extraction,according to the manufacturer’s standard protocol.RNA concentrations were determined by the absorbance at260nm.The quality of total RNA was estimated based on the ratio of the optical densities from RNA samples measured at260and280nm.2.6.Microarray analysisFive micrograms of the total RNA extracted from nema-todes exposed to DEHP and the control was used for reverse and in vitro transcription followed by application to a GeneChip®C.elegans Genome Array(Affymetrix,Santa Clara,CA, USA),which contains22,500probe sets against22,150unique C.elegans transcripts.The arrays were scanned with the GeneChip scanner3000(Affymetrix),controlled by GeneChip Operating Software(GCOS,Affymetrix).2.7.Semi-quantitative reverse transcription-polymerase chain reactionThe two-step reverse transcription-polymerase chain reac-tion(RT-PCR)method was used with RT Premix(Bioneer Co., Seoul,Korea)and PCR Premix kits(Bioneer Co.),using a PTC-100thermal cycler(MJ Research,Lincoln,MA,USA). The primers were designed on the basis of the sequences retrieved from the C.elegans database(). Actin mRNA was used for expression-level normalization of the studied genes.The PCR products were separated through electrophoresis on1.5%agarose gel(Promega,Madison,WI, USA)and were visualized with ethidium bromide(Bioneer Co.).All the tests were replicated at least three times,and the relative densities of each band were determined with the use of a Kodak EDAS290image analyzer(Kodak,Rochester,NY, USA),with a TFX-20.M UV transilluminator(Vilber Lourmat, Marne la Vallee,France).2.8.Detection of greenfluorescence protein transgenic C. elegansThe transgenic strains(hsp-16.2::gfp and hsp-16.48::gfp)of C.elegans were developed as previously described by Hong et al.(2004).Transgenic C.elegans were incubated for24h with 2mg/l of DEHP,as well as,with soil from landfill sites,and the fluorescence signal was examined from20independent trans-genic worms per treatment.Fluorescence was observed using a Leica DM IRB microscope(Leica,Wetzlar,Germany),and the image was taken using a Leica DC300FX camera(Leica). Levamisole(Sigma–Aldrich Chemical)treatment(2mM)was used to take pictures of the live worms.2.9.Analysis of DEHP from soil of Korean landfill sitesSoil was collected from Korean landfill sites,namely Jeonju (J),Gwangju(G)and Mokpo(M).Reference soil(R)was sampled from the clean area.Soil samples were dried in the air at room temperature.DEHP was extracted using the Pres-surized Solvent Extraction(SPE)method and was analyzed using6890Gas Chromatography–5973N Mass Spectroscopy systems(Agilent,Santa Clara,CA,USA).2.10.Data analysisMedian lethal concentration(LC50)were derived through Probits analysis.The statistical differences between the con-trol and treated worms were determined with the aid of the parametric t-test.3.ResultsAcute toxicity of DEHP on C.elegans was investigated using mortality as endpoint(Table1). Twenty-four-hour LC50of DEHP in C.elegans was estimated as22.55mg/l.Based on the results of the acute toxicity test,three concentrations corresponding to 1/1000,1/100,and1/10of the24h LC50were selected for laboratory sublethal exposure conditions,that is, 0.02,0.2and2mg/l.The changes in the worms’body lengths and in the number of eggs per worm were investigated as a growth and a reproduction indicator,respectively(Fig.1).The worms,which had been exposed to0.02and0.2mg/l of DEHP,showed a decrease in their body length,as well as,in the number of eggs per worm.Suitability of the DNA microarray technique for the ecotoxicological approach was investigated in C. elegans exposed to0.2mg/l of DEHP.DEHP-induced genes expression was screened using C.elegans Genome Arrays.Major up-and down-regulated known genes by Table1Estimation of24h LC10,LC50and LC90of DEHP in C.elegans24-h LC(mg/l)Interval of confidence(95%) LC10 3.6500.016<LC10<11.33LC5022.55 4.200<LC50<56.63LC90139.455.75<LC90<5611LC means lethal concentration.J.-Y.Roh et al./Toxicology237(2007)126–133129Fig.1.Growth and reproduction indicators examined in the young adult of Caenorhabditis elegans exposed to DEHP for24h.Growth was assessed by measuring the length of the worms that had been killed by the heat through microscopy,with a scaled lens in each sample.Reproduction was assessed by counting the eggs of each worm through the microscopic inspection of the transparent C.elegans body in each sample.DEHP exposure in C.elegans were shown in Table2.For stress-related gene expression profiling analysis,based on the result from the microarray and what is found in the literature,we investigated alteration on the gene expression of heat shock proteins(hsp-16.1,hsp-16.2, hsp-16.48,hsp-70),metallothioneins(mt-1,mt-2),vitel-logenins(vit-2,vit-6),xenobiotic metabolism