Enhancing the systemic distribution of a bioactive compound by exploiting the vascular transport system of a plant presents a means of reducing both the volume and frequency of pesticide/fungicide application. The foliar uptake of the glucose–fipronil conjugate N-[3-cyano-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-[(trifluoromethyl)sulfinyl]-1H-pyrazol-5-yl]-1-(β-d-glucopyranosyl)-1H-1,2,3-triazole-4-methanamine (GTF) achieved in castor bean (Ricinus communis) and its transport via the phloem are known to be mediated by monosaccharide transporter(s) [MST(s)], although neither the identity of the key MST(s) involved nor the mechanistic basis of its movement have yet to be described. On the basis of homology with Arabidopsis thaliana sugar transporters, the castor bean genome was concluded to harbor 53 genes encoding a sugar transporter, falling into the eight previously defined subfamilies INT, PMT, VGT, STP, ERD6, pGlucT, TMT, and SUT. Transcriptional profiling identified the product of RcSTP1 as a candidate for mediating GTF uptake, because this gene was induced by exposure of the plant to GTF. When RcSTP1 was transiently expressed in onion epidermis cells, the site of RcSTP1 deposition was shown to be the plasma membrane. A functional analysis based on RcSTP1 expression in Xenopus laevis oocytes demonstrated that its product has a high affinity for GTF. The long-distance root-to-shoot transport of GTF was enhanced in a transgenic soybean chimera constitutively expressing RcSTP1.Keywords: glucose−fipronil; monosaccharide transporter; Ricinus communis; transgenic soybean composite plants; Xenopus oocyte;

Nanopesticides have been increasingly used in agriculture. To improve the uptake of the target organisms for nanopesticides, we designed a dual-ligand nanopesticide based on gold nanoparticles (Au NPs) as a carrier. Herein, the novel structure and properties of Au NPs conjugated with D-glucose (Glc) and rotenone (R) were characterized by UV-visible and FTIR spectroscopy, transmission electron microscopy (TEM), dynamic light scattering (DLS), and zeta potential. The uptake and biodistribution of Glc-Au NPs, R-Au NPs, and Glc-Au NP-R were tested in tobacco BY-2 cells using flow cytometry, inductively coupled plasma optical emission spectroscopy (ICP-OES), confocal laser scanning microscopy (CLSM), and TEM. The cellular uptake of R-Au NP-Glc was increased by 1.8 folds as compared to that of R-Au NPs. Moreover, R-Au NPs-Glc could be actively transported into cells by hexose transporters and this uptake of the nanopesticides was an energy-dependent process. These results suggest that Glc is a promising ligand for the uptake and delivery of nanopesticides, and a well-designed system can enhance pesticidal bioavailability in agricultural scenarios.

The acetylcholinesterase 2 (AChE2) cloned from Plutella xylostella was first successfully expressed in methylotrophic yeast Pichia pastoris GS115. One transformant with high-level expression of the recombinant AChE (rAChE, 23.2 U mL−1 in supernatant) was selected by plating on increasing concentrations of antibiotic G418 and by using a simple and specific chromogenic reaction with indoxyl acetate as a substrate. The maximum production of rAChE reached about 11.8 mg of the enzyme protein per liter of culture. The rAChE was first precipitated with ammonium sulfate (50% saturation) and then purified with procainamide affinity column chromatography. The enzyme was purified 12.1-fold with a yield of 22.8% and a high specific activity of 448.3 U mg−1. It was sensitive to inhibition by methamidophos and pirimicarb, the calculated 50% inhibitory concentration (IC50) values of the two pesticides were 0.357 and 0.888 mg L−1, respectively, and the calculated 70% inhibitory concentration (IC70) values were 0.521 and 0.839 mg L−1, respectively. The results suggested that it has a potential application in the detection of pesticide residues.

