De novo synthesis of a novel hapten and development of a monoclonal antibody-based immunoassay for the detection of dichlorvos and trichlorfon.

The absence of high-affinity antibodies has hindered the development of satisfactory immunoassays for dichlorvos (DDVP) and trichlorfon (TCP), two highly toxic organophosphorus pesticides. Herein, the de novo synthesis of a novel anti-DDVP hapten was introduced. Subsequently, a specific anti-DDVP monoclonal antibody (Mab) was produced with satisfying affinity to DDVP (IC50: 12.4 ng mL-1). This Mab was highly specific to DDVP, and TCP could readily convert into DDVP under mild alkaline conditions. Leveraging this insight, an indirect competitive ELISA was successfully developed for simultaneous detection of DDVP and TCP. The limit of detection in rice, cabbage and apple for DDVP /TCP was found to be 12.1/14.6 µg kg-1, 7.3/8.8 µg kg-1 and 6.9/8.3 µg kg-1, respectively. This study not only provides an effective strategy for producing a high-quality anti-DDVP Mab but also affords a reliable and cost-effective tool suitable for high-throughput detection of DDVP and TCP in food samples.

Study on the acute toxicity of trichlorfon and its breakdown product dichlorvos to goldfish (Carassius auratus) based on 1H NMR metabonomics.

Trichlorfon, one of the most widely used organophosphate insecticides, is commonly employed in aquaculture and agriculture to combat parasitic infestations. However, its inherent instability leads to rapid decomposition into dichlorvos (DDVP), increasing its toxicity by eightfold. Therefore, the environmental effects of trichlorfon in real-world scenarios involve the combined effects of trichlorfon and its degradation product, DDVP. In this study, we systematically investigated the degradation of trichlorfon in tap water over time using HPLC and LC-MS/MS analysis. Subsequently, an experiment was conducted to assess the acute toxicity of trichlorfon and DDVP on goldfish (Carassius auratus), employing a 1H NMR-based metabolic approach in conjunction with serum biochemistry, histopathological inspection, and correlation network analysis. Exposure to trichlorfon and its degradation product DDVP leads to increased lipid peroxidation, reduced antioxidant activity, and severe hepatotoxicity and nephrotoxicity in goldfish. Based on the observed pathological changes and metabolite alterations, short-term exposure to trichlorfon significantly affected the liver and kidney functions of goldfish, while exerting minimal influence on the brain, potentially due to the presence of the blood-brain barrier. The changes in the metabolic profile indicated that trichlorfon and DDVP influenced several pathways, including oxidative stress, protein synthesis, energy metabolism, and nucleic acid metabolism. This study demonstrated the applicability and potential of 1H NMR-based metabonomics in pesticide environmental risk assessment, providing a feasible method for the comprehensive study of pesticide toxicity in water environments.

Degradation, migration, and removal of trichlorfon on harvested apples during storage at room temperature.

Dissipation of an organophosphorus pesticide, trichlorfon, in natural and waxed apples during storage was studied. The results showed that the trichlorfon content in natural and waxed apples decreased by 85% and 64%, respectively, during storage. The morphology of the surface film was dense and regular, which resulted in a higher water vapor resistance and a lower respiration rate in the waxed apples. This indicates that waxing affected the dissipation of pesticide residues in the apple storage environment, increasing food safety risk. Ozone was used to remove the residual pesticides on the apple surface. The trichlorfon degradation rate reached 73%. The ozone treatment had no effect on the surface color of the apple, which means that the fruit can be pre-treated with ozone prior to waxing or storage.

Quinolone and Organophosphorus Insecticide Residues in Bivalves and Their Associated Risks in Taiwan.

