Discovery of novel amide derivatives as potent quorum sensing inhibitors of Pseudomonas aeruginosa.

With the increasing reports of antibiotic resistance in this species, Pseudomonas aeruginosa is a common human pathogen with important implications for public health. Bacterial quorum sensing (QS) systems are potentially broad and versatile targets for developing new antimicrobial compounds. While previous reports have demonstrated that certain amide compounds can inhibit bacterial growth, there are few reports on the specific inhibitory effects of these compounds on bacterial quorum sensing systems. In this study, thirty-one amide derivatives were synthesized. The results of the biological activity assessment indicated that A9 and B6 could significantly inhibit the expression of lasB, rhlA, and pqsA, effectively reducing several virulence factors regulated by the QS systems of PAO1. Additionally, compound A9 attenuated the pathogenicity of PAO1 to Galleria mellonella larvae. Meanwhile, RT-qPCR, SPR, and molecular docking studies were conducted to explore the mechanism of these compounds, which suggests that compound A9 inhibited the QS systems by binding with LasR and PqsR, especially PqsR. In conclusion, amide derivatives A9 and B6 exhibit promising potential for further development as novel QS inhibitors in P. aeruginosa.

Bromine Atoms Decorated Pyene-based Covalent Organic Frameworks for Accelerated Photocatalytic H2 Production.

Designing materials with low exciton binding energy is an efficient way of improving the hydrogen production performance of COFs（Covalent Organic Frameworks. Here, it is demonstrated that the strategy of decorating bromine atoms on Pyene-based COFs can achieve elevated photocatalytic H2 evolution rates (HER = 13.61 mmol g-1 h-1 ). Low-temperature fluorescence and time-resolved fluorescence spectroscopy (TRPL) indicate that the introduction of bromine atoms can significantly suppress charge recombination. DFT (Density Functional Theory) calculation clarified that the C atoms adjacent to Br are the active sites with a reduced energy barrier in the process of formatting H intermediate species (H*). The modification strategy of Br atoms in COF furnishes a new medium for exploiting exquisite photocatalysts.

Formulation and Characterization of Matrine Oil Dispersion to Improve Droplet Wetting and Deposition.

The unreasonable use of chemical pesticides has caused serious damage to crops and the ecological environment. The botanical pesticide matrine has attracted attention as an environmentally friendly pesticide. Compared with traditional spraying methods, unmanned aerial vehicle (UAV) spraying has the advantages of safety, rapidity, uniform droplets, low dosages, and no terrain or crop restrictions. In this study, matrine OD was prepared according to the application requirements of flight prevention preparations using three different emulsifiers. The stability, wettability, particle size and distribution, and spraying performance of matrine OD were studied. The results indicated that when the amount of emulsifier was 8%, the three types of matrine OD had good stability. The stability, wettability, particle size and distribution, and spray performance of the suspension prepared using emulsifier VO/03 were better than the other two emulsifiers. Therefore, matrine OD prepared using 8% VO/03 could be used for ultra-low-volume sprays and aerial applications. In this study, we provide a theoretical basis and technical guidance to develop pesticide formulations for aerial applications.

A pH-responsive MOF-functionalized hollow mesoporous silica controlled herbicide delivery system exhibits enhanced activity against ACCase-herbicide-resistant weeds.

