Development of Analytical Methods to Analyze Pesticide Residues.

Pesticides are compounds applied on crops to eliminate or control pests, diseases and weeds and it is known that their use provides unquestionable benefits in increasing agricultural production [...].

Impact of drinking water treatment processes on the residues of plant protection products for consumer and aquatic risk assessment: theoretical and experimental studies.

Pesticides residues can occur in ground and surface waters, and potentially react with chemicals used for water disinfection treatments, such as chlorine. This can lead to the formation of unknown reaction products, which can be more toxic and/or persistent than the active substances themselves, and therefore become a potential risk for human health and environment. Thus, in the framework of the EU Regulation 1107/2009, the identification of these by-products and their potential risk should be assessed. Within the European Food Risk Assessment (EU-FORA) Fellowship Programme, the fellow studied the behaviour of herbicides belonging to the families of imidazolinones and sulfonylureas in waters treated with chlorine disinfectants. Due to their physicochemical properties, these herbicides are susceptible of reaching natural waters. In fact, some of them have been detected in water monitoring programmes. During the experimental part of the present work programme, reactions between the active substances and the most used chlorine disinfecting reactants (hypochlorite and chloramines) were performed. Degradation kinetic parameters such as half-lives and degradation constants were calculated. Results showed that herbicide degradation was both pH and chlorine/chloramines concentration dependent. In order to identify the degradation by-products, high-resolution mass spectrometry experiments were performed, and a possible route of formation of these compounds was proposed. Finally, their risk assessment was carried out by using tox/ecotoxicological properties determined by QSAR methodology and FOCUS modelling for hazard and exposure assessment, respectively. These results will contribute to the definition of a risk assessment scheme for pesticides by-products potentially occurring in drinking water.

A study using QSAR/QSPR models focused on the possible occurrence and risk of alloxydim residues from chlorinated drinking water, according to the EU Regulation.

Any active substance with phytosanitary capacity intended to be marketed in Europe must pass exhaustive controls to assess its risk before being marketed and used in European agriculture. Since the implementation of Regulation (EC) No 1107/2009, agrochemical companies have been obliged to study the formation of pesticide transformation products (TPs) during the treatment of drinking water containing pesticide residues. However, there is no consensus on how to address this requirement. In this research work, the open literature collection on alloxydim was used to propose potential chlorination paths from alloxydim isomers. Furthermore, several QSAR/QSPR models have been used to fill the of knowledge gap relative to some key parameters in the physico-chemical, environmental and ecotoxicological areas of potential alloxydim TPs from chlorinated water for which little information exists. In this way, it has been possible to estimate the state of aggregation of these TPs (they exist mainly as liquids) as well as their ease of transit between the different phases, to predict their possible behaviour in the three environmental compartments (e.g., thermophysical properties point to a change in their evolution with respect to the parent alloxydim isomers) and to anticipate their potential risk to human and animal health (e.g., all of them cause developmental toxicity). These and other results highlight that the hazards of several TPs, i.e., both chlorinated and nonchlorinated from parent alloxydim or from those obtained after cleavage of the N - O bond and the subsequent reaction with chlorine, should be seriously considered. The obtained results reopen the debate on the implications of the use of QSAR/QSPR models for pesticide risk assessment in the legislative framework.

Analysis of Pesticide Residues by QuEChERS Method and LC-MS/MS for a New Extrapolation of Maximum Residue Levels in Persimmon Minor Crop.

According to EU guidance SANCO/7525/VI/95 Rev. 10.3, residue data extrapolation from a surrogate major crop to a minor crop can be used for setting maximum residue levels (MRLs) with a reduced number of residue trials and representative selected pesticides. In this work, a QuEChERS method (citrate-buffered version and PSA with MgSO4 clean-up) and LC-ESI-MS/MS for the determination of boscalid, pyraclostrobin, fludioxonil, fluopyram and tebuconazole in persimmon was developed and validated according to EU Commission guidelines and afterwards used for the determination of residues in four field trials. Residue levels at harvest for each pesticide ranged between 0.347 and 0.028 mg/kg. After comparing EFSA residue data on apples, as the surrogate major crop, and conducting a consumer risk assessment, a proposal of residue data extrapolation to set MRLs in persimmons was performed. The results showed that pesticide residues in persimmons at harvest were consistently lower than residues in apples when substances were applied according to the same critical GAP. MRLs were set at 0.5 mg/kg for fludioxonil, 0.6 mg/kg for boscalid, 0.3 mg/kg for tebuconazole, 0.4 mg/kg for fluopyran and 0.3 mg/kg for pyraclostrobin. The ratio of the MRLs for apple/persimmon varied between 2.5 for boscalid and 1.25 for fluopyram, suggesting that residue extrapolation can be feasible, promoting the process of pesticide registration for minor crops and the settlement of MRL.