Phloem-mobile insecticides are efficient for piercing and sucking insect control. Introduction of sugar or amino acid groups to the parent compound can improve the phloem mobility of insecticides, so a glycinergic–fipronil conjugate (GlyF), 2-(3-(3-cyano-1-(2,6-dichloro-4-(trifluoromethyl)phenyl)-4-((trifluoromethyl)sulfinyl)-1H-pyrazole-5-yl)ureido) acetic acid, was designed and synthesized. Although the “Kleier model” predicted that this conjugate is not phloem mobile, GlyF can be continually detected during a 5 h collection of Ricinus communis phloem sap. Furthermore, an R. communis seedling cotyledon disk uptake experiment demonstrates that the uptake of GlyF is sensitive to pH, carbonyl cyanide m-chlorophenylhydrazone (CCCP), temperature, and p-chloromercuribenzenesulfonic acid (pCMBS) and is likely mediated by amino acid carrier system. To explore the roles of amino acid transporters (AATs) in GlyF uptake, a total of 62 AAT genes were identified from the R. communis genome in silico. Phylogenetic analysis revealed that AATs in R. communis were organized into the ATF (amino acid transporter) and APC (amino acid, polyaminem and choline transporter) superfamilies, with five subfamilies in ATF and two in APC. Furthermore, the expression profiles of 20 abundantly expressed AATs (cycle threshold (Ct) values <27) were analyzed at 1, 3, and 6 h after GlyF treatment by RT-qPCR. The results demonstrated that expression levels of four AAT genes, RcLHT6, RcANT15, RcProT2, and RcCAT2, were induced by the GlyF treatment in R. communis seedlings. On the basis of the observation that the expression profile of the four candidate genes is similar to the time course observation for GlyF foliar disk uptake, it is suggested that those four genes are possible candidates involved in the uptake of GlyF. These results contribute to a better understanding of the mechanism of GlyF uptake as well as phloem loading from a molecular biology perspective and facilitate functional characterization of candidate AAT genes in future studies.

•Two Novel fluorescent glycosyl labels have been synthesized in mild conditions.•Fluorescent glycosyl labels feature azide groups.•Three fluorescent glycosyl conjugates were synthesized and studied their imaging in plant.Two novel, biocompatible and highly stereoselective fluorescent glycosyl labels featuring azide groups have been successfully synthesized. The functional groups provide convenient routes for fluorescent glycosyl conjugates via click chemistry. Three fluorescent glycose-conjugates were synthesized. One fluorescent conjugate was used to trace the translocation in castor seedlings. The developed fluorescent labels for biomolecules are likely to provide an important method for investigating various biological events. We believe that the probes obtained are useful for studies on biological processes.

Understanding and visualizing the biodistribution of agricultural chemicals inside cells and living plants is very important for enhancing targeting and changing the application approaches of chemicals. Here, a novel material was synthesized through 2,4-dichlorophenoxyacetic acid functionalized small gold nanoparticles (2,4-D-MP-Au NPs). The successful modification of Au NPs (4.46 ± 0.70 nm) was ascertained by UV-vis, TEM, FTIR and XPS. TGA data revealed about 1197 molecules of 2,4-D were coupled to the surface of one Au nanoparticle, which was sufficient for bioapplications. The optical imaging of 2,4-D-MP-Au NPs inside BY-2 cells was directly examined, revealing that 2,4-D-MP-Au NPs could be internalized in BY-2 cells by the two-photo microscopy and endocytosis, as the internalization mechanism was energy dependent for 2,4-D-MP-Au NPs. Furthermore, the biodistribution of 2,4-D-MP-Au NPs in Ricinus cotyledons was measured, revealing that 2,4-D-MP-Au NPs could enter into mesophyll cells of Ricinus cotyledons; the cell recognition was enhanced after 2,4-D conjugated Au NPs. These results indicate that the conjugates have great potential for applications on bioimaging and biolabeling for agricultural chemicals in plant physiology.