BACKGROUND: Weeds grow aggressively in agricultural fields, leading to reduced crop yields and an inability to meet the growing demand for food. Herbicides are currently the most effective method for weed control. However, the overuse of herbicides has resulted in the evolution of resistance mutants and has caused environmental pollution. Therefore, new technologies are urgently required to address this global challenge. RESULTS: We report a copper-benzene-1,4-dicarboxylate metal organic framework (Cu-BDC MOF)-functionalized carboxyl hollow mesoporous silica (HMS-COOH) delivery system for the pH-controlled release of the acetyl-CoA carboxylase (ACCase)-inhibiting herbicide quizalofop-p-ethyl (QE). The delivery system (QE@HMS@Cu-BDC) enabled the efficient control of barnyard grasses that are susceptible and resistant to ACCase-inhibiting herbicides, which showed 93.33% and 88.33% FW control efficacy at 67.5 g ha-1 , respectively. With the lowest pH value (3), QE and copper ion were released slowly to total 70.30% and 78.55% levels (respectively) from QE@HMS@Cu-BDC after 89 h. QE@HMS@Cu-BDC showed better absorption, conduction, transportation and ACCase activity inhibition performance than that of QE emulsifiable concentrate (EC) in both susceptible and ACCase-herbicide resistant barnyard grasses. In addition, with the safener effect of carrier HMS@Cu-BDC and the aid of the safener fenchlorazole-ethyl (FE), the application of QE@HMS@Cu-BDC was shown to mitigate the damage caused by QE to rice plants. CONCLUSION: This work found that the new material HMS-COOH@Cu-BDC can be used to mitigate herbicide-induced oxidative stress and improve rice plant safety. Futhermore, the QE@HMS-COOH@Cu-BDC constructed in this research might be used as an efficient nanopesticide formulation for weed controls in paddy rice fields. © 2023 Society of Chemical Industry.

Preparation of Thifluzamide Polylactic Acid Glycolic Acid Copolymer Microspheres and Its Effect on the Growth of Cucumber Seedlings.

The polylactic acid-glycolic acid copolymer (PLGA) has been proven to be applicable in medicine, but there is limited research on its application and safety in the agricultural field. In this paper, thifluzamide PLGA microspheres were prepared via phacoemulsification and solvent volatilization, using the PLGA copolymer as the carrier and thifluzamide as the active component. It was found that the microspheres had good slow-release performance and fungicidal activity against Rhizoctonia solani. A comparative study was conducted to show the effect of thifluzamide PLGA microspheres on cucumber seedlings. Physiological and biochemical indexes of cucumber seedlings, including dry weight, root length, chlorophyll, protein, flavonoids, and total phenol content, indicated that the negative effect of thifluzamide on plant growth could be mitigated when it was wrapped in PLGA microspheres. This work explores the feasibility of PLGA as carriers in fungicide applications.

Self-Assembled Sphere Covalent Organic Framework with Enhanced Herbicidal Activity by Loading Cyhalofop-butyl.

Nanopesticides are considered to be a novel and efficient kind of tool for controlling pests in modern agriculture. Covalent organic frameworks (COFs), with high surface areas, ordered structures, and rich functional groups for loading pesticides, are a class of promising carrier materials that can be used to develop efficient nanopesticide delivery systems. However, until now, only a strong ionic interaction between the pesticide and COF can be utilized to achieve the combination between the pesticide and COF. On the basis of this method, charged pesticide molecules are the only choice for COF-based nanopesticides, which limits the exploitation. The way to load the uncharged pesticide molecules into COF still needs to be explored. Herein, in this research, we provided a commonly mild and high-efficacy strategy for loading an uncharged pesticide molecule into COF. The herbicide cyhalofop-butyl (CB), as a neutral model pesticide molecule, was loaded into the sphere COF (SCOF, a model COF synthesized at room temperature) without any ionic interaction via the host-guest strategy. The loading capacity of CB into SCOF (CB@SCOF) was determined at 57% (w/w). Smaller CB@SCOF particles (150-200 nm) can efficiently enter the weed leaves and stems, enhancing the accumulation of the effective concentration in weeds, thus increasing herbicidal activity, in comparison to CB emulsifiable (EC, micrometer scale). Furthermore, CB@SCOF had a solubilization effect for CB in water and can improve the photostability of CB. Thus, the CB-loaded COF nanosphere showed excellent herbicidal activities against the target weeds Echinochloa crus-galli and Leptochloa chinensis compared to commercial CB EC. In conclusion, this study also provides a mild and high-efficacy pesticide loading strategy for COFs. The constructed efficient delivery system and pesticide formulation containing herbicidal COF nanospheres exhibit great potential applications for controlling weeds in sustainable agriculture.

A Mini Review on Natural Safeners: Chemistry, Uses, Modes of Action, and Limitations.