Computational-Based Study of QuEChERS Extraction of Cyclohexanedione Herbicide Residues in Soil by Chemometric Modeling.

Assessment of two buffered QuEChERS (quick, easy, cheap, effective, rugged, and safe) versions (i.e., citrate and acetate) modified by including methanol to recover the residues of three cyclohexanedione oxime (CHD) herbicides and three of their byproducts from agricultural soil was performed. In this context, a full second-order face-centered factorial experimental design was developed to quantify the influences of the main five variables (i.e., extraction time, water content, soil weight, and extraction solvent volume and composition) on the target compound recoveries. The fitting equations satisfactorily described the extraction process behavior. The mathematical models also showed the most influencing independent variables (i.e., extraction solvent composition and soil weight). Handling simpler expressions was possible with the acetate QuEChERS but not with the citrate QuEChERS. The recoveries of the CHD residues were close to 100% after performing the extraction under suitable conditions. Furthermore, dispersive solid-phase extraction (dSPE) clean-up steps were assessed to reduce the matrix effect in mass spectrometry. In this sense, the citrate QuEChERS in combination with the PSA + C18 clean-up step was the best option for the extraction of CHD residues.

Considerations of nano-QSAR/QSPR models for nanopesticide risk assessment within the European legislative framework.

The European market for pesticides is currently legislated through the well-developed Regulation (EC) No. 1107/2009. This regulation promotes the competitiveness of European agriculture, recognizing the necessity of safe pesticides for human and animal health and the environment to protect crops against pests, diseases and weeds. In this sense, nanotechnology can provide a tremendous opportunity to achieve a more rational use of pesticides. However, the lack of information regarding nanopesticides and their fate and behavior in the environment and their effects on human and animal health is inhibiting rapid nanopesticide incorporation into European Union agriculture. This review analyzes the recent state of knowledge on nanopesticide risk assessment, highlighting the challenges that need to be overcame to accelerate the arrival of these new tools for plant protection to European agricultural professionals. Novel nano-Quantitative Structure-Activity/Structure-Property Relationship (nano-QSAR/QSPR) tools for risk assessment are analyzed, including modeling methods and validation procedures towards the potential of these computational instruments to meet the current requirements for authorization of nanoformulations. Future trends on these issues, of pressing importance within the context of the current European pesticide legislative framework, are also discussed. Standard protocols to make high-quality and well-described datasets for the series of related but differently sized nanoparticles/nanopesticides are required.

Assessing the Effects of Alloxydim Phototransformation Products by QSAR Models and a Phytotoxicity Study.

Once applied, an herbicide first makes contact with leaves and soil. It is known that photolysis can be one of the most important processes of dissipation of herbicides in the field. However, degradation does not guarantee detoxification and can give rise to byproducts that could be more toxic and/or persistent than the active substance. In this work, the photodegradation of alloxydim herbicide in soil and leaf cuticle surrogates was studied and a detailed study on the phytotoxicity of the main byproduct on sugar beet, tomato, and rotational crops was performed. Quantitative structure⁻activity relationship (QSAR) models were used to obtain a first approximation of the possible ecotoxicological and environmental implications of the alloxydim and its degradation product. The results show that alloxydim is rapidly degraded on carnauba and sandy loam soil surfaces, two difficult matrices to analyze and not previously studied with alloxydim. Two transformation products that formed in both matrices were identified: alloxydim Z-isomer and imine derivative (mixture of two tautomers). The phytotoxicity of alloxydim and the major byproduct shows that tomato possesses high sensitivity to the imine byproduct, while wheat crops are inhibited by the parent compound. This paper demonstrates the need to further investigate the behavior of herbicide degradation products on target and nontarget species to determine the adequate use of herbicidal products to maximize productivity in the context of sustainable agriculture.

