Residual estimation of spirotetramat and its metabolites in chilli and soil by LC-MS/MS.

Two applications of spirotetramat were done to study the dissipation and persistence of spirotetramat and its four different metabolites in chilli and soil at 10 days interval. Total spirotetramat residues were estimated by LC-MS/MS instrument. The mean initial deposits of total spirotetramat after application of spirotetramat 15.31 OD @ 60 (X dose), 75 (1.25 × dose) and 120 (2 × dose) g a.i. ha-1 on green chilli were found to vary from 0.38 to 0.83 mg kg-1 during the initial year. Spirotetramat and its metabolite residues in green chilli were found to be below limit of quantification (0.01 mg kg-1) after 15 days of application. The spirotetramat cis enol (the major metabolite) was formed in both the soil and the plant. The residues of spirotetramat-monohydroxy were below LOQ irrespective of any substrate during the estimation. In soil, the total initial spirotetramat deposits for the 1st year were found 0.09 for X dose, 0.12 for 1.25 × dose and 0.20 mg kg-1 for 2 × dose. After 3 days for both X and 1.25 × doses and 5 days for 2 × dose, the total spirotetramat residues were below LOQ. The spirotetramat's half-life values have been determined to be between 3.19 and 3.93 days and 1.00 and 1.59 days, respectively, in soil and green chilli fruits. One day waiting period is proposed for the safe consumption of green chilli when the spirotetramat was applied irrespective of the dose.

Quantification of pendimethalin residue in green fodder and silage of winter cereals using gas chromatography-tandem mass spectrometry.

Use of agro-chemicals in forage crops is restricted due to the fear of direct toxicity to livestock and risk of pesticide residue accumulation in the food chain. Wheat and barley can be used as green fodder and silage, and herbicide residue estimation in green fodder and silage is important for ensuring the safety of dairy cattle. A field experiment was conducted for two years to study pendimethalin residues in the green fodder and silage of wheat and barley. In both cereal crops, pendimethalin (1.125 kg a.i./ha) was applied as pre-emergence along with an unsprayed control. Pendimethalin residues in fodder, silage, and soil were estimated using gas chromatography-tandem mass spectrometry (GC-MS/MS). At harvest, pendimethalin residues in fodder and silage of wheat and barley were below the limit of quantification (<0.01 mg/kg) during both crop seasons. Pendimethalin can be safely used for weed control in winter cereals grown for fodder and silage.

Dissipation kinetic study of fenazaquin in chilli and soil by LC-MS/MS.

Dissipation and persistence of fenazaquin residues in chilli and soil were studied for 2 years following two applications of fenazaquin at 10-day interval. The limit of detection and limit of quantification were 0.003 and 0.01 mg kg-1. The mean initial deposits of fenazaquin 10 EC on green chilli fruits were found to be 0.74, 1.17, and 1.79 mg kg-1 after the application @ 125 (X dose), 156.25 (1.25X dose), and 250 (2X dose) g a.i. ha-1, respectively, during the first year followed by 0.78, 1.20, and 1.70 mg kg-1, respectively, during the next year. The mean initial deposits in soil were found to be 0.18, 0.25, and 0.44 mg kg-1 for the X, 1.25X, and 2X doses, respectively, during the first year and 0.19, 0.22, and 0.39 mg kg-1, respectively, during second year. The residues of fenazaquin in green chilli dissipated above 96% at 20 days in the three different doses whereas in red chilli, the residues were present on 25 days and at maturity, residues were below the limit of quantification (LOQ). In soil, the residues dissipated below the LOQ at 15 days for X and 1.25X while 20 days for the 2X dose, respectively. The half-life values in green chilli fruits and soil for the fenazaquin were found to be in the range of 3.22-3.93 days and 2.41-3.35 days, respectively. The waiting period was calculated to be 3, 5, and 8 days for green chilli after the application of fenazaquin at 25, 156.25, and 250 g a.i. ha-1, respectively.

Validation of simple and efficient analytical methodology for estimation of forchlorfenuron residues.