Herbicide injury is a common problem during the application of herbicides in practice. However, applying herbicide safeners can avoid herbicide damage. Safeners selectively protect crops against herbicide injury without affecting the biological activity of herbicides against the target weeds. However, after long-term application, commercial safeners were found to pose risks to the agricultural ecological environment. Natural safeners are endogenous compounds from animals, plants, and microbes, with unique structures and are relatively environment-friendly, and thus can address the potential risks of commercial safeners. This paper summarizes the current progress of the discovery methods, structures, uses, and modes of action of natural safeners. This study also concludes the limitations of natural safeners and prospects the future research directions, offering guidance for the practical application of natural safeners to prevent herbicide injury. This study will also guide the research and development of corresponding products.

Phenyl imidazolidin-2-ones antagonize a ß-adrenergic-like octopamine receptor in diamondback moth (Plutella xylostella).

BACKGROUND: The diamondback moth (Plutella xylostella) is one of the most destructive lepidopteran pests on cruciferous vegetables. However, resistance has emerged to current chemical and biological insecticides used for P. xylostella control, indicating the necessity of screening new targets on P. xylostella, and finding new insecticides against P. xylostella. In particular, octopamine receptors are representative G protein-coupled receptors found only in invertebrates and are potential targets for identifying novel insecticides. RESULTS: A ß-adrenergic-like octopamine receptor gene (PxOA2B1) was cloned, and its pharmacological characteristics in P. xylostella were studied. The results demonstrated that octopamine could activate the PxOA2B1 receptor, with a half-maximal effective concentration (EC50 ) of 49.5 nm. Amitraz, an insecticide and acaricide, and its metabolite (N-2,4-dimethylphenyl-N'-methylformamidine; DPMF) were also found to act as PxOAB1R agonists. We synthesized phenyl imidazolidin-2-one derivatives 3a-h using DPMF as the lead compound, and compounds 3a-h showed similar antagonist activities as phentolamine, mianserin and chlorpromazine. In particular, 3d, with an EC50 of 25.2 nm, showed very similar antagonist activity to mianserin. CONCLUSION: This research found that PxOAB1R might be a potential target for P. xylostella control. Phenyl imidazolidin-2-ones could be novel potential antagonists targeted at octopamine receptors and would be useful tools for the design and development of novel insecticides. © 2021 Society of Chemical Industry.

Novel phenylpyrimidine derivatives containing a hydrazone moiety protect rice seedlings from injury by metolachlor.

One strategy for solving the phytotoxicity of herbicides is to apply herbicide safeners that can efficiently alleviate the injuries of agricultural crops caused by herbicides. When metolachlor, a chloroacetamide herbicide, is applied with paddy rice, for example, the mechanisms associated with metolachlor and its residue negatively impact on the growth and yields of rice. To identify novel high-activity herbicide safener candidates for metolachlor, a series of (E)-4-(2-substituted hydrazinyl)-6-chloro-2-phenyl pyrimidines were synthesized and their structures were confirmed using IR (infrared radiation), 1H NMR, 13C NMR, and HRMS (high resolution mass spectrometry). The herbicide safener activities were then evaluated via primary tests. Compounds 3i and 3t were found to have the best herbicide activity on plant height. These compounds were then further screened for their activities at lower concentrations and showed better or similar activities compared to the positive control fenclorim, a commercial herbicide safener. The compounds 3i and 3t significantly enhanced glutathione S-transferase (GST) activity related with the herbicide safener activity in both shoots and roots tissues. Moreover, a qPCR (Real-time quantitative polymerase chain reaction) analysis found that the 3i and 3t treatments enhanced the expressions of OsGSTU3, OsGsTU39, and OsGSTF5. Finally, the results of an acute toxicity assessment with zebrafish (Danio rerio) embryos using treatments 3i and 3t indicated they are relatively safe to aquatic organisms.

Assessing the toxicity of three "inert" herbicide safeners toward Danio rerio: Effects on embryos development.