Computational Study of the Structure and Degradation Products of Alloxydim Herbicide.

Density functional theory calculations allowed us to study alloxydim herbicide and to identify the most stable conformers, the factors that governs their stability, and the interconversion mechanisms among the most relevant conformers. The degradation chain involves, as a first step, the cleavage of the N-O bond and the formation of a stable intermediate difficult to characterize experimentally. The study performed also allowed us to identify the properties of this elusive intermediate and to determine that the dominant fragmentation process in the gas phase is the homolytic fragmentation. Stability of alloxydim conformers and homolytic fragments were also assessed in the water phase. Computed IR spectra were consistent with those observed experimentally.

Photolysis of clethodim herbicide and a formulation in aquatic environments: Fate and ecotoxicity assessment of photoproducts by QSAR models.

The photochemical fate of the herbicide clethodim in natural waters was investigated under simulated and natural sunlight radiation. This herbicide exhibited a rapid degradation rate in simulated aquatic environment with half-lives ranged from 27.9min to 4.6h. The commercial formulation of clethodim showed a faster degradation with half-lives from 19.3min to 1.4h. It has also been demonstrated that the photolytic behavior of clethodim was affected by the water composition and the radiation intensity. Nine major photoproducts were identified and their distribution was dependent on the experimental conditions. Photodegraded solutions of clethodim were shown to be more toxic to the bacteria Vibrio fischeri than the herbicide itself, reaching the maximum toxicity when the herbicide is completely degraded. QSAR analysis of the fate, ecotoxicological and physicochemical endpoints of the degradation products provided positive alerts for several identified by-products. Environmental fate and transport estimates showed that all photoproducts, unlike the active substance, are potential leachers. Moreover, predicted vapor pressures suggested that dermal contact and ingestion are the most probable exposure routes for workers and general population to both clethodim and its photoproducts. These results highlight the importance of the degradation products in attaining a complete knowledge of the fate and behavior of an herbicide in the environment. To our knowledge, this is the first study to report a detailed QSAR study on clethodim photoproducts under environmental conditions. These results provide a very valuable information that will guide further experimental studies leading to a better pesticide risk assessment.

Quantum chemistry in environmental pesticide risk assessment.

The scientific community and regulatory bodies worldwide, currently promote the development of non-experimental tests that produce reliable data for pesticide risk assessment. The use of standard quantum chemistry methods could allow the development of tools to perform a first screening of compounds to be considered for the experimental studies, improving the risk assessment. This fact results in a better distribution of resources and in better planning, allowing a more exhaustive study of the pesticides and their metabolic products. The current paper explores the potential of quantum chemistry in modelling toxicity and environmental behaviour of pesticides and their by-products by using electronic descriptors obtained computationally. Quantum chemistry has potential to estimate the physico-chemical properties of pesticides, including certain chemical reaction mechanisms and their degradation pathways, allowing modelling of the environmental behaviour of both pesticides and their by-products. In this sense, theoretical methods can contribute to performing a more focused risk assessment of pesticides used in the market, and may lead to higher quality and safer agricultural products. © 2017 Society of Chemical Industry.

Photodegradation behaviour of sethoxydim and its comercial formulation Poast® under environmentally-relevant conditions in aqueous media. Study of photoproducts and their toxicity.

Photolysis is an important route for the abiotic degradation of many pesticides. However, the knowledge of the photolytic behaviour of these compounds and their commercial formulations under environmentally-relevant conditions are limited. The present study investigated the importance of photochemical processes on the persistence and fate of the herbicide sethoxydim and its commercial formulation Poast® in aqueous media. Moreover, the effect of important natural water substances (nitrate, calcium, and ferric ions) on the photolysis of the herbicide was also studied. The results showed that additives existing in the commercial formulation Poast® accelerated the rate of photolysis of sethoxydim by a factor of 3. On the contrary, the presence of nitrate and calcium ions had no effect on the photodegradation rate while ferric ions resulted in an important decrease in the half-life of sethoxydim possibly due to the formation of a complex. Different transformation products were identified in the course of sethoxydim irradiation and the effect of experimental conditions on their concentrations was investigated. Finally, Microtox® test revealed that aqueous solutions of sethoxydim photoproducts increased the toxicity to the bacteria Vibrio fischeri.

