Toxic evaluation of Proclaim Fit® on adult and larval worker honey bees.

Impacts to honey bees due to exposure to agricultural pesticides is one of the most serious threats to the beekeeping industry. Our research evaluated toxicity of the formulated insecticides Lufenuron+Emamectin benzoate (Proclaim Fit®) on the European honey bee Apis mellifera L. at field-realistic concentration (worst-case scenario). Newly emerged (≤24-h old) and forager (unknown age) worker bees were treated with the field recommended concentration of Proclaim Fit® using three routes of exposure including residual contact, oral, and spray within the laboratory. We also assessed the effects of Proclaim Fit® on the specific activity of some well-known detoxifying enzymes including α-esterase, ß-esterase, and Glutathione S-transferase (GST) in the honey bees. In addition, toxicity of the formulation was tested on 4th instar larvae within the hive. Based on estimated median survival times (MSTs), Proclaim Fit® was highly toxic to the bees, especially when applied as spray. According to our estimated relative median potency (RMP) values, newly emerged bees were 1.72× more susceptible than foragers to Proclaim Fit® applied orally. Enzyme assays revealed the considerable involvement of the enzymes, especially GST and α-esterase, in detoxification of the Proclaim Fit®, but their activities were significantly influenced by route of exposure and age of bee. Notably, Proclaim Fit® was highly toxic to 4th instar honey bee larvae. Our results generally indicate a potent toxicity of Proclaim Fit® toward honey bees. Therefore, its application requires serious consideration and adherence to strict guidelines, especially during the flowering time of crops.

Monophosphoramide derivatives: synthesis and crystal structure, theoretical and experimental studies of their biological effects.

Synthesizing new chemical compounds and studying their biological applications have been important issues in scientific research. In this investigation, we synthesized and characterized ten new N-acetyl phosphoramidate compounds and explored the crystal structure of three others. Furthermore, not only were some kinetic inhibition parameters measured, like IC50, Ki, kp, KD for 7 compounds on human acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), but also their hydrophobic parameter was determined by shake-flask technique. All compounds (number 1-10) were investigated for anti-bacterial activity against three Gram-positive and three Gram-negative bacteria, while chloramphenicol was used as a standard antibiotic. In order to find new insecticide, toxicities of 13 acephate (Ace)-derived compounds (number 20-32) were bioassayed on third larval instar of elm leaf beetle and Xanthogaleruca luteola. Additionally, screening in vivo tests revealed that two compounds had had the greatest insecticidal potential in comparison with others. It means these ones inhibited AChE (with mixed mechanisms) and general esterase more than the rest. According to ChE-QSAR models, the inhibitory potency for enzyme and bacteria is directly influenced by the electronic parameters versus structural descriptors. AChE-QSPR model of fluorescence assay indicated that the inhibitory power of AChE is primarily influenced by a set of electronic factors with the priority of: EHB > PL > δ(31P) versus structural descriptor (SA and Mv). Synthesizing new chemical compounds and studying their biological applications have been important issues in scientific research. Toxicities of 13 acephate (Ace)-derived compounds (number 20-32) were bioassayed on third larval instar of elm leaf beetle and Xanthogaleruca luteola. Insect-QSAR equations of these compounds, based on MLR and PCA, showed that non-descriptor net charge nitrogen atom (which was affected by the polarization of N-H group) had the greatest effect on insecticidal potential.

Synthesis, crystal structure, cholinesterase inhibitory activity, evaluation of insecticidal activities, and computational studies of new phosphonic acids.

In an attempt to obtain the modified and novel insecticides with low human toxicity, a series of novel mono-, bis-, and tetraphosphonic acid derivatives were designed and characterized by infrared, 1H, 13C, and 31P NMR spectroscopy and X-ray crystallography. The inhibitory effects of the synthesized compounds were evaluated using the in vitro Ellman method on human and insect acetylcholinesterase (AChE). Some of these compounds, which had low human and high insect toxicity, were chosen to assess the killing effects (in vivo) on third larval instar of elm leaf beetle (X. luteola). In vivo and in vitro evidence has revealed that bisphosphonic acids, containing hydrophobic systems, have a good selectivity of insect AChE inhibition. In the present study, docking results showed that bisphosphonic acids had lower binding energy and higher inhibition compared with tetraphosphonic acids due to the type of their topology and the ability of their hydrogen to interact with the catalytic triad (the main active site of the enzyme). Additionally, the QSAR results demonstrated that the major effecting factors on the insecticidal activity of the subject compounds are the hydrophobicity, size, shape, and ability to form a hydrogen bond. The present study can be helpful in the development of new insecticides.