enzymes (cyp35a2,gst-4),tumor suppressor and apoptosis pro-teins(cep-1,ape-1),and antioxidant enzymes(sod-1, ctl-2)in C.elegans by DEHP exposure.The stress-related gene expression profile was inves-tigated in the young C.elegans adults exposed to DEHP for24h(Fig.2).Expression of hsp-16.1and hsp-16.2decreased at0.02and0.2mg/l of DEHP expo-sure.Expression of mt-2increased at DEHP-treated C.elegans.However,due to high data variation,sta-tistical significance was not observed.Expression of cyp35a2and gst-4,phase I and phase II of xenobiotic metabolism enzymes,was increased by DEHP expo-sure in a concentration-dependent manner.Expression of genes related to vitellogenes,apoptosis,or antioxidant enzymes was not changed by DEHP exposure.As shown in Fig.3,the hsp-16.2and hsp-16.48 gene expression levels were assayed using gfp-basedTable2Screening of DEHP induced gene expression using C.elegans whole genome microarrayUp-regulated genes Down-regulated genesWormbase no.Sequence description Flod change Wormbase no.Sequence description Flod change K02D7.3Cuticular collagen17Y71D11A.5Ion channel protein0.03C39E9.3Collagen 6.4W03G1.7Sphingomyelin phosphodiesterase0.03R08F11.7Peroxidase 6.2F36D3.9Cysteine protease0.07B0365.6C-type lectin domain 5.8C06A8.9Glutamate receptor0.07CEC2033Major sperm protein 5.0F21F8.2Protease0.08K08E7.9Multidrug resistance protein 4.2C58111GTP-binding protein0.08C33A12.6UDP-glucuronysltransferase 3.6C52B9.9AMP-binding protein0.1K09F5.2vit-1 3.0C37H5.1Annexin0.13F09G2.3Permease 2.9C08H9.1Serine carboxypeptidase0.15T11F9.3Zinc metalloprotease 2.9T07G12.1Calmodulin0.16F34H10.1Ubiquitin/ribosomal protein 2.7K08C7.5Flavin-containing monoxygenase0.16C03G6.15Cytochrome P450 2.7F53B7.2G-protein coupled receptor0.18F28A12.4Peptidase 2.6F48E3.7Acetylcholine receptor0.18F22D6.10Collagen 2.6Y46H3A.3Heat shock protein0.19ZK1248.17Major sperm protein 2.6C42C1.2Protein phosphatase0.19F08A8.3Acyl-coenzyme A oxidase 2.5E02H9.5Glycosyl hydrolase0.23R07B1.3Membrane glycoprotein 2.5T27E4.8Heat shock protein HSP16-10.28W06D12.3Fatty acid desaturase 2.4T27E4.9Heat shock protein0.3K08B4.3Glucuronosyltransferase 2.4K03A1.4Calmodulin calcium-binding sites0.31F44G3.2Arginine kinase 2.4AU113943Carbonic anhydrase0.31130J.-Y.Roh et al./Toxicology 237(2007)126–133Fig.2.Stress-related gene expression profiling in the young adult of C.elegans exposed to DEHP for 24h.Stress related gene mRNA was amplified by RT-PCR method using the primers designed on the basis of the sequences retrieved from the C.elegans database ( ).Actin mRNA was used for expression-level normalization of the studied genes.The PCR products were separated through electrophoresis on 1.5%agarose gel and were visualized with ethidium bromide.All the tests were replicated at least threetimes.Fig.3.Response of hsp-16.2::gfp and hsp-16.48::gfp transgenic C.elegans exposed to DEHP for 24h.Transgenic C.elegans were incubated for 24h with 2mg/l of DEHP and the fluorescence signal was examined from 20independent transgenic worms per treatment.Fluorescence was observed using a Leica DM IRB microscope,and the image was taken using a Leica DC 300FX camera.reporter transgenic nematodes.Transgenic nematodes were exposed to 2mg/l of DEHP and the fluorescence signals from both gfp transgenic lines increased after DEHP exposure.The response of hsp-16.48::gfp,how-ever,was greater than that of hsp-16.2::gfp.Concomitantly with bioassay using C.elegans ,the analysis of DEHP was conducted on three Korean landfill sites (Table 3).The contamination levels of DEHP in soil from landfill sites were between 6and 20mg/kg soil.DEHP was not detected in the soil from the referencesite.Fig.4.Mortality,growth and reproduction indicators (A),stress-related gene expression profiling measured in the young adult of C.elegans exposed for 24h to DEHP contaminated soil from Korean landfill sites (B).Response of hsp-16.2::gfp and hsp-16.48::gfp transgenic C.elegans exposed for 24h to DEHP contaminated soil from Korean landfill sites (C).The soils were named as R,G,M,J (R:reference site soil,G:Gwangju landfill soil,M:Mokpo landfill soil,J:Jeonju landfill soil).J.