Recent studies indicated that 'inert ingredients' exert negative effects on the environment. Herbicide safeners are classed as 'inert ingredients', which increase the selectivity and detoxification of herbicides. However, little attention has been focused on the environmental behavior of herbicide safeners. AD-67 (AD), fenclorim (FM), and flurazole (FZ) are three commercially available herbicide safeners. In this study, zebrafish embryos were used as a model to investigate the potential developmental toxicity of these three safeners. The results showed that AD, FM, and FZ 96 h-LC50 values were 2.52, 1.26, and 2.01 mg/L, respectively. Significant decreased body lengths were observed in embryos after 96 h of exposure to 0.2 mg/L FM and FZ. Hatching rates significantly increased at 24 h and decreased at 96 h in all treatment groups (0.02 mg/L AD, 0.2 mg/L AD, 0.02 mg/L AD, 0.2 mg/L FZ, as well as 0.01 and 0.1 mg/L FM). No significant (p > 0.05) changes in heartbeat numbers (60 s), but clearly increased malformation rates were observed in response to safeners exposure. Furthermore, embryos showed signs of oxidative stress, such as decreased activities of superoxide dismutase, altered activities of glutathione reductase and catalase and cell apoptosis. The gene transcription related to body malformation (egf, krt 17, and tbx 16) and cell apoptosis (bcl 2 and bax) changed in treated groups. These genes have been connected to potential toxicological mechanisms. These results indicate that the herbicide safeners AD, FM, and FZ induced developmental toxicities in zebrafish embryos. This study is the first report of the toxicity of safeners in the development of zebrafish embryos. The results may be useful for assessing the risk of herbicides safeners in the aquatic ecosystem.

Developmental toxicity and transcriptome analysis of zebrafish (Danio rerio) embryos following exposure to chiral herbicide safener benoxacor.

Benoxacor, a chiral herbicide safener for S-metolachlor, has been detected in streams. However, the potential risk this poses to aquatic ecosystems is not clear. This study used zebrafish (Danio rerio) embryos as a model to assess the enantioselective toxicity of benoxacor and its effects on biological activity and development from 2 h to 96 h post-fertilization (hpf). Results showed that benoxacor had negative effects on hatchability, malformations, and mortality. Compared to either individual enantiomer, embryos exposed to Rac-benoxacor had higher acute and developmental toxicities, glutathione S-transferase (GST) and glutathione peroxidase (GPx) enzyme activities, and nrf 2 expression levels. They also had lower superoxide dismutase (SOD), catalase (CAT), and glutathione reductase (GR) enzyme activity and krt 17, tbx 16, osx, cat, bcl 2, bax, and ifn expression levels. High-throughput RNA sequencing revealed that Rac-benoxacor had a greater effect on gene regulation than either enantiomer. Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses demonstrated that changes in oxidoreductase activity, cellular lipid metabolic process, and catalytic activity related genes may be due to the enantioselective effects of benoxacor isomers. These results suggest that the ecotoxicology data and safety knowledge about the effects of chiral benoxacor on zebrafish should be considered in future environmental risk evaluation.

New Lead Discovery of Herbicide Safener for Metolachlor Based on a Scaffold-Hopping Strategy.

The use of herbicide safeners can significantly alleviate herbicide injury to protect crop plants and expand the application scope of the existing herbicides in the field. Sanshools, which are well known as spices, are N-alkyl substituted compounds extracted from the Zanthoxylum species and have several essential physiological and pharmacological functions. Sanshools display excellent safener activity for the herbicide metolachlor in rice seedlings. However, the high cost of sanshools extraction and difficulties in the synthesis of their complicated chemical structures limit their utilization in agricultural fields. Thus, the present study designed and synthesized various N-alkyl amide derivatives via the scaffold-hopping strategy to solve the challenge of complicated structures and find novel potential safeners for the herbicide metolachlor. In total, 33 N-alkyl amide derivatives (2a-k, 3a-k, and 4a-k) were synthesized using amines and saturated and unsaturated fatty acids as starting materials through acylation and condensation. The identity of all the target compounds was well confirmed by 1H-NMR, 13C-NMR, and high-resolution mass spectrometry (HRMS). The primary evaluation of safener activities for the compounds by the agar method indicated that most of the target compounds could protect rice seedlings from injury caused by metolachlor. Notably, compounds 2k and 4k displayed excellent herbicide safener activities on plant height and demonstrated relatively similar activities to the commercialized compound dichlormid. Moreover, we showed that compounds 2k and 4k had higher glutathione S-transferase (GST), superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and polyphenol oxidase (PPO) activities in rice seedlings, compared to the metolachlor treatment. In particular, 2k and 4k are safer for aquatic organisms than dichlormid. Results from the current work exhibit that compounds 2k and 4k have excellent crop safener activities toward rice and can, thus, be promising candidates for further structural optimization in rice protection.