Development of a rapid and direct method for the determination of organic acids in peach fruit using LC-ESI-MS.

An accurate, simple and rapid liquid chromatography mass spectrometry method for the determination of organic acids in peach fruit has been developed. Direct injection and sample clean-up with a mixed-mode sorbent was compared. The best results for the determination of gluconic, oxalic, malic, citric and fumaric acids were obtained with only a simple dilution and filtration step, and nylon filters should be avoided since some organic acids are retained by them. It is the first time that gluconic acid has been determined in peach fruit. Different parameters involved in the separation and detection process have been optimized. Since matrix effects were observed in the peach commodity, organic acids were quantified by the standard addition method. All validation parameters of the method were found acceptable of all organic acids. Finally, the method was successfully applied to the analysis of samples of peach from two cultivars.

Biopesticides in the framework of the European Pesticide Regulation (EC) No. 1107/2009.

BACKGROUND: The European Pesticide Regulation (EC) No. 1107/2009 encourages the use of less harmful active substances. Two main concerns involve the application of cut-off criteria for pesticides without losing tools for future agriculture (especially for minor uses) and the implementation of zonal evaluations. Biopesticides are considered to have lower risks than synthetic pesticides; consequently, there is strong interest for their use in integrated pest management practices. RESULTS: This paper provides an analysis of the current European situation, starting with the first attempts to regulate the use of plant protection products and focusing on the implications of the new legislative criteria for biopesticides. CONCLUSION: It is important to be aware that biopesticides are still pesticides and fall under the same regulations as their synthetic counterparts. Although manufacturers are still reluctant to commit to such alternatives due to difficulties with approval and registration, biopesticides could be alternatives for traditional plant protection products, either as a base for the synthesis of new products or integrated with traditional plant protection products. In addition, biopesticides have to be used only as indicated on the label, which provides critical information about how to safely handle and use plant protection products.

Smile esthetics from odontology students' perspectives.

OBJECTIVES: To analyze the perception of smile esthetics and its alterations in dental degree students; to determine whether there are differences in that perception among students in different study years on those courses and between genders; and to determine if the circumstance of having received prior orthodontic treatment could influence that perception. MATERIAL AND METHODS: Students (n = 192) in different study years of the dental degree course at the University of Valencia, Spain, analyzed two photographs of a patient in which, by means of computer software, midline diastema, upper and lower midlines, crown length of the maxillary right central incisor, occlusal cant, and "gummy" smile were altered. Students assessed the photographs on a scale from 1 to 10. Statistical analyses for assessing each group's level of perception were carried out. RESULTS: After checking the validity of the study, it was observed that the students' ability to detect alterations in smile esthetics did not improve over their degree courses, given that the differences do not present a linear development. There were no differences between genders and between those who had or had not undergone an orthodontic treatment. CONCLUSIONS: There are no statistically significant differences between the results of students in different study years or between genders. The circumstance of having undergone prior orthodontic treatment is not a determining factor in the ability to perceive such anomalies.

Aqueous photodegradation of sethoxydim herbicide: Qtof elucidation of its by-products, mechanism and degradation pathway.

The photochemical fate of sethoxydim herbicide in water was studied under simulated sunlight radiation (Suntest apparatus). This compound exhibited a rapid degradation rate in water with a half-life of approximately 1h, which is in accordance with the high calculated value of the quantum yield (Φ=0.26). Consequently, the photolysis of sethoxydim should be considered as an efficient route of sethoxydim removal in aqueous media. During the photodegradation studies, ten by-products were detected, and their identification was accomplished using high-performance liquid chromatography (HPLC) coupled with quadrupole time-of-flight mass spectrometry (Qtof) and electrospray ionization in positive mode (ESI+). A detailed and exhaustive study of the mass spectra of the precursor ions and their MS/MS fragmentation patterns allowed for their identification. The photodegradation products resulted from the cleavage and isomerization of the NO bond of the oxime, oxidation of the sulfur atom, oxidative CS bond cleavage and Beckmann rearrangement followed by intramolecular cyclization. Additionally, the reactive species involved and a plausible mechanism for the photodegradation of sethoxydim in water are discussed. Finally, based on the identified products, a rational pathway for the photodegradation of sethoxydim in water is proposed.