-Y.Roh et al./Toxicology237(2007)126–133131Table3Analysis of DEHP on soil from Korean landfill sites(number=3; mean±standard error of the mean)R a G b M c J dDEHP(mg/kg)ND e 6.27±0.6520.05±1.3410.73±1.37a R:Reference soil site.b G:Gwangju landfill site.c M:Mokpo landfill site.d J:Jeonju landfill sites.e ND:non detected.Fig.4shows the response of C.elegans on mortal-ity,growth,reproduction,stress-related gene expression and the response of transgenic C.elegans exposed to soil from Korean landfill sites for24h.Increase in mortality occurred in the most severe extent(about30%)in soil from the M landfill site,which showed the highest DEHP contaminated level among the three sites(20mg/kg). Growth parameters did not change in the worm exposed to soil from the landfill site.The number of eggs per worm slightly increased in the nematode that had been exposed to soil from the J landfill site.Differently from that in Fig.2,worms that had been exposed to soil from landfill sites did not show any statistically significant change in studied stress-related gene expression.Theflu-orescence signals from both gfp transgenic lines slightly increased in transgenic nematode exposed to soil from the landfill site.4.DiscussionsC.elegans is an attractive animal model for the study of the ecotoxicological relevance of chemical-induced molecular-level responses(Menzel et al.,2005;Reichert and Menzel,2005).In this study,the utility of molecu-lar parameters,such as stress-related gene expression, as biomarkers in C.elegans were investigated to iden-tify specific and sensitive tools to develop a screening system for ecotoxicity monitoring.And the possible use of C.elegans as a sentinel organism in the monitoring of soil ecosystem health was also tested by conduct-ing the experiment on the exposure of nematode tofield soil.Although DEHP is a widely used environmental compound,little study has been performed on its eco-toxicological properties.Twenty-four-hour LC50data (Table1)suggest that DEHP has a relatively higher potential of acute toxicity to C.elegans than previ-ously studied metals have(Roh et al.,2006).Mortality is a reliable ecotoxicological endpoint.However,such a high level of exposure hardly occurred in the real environment.More sensitive indicators,physiological-level alterations,such as growth,reproduction,feeding, movement,or behavior,have been used as endpoints for chemical-induced toxicity testing in C.elegans(Dhawan et al.,2000;Anderson et al.,2001,2004;Kohra et al., 2002;Tominaga et al.,2003).The effects of xenobiotics on the growth and reproduction of the test organisms are broadly accepted test parameters,and were found to be more sensitive indicators of toxicity than lethality,as also shown in this study(Fig.1).The decreases in body length and egg number per worm observed after24h of DEHP exposure may induce alteration in the growth and reproduction of the nematode population in the long term.However,due to the low concentration of xenobi-otics in the environment,it is hard tofind a correlation between the occurrence of contaminants and a physio-logical effect of a test organism in the environment,even when using reliable ecotoxicological endpoints,such as,growth or reproduction,which emphasizes the need for understanding the sublethal effects at the biochem-ical and molecular levels where the toxicant-induced responses are initiated.The effect of DEHP to the C.elegans whole genome gene expression was ing concentra-tion corresponding to the1/100of LC50value,which was0.2mg/l of DEHP,it was possible to show that a strong and differential gene induction or repression was detectable in response to DEHP exposure;they were cyp superfamily,hsp,vit,multidrug-resistance protein, etc.(Table2).In case of some genes,such as C-type lectin,collagene,and major sperm protein genes,it is unclear what physiological meaning the over-expression has.Stress-related genes were selected from the array results and previously reported literature.Expression of selected stress-related genes to DEHP exposure was investigated at three different sublethal concentrations, using the semi-quantitative RT-PCR method.It is obvi-ous that members of the cyp family,gst,hsp,and vit genes should be included in a selection of stress-related genes.Among the group of genes,which encode proteins belonging to different metabolic pathways,the met-allothionein,apoptosis,and antioxidant enzyme genes were also included in the stress-related gene selection. As screened in the microarray result,the expression of hsp-16.1and hsp-16.2genes was decreased by DEHP exposure.DEHP exposure led to increases in the expres-sion of some stress-related genes tested,including mt-2, cyp35a2and gst-4.In particular,it has been reported that almost all cyp35forms in C.elegans are moder-ately or strongly inducible by different xenobiotics in a cyp450gene-expression screening experiment(Menzel et al.,2001,2005).Increase in stress-related gene expres-sions occurred concomitantly with this deterioration on132J.