Synthesis, crystal structure, insecticidal activities, molecular docking and QSAR studies of some new phospho guanidines and phospho pyrazines as cholinesterase inhibitors.

Novel phospho guanidine and phospho pyrazine derivatives were synthesized and characterized by 31P, 13C, 1HNMR and IR spectroscopy to obtain novel and human-safe insecticides. Compound 35 [(C4H4N2NH)2P(O)(C6H6)] was investigated by X-ray crystallography. The inhibitory effects of synthesized compounds were evaluated on human and insect acetylcholinesterase (AChE) using in vitro Ellman method. A few of these compounds, which had low human toxicity, were selected for assessing the killing effects (in vivo) on the elm leaf beetle (X.luteola). The in vitro and in vivo results indicated that compounds bearing both phosphoryl groups and aromatic systems were found to possess a good selectivity for the inhibition of insect AChE over human AChE; up to 550-fold selectivity was achieved for compound 19. Docking studies were performed to explain reasons for the selective behavior of AChE inhibitors. Additionally, the quantitative structure-activity relationship (QSAR) and density functional theory (DFT) results of AChEs demonstrated that the size, shape, dipole moment, and ability to form hydrogen bond played the main role in both models. In addition, the aromatic π - π interactions and charge of the amide nitrogen had a major effect on insecticidal activity of the compounds. The present research can be helpful to gain a better understanding of the interactions between the insect AChE and its inhibitors and introduces compounds which are capable of becoming human-safe insecticides.

Characterization of acetylcholinesterase from elm left beetle, Xanthogaleruca luteola and QSAR of temephos derivatives against its activity.

Insect acetylcholinesterase (AChE) is the principal target for organophosphate (OP) and carbamate (CB) insecticides. In this research, an AChE from third instar larvae of elm left beetle, Xanthogaleruca luteola was purified by affinity chromatography. The enzyme was purified 75.29-fold with a total yield of 8.51%. As shown on denaturing SDS-PAGE, the molecular mass of purified AChE was 70kDa. The enzyme demonstrated maximum activity at pH7 and 35°C. Furthermore, a series of temephos (Tem) derivatives with the general structure of P(O)XP(O) (1-44) were prepared, synthesized and characterized by 31P, 13C, 1H NMR and FT-IR spectral techniques. The toxicity of 36 new Tem derivatives was screened on the third instar larvae and the compound compound 1,2 cyclohexane-N,N'-bis(N,N'-piperidine phosphoramidate) exhibited the highest insecticidal potential. The method of kinetic analysis is applied in order to obtain the maximum velocity (Vmax), the Michaelis constant (Km) and the parameters characterizing the inhibition type for inhibitors with >75% mortality in preliminary bioassay. The inhibition mechanism was mixed and inhibitory constant (Ki) was calculated as 4.70µM-1min-1 for this compound. Quantitative structure-activity relationship (QSAR) equations of these compounds indicated that the electron orbital energy has major effect on insecticidal properties.

Effects of 4-hexylresorcinol on the phenoloxidase from Hyphantria cunea (Lepidoptera: Arctiidae): In vivo and in vitro studies.

Insecticidal effects of 4-hexylresorcinol, a phenoloxidase (PO) inhibitor, were determined on Hyphantria cunea (Drury) under laboratory conditions. The LC50 for the 15-d-old larvae was estimated to be 2.95 g/L after 96 h exposure. The activities of glutathione S-transferase (GST) and PO showed a decrease in larvae treated with 4-hexylresorcinol, and the IC50 of GST and PO were estimated to be 0.8 and 0.43 g/L, respectively, 24 h after treatment. The PO from the hemolymph of fall webworm was purified by ammonium sulfate precipitation, gel-filtration, and ion-exchange chromatography, and then enzymatic characteristics and the mechanism of inhibition were determined using L-dihydroxyphenylalanine (L-DOPA) as the substrate. The purified PO showed a single band on SDS-PAGE with a molecular weight of about 70 kDa. The optimum pH for PO activity was observed at pH 7.0, optimum temperature was found to be 45 °C, and PO activity was strongly inhibited by Zn(2+) . IC50 values were estimated to be 8.2, 19.14, and 24.04 µmol/L for 4-hexylresorsinol, kojic acid, and quercetin, respectively. The inhibitory potencies (i.e., I50 of each compound/I50 of 4-hexylresorcinol) of kojic acid and quercetin on H. cunea PO were estimated to be 1.87 and 2.89, respectively. 4-hexylresorcinol was determined to be a competitive inhibitor, and kojic acid and quercetin were determined to be mixed inhibitors. PO is one of the most important enzymes in an insect's immune system, and the use of PO inhibitors seems to be a promising approach for pest control due to their potential safety for humans.