A series of novel phenyl pyrazole inner salt derivatives based on fipronil were designed and synthesized in the search for dual-target insecticides. These compounds were designed to target two families of nicotinic acetylcholine receptors and γ-aminobutyric acid receptors. The insecticidal activities of the new compounds against diamondback moth (Plutella xylostella) were evaluated. The results of bioassays indicated that most of the inner salts showed moderate to high activities, of which the phenyl pyrazole inner salts containing quinoline had excellent biological activity. Previous structure–activity relationship studies revealed that a suitable structure of the quaternary ammonium salts was critical for the bioactivity of phenyl pyrazole inner salts, which contribute to exposing the cationic nitrogen to bind to the receptor (for instance, nicotinic acetylcholine receptors) and possibly interact with the receptor via hydrogen bonding and cooperative π–π interaction. The present work demonstrates that the insecticidal potency of phenyl pyrazole inner salts holds promise for the development new dual-target phenyl pyrazole insecticides.

Six monosaccharide derivatives of rotenone were designed and synthesized to assess whether rotenone could become phloem mobile by the addition of a monosaccharide group. Phloem mobility experiments showed that only d-glucose conjugates exhibit phloem transport properties in castor bean (Ricinus communis L.) seedlings. Two d-glucose conjugates, 2-O-β-d-glucopyranosyldemethylrotenone and 6′-O-β-d-glucopyranosyldalpanol, had significantly obtained systemicity compared with that of rotenone, and 6′-O-β-d-glucopyranosyldalpanol was more mobile than 2-O-β-d-glucopyranosyldemethylrotenone. Coupling with a monosaccharide core is a reasonable method for conferring phloem mobility on insecticides, but phloem mobility is also affected by the parent molecule and the position of the monosaccharide.

In our previous work, a glucose–fipronil (GTF) conjugate at the C-1 position was synthesized via click chemistry and a glucose moiety converted a non-phloem-mobile insecticide fipronil into a moderately phloem-mobile insecticide. In the present paper, fipronil was introduced into the C-2, C-3, C-4, and C-6 positions of glucose via click chemistry to obtain four new conjugates and to evaluate the effects of the different glucose isomers on phloem mobility. The phloem mobility of the four new synthetic conjugates and GTF was tested using the Ricinus seedling system. The results confirmed that conjugation of glucose at different positions has a significant influence on the phloem mobility of GTF conjugates.

The ability to visualize the movement of glycosyl insecticides contributes to learning their translocation and distribution in plants. In our present work, a novel fluorescent glucose–fipronil conjugate N-[3-cyano-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-[(trifluoromethyl)sulfinyl]-1H-pyrazol-5-yl]-1-(2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-β-d-glucopyranosyl)-1H-1,2,3-triazole-4-methanamine (2-NBDGTF), was obtained via click chemistry. Disk uptake experiments showed that an active carrier-mediated system was involved in the 2-NBDGTF uptake process. Meanwhile, 2-NBDGTF exhibited comparable phloem mobility with GTF in castor bean seedlings. Visualization of 2-NBDGTF uptake and transport experiment showed that this fluorescent glucose–fipronil conjugate could be loaded into sieve tubes after transiting through epidermal cells and mesophyll cells and then translocate from cotyledon to hypocotyl via phloem in castor bean seedlings. The results above determined that it is a promising fluorescence tagging approach for revealing the activities of glycosyl insecticides and 2-NBDGTF is a reasonable and feasible fluorescent surrogate of GTF for tracing the distribution of glucose–fipronil conjugates.

Mobile glucose–pesticide conjugates in the phloem are often restricted by decreases in biological activity. However, plants can bioactivate endogenous glucosides, which are assumed as able to bioactivate exogenous conjugates. In this study, four glycosidic bonds (O-, S-, N-, and C-glycosidic bonds) of glucose–pesticide conjugates were designed and synthesized, and then metabolism assays were carried out in vitro and in vivo. Results showed that β-glucosidases played a role in the hydrolysis of O-glycosidic bond conjugates in Ricinus communis L. The liberated aglycons possessed insecticidal activities against Plutella xylostella L. and Spodoptera litura F. These results could help establish methods of circumventing the mutual exclusivity of phloem mobility and biological activity by hydrolyzing endogenous β-glucosidases.