Bivalves, such as freshwater clams (Corbicula fluminea) and hard clams (Meretrix lusoria), are the most extensive and widely grown shellfish in land-based ponds in Taiwan. However, few studies have examined the contamination of bivalves by quinolone and organophosphorus insecticides. Thus, we adapted an established procedure to analyze 8 quinolones and 12 organophosphorus insecticides using liquid and gas chromatography-tandem mass spectrometry. Surveys in Taiwan have not noted high residual levels of these chemicals in bivalve tissues. A total of 58 samples of freshwater or hard clams were obtained from Taiwanese aquafarms. We identified 0.03 mg/kg of enrofloxacin in one freshwater clam, 0.024 mg/kg of flumequine in one freshwater clam, 0.02 mg/kg of flumequine in one hard clam, 0.05 mg/kg of chlorpyrifos in one freshwater clam, 0.03 mg/kg of chlorpyrifos in one hard clam, and 0.02 mg/kg of trichlorfon in one hard clam. The results indicated that 5.17% of the samples had quinolone insecticide residues and 5.17% had organophosphorus residues. However, the estimated daily intake (EDI)/acceptable daily intake quotient (ADI) indicated no significant risk and no immediate health risk from the consumption of bivalves. These results provide a reference for the food-safety screening of veterinary drugs and pesticides in aquatic animals. Aquatic products should be frequently screened for residues of prohibited chemicals to safeguard human health.

Fluorescence assay for three organophosphorus pesticides in agricultural products based on Magnetic-Assisted fluorescence labeling aptamer probe.

There has been increasing recent concern about the agricultural use of organophosphorus pesticides. A rapid and sensitive fluorescence assay for the detection of three organophosphorus pesticides has therefore been developed using 6-carboxy-fluorescein labeling aptamer as the probe and functionalized magnetic nanoparticles as the separation carrier. The aptamer hybridized with complementary DNA conjugated on the surface of the magnetic nanoparticles to form a magnetic aptamer-complementary DNA complex. Upon introducing the target organophosphorus pesticide, the aptamer departed from the complementary DNA, resulting in the fluorescence signal. Under optimized conditions, the limits of detection (LODs, S/N = 3) for trichlorfon, glyphosate, and malathion were 72.20 ng L-1, 88.80 ng L-1, and 195.37 ng L-1, respectively. The method was applied for the detection of trichlorfon, glyphosate, and malathion in spiked lettuce and carrot samples. The recoveries were in the range of 79.4%-118.7%, which were in good agreement with those obtained by gas chromatography, and the relative standard deviations were also acceptable. The method therefore has high sensitivity, so provides a means for the detection of multiple organophosphorus pesticides.

Bioremediation of cadmium-trichlorfon co-contaminated soil by Indian mustard (Brassica juncea) associated with the trichlorfon-degrading microbe Aspergillus sydowii: Related physiological responses and soil enzyme activities.

Soil co-contaminated with heavy metals and organics is often difficult to remediate. In this study, pot experiments were conducted to investigate the concurrent removal of cadmium (Cd, two levels: CdL [10 mg kg-1] and CdH [50 mg kg-1]) and trichlorfon (TCF, 100 mg kg-1) from co-contaminated soil by comparing the following remediation methods: natural remediation (NR), soil inoculated with Aspergillus sydowii (AS), soil planted with Brassica juncea (BJ), and soil planted with B. juncea and inoculated with A. sydowii (BJ-AS). The physiological responses of B. juncea and soil enzyme activities after remediation were also studied. B. juncea grew well in co-contaminated soil at both Cd levels. The biomass and chlorophyll content of B. juncea in CdH soil were lower than those in CdL soil, whereas the malondialdehyde content and activities of catalase, peroxidase and superoxide dismutase of B. juncea in CdH soil were higher than those in CdL soil. Cd accumulation in B. juncea was high in CdH soil, whereas high Cd removal efficiency was observed in CdL soil. TCF could be thoroughly degraded within 35 days in NR at both Cd-level soils. AS, BJ and BJ-AS promoted TCF degradation and enhanced the activities of catalase, urease, sucrase and alkaline phosphatase in soil compared with the NR. BJ-AS showed the highest phytoextraction ratio (3.32% in CdL and 1.34% in CdH soil) and TCF degradation rate (half-life of 2.18 and 2.37 days in CdL and CdH soil, respectively). These results demonstrate that BJ-AS could effectively remove Cd and TCF from soil and is thus a feasible technology for the bioremediation of these co-contaminated soil.

Web-based Korean maximum residue limit evaluation tools: an applied example of maximum residue limit evaluation for trichlorfon in fishery products.

