Production of Antibodies and Development of Specific Polarization Fluoroimmunoassay for Acetochlor

Specific polyclonal antibodies towards acetochlor (2-chloro- N -(ethoxymethyl)- N -(2-ethyl-6-methylphenyl)acetamide) were obtained from rabbits immunized against a 3-mercaptopropionic acid derivative of acetochlor, covalently attached to bovine serum albumin. A polarization fluoroimmuoassay (PFIA) based on these antibodies was developed and optimized to detect acetochlor in water samples. The optimized PFIA had a detection limit of 9 µg/L, linear working range from 50 to 5500 µg/L and within-assay coefficient of variation less than 4%. Cross-reactivity studies demonstrated that these antibodies are capable of specific detection of acetochlor amongst structurally related chloroacetanilide herbicides. Assay cross-reactivity values were: alachlor 0%, metolachlor 2.4%, propachlor 0%, butachlor 0.2% and dimethachlor 0.5%. Five organic solvents commonly used in sample extraction were evaluated for their effect on acetochlor PFIA performance, and methanol and ethanol were found to be compatible with the assay up to 10% v/v.     

扑草净、乙草胺混配除草剂的HPLC 分析方法

建立了正相高效液相色谱法定量分析二元混配除草剂"昆明6号"中扑草净和乙草胺的方法。采用ShimPackCLC CN(150mm×6.0mm)5μm柱,流动相为正己烷/氯仿(87∶13,V/V),λ=264nm,线性相关系数为0 9997～0 9999。扑草净和乙草胺的加料回收率分别为99 0%和101 0%,RSD分别为0 52%和0 60%。     

An Enzyme-Linked Immunosorbent Assay (Elisa) for the Detection of Acetochlor

Competitive enzyme-linked immunosorbent assays (ELISAs) were developed in hapten-homologous and hapten-heterologous formats for the detection of the chloroacetanilide herbicide acetochlor. ELISA systems were devised using antibodies generated against acetochlor conjugated to carrier proteins through a thioether moiety replacing the chlorine atom in the parent structure, while haptens modified both on the chloroacetyl moiety and on the ethoxymethyl group of acetochlor have been used for coating antigens. The optimized ELISA systems allowed the detection of acetochlor 0.2-65 µg/L, and cross-reactivity studies revealed high specificity of the immunoassay: only four (propisochlor, butachlor, alachlor and metolachlor) among 18 structurally related acetanilide herbicides, fungicides and intermediates showed significant (> 1%) cross-reactivity, with even the highest value (propisochlor) being below 10%. Assay performance was not affected detrimentally by methanol up to 10% (v/v) and ethanol up to 5% (v/v). Assay performance was tested by measuring acetochlor concentration in water samples and compared favorably ( r 2 = 0.976) with those detected by gas chromatographic method coupled with mass spectrometry (GC-MS) using solid-phase microextraction (SPME) for sample preparation.     

固相微萃取-气相色谱-质谱联用测定水中的除草剂乙草胺

建立了固相微萃取技术与气相色谱-质谱联用快速检测水中除草剂乙草胺的方法。对萃取时间、搅拌速度及离子强度等实验条件进行了优化。该方法的线性范围为0．2—20μg／L；检出限为0．018μg／L；相对标准偏差为4．6％。该法可直接用于田间地面水中乙草胺的测定。     

气相色谱-质谱法测定甘蔗中除草剂残留量

提出了采用QuEChERS法提取净化,气相色谱-质谱法同时测定甘蔗中除草剂莠去津、乙草胺和莠灭净的残留量。采用DB-5MS毛细管柱分离,电子轰击离子源选择离子检测模式进行质谱测定。莠去津、乙草胺和莠灭净的检出限(3S/N)分别为0.001,0.002,0.001mg.kg-1。在3个浓度水平上对方法的回收率作了测定,测得回收率在88.0%～108.0%,相对标准偏差(n=6)在1.1%～6.9%之间。     

Rapid determination of amide herbicides in environmental water samples with dispersive liquid–liquid microextraction prior to gas chromatography–mass spectrometry

This paper describes a novel, simple and environmentally friendly method for rapid determination of the amide herbicides metoalchlor, acetochlor, and butachlor. It is based on dispersive liquid-liquid microextraction and gas chromatography–mass spectrometry. Factors that may influence the enrichment efficiency, such as type and volume of extraction solvent, type and volume of dispersive solvent, extraction time, and content of NaCl, were investigated and optimized in detail. Under the optimum conditions, the limits of detection of metoalchlor, acetochlor, and butachlor were 0.02, 0.04, and 0.003 μg L⁻¹, respectively. The experimental results indicated that there was linearity over the range 0.1–50 μg L⁻¹ and good reproducibility with relative standard deviations over the range 1.6–3.0% (n = 5). The proposed method has been applied for the analysis of real-world water samples, and satisfactory results were achieved. Average recoveries of spiked herbicides were in the range 80.3–108.8%. All of these indicated that the developed method would be an efficient method for simultaneous determination of the three herbicides in environmental water samples.     