Based on the discovery of thymine as an ecdysteroid agonist, a series of 1,4-disubstituted diacylhydrazine derivatives with a thymine moiety were designed and synthesized. The activities of these compounds against Spodoptera litura (Fabricius) were evaluated by the insect immersion method. Results showed that compound 2h with an N-cyclohexylmethyl substituent exhibits the most potent agonist activity with a median lethal concentration of 23.21 μg/mL. This compound also caused malformation of molting larvae and adults. Compound 2h was further demonstrated as an ecdysteroid agonist by reporter gene assay on the Spodoptera frugiperda cell line (Sf9 cells). A molecular docking study indicated that hydrophobic interactions and the formation of hydrogen bonds between the compounds and the ecdysone receptor play critical roles in promoting the binding affinity of the compound. The structure of compound 2h may serve as a favorable template for the development of new ecdysteroid agonists with a pyrimidinedione moiety.

To test the effect of adding different monosaccharide groups to a non-phloem-mobile insecticide on the phloem mobility of the insecticide, a series of conjugates of different monosaccharides and fipronil were synthesized using the trichloroacetimidate method. Phloem mobility tests in castor bean (Ricinus communis L.) seedlings indicated that the phloem mobility of these conjugates varied markedly. l-Rhamnose–fipronil and d-fucose–fipronil displayed the highest phloem mobility among all of the tested conjugates. Conjugating hexose, pentose, or deoxysugar to fipronil through an O-glycosidic linkage can confer phloem mobility to fipronil in R. communis L. effectively, while the −OH orientation of the monosaccharide substantially affected the phloem mobility of the conjugates.

Some compounds containing glucose are absorbed via the monosaccharide transporters of the plasma membrane. A glucose-fipronil conjugate, N-[3-cyano-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-[(trifluoromethyl)sulfinyl]-1H-pyrazol-5-yl]-1-(β-d-glucopyranosyl)-1H-1,2,3-triazole-4-methanamine (GTF), has been synthesized in our previous work. GTF exhibits moderate phloem mobility in Ricinus communis. In the current paper, we demonstrate that the uptake of GTF by Ricinus seedling cotyledon discs is partly mediated by an active carrier system (Km1 = 0.17 mM; Vmax1 = 2.2 nmol cm–2 h–1). Four compounds [d-glucose, sucrose, phloridzin, and carbonyl cyanide m-chlorophenylhydrazone (CCCP)] were examined for their effect on GTF uptake. Phloridzin as well as CCCP markedly inhibit GTF uptake, and d-glucose weakly competes with it. The phloem transport of GTF in Ricinus seedlings is found to involve an active carrier-mediated mechanism that effectively contributes to the GTF phloem loading. The results prove that adding a glucose core is a reasonable and feasible approach to confer phloem mobility to fipronil by utilizing plant monosaccharide transporters.

A new C-methylated flavone glycoside, 5,7-dihydroxy-3-methoxy-6-C-methylflavone 8,4′-di-O-β-d-glucopyranoside (1), was isolated from the twigs and leaves of Picea neoveitchii Mast, together with eight known compounds, 5,7,8,4′-tetrahydroxy-3-methoxy-6-methylflavone 8-O-β-d-glucopyranoside (2) kaempferol 3,4′-di-O-β-d-glucopyranoside (3), apigenin 7-O-β-d-glucopyranoside (4), tiliroside (5), massonianoside B (6), umbeliferone 7-O-β-d-glucopyranoside (7), dihydroconiferin (8) and gleditschiaside A (9). Their structures were elucidated on the basis of analyses of spectroscopic data. Compound 1 showed moderate antifungal activity against tested plant pathogens (Pyricularia grisea (Cooke) Sacc., Sclerotium rocfsii Sacc. and Alternaria mali Roberts), however, compounds 2 and 5 had obvious inhibitory effect against S. rocfsii and A. mali, respectively. Compounds 1, 2, 3 and 9 also exhibited potent cytotoxicity against Spodoptera litura Fabricius cells.A new C-methylated flavone glycoside, 5,7-dihydroxy-3-methoxy-6-C-methylflavone 8,4′-di-O-β-d-glucopyranoside (1), together with seven known compounds (2–9) were isolated from the twigs and leaves of Picea neoveitchii Mast. The cytotoxicities of the isolated compounds on Spodoptera litura Fabricius cell were evaluated by MTT assay and their antifungal activities were evaluated by disc diffusion method. Three of these compounds were isolated from the Pinaceae family for the first time.