A simple and efficient analytical methodology was standardised for the estimation of forchlorfenuron residues in rice leaves, grains, husk, straw and soil using LC-MS/MS. The methodology was validated in terms selectivity, sensitivity, linearity, limit of detection, limit of quantification, matrix effect, repeatability and reproducibility. The limit of detection and limit of quantification of forchlorfenuron in rice leaves, grains, straw, husk and soil were worked out to be 0.003 and 0.01 mg kg-1, respectively. Recoveries ranged between 81.54 and 99.05% with RSDr below 4.57 and RSDR below 4.19%. This validated method was applied for the determination of forchlorfenuron residues in paddy leaves after foliar application. The residues persisted for 0, 3 and 7 days after application of forchlorfenuron 0.12% EC @ 1.25, 2.50 and 5.00 ppm a.i. The half-life values were estimated to be 1.91 and 2.11 days for 2.50 and 5.00 ppm a.i. concentration. The residues were found to be below the limit of quantification of 0.01 mg kg-1 in straw, grain, husk and soil samples collected at harvest time.

Dissipation and risk assessment of fluopyram and trifloxystrobin on onion by GC-MS/MS.

The dissipation and risk assessment studies on fluopyram, trifloxystrobin and their metabolites were carried out on onion under field conditions after two treatments of fluopyram 250 g/L + trifloxystrobin 250 g/L SC @ 150 and 300 g a.i. ha-1. The onion bulb samples were collected at 0, 3, 7, 14, and 21 days after second spray to study the pattern of dissipation using QuEChERS methodology for processing and analysis on GC-MS/MS. The total initial residues of fluopyram (fluopyram + fluopyram benzamide) in immature onion bulb were 2.14 and 4.93 mg kg-1, at single and double dose, respectively. The residues of 0.02 and 0.06 mg kg-1 persisted in the mature onion bulb collected at the harvest (30 days after treatment). The total initial residues of trifloxystrobin (trifloxystrobin + CGA 321113) in immature onion bulb were 0.65 and 1.97 mg kg-1, at single and double dose, respectively, which reached < LOQ and 0.06 mg kg-1 at the respective doses at the harvest time. Dissipation of fluopyram followed second-order kinetics with DT50 values of 1.83 and 1.74 days, whereas trifloxystrobin followed first-order kinetics with DT50 values of 4.73 and 4.78 days, at single and double dose respectively. Risk assessment in terms of hazard quotient was done to estimate the risk that can occur due to application of this combination pesticide. It was observed that even the spray at the double recommended dose could not have dietary risks on the consumers.

Nanoscale organization of Nicastrin, the substrate receptor of the γ-secretase complex, as independent molecular domains.

Alterations in the canonical processing of Amyloid Precursor Protein generate proteoforms that contribute to the onset of Alzheimer's Disease. Modified composition of γ-secretase or mutations in its subunits has been directly linked to altered generation of Amyloid beta. Despite biochemical evidence about the role of γ-secretase in the generation of APP, the molecular origin of how spatial heterogeneity in the generation of proteoforms arises is not well understood. Here, we evaluated the localization of Nicastrin, a γ-secretase subunit, at nanometer sized functional zones of the synapse. With the help of super resolution microscopy, we confirm that Nicastrin is organized into nanodomains of high molecular density within an excitatory synapse. A similar nanoorganization was also observed for APP and the catalytic subunit of γ-secretase, Presenilin 1, that were discretely associated with Nicastrin nanodomains. Though Nicastrin is a functional subunit of γ-secretase, the Nicastrin and Presenilin1 nanodomains were either colocalized or localized independent of each other. The Nicastrin and Presenilin domains highlight a potential independent regulation of these molecules different from their canonical secretase function. The collisions between secretases and substrate molecules decide the probability and rate of product formation for transmembrane proteolysis. Our observations of secretase nanodomains indicate a spatial difference in the confinement of substrate and secretases, affecting the local probability of product formation by increasing their molecular availability, resulting in differential generation of proteoforms even within single synapses.

Validation of QuEChERS Method Coupled with LC-MS/MS for Determination of Thiamethoxam and its Metabolites in Wheat and Soil.