High chlorogenic and neochlorogenic acid levels in immature peaches reduce Monilinia laxa infection by interfering with fungal melanin biosynthesis.

Chlorogenic acid (CGA) and its isomer, neochlorogenic acid (NCGA), were found to be the major phenolic compounds in the flesh and peel of three peach cultivars. Their concentrations are especially high in immature fruits (CGA, 151-548 mg/kg; NCGA, 85-380 mg/kg), whose resistance to the brown rot fungus, Monilinia laxa , is very high. The concentrations of these two phenolic compounds decline in maturing fruits (CGA, 77-181 mg/kg; NCGA, 30-82 mg/kg), and this decline is associated with a concomitant increase in susceptibility to brown rot infection. Other phenolic compounds found in the same HPLC chromatograms at 340 nm from each peach extract at varying sampling dates in each of the three peach cultivars were not correlated with the incidence of brown rot and appeared only in some cultivars. The incidence of brown rot for each cultivar at each sampling date was significantly negatively correlated with the NCGA (r > -0.85) and CGA (r > -0.90) contents. At concentrations that are similar to those in peach fruit, CGA does not inhibit spore germination or mycelial growth of M. laxa in culture but markedly inhibits the production of melanin-like pigments in the mycelia of M. laxa in culture (42% melanin reduction). Accordingly, we propose that the high concentrations of CGA and NGA in immature fruits might contribute to their reduced susceptibility or increased resistance to brown rot infection by interfering with fungal melanin production.

Indirect photodegradation of clethodim in aqueous media. byproduct identification by quadrupole time-of-flight mass spectrometry.

Aqueous photolysis of clethodim herbicide in the presence of natural substances such as humic acids (HA), nitrate, and Fe(III) ions has been investigated. The photodegradation rate of clethodim was retarded in the presence of HA compared to ultrapure water, while nitrate ions had no effect. On the other hand, water containing different concentrations of Fe(III) ions enhanced degradation of this herbicide. Clethodim transformation gave rise to the formation of nine byproducts, some of them, to the best of our knowledge, described for the first time in this work. The identification of these photoproducts has been accomplished by coupling liquid chromatography to quadrupole time-of-flight mass spectrometry. The main transformation reactions observed for clethodim were photoisomerization to the Z-isomer, S-oxidation of E- and Z-clethodim isomers giving rise to sulfoxide diastereoisomers, reduction of the oxime moiety to yield clethodim imine, oxidative cleavage of the C-S bond, and S-oxidation of clethodim imine leading to the formation of imine ketone and imine sulfoxide.

Analysis of competing risks of causes of death and their variation over different time periods in Hodgkin's disease.

PURPOSE: Hodgkin's disease is considered a model of curable illness. However, long-term studies show excessive mortality in relation to the general population. We studied the various causes of death by use of competing risks and their evolution over the years. EXPERIMENTAL DESIGN: All patients diagnosed with Hodgkin's disease at our institution between 1967 and 2003 were included. The competing risks of causes of death and their vital situation were examined in three time periods: cohort A with patients treated before 1980, cohort B with patients treated from 1981 to 1986, and cohort C with patients treated from 1986 onwards. RESULTS: We studied 534 patients, with a median follow-up time of 9.1 years for the whole cohort. The 5-year, 15-year, and 20-year Kaplan-Meier survival estimates for all patients were 81%, 72%, and 65%, respectively. At the close of the study, 337 (63.1%) were alive and 170 (31.8%) patients had died. The most common cause of death was the progression of Hodgkin's disease, followed by deaths due to a second tumor. Survival was significantly worse in the first period than in the other two (P < 0.001), and in the three periods, the main cause of death was tumor progression. CONCLUSIONS: The progression of Hodgkin's disease is the main cause of death. Over time, a reduction in death related to infection and the acute toxicity of treatment was seen. A lot of patients still die for reasons linked to delayed side effects of radiotherapy, such as second tumors and heart disease, which is important to plan preventive activities and clinical research.