To ensure public safety against veterinary drug residues in food products from animal sources, maximum residue limits (MRLs) should be established by scientific evidence and a transparent estimation process. The Joint Food and Agriculture Organization (FAO)/World Health Organization (WHO) Expert Committee on Food Additives (JECFA) developed an Excel workbook-based tool for MRLs evaluation in 2003. In this study, we developed a web-based tool for MRL evaluation, called Korean MRL evaluation tools (KMET). While KMET used algorithms of JECFA workbook, it added some databases (e.g., Korean food consumption database) and provided additional functions (e.g., selection of target marker residue). Web-based KMET enabled regulatory policy makers to update the database. All input data and output results related to MRL evaluation based on residue depletion and food consumption datasets were archived and provided overall processes from the initial depletion data entry to MRL establishment with user-friendly interface. Our results demonstrated the stepwise processes whereby MRL for trichlorfon in the muscle of Paralichthys olivaceus was established with functional descriptions of KMET. MRL for trichlorfon derived from KMET was proposed and notified by the Ministry of Food and Drug Safety in 2018.

Development of direct competitive biomimetic immunosorbent assay based on quantum dot label for determination of trichlorfon residues in vegetables.

A direct competitive biomimetic immunosorbent assay method based on molecularly imprinted polymer was developed for the determination of trichlorfon. A CdSe/ZnS quantum dot label was used as the marker. The hydrophilic imprinted film was synthesized directly on the surface of a 96-well plate, and characterized by Fourier-transform infrared spectroscopy and thermo-gravimetric analyses. The method exhibited high stability, selectivity, and sensitivity. Under optimal conditions, the limits of detection and sensitivity of the biomimetic immunosorbent assay method were 9.0 µg L-1 and 5.0 mg L-1 (0.1 mg kg-1 and 62.5 mg kg-1 for vegetable sample), respectively. Low cross-reactivity values of 19.2% and 15.6% were obtained for the structural analogues. Spinach and rape samples spiked with trichlorfon were extracted and determined by this method with recoveries ranging from 83.6% to 91.1%. The method was applied for the detection of trichlorfon residues in leek and cucumber samples, and results correlated well with those obtained using GC.

Dissipation kinetics, safety evaluation, and preharvest interval assessment of trichlorfon application on rice.

Nowadays, there is an urgent need for the investigation of the field dissipation and assessment of the preharvest interval for trichlorfon residues on rice. To protect consumers from potential health risks, this study can provide references for the safe application of trichlorfon in the rice fields. Results of the field dissipation study showed that the dissipation dynamic equations of trichlorfon were based on the first-order reaction dynamic equations and that the dissipation rates vary among rice plant, brown rice, rice bran, soil, and water. The 2-year field trials conducted in Yangzhou and Xiaogan suggested the interval of each application for trichlorfon on rice to be at least 7 days when 80 % trichlorfon SP was sprayed with a dose ranges between 80 and 160 a.i g/667 m(2). Additionally, the preharvest interval of the last application should be at least 15 days to ensure the amounts of residues below the maximum residue limits of trichlorfon on brown rice (0.1 mg/kg).

Development of molecularly imprinted electrochemical sensors based on Fe3O4@MWNT-COOH/CS nanocomposite layers for detecting traces of acephate and trichlorfon.

In this study, we developed a novel biomimetic electrochemical sensor sensitized with a Fe3O4@carboxyl-functionalized multiwalled carbon nanotube/chitosan nanocomposite layer using a molecularly imprinted film as a recognition element for the rapid detection of acephate and trichlorfon. The performance of the imprinted sensor was investigated using cyclic voltammetry and differential pulse voltammetry, and the results indicated that the sensor exhibited fast responses to both acephate and trichlorfon. The imprinted sensor had good linear current responses to acephate and trichlorfon concentrations in the ranges from 1.0 × 10(-4) to 1.0 × 10(-10) M and 1.0 × 10(-5) to 1.0 × 10(-11) M, respectively. Under optimal conditions, the imprinted sensor had low limits of detection (signal to noise ratio, S/N = 3) of 6.81 × 10(-11) M for acephate and 8.94 × 10(-12) M for trichlorfon. The developed method was successfully applied to detect acephate and trichlorfon spiked in fortified kidney bean and cucumber samples with good recoveries ranging from 85.7% to 94.9% and relative standard deviations of 3.46-5.18%.