-Y.Roh et al./Toxicology237(2007)126–133the physiological level,which suggests that the increase in the expression of those genes may not be considered as a homeostatic response but as toxicity that might lead to physiological consequences(Figs.1and2).C.elegans also offers the advantage of transgenetic approaches,which may allow the development of a sensi-tive biosensor for environmental monitoring(Stringham and Candido,1994;Jones et al.,1996;Chu et al.,2005). In our previous study(Roh et al.,2006),gfp trans-genic nematode,incorporated full-length hsp-16.2and hsp-16.48genes were developed,since hsps are thought to play roles in various stress conditions.As shown in their response to the exposure to four metals(Roh et al.,2006),semi-quantitatively assayed using gfp-based reporter hsp-16.2and hsp-16.48transgenic nematodes were not very sensitive towards DEHP exposure.Even though,transgenic nematode seems to have a consider-able potential as a biosensor for toxicity monitoring,to develop a sensitive biosensor using transgenic C.ele-gans,the responses of a broad range of stress-related gene promoters to various classes of chemicals should be screened and validated with environmentally relevant low-concentration samples.Bioassay methods for soil toxicity monitoring have also been developed and frequently used in C.ele-gans(Peredney and Williams,2000;Boyd and Williams, 2003b).As a soil-dwelling organism,C.elegans has a potential as a bioindicator for the detection of soil con-tamination because of its abundance in soil ecosystems and its sensitivity to different kinds of stresses,which was tested in this study using the same endpoints used for laboratory K-media exposure condition(Fig.4).Increase in egg number per worm in the nematode exposed to soil from landfill sites suggests a mixture of contaminants, including DEHP,might have a stimulatory effect on the reproduction of the nematode.Expression of stress-related genes did not affect C.elegans exposed to soil from the landfill site,which were different from the response of nematodes exposed to DEHP prepared in K-media in laboratory conditions.The common feature of responses to DEHP of C.elegans between labora-tory andfield conditions was the increase in gst-4gene expression.But,as gst is known to use a broad range of substrates,it is difficult to consider it as to DEHP-specific biomarker.Increase influorescence signal of hsp-16.2::gfp and hsp-16.48::gfp transgenic nematodes might be interpreted as a response of heat-shock protein upon exposure to mixture contaminants.The experiment with the soil from the landfill site suggests that the poten-tial of the C.elegans biomarker identified in laboratory conditions should be calibrated and validated for their use in situ.The data obtained from this study can comprise an important contribution to the knowledge of the toxi-cology of DEHP in C.elegans,about which little data are available.A link or correlation between a vali-dated toxicity endpoint(e.g.,growth and reproduction) and upstream-induced gene expression is interest-ing,particularly for ecotoxicological purposes.Direct experimental demonstrations of the wider relationships between molecular-/biochemical level effects and their subsequent consequences at higher levels of biologi-cal organization are needed in order to establish causal relationships.The characterization of the causal rela-tionships between the molecular level responses and the effects at higher biological levels will help to define the sublethal hazards of chemicals in C.elegans. AcknowledgementThis work was accomplished through the generous support of the Korea Research Foundation Grant funded by the Korean Government(MOEHRD)(Grant No. KRF-2005-204-D00009).Appendix A.Supplementary dataSupplementary data associated with this arti-cle can be found,in the online version,at doi:10.1016/j.tox.2007.05.008.ReferencesAnderson,G.L.,Boyd,W.A.,Williams,P.L.,2001.Assessment of sublethal endpoints for toxicity testing with the nematode Caenorhabditis elegans.Environ.Toxicol.Chem.20,833–838. Anderson,G.L.,Cole,R.D.,Williams,P.L.,2004.Assessing behav-ioral toxicity with Caenorhabditis elegnas.Environ.Toxicol.Chem.23,1235–1240.ASTM,2001.Standard Guide for Conducting Laboratory Soil Toxi-city Test with the Nematocde Caenorhabditis elegans.E2172-01.American Society for Testing and Materials,West Conshohocken, PA.ATDSR,2002.Toxicological Profile for Di-(2-ethylhexyl)phthalate (DEHP).Department of Health and Human Services,Public Health Service,Agency for Toxic Substances and Disease Registry, Atlanta.Bettinger,J.C.,Carnell,L.,Davies,A.G.,McIntire,S.L.,2004.The use of Caenorhabditis elegans in molecular neuropharmacology.Int.Rev.Neurobiol.62,195–212.Boyd,W.A.,Williams,P.L.,2003a.Availability of metals to the nematode Caenorhabditis elegans:toxicity based on total concen-trations in soil and extracted fractions.Environ.Toxicol.Chem.22,1100–1106.Boyd,W.A.,Williams,P.L.,parison of the sensitivity of three nematode species to cooper and their utility in aquatic and soil toxicity tests.Environ.Toxicol.Chem.22,2768–2774.。