BACKGROUND: Thiamethoxam, a neonicotinoid insecticide, has been widely accepted for the control of sucking and certain chewing pests. Through different routes, i.e., wind drift, leaching, and surface runoff, it can reach non-target areas, which include humans as well. Therefore the fate of thiamethoxam in food grains and soil is of prime importance, entailing a need for pesticide use to be subject to steady observation. OBJECTIVE: The review aimed to validate the Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) method for the detection of thiamethoxam and its metabolites in wheat leaves, grain, straw, and soil using liquid chromatography-tandem mass spectrometry (LC-MS/MS). METHOD: The QuEChERS method allows sample extraction by acetonitrile followed by cleanup with a primary secondary amine sorbent, magnesium sulphate, and graphitized carbon black. The method was validated in terms of selectivity, linearity, LOD, LOQ, matrix match, accuracy, and precision. The estimation of residues was done by LC-MS/MS. RESULTS: The recovery of thiamethoxam and its metabolites in wheat and soil samples was in the range of 81.22-98.14%. The LOQ for thiamethoxam and its metabolites was 0.01 mg/kg. The matrix effect values were in the acceptable range of -20 to +20%. CONCLUSIONS: The method allows a simple and fast extraction of thiamethoxam and its metabolites from wheat leaves, grain, straw, and soil. It could be helpful in the evaluation of regulatory and food safety decisions regarding the use of neonicotinoids. HIGHLIGHTS: The method was validated according to the European Commission SANTE/12682/2019 guidelines. An accurate, simple, and sensitive analytical method was validated for thiamethoxam and its metabolites.

Bioremediation of organophosphorus pesticide phorate in soil by microbial consortia.

Microbial consortia isolated from aged phorate contaminated soil were used to degrade phorate. The consortia of three microorganisms (Brevibacterium frigoritolerans, Bacillus aerophilus and Pseudomonas fulva) could degrade phorate, and the highest phorate removal (between 97.65 and 98.31%) was found in soils inoculated with mixed cultures of all the three bacterial species. However, the mixed activity of any of two of these bacteria was lower than mixed consortia of all the three bacterial species. The highest degradation by individual mixed consortia of (B. frigoritolerans+B.aerophilus, B. aerophilus+P. fulva and B. frigoritolerans+P. fulva) appeared in soil between (92.28-94.09%, 95.45-97.15% and 94.08-97.42%, respectively). Therefore, inoculation of highly potential microbial consortia isolated from in situ contaminated soil could result in most effective bioremediation consortia for significantly relieving soils from phorate residues. This much high phorate remediation from phorate contaminated soils have never been reported earlier by mixed culture of native soil bacterial isolates.

Bioefficacy and persistence of acephate in mungbean Vigna radiata (L.) Wilczek.

The bioefficacy, persistence and metabolism of foliar application of acephate 75 SP at the respective recommended and double doses of 1500 and 3000 g a.i. ha(-1) were studied on kharif mungbean crop. Acephate gave a very effective control of the pod borer, Helicoverpa armigera (Hubner) at the tested doses on mungbean crop throughout the cropping season, besides recording lower percent pod damage and higher grain yield. The recommended dose of acephate also recorded higher net returns and thus indicating its superiority. Persistence of acephate in mungbean leaves and soil was studied following applications of acephate @ 1500 and 3000 g a.i. ha(-1) at 30 days after sowing. Residues of acephate in mungbean leaves and soil were estimated using gas liquid chromatograph (GLC) and confirmed on gas chromatograph-mass spectrometry (GC-MS). Half-life periods for acephate in mungbean leaves were observed to be 2.98 and 3.27 days at single and double the application rates, respectively. Residues of acephate dissipated below its limit of quantification (LOQ) of 0.05 mg kg(-1) after 20 and 25 days at single and double the application dosage, respectively. Similarly, half-life periods for acephate in mungbean soil were observed to be 1.86 and 1.94 days at single and double the application rates, respectively. Residues of acephate dissipated below its LOQ of 0.05 mg kg(-1) after 10 and 15 days at single and double the application dosage, respectively.

Estimation and risk assessment of flubendiamide on fodder berseem clover (Trifolium alexandrinum L.) by QuEChERS methodology and LC-MS/MS.