Monitoring water-polluting pesticides in Hungary

A 5-year survey of pesticide active ingredients and residues in Hungarian surface water samples was carried out within the framework of a national monitoring program. Based on physicochemical and ecotoxicological properties of currently registered pesticide active ingredients, a range of analytes was selected to cover compounds that potentially contaminate surface waters due to their solubility properties or mode of use. Target analytes thus included acetochlor, atrazine, carbofuran, diazinon, fenoxycarb, metribuzin, phorate, prometryn, terbutryn, and trifluralin. During the sampling campaign these pesticides were monitored in Hungarian surface waters including streams, rivers and lakes. Samples were obtained annually in two runs: before and after pesticide sprayings in spring and early summer. Samples were prepared for analysis by solid-phase extraction and solid-phase microextraction. Target analytes were monitored by gas chromatography - mass spectrometry, using electron impact and chemical ionization techniques. Spatial distribution monitoring of the surface water pollutants indicated two heavily contaminated point sources, as well as a wide range of non-point contamination. One or more pesticide active ingredients above the detection limit of the instrumental method used were measured in 209 samples, giving the result that 59% of the samples collected during the sampling campaign contained pesticide residues.     

Development of a model for the rational design of molecular imprinted polymer: Computational approach for combined molecular dynamics/quantum mechanics calculations

A new rational approach for the preparation of molecularly imprinted polymer (MIP) based on the combination of molecular dynamics (MD) simulations and quantum mechanics (QM) calculations is described in this work. Before performing molecular modeling, a virtual library of functional monomers was created containing forty frequently used monomers. The MD simulations were first conducted to screen the top three monomers from virtual library in each porogen-acetonitrile, chloroform and carbon tetrachloride. QM simulations were then performed with an aim to select the optimum monomer and progen solvent in which the QM simulations were carried out; the monomers giving the highest binding energies were chosen as the candidate to prepare MIP in its corresponding solvent. The acetochlor, a widely used herbicide, was chosen as the target analyte. According to the theoretical calculation results, the MIP with acetochlor as template was prepared by emulsion polymerization method using N,N-methylene bisacrylamide (MBAAM) as functional monomer and divinylbenzene (DVB) as cross-linker in chloroform. The synthesized MIP was then tested by equilibrium-adsorption method, and the MIP demonstrated high removal efficiency to the acetochlor. Mulliken charge distribution and ¹H NMR spectroscopy of the synthesized MIP provided insight on the nature of recognition during the imprinting process probing the governing interactions for selective binding site formation at a molecular level. We think the computer simulation method first proposed in this paper is a novel and reliable method for the design and synthesis of MIP.     

凝胶渗透色谱-气相色谱-质谱测定花生中乙草胺的残留量

建立了以凝胶渗透色谱(GPC)净化和气相色谱-质谱(GC-MS)技术快速测定花生中乙草胺残留量的方法.经乙腈提取,共提物中的油脂和色素经GPC去除,目标农药采用GC-MS-SIM方式进行定性和定量分析.方法的回收率90%～120%;相对标准偏差2.5%～10%.方法定量限0.005 μg/g.     

Interaction of acetochlor and atrazine with natural freshwater biofilms grown on polycarbonate substrate in lake Velence (Hungary)

Biofilms were colonized in Lake Velence, Hungary, on polycarbonate film bands. After 4 weeks of colonization, the biofilms were transported into the laboratory and were kept in temperature-controlled aquaria for 2 subsequent weeks. The aquaria were filled with herbicide-spiked lake water (containing 12.5 μg/dm³ acetochlor and 12.5 μg/dm³ atrazine). The degradation of these herbicides was investigated by measuring their concentration in the water phase in the presence and absence of biofilms. The presence of biofilms resulted in a considerable reduction of acetochlor (45%) and atrazine (32%) in the water phase; however, the photodegradation of herbicides was relatively negligible. Inasmuch as only a small amount of herbicides was accumulated in the biofilms, it can be concluded that the biodegradation is responsible for the reduced concentration of herbicides in the water phase, and this process plays the decisive role in the decomposition of the contaminants. On the other hand, these herbicides can inhibit the colonization of algae, but simultaneously, they influence the metabolic processes of algae and bacteria, resulting in an enhanced production of extracellular polymer matrix.