Degradation, residual determination, and cholinesterase activity of triclorfon in Piaractus mesopotamicus Holmberg (PACU) 1887.

The aim of this study was to determine the no-observable-adverse-effect concentration (NOAEC) for trichlorfon, an antiparasitic agent used in aquaculture, in Piractus mesopotamicus (pacu) using acetylcholinesterase (AChE) activity as an end point. Fish were exposed 24 h/d for 15 d to different concentrations of trichlorfon in tanks of water for which a curve of dissipation was previously determined. Analysis of trichlorfon in water and fish plasma using gas chromatography with electron capture detection (GC-ECD) enabled measurement of limit of detection (LOD) and limit of quantification (LOQ), respectively, to be 3 and 10 ppb. Thirty-six hours after trichlorfon dilution in water, the concentration was below the LOD, and data showed that plasma concentrations did not exceed the LOQ. Apart from the 6.25 µg/L, all concentrations of trichlorfon significantly inhibited plasma and brain AChE activity compared to controls. The AChE activity levels returned to control values in 7 d. These data may be useful to determine the concentration of trichlorfon that destroys parasites without producing adverse effects in fish.

Study of a molecularly imprinted solid-phase extraction coupled with high-performance liquid chromatography for simultaneous determination of trace trichlorfon and monocrotophos residues in vegetables.

BACKGROUND: Organophosphate pesticide residues are harmful to human health because of their potential mutagenic and carcinogenic properties. Therefore, it is of great importance to development an accurate and reliable analytical method to prevent their uncontrolled effects on environmental pollution and human health. RESULTS: This study reports a new method of molecularly imprinted solid-phase extraction coupled with high-performance liquid chromatography (MISPE-HPLC) for simultaneous determination of two organophosphate pesticides residues. Two types of molecularly imprinted polymers (MIPs) were prepared using the trichlorfon and monocrotophos as the template molecule, respectively, methacrylic acid as the functional monomer, and ethylene glycol dimethacrylate as the cross-linker. The recognition ability and adsorption-desorption dynamic of each imprinted polymer toward the trichlorfon or monocrotophos were characterised. Using the mixture of trichlorfon-MIP and monocrotophos-MIP (20:80, wt/wt) as solid-phase extraction sorbent, the factors affecting the pre-concentration on the analytes and the sensitivity of the MISPE-HPLC method were optimised. Under optimal condition, the linear range was 0.005-1.0 mg L⁻¹. The limit of detection was 4.2 µg g⁻¹ for trichlorfon, and 1.2 ng g⁻¹ for monocrotophos. The peak area precision [Relative standard deviation (RSD)] for three replicates was 2.9-4.5%. The blank rape and cauliflower samples spiked with trichlorfon and monocrotophos at 0.05 and 0.005 µg g⁻¹ levels were extracted and determined by this method with recoveries ranging from 88.5% to 94.2%. Moreover, this method was successfully applied to the quantitative detection of the trichlorfon and monocrotophos residues in leek samples. CONCLUSION: With good properties of high sensitivity, simple pre-treatment and low cost, this MISPE-HPLC method could provide a new tool for the rapid determination of multi-pesticide residues in the complicated food samples.

Study on an electrochromatography method based on organic-inorganic hybrid molecularly imprinted monolith for determination of trace trichlorfon in vegetables.