The residues of flubendiamide from berseem were extracted and cleaned up by the QuEChERS approach and analysed by liquid chromatography/mass spectrometry (LC-MS/MS). The dissipation studies on berseem were carried out by application of flubendiamide at five different dosages i.e. 24, 36, 48, 72 and 96 g a.i. ha(-1). The residues reached below determination limit (BDL) of 0.01 mg kg(-1) in 7, 10, 10, 15 and 15 days for 24, 36, 48, 72 and 96 g.a.i. ha(-1)dosages, respectively. Half-life (t1/2) of flubendiamide on berseem was observed to be 1.08, 1.88, 1.94, 2.26 and 2.27 days, respectively, at 24, 36, 48, 72 and 96 g a.i. ha(-1). Theoretical maximum residue contributions (TMRC) values reached below the maximum permissible limit (MPI) for 3, 5 and 7 days in treatments at 24, 36 and 48 g.a.i. ha(-1), respectively, whereas in case of higher dose treatments i.e. 72 and 96 g a.i. ha(-1), the residues of flubendiamide reached below MPI after 10 days of treatment. These data could provide guidance for the proper and safe use of this pesticide on berseem in India.

Dissipation pattern and risk assessment studies of triazophos residues on capsicum (Capsicum annuum L.) using GLC-FPD and GC-MS.

The present study was carried out to observe the dissipation pattern of triazophos on capsicum and risk assessment of its residues on human beings and to suggest a waiting period for the safety of consumers. Following two applications of triazophos (Truzo 40 EC) at 500 and 1000 g a.i. ha(-1), the average initial deposits were found to be 3.61 and 6.26 mg kg(-1), respectively. These residues dissipated below the limit of quantification (LOQ) of 0.05 mg kg(-1) in 10 and 15 days at the recommended and double the recommended dosages, respectively. The calculated values of half-life were 2.31 and 2.14 days at recommended and double the recommended dosages, respectively. Theoretical maximum residue contribution (TMRC) values were found to be 28.8 and 41.6 µg person(-1) day(-1) at 500 and 1000 g a.i. ha(-1), respectively, and found to be below the maximum permissible intake on capsicum fruit on the 7th day. Therefore, a waiting period of 7 days is suggested for consumption of capsicum sprayed with triazophos at the recommended dosages.

Estimation and Validation of Propanil Residues in Rice and Soil Samples by Gas Liquid Chromatography with Electron Capture Detector.

The purpose of this analysis was to develop an efficient analysis for the estimation of residues of propanil in rice grain, husk, straw and soil samples. A liquid-liquid partitioning method was used for the extraction of propanil residues from the rice grains and its contents. For this, representative 10 g samples of blended rice grain, husk, straw and soil were soaked in acetone for 24 h, and the contents then partitioned two times into 100 and 50 mL dichloromethane and one time with 100 mL hexane. The combined organic layers were collected and were concentrated to 10 mL acetone using a rotary vacuum evaporator at 40°C. The extract was then subjected to cleanup by dispersive solid phase extraction. The final extract was injected onto a GLC column, where the propanil residues were determined by electron capture detector. The percentage recoveries were ranged from 84.9 % to 98.3 % when samples were spiked at the levels of 0.05, 0.25 and 0.50 mg/kg. The limits of quantification and detection were 0.05 and 0.017 mg/kg, respectively.

Isolation and evaluation of potent Pseudomonas species for bioremediation of phorate in amended soil.

Use of phorate as a broad spectrum pesticide in agricultural crops is finding disfavor due to persistence of both the principal compound as well as its toxic residues in soil. Three phorate utilizing bacterial species (Pseudomonas sp. strain Imbl 4.3, Pseudomonas sp. strain Imbl 5.1, Pseudomonas sp. strain Imbl 5.2) were isolated from field soils. Comparative phorate degradation analysis of these species in liquid cultures identified Pseudomonas sp. strain Imbl 5.1 to cause complete metabolization of phorate during seven days as compared to the other two species in 13 days. In soils amended with phorate at different levels (100, 200, 300 mg kg(-1) soil), Pseudomonas sp. strain Imbl 5.1 resulted in active metabolization of phorate by between 94.66% and 95.62% establishing the same to be a potent bacterium for significantly relieving soil from phorate residues. Metabolization of phorate to these phorate residues did not follow the first order kinetics. This study proves that Pseudomonas sp. strain Imbl 5.1 has huge potential for active bioremediation of phorate both in liquid cultures and agricultural soils.

Brevibacterium frigoritolerans as a Novel Organism for the Bioremediation of Phorate.