Synthesis of Novel 6-Aryloxy-4-chloro-2-phenylpyrimidines as Fungicides and Herbicide Safeners.

Fenclorim is a commercial herbicide safener with fungicidal activity used for chloroacetanilide herbicides, which might be suitable as a lead compound for screening novel fungicides. However, little has been reported so far on the structure-activity relationship of fungicidal activities of fenclorim or its analogues. Here, a series of 4-chloro-6-substituted phenoxy-2-phenylpyrimidine derivatives was synthesized by a substructure splicing route using fenclorim as a lead compound. The structures of synthesized derivatives were characterized by 1H NMR, 13C NMR, and HRMS. Their fungicidal and herbicide safening activities were then evaluated. The results revealed that compound 11 had the best fungicidal activity against Sclerotinia sclerotiorum and Thanatephorus cucumeris, which was better than that of the control pyrimethanil. Moreover, compounds 3, 5, and 25 exhibited excellent safening activities against fresh weight, plant height, and root length, respectively. Such activities were significantly improved when compared to fenclorim. In summary, these findings look promising for the preparation of new fungicides and herbicide safeners based on the structure of fenclorim.

Direct Enantiomeric Separation and Determination of Hexythiazox Enantiomers in Environment and Vegetable by Reverse-Phase High-Performance Liquid Chromatography.

In the present study, the direct enantiomeric separation of hexythiazox enantiomers on Lux cellulose-1, Lux cellulose-2, Lux cellulose-3, Lux cellulose-4, Lux amylose-1 and Chirapak IC chiral columns were carefully investigated by reverse-phase high-performance liquid chromatography (RP-HPLC). Acetonitrile/water and methanol/water were used as mobile phase at a flow rate of 0.8 mL·min-1. The effects of chiral stationary phase, temperature, thermodynamic parameters, mobile phase component and mobile phase ratio on hexythiazox enantiomers separation were fully evaluated. Hexythiazox enantiomers received a baseline separation on the Lux cellulose-3 column with a maximum resolution of Rs = 2.09 (methanol/water) and Rs = 2.74 (acetonitrile/water), respectively. Partial separations were achieved on other five chiral columns. Furthermore, Lux amylose-1 and Chirapak IC had no separation ability for hexythiazox enantiomers when methanol/water was used as mobile phase. Temperature study indicated that the capacity factor (k) and resolution factor (Rs) decreased with column temperature increasing from 10 °C to 40 °C. The enthalpy (ΔH) and entropy (ΔS) involved in hexythiazox separation were also calculated and demonstrated the lower temperature contributed to better separation resolution. Moreover, the residue analytical method for hexythiazox enantiomers in the environment (soil and water) and vegetable (cucumber, cabbage and tomato) were also established with reliable accuracy and precision under reverse-phase HPLC condition. Such results provided a baseline separation method for hexythiazox enantiomers under reverse-phase conditions and contributed to an environmental and health risk assessment of hexythiazox at enantiomer level.

Design, Synthesis, and Biological Activity of Novel Heptacyclic Pyrazolamide Derivatives: A New Candidate of Dual-Target Insect Growth Regulators.

Insect growth regulators (IGRs) can cause abnormal growth and development in insects, resulting in incomplete metamorphosis or even death of the larvae. Ecdysone receptor (EcR) and chitinase in insects play indispensable roles in the molting process. Ecdysone analogues and chitinase inhibitors are considered as potential IGRs. In order to find new and highly effective IGR candidates, based on the structure-activity relationship and molecular docking results of the active compound 6i (3-(tert-butyl)-N-(4-(tert-butyl)phenyl)-1-phenyl-1H-pyrazole-5-carboxamide) discovered in our previous work, we changed the t-butyl group on the pyrazole ring into heptacycle to enhance the hydrophobicity. Consequently, a series of novel heptacyclic pyrazolamide derivatives were designed and synthesized. The bioassay results demonstrated that some compounds showed obvious insecticidal activity. Especially, D-27 (N-(4-(tert-butyl)phenyl)-2-phenyl-2,4,5,6,7,8-hexahydrocyclohepta[c]pyrazole-5-carboxamide) showed good activities against Plutella xylostella (LC50, 51.50 mg·L-1) and Mythimna separata (100% mortality at 2.5 mg·L-1). Furthermore, protein validation indicated that D-27 acts not only on the EcR but also on chitinase Of ChtI. Molecular docking and molecular dynamics simulation explained the vital factors in the interaction between D-27 and receptors. D-27 may be a new lead candidate with a dual target in which Of ChtI shall be the main one. This work created a new starting point for discovering a novel type of IGRs.