植物组织中重金属含量测定思考题植物组织中重金属含量测定是环境监测和食品安全评估中的重要任务。

重金属是指相对密度大于5g/cm³的金属元素，如铅、镍、汞、铬等。

它们主要来自工业废水、农业污水、废弃物排放和大气沉降等人类活动的排放。

重金属的富集和累积会对土壤、水体和植物造成污染，进而对人类健康产生潜在威胁。

因此，准确测定植物组织中重金属含量非常重要。

测定植物组织中重金属含量的方法有很多种，下面将介绍常用的几种方法。

1. 原子吸收光谱法（AAS， Atomic Absorption Spectroscopy）原子吸收光谱法是一种常用的测定重金属含量的方法。

它利用重金属原子在特定波长的光束照射下吸收能量并使原子转换为高能态，测定原子吸收能量的大小来计算重金属的浓度。

此方法准确可靠，且对样品处理的要求较为简单，适用于不同植物组织的重金属含量测定。

2. 电感耦合等离子体质谱（ICP-MS，Inductively Coupled Plasma Mass Spectrometry）电感耦合等离子体质谱是一种高灵敏度、高准确度的测定重金属含量的方法。

它通过将样品离子化形成等离子体，进而利用质谱仪测定各种离子的质量和相对丰度，从而计算出重金属的含量。

ICP-MS方法灵敏度高，可同时测定多种重金属，并对样品的基体干扰有较好的抑制能力。

3. 石墨炉原子吸收光谱法（GF-AAS，Graphite Furnace AtomicAbsorption Spectroscopy）石墨炉原子吸收光谱法是一种测定重金属含量的敏感方法。

它通过将样品中的重金属原子谱线放大，在石墨炉中加热样品使其原子转化为气态，然后利用原子吸收光谱法测定气态原子对特定波长光的吸收程度，从而计算出重金属的含量。

该方法对样品处理要求严格，但具有较高的灵敏度和准确度。

4. 电感耦合等离子体发射光谱法（ICP-OES，Inductively Coupled Plasma Optical Emission Spectroscopy）电感耦合等离子体发射光谱法是一种广泛应用于重金属含量测定的方法。