BACKGROUND: Organophosphorus pesticides have been widely used in agricultural production. However, the wide use of organophosphate also results in pesticide residues on the plant, which are harmful to human health because of their potential mutagenic and carcinogenic properties. Therefore, it is vital to develop a sensitive and effective analysis method to control pesticide residues. RESULTS: In this study, a novel molecularly imprinted capillary monolithic column was prepared using trichlorfon as the template molecule by combining non-hydrolytic sol-gel process with a molecular imprinting technique. The resulting material was characterized by scanning electron microscopy and Fourier transform infrared. Under capillary electrochromatography, the effects of voltage, pH, ACN content and concentration of buffer solution on the electro-osmotic flow (EOF) of imprinted capillary monolithic column were evaluated in detail. Using this prepared material as stationary phase for capillary electrochromatography, a novel method of molecularly imprinted capillary electrochromatography (MICEC) for the detection of trace trichlorfon residues in vegetables was developed. Under optimal conditions, appreciable sensitivity was achieved with a LOD (S/N = 3) of 92.5 µg kg(-1) and method quantitation limit (MQL) of 305.3 µg kg(-1), respectively. The linear ranges of the calibration graph were 0.1 µg L(-1) to 10 mg L(-1). The peak area precision (RSD) for five replicate extractions of 0.01 mg L(-1) trichlorfon standard aqueous solution was 4.5%. To evaluate the accuracy of this method, the blank cucumber and cauliflower samples spiked with trichlorfon were extracted and analyzed by this method with good recoveries, ranging from 80.2% to 95.8%. Moreover, this method was successfully applied to the quantitative detection of the trichlorfon residues in leek samples. CONCLUSION: With good properties of high sensitivity and simple pre-treatment, this MICEC method could provide a new tool for the rapid determination of trace trichlorfon residue in complex food samples.

Surface-enhanced Raman spectroscopy studies of organophosphorous model molecules and pesticides.

This work reports the analytical application of surface-enhanced Raman spectroscopy (SERS) in the trace analysis of organophosphorous pesticides (trichlorfon and glyphosate) and model organophosphorous compounds (dimethyl methylphosphonate and o-ethyl methylphosphonothioate) bearing different functional groups. SERS measurements were carried out using Ag nanocubes with an edge square dimension of ca. 100 nm as substrates. Density functional theory (DFT) with the B3LYP functional was used for the optimization of ground state geometries and simulation of Raman spectra of the organophosphorous compounds and their silver complexes. Adsorption geometries and marker bands were identified for each of the investigated compound. Results indicate the usefulness of SERS methodology for the sensitive analyses of organophosphorous compounds through the use of vibrational spectroscopy.

Study of an online molecularly imprinted solid phase extraction coupled to chemiluminescence sensor for the determination of trichlorfon in vegetables.

This study reports a new online molecularly imprinted solid phase extraction coupled to chemiluminescence for the determination of trichlorfon. This molecularly imprinted polymer (MIP) was prepared through bulk polymerization, in which methacrylic acid (MAA) was used as the functional monomer and ethylene glycol dimethacrylate (EGDMA) as the cross-linker. This novel functionalized material was characterized by FT-IR spectra and adsorption, and it exhibited good recognition and selective ability and fast adsorption-desorption dynamics toward trichlorfon. The factors affecting preconcentration of the analytes and sensitivity of the method are discussed in detail. Under the optimal condition, the linear range of the calibration graph was between 0.02 and 1.0 ng L(-1), and the detection limit was 1 × 10(3) ng L(-1). The blank cucumber samples spiked with trichlorfon at three levels were extracted and determined by the presented method with recoveries ranging from 83.5 to 94.5%, and the results were correlated well with those obtained using gas chromatography. Moreover, this developed method was successfully applied to the quantitative detection of trichlorfon residues in leek samples.

The dissipation rates of trichlorfon and its degradation product dichlorvos in cabbage and soil.

The residual levels and dissipation rate of trichlorfon, and its degradation product, dichlorvos, in cabbage crops and the soil in which these were grown, were determined by gas chromatography at two geographically distant experimental sites, one in Kunming and one in Beijing, China. Trichlorfon was applied at two dosages (900 g ai ha(-1) and 1350 g ai ha(-1)). Maximum final residues of trichlorfon in soil and cabbage were 1.23 mg kg(-1) and 1.81 mg kg(-1) respectively at Kunming, and 0.35 mg kg(-1) and 0.70 mg kg(-1) respectively at Beijing. However, the final residues of dichlorvos in both cabbage and soil was only 0.04 mg kg(-1) at Kunming, and only 0.03 mg kg(-1), or "not detectable", at Beijing. The mean half-life of trichlorfon in cabbage was 1.80 d with a dissipation rate of 90% over 5 d, while that in soil was 3.05 d with a dissipation rate of 90% over 14 d at one experimental site. The dissipation rates of trichlorfon and its degradation product dichlorvos at the two experimental sites were different, suggesting that degradation of these pesticides was affected by local soil characteristics and climate. When applied at both the recommended dosage and at 1.5 times this, no detectable residues of either trichlorfon or dichlorvos were found in soil or cabbage at harvest. Although trichlorfon can easily degrade into dichlorvos, which is highly toxic to humans and other animals, the observed low residual levels of dichlorvos suggest that trichlorfon is safe when applied at the recommended dosage.