Phorate, an organophosphorus insecticide, has been found effective for the control of various insect pests. However, it is an extremely hazardous insecticide and causes a potential threat to ecosystem. Bioremediation is a promising approach to degrade the pesticide from the soil. The screening of soil from sugarcane fields resulted in identification of Brevibacterium frigoritolerans, a microorganism with potential for phorate bioremediation was determined. B. frigoritolerans strain Imbl 2.1 resulted in the active metabolization of phorate by between 89.81% and 92.32% from soils amended with phorate at different levels (100, 200, 300 mg kg(-1) soil). But in case of control soil, 33.76%-40.92% degradation were observed. Among metabolites, sulfone was found as the main metabolite followed by sulfoxide. Total phorate residues were not found to follow the first order kinetics. This demonstrated that B. frigoritolerans has potential for bioremediation of phorate both in liquid cultures and agricultural soils.

Dissipation pattern and risk assessment of flubendiamide on chili at different agro-climatic conditions in India.

Dissipation pattern and risk assessment of flubendiamide and its metabolite (desiodo flubendiamide) on chili were studied at four different agro-climatic locations of India at the standard and double dose at 60 and 120 g a.i. ha(-1) at 10 days interval. Quantification of residues was done on a high-performance liquid chromatograph (HPLC) with a photo diode array detector. The limit of quantification (LOQ) of this method was found to be 0.01 mg kg(-1) while limit of detection (LOD) being 0.003 mg kg(-1). Residues of flubendiamide were found to be below the determination limit in 15 days at both the dosages in all locations. Half-life of flubendiamide when applied at 60 and 120 g a.i. ha(-1) ranged from 0.85 to 1.80 and from 0.95 to 2.79 days, respectively. On the basis of data generated under the All India Network Project on Pesticide Residues, a preharvest interval (PHI) of 1 day has been recommended and the flubendiamide 480 SC has been registered for use on chili in India by the Central Insecticide Board and Registration Committee, Ministry of Agriculture, Government of India. The maximum residue limit (MRL) of flubendiamide on chili has been fixed by the Food Safety Standard Authority of India, Ministry of Health and Family Welfare, Government of India, as 0.02 µg g(-1) after its risk assessment.

Analytical method for determination of fipronil and its metabolites in vegetables using the QuEChERS method and gas chromatography/mass spectrometry.

An easy, simple, and efficient analytical method was validated for the determination of residues of fipronil and its metabolites in different vegetables comprising brinjal, cabbage, capsicum, cauliflower, okra, and tomato. The Quick, Easy, Cheap, Rugged and Safe method was used for the extraction and cleanup of fipronil and its metabolites on these vegetables. Final clear extracts of ethyl acetate were concentrated under vacuum and reconstituted into acetone, and residues were determined by GC/MS with an Rtx-5 capillary column. The retention times of desulfinyl, sulfide, fipronil, sulfone, and amide were 11.8, 15.8, 16.4, 19.7, and 24.1 min, respectively. The average recoveries obtained for fipronil and its metabolites at different spiking levels were determined to be above 85% in all samples. The repeatability and reproducibility in different matrixes ranged from 1.36 to 5.42 and 1.75 to 4.53%, respectively, for fipronil and its metabolites. The LOQ of this method was 0.01 mg/kg and LOD was 0.003 mg/kg.

Effect of dehydrogenase, phosphatase and urease activity in cotton soil after applying thiamethoxam as seed treatment.

Soil enzymes are indicators of microbial activities in soil and are often considered as an indicator of soil health and fertility. They are very sensitive to the agricultural practices, pH of the soil, nutrients, inhibitors and weather conditions. To understand the effect of an insecticide, thiamethoxam, on different soil enzyme activities, the experiments were conducted at cotton experimental fields of Punjab Agricultural University, Ludhiana. The results here were presented to understand the impact of thiamethoxam on soil enzyme activities. Thiamethoxam was applied as seed treatment to control the pest. Soil from three localities, i.e. soil in which seed was treated with recommended dose at 2.1 g a.i. kg(-1), soil in which seed was treated with four times recommended dose at 8.4 g a.i. kg(-1) and from the control field, were tested for different enzyme activities. Phosphatase and dehydrogenase activities were high in control soil in comparison to control soil while no effect of this insecticide on urease activity. Thiamethoxam had inhibitory effects on dehydrogenase and phosphatase activities. Therefore, it can be attributed that agricultural practices, weather conditions and use of thiamethoxam might be responsible for the different level of enzyme activities in soil.