ABCC2 participates in the resistance of Plutella xylostella to chemical insecticides.

The ABCC2 protein of Plutella xylostella is an important target of Cry1A toxins from Bacillus thuringiensis (Bt), but whether this protein is involved in the resistance of P. xylostella to other insecticides remains unclear. In this study, the abcc2 gene of P. xylostella was cloned and the expression levels of Pxabcc2 in susceptible and resistant strains were investigated. ABCC2 was found to be expressed 3.2-6.7-fold higher in the resistant strain than in the susceptible strain; in the surviving P. xylostella, ABCC2 levels were significantly higher when treated with indoxacarb, avermectin, and beta-cypermethrin. We constructed a stable ABCC2-expressing HEK-293 cell line to reveal the contribution of ABCC2 to insecticide resistance. The avermectin and chlorfenapyr sensitivities of the stably-transfected cell line were significantly lower than those of the control cells. The intracellular avermectin concentration was significantly lower in the stably-transfected cell line than in the control cells after four hours of exposure. This study shows that up-regulated ABCC2 expression is related to insecticide resistance in P. xylostella. Moreover, we used RNA interference technology to reduce ABCC2 levels in P. xylostella. Down-regulating ABCC2 expression did not significantly affect avermectin or chlorfenapyr resistance in P. xylostella. We speculate that increased ABCC2 expression can enhance metabolic resistance in P. xylostella. This study also provides new insights into cross-resistance between B. thuringiensis toxins and chemical insecticides.

Pharmacological Properties of the Type 1 Tyramine Receptor in the Diamondback Moth, Plutella xylostella.

Tyramine receptors (TARs) can be activated by tyramine (TA) or octopamine (OA) and have been shown to be related to physiological regulation (e.g., gustatory responsiveness, social organization, and learning behavior) in a range of insect species. A tyramine receptor gene in Plutella xylostella, Pxtar1, was cloned and stably expressed in the HEK-293 cell line. Pharmacological properties and expression profile of Pxtar1 were also analyzed. Tyramine could activate the PxTAR1 receptor, increasing the intracellular Ca2+ concentration ((Ca2+)i) at an EC50 of 13.1 nM and reducing forskolin (10 µM)-stimulated intracellular cAMP concentration ((cAMP)i) at an IC50 of 446 nM. DPMF (a metabolite of amitraz) and L(-)-carvone (an essential oil) were found to act as PxTAR1 receptor agonists. Conversely, yohimbine and mianserin had significant antagonistic effects on PxTAR1. In both larvae and adults, Pxtar1 had the highest expression in the head capsule and expression of Pxtar1 was higher in male than in female reproductive organs. This study reveals the temporal and spatial differences and pharmacological properties of Pxtar1 in P. xylostella and provides a strategy for screening insecticidal compounds that target PxTAR1.

Dissipation Dynamic and Final Residues of Oxadiargyl in Paddy Fields Using High-Performance Liquid Chromatography-Tandem Mass Spectrometry Coupled with Modified QuEChERS Method.