重金属检测技术在环境水质分析中的应用探讨水是生命之源，对于人类的生存和发展至关重要。

然而，随着工业化和城市化进程的加速，重金属污染成为了水环境面临的严峻问题之一。

重金属具有毒性、持久性和生物累积性，对生态系统和人类健康构成严重威胁。

因此，准确、灵敏地检测水中的重金属含量对于评估水质状况、保障水安全具有重要意义。

一、常见的重金属污染物及其危害在环境水质中，常见的重金属污染物包括汞（Hg）、镉（Cd）、铅（Pb）、铬（Cr）、砷（As）等。

这些重金属污染物来源广泛，如工业废水排放、矿山开采、农业化学品使用等。

汞是一种具有强烈神经毒性的重金属，能够损害中枢神经系统，导致认知障碍、运动失调等症状。

镉被人体摄入后，会在肾脏中累积，引发肾衰竭等疾病。

铅会影响儿童的智力发育和神经系统功能，对成年人则可能导致心血管疾病和贫血。

铬具有致癌性，长期接触可能导致肺癌等疾病。

砷则会引起皮肤病变、心血管疾病和多种癌症。

二、重金属检测技术的分类（一）原子吸收光谱法（AAS）原子吸收光谱法是一种基于原子对特定波长光的吸收来定量分析重金属的技术。

它具有灵敏度高、选择性好、分析速度快等优点。

火焰原子吸收光谱法（FAAS）适用于较高浓度的重金属检测，而石墨炉原子吸收光谱法（GFAAS）则能够检测更低浓度的重金属。

（二）原子荧光光谱法（AFS）原子荧光光谱法利用原子在特定条件下发射的荧光强度来测定重金属含量。

该方法具有灵敏度高、干扰少等优点，特别适用于砷、汞等元素的检测。

（三）电感耦合等离子体发射光谱法（ICPOES）电感耦合等离子体发射光谱法可以同时测定多种元素，具有检测范围宽、精密度高的特点。

它能够快速准确地分析水中多种重金属的含量。

（四）电感耦合等离子体质谱法（ICPMS）ICPMS 是一种高灵敏度、高分辨率的检测技术，可以检测极低浓度的重金属。

但仪器设备昂贵，运行成本较高。

（五）电化学分析法电化学分析法包括极谱法、伏安法等，通过测量电化学反应中的电流、电位等参数来确定重金属的浓度。

重金属检验报告1. 引言重金属是一类具有高密度和毒性的金属元素，存在于环境中的许多不同来源中，如工业废水、农药和污染的土壤等。

这些重金属对人类健康和环境具有潜在危害，因此对其进行检测和监测非常重要。

本报告旨在提供一份重金属检验的详细分析报告，包括检测方法、样品信息、结果分析和结论。

2. 方法和材料2.1 检测方法本次重金属检验采用原子吸收光谱法（Atomic Absorption Spectroscopy, AAS）进行分析。

该方法通过测量重金属元素在可见光范围内对特定波长的吸收程度来确定其存在的浓度。

2.2 样品信息样品名：地下水样品来源：位于某工业区附近的监测井样品采集日期：2021年5月10日3. 结果分析3.1 检测结果以下是地下水样品中各种重金属的检测结果（单位：ppm）：重金属元素检测结果铅（Pb）0.03镉（Cd）0.01铬（Cr）0.05汞（Hg）0.002镍（Ni）0.023.2 结果解读根据国家相关标准，地下水中重金属元素的允许浓度限值如下：重金属元素允许浓度限值（ppm）铅（Pb）0.05镉（Cd）0.01铬（Cr）0.05汞（Hg）0.01镍（Ni）0.02根据检测结果可以看出，地下水中铅、镉、汞和镍的浓度均在标准限值范围内，符合相关规定要求。

然而，地下水中铬的浓度略高于限值，需要进一步监测和评估是否存在潜在风险。

4. 结论根据对地下水样品的重金属检测结果进行分析，可以得出以下结论：1.地下水中铅、镉、汞和镍的浓度均在国家允许浓度限值范围内，属于合格水质。

2.地下水中铬的浓度略高于国家允许浓度限值，需要进一步监测和评估是否存在潜在风险。

3.对于铅、镉、汞和镍浓度符合标准的地下水样品，继续加强监测和控制是保证水源安全和环境健康的重要措施。

建议进一步进行环境评估，制定相应的控制策略，确保地下水资源的可持续利用和环境的安全性。

以上就是本次重金属检验的详细报告。

注意：本报告仅供参考，结果可能受样品数量和采集地点等因素的影响。