Simultaneous determination of residues of trichlorfon and dichlorvos in animal tissues by LC-MS/MS.

Trichlorfon is a thermally unstable insecticide that can be easily decomposed to dichlorvos at high temperatures. In this study, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous determination of the residues of trichlorfon and dichlorvos in animal tissues. Trichlorfon and dichlorvos in animal tissues were extracted with dichloromethane, homogenized/shaken, concentrated, and determined by LC-MS/MS. The two insecticides can be effectively separated and individually determined. The limits of detection (LODs) of trichlorfon and dichlorvos were 0.04 and 0.07 microg kg(-1), respectively; the limits of quantification (LOQs) of the two insecticides were both 5 microg kg(-1). The average recoveries for three spiked blank samples at 10, 20 and 40 microg kg(-1) were in the range 85-106%, with relative standard deviations (RSDs) below 10.6%, which falls in a reasonable range for the analysis of the two insecticides in animal-derived foods. The method is fit-for-purpose for the simultaneous determination of residues of trichlorfon and dichlorvos in animal tissues.

High performance gold nanorods and silver nanocubes in surface-enhanced Raman spectroscopy of pesticides.

The behavior of Au nanorods and Ag nanocubes as analytical sensors was evaluated for three different classes of herbicides. The use of such anisotropic nanoparticles in surface-enhanced Raman scattering (SERS) experiments allows the one to obtain the spectrum of crystal violet dye in the single molecule regime, as well as the pesticides dichlorophenoxyacetic acid (2,4-D), trichlorfon and ametryn. Such metallic substrates show high SERS performance at low analyte concentrations making them adequate for use as analytical sensors. Density functional theory (DFT) calculations of the geometries and vibrational wavenumbers of the adsorbates in the presence of silver or gold atoms were used to elucidate the nature of adsorbate-nanostructure bonding in each case and support the enhancement patterns observed in each SERS spectrum.

Determination of organophosphorus pesticides by capillary electrophoresis-inductively coupled plasma mass spectrometry with collective sample-introduction technique.

A new method for the determination of organophosphorus pesticides using CE-ICP-MS with collective sample-introduction technique has been developed in this study. The method has been successfully used to separate and determine dimethoate, trichlorfon and glyphosate with an RSD of < 5% for migration times (n = 6) and < 4% for peak areas (n = 6). The experimental results showed that the collective sample-introduction considerably reduced the makeup volume and the dilution of analyte, and eventually resulted in a much lower detection limit and a much better electrophoretic resolution. The peak widths and the detection limits of dimethoate, trichlorfon and glyphosate obtained with this method are 15-17 s and 0.05-0.07 microg/mL (as compound), respectively. Using this method, we have successfully separated and determined dimethoate, trichlorfon and glyphosate in vegetable sample with a recovery of 90-96%.

Nucleic acid sensor for insecticide detection.

Nucleic acid sensor based on polyaniline (PANI) has been fabricated by covalently immobilizing double stranded calf thymus (dsCT) DNA onto perchlorate (ClO(-) (4))-doped PANI film deposited onto indium-tin-oxide (ITO) glass plate using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) chemistry. These dsCT-DNA-PANI-ClO(4)/ITO and PANI-ClO(4)/ITO electrodes have been characterized using square wave voltammetry, electrochemical impedance, scanning electron microscopy (SEM) and Fourier-transform-infrared (FTIR) measurements. This disposable dsCT-DNA-PANI-ClO(4)/ITO bioelectrode, stable for about 4 months, can be used to detect cypermethrin (0.005 ppm) and trichlorfon (0.01 ppm) in 30 and 60 s, respectively.