Oxadiargyl, which binds to the protoporphyrinogen oxidase IX to exhibit herbicide activity, is mainly used in the prevention of certain perennial broadleaved and grass weeds during the preemergence of rice in paddy fields. However, oxadiargyl affects the germination and seedling growth of rice, causing damage to the plant and reducing rice yield. Hence, monitoring fate and behaviour of oxadiargyl in rice paddy fields is of great significance. A modified quick, easy, cheap, effective, rugged, and safe (QuEChERS) sample preparation method coupled with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was established in paddy water, paddy soil, rice straw, paddy hull, and brown rice. We validated this method for the first time in the analysis of the dissipation dynamic and residues of oxadiargyl over two years (2015⁻2016) at three sites in China. The average recoveries of oxadiargyl ranged from 76.0 to 98.8%, with relative standard deviations of 3.5⁻14.0%. The dissipation curves for paddy soil fit to a first-order kinetic equation, revealing that oxadiargyl degraded rapidly in paddy soil with half-lives (t1/2) of 4.5⁻7.6 days. The final oxadiargyl residues in all samples remained below the detection limit and the maximum residue limit in China (0.02 mg kg-1) and Japan (0.05 mg kg-1) during the harvesting dates and were not detected in rice straw.

Discovery of a Manduca sexta Allatotropin Antagonist from a Manduca sexta Allatotropin Receptor Homology Model.

Insect G protein coupled receptors (GPCRs) have important roles in modulating biology, physiology and behavior. They have been identified as candidate targets for next-generation insecticides, yet these targets have been relatively poorly exploited for insect control. In this study, we present a pipeline of novel Manduca sexta allatotropin (Manse-AT) antagonist discovery with homology modeling, docking, molecular dynamics simulation and structure-activity relationship. A series of truncated and alanine-replacement analogs of Manse-AT were assayed for the stimulation of juvenile hormone biosynthesis. The minimum sequence required to retain potent biological activity is the C-terminal amidated octapeptide Manse-AT (6-13). We identified three residues essential for bioactivity (Thr4, Arg6 and Phe8) by assaying alanine-replacement analogs of Manse-AT (6-13). Alanine replacement of other residues resulted in reduced potency but bioactivity was retained. The 3D structure of the receptor (Manse-ATR) was built and the binding pocket was identified. The binding affinities of all the analogs were estimated by calculating the free energy of binding. The calculated binding affinities corresponded to the biological activities of the analogs, which supporting our localization of the binding pocket. Then, based on the docking and molecular dynamics studies of Manse-AT (10-13), we described it can act as a potent Manse-AT antagonist. The antagonistic effect on JH biosynthesis of Manse-AT (10-13) validated our hypothesis. The IC50 value of antagonist Manse-AT (10-13) is 0.9 nM. The structure-activity relationship of antagonist Manse-AT (10-13) was also studied for the further purpose of investigating theoretically the structure factors influencing activity. These data will be useful for the design of new Manse-AT agonist and antagonist as potential pest control agents.

Pharmacological characterization of a ß-adrenergic-like octopamine receptor in Plutella xylostella.

The ß-adrenergic-like octopamine receptor (OA2B2) belongs to the class of G-protein coupled receptors. It regulates important physiological functions in insects, thus is potentially a good target for insecticides. In this study, the putative open reading frame sequence of the Pxoa2b2 gene in Plutella xylostella was cloned. Orthologous sequence alignment, phylogenetic tree analysis, and protein sequence analysis all showed that the cloned receptor belongs to the OA2B2 protein family. PxOA2B2 was transiently expressed in HEK-293 cells. It was found that PxOA2B2 could be activated by both octopamine and tyramine, resulting in increased intracellular cyclic AMP (cAMP) levels, whereas dopamine and serotonin were not effective in eliciting cAMP production. Further studies with series of PxOA2B2 agonists and antagonists showed that all four tested agonists (e.g., naphazoline, clonidine, 2-phenylethylamine, and amitraz) could activate the PxOA2B2 receptor, and two of tested antagonists (e.g., phentolamine and mianserin) had significant antagonistic effects. However, antagonist of yohimbine had no effects. Quantitative real-time polymerase chain reaction analysis showed that Pxoa2b2 gene was expressed in all developmental stages of P. xylostella and that the highest expression occurred in male adults. Further analysis with fourth-instar P. xylostella larvae showed that the Pxoa2b2 gene was mainly expressed in Malpighian tubule, epidermal, and head tissues. This study provides both a pharmacological characterization and the gene expression patterns of the OA2B2 in P. xylostella, facilitating further research for insecticides using PxOA2B2 as a target.