Thiamethoxam degradation by Pseudomonas and Bacillus strains isolated from agricultural soils.

Twelve bacterial species were evaluated to know the degradation pattern of thiamethoxam in liquid medium. All the bacterial species could actively degrade phorate in a mineral salt medium containing phorate (50 µg ml(-1)) as sole carbon source. As these species have ability to degrade, we used these for the degradation of thiamethoxam--a neonicoitinoids. Screening of 12 active phorate-metabolizing bacterial species resulted in selection of Bacillus aeromonas strain IMBL 4.1 and Pseudomonas putida strain IMBL 5.2 causing 45.28 and 38.23 % thiamethoxam (50 µg ml(-1)) reduction, respectively, in 15 days as potential thiamethoxam degrading species. These two bacterial species grew optimally at 37 °C under shake culture conditions in MSMT medium raised with initial pH of 6.0-6.5 and use of these optimum cultural conditions resulted in improved thiamethoxam degradation by these bacterial species. These species caused maximum thiamethoxam degradation only in the presence of thiamethoxam as sole source of carbon and energy and the same was reduced in the presence of easily metabolize able carbon (C0 and C1) and nitrogen ((N0, N1 and N2) sources. This could be attributed to involvement of repressible metabolic pathways, reactions of which are inhibited by the presence of easily available nutrients for growth. Besides above, qualitative analysis of thiamethoxam residues by gas liquid chromatography revealed complete metabolization of thiamethoxam without detectable accumulation of any known thiamethoxam metabolites.

Persistence and risk assessment of cypermethrin residues on chilli (Capsicum annuum L.).

The study was conducted to observe the persistence pattern and risk assessment of cypermethrin in chilli fruits following three applications of cypermethrin (Super fighter 25 EC) at 50 and 100 g a.i. ha(-1) at 10-day interval. Residues of cypermethrin in chilli were estimated by gas liquid chromatography (GLC) and were confirmed by gas chromatography-mass spectrometry (GC-MS). The average initial deposits of cypermethrin in chilli fruits were found to be 1.46 and 3.11 mg kg(-1), at recommended and double the recommended dosages, respectively, following third application of the insecticide. Half-life periods for cypermethrin were found to be 4.43 and 4.70 days at recommended and double the recommended dosages, respectively. Residues of cypermethrin declined below its limit of quantification (LOQ) of 0.05 mg kg(-1) after 25 days at both the application dosages. Theoretical maximum residue contribution (TMRC) values were calculated from the residue data generated and were found to be below maximum permissible intake (MPI) even on 0 day. Therefore, according to our risk assessment studies, a waiting period of 1 day is suggested for consumption of chilli sprayed with cypermethrin at the recommended dosages.

Dissipation behavior and risk assessment of acephate in brinjal using GLC with FPD.

A study was conducted to observe the persistence, dissipation behavior, and risk assessment of acephate on brinjal fruit. Brinjal crop was sprayed with acephate 75 SP at 560 and 1120 g a.i. ha(-1) at fruiting stage followed by another application at 10-day interval. After sampling, the samples were extracted and cleaned up using quick, easy, cheap, effective, rugged, and safe (QuEChERS) technique, and the residues of acephate were analyzed with gas chromatography using flame photometric detector (FPD). The average initial deposits of acephate on brinjal fruits were found to be 2.54 and 4.07 mg kg(-1) following application of insecticide at 560 and 1120 g a.i. ha(-1), respectively. Residues of acephate reached below determination level of 0.10 mg kg(-1) after 7 days at recommended dosages and after 10 days at double the recommended dosages. The half-life of acephate was found to be 1.55 and 1.52 days, respectively, at 560 and 1120 g a.i. ha(-1). For risk assessment studies, theoretical maximum residue contributions (TMRC) were calculated and compared with maximum permissible intake (MPI). It was observed that TMRC values reached below MPI in 0-day samples at both recommended and double the recommended dosages. Therefore, it was concluded that if waiting period of 1 day is observed, there will be much reduced risk to consumers and the insecticide could be safely used for the protection of brinjal crop from insect pests.