An unprecedented dimeric triterpenoid, designated dibelamcandal A, with a six-membered ring linking two iridal type triterpenoid nuclei, was isolated from the rhizome of Belamcanda chinensis. Its structure was determined by extensive spectroscopic measurements, including IR, ESI-MS, HR-ESI-MS, and 1D and 2D NMR. It demonstrated significant molluscicide activity against Pomacea canaliculata.

Phloem-mobile insecticides are preferred to achieve economically useful activity. However, only a few phloem-mobile synthetic insecticides are available. One approach to converting nonmobile insecticides into phloem-mobile types is introducing sugar to the parent compound. To test whether the addition of a glucose group to a non-phloem-mobile insecticide enables conversion into phloem-mobile, N-[3-cyano-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-[(trifluoromethyl)sulfinyl]-1H-pyrazol-5-yl]-1-(β-d-glucopyranosyl)-1H-1,2,3-triazole-4-methanamine (GTF) was prepared through click chemistry. A phloem-mobility test in Ricinus communis L. seedlings confirmed that GTF was mobile in the sieve tubes. Although GTF exhibited lower insecticidal activity against the third-instar larvae of Pzlutella xylostella than fipronil did, it can be reconverted into fipronil in adult plants of castor bean, thereby offsetting the decrease of insecticidal activity. Therefore, the presence of a glucose core confers phloem mobility to fipronil.

Four flavonoids, 5,7,4′-trihydroxy-3,8,-dimethoxy-6-C-methylflavone (1), 5,8,4′-trihydroxy-3,7-dimethoxy-6-C-methylflavone (2), 7-methoxy-6-C-methylkaempferol (3) and kaempferol-7-O-(2″-E-p-coumaroyl)-α-l-arabinofuranoside (4), together with 15 known compounds, were isolated from the twigs and leaves of Picea neoveitchii Mast. Their structures were elucidated on the basis of analyses of spectroscopic data. Compound 4 showed strong anti-fungal activity against Fusarium oxysporum whereas compounds 1–4 were all active against Rhizoctonia solani.Graphical abstractFlavonoids, 5,7,4′-trihydroxy-3,8-dimethoxy-6-C-methylflavone (1), 5,8,4′-trihydroxy-3,7-dimethoxy-6-C-methylflavone (2), 7-O-methyl-6-C-methylkaempferol (3), kaempferol-7-O-(2″-E-p-coumaroyl)-α-l-arabinofuranoside (4), were isolated from Picea neoveitchii. Their anti-fungal activities were evaluated by the disc diffusion method.Research highlights► Picea neoveitchii was phytochemically studied for the first time. ► Nineteen compounds were isolated. ► Four flavonoids were characterized as new compounds. ► Antifungal activities were evaluated against F. oxysporum and Rhizoctonia solani.

Novel amphiphilic chitosan derivatives N-(octadecanol-1-glycidyl ether)-O-sulfate chitosan (NOSCS) with octadecanol glycidyl ether as hydrophobic groups and sulfate as hydrophilic groups were synthesized successfully. The NOSCS were characterized with 1H NMR, FT-IR, and their critical micellar concentrations (CMCs) were found to be 3.55 × 10−3 to 5.50 × 10−3 mg/mL. The degree of substitution (DS) of octadecanol glycidyl ether grafted on chitosan was ranging from 0.6% to 7.1%. NOSCS could form polymeric micelles with the size of 167.7–214.0 nm by self-assembly in aqueous solution. Rotenone, a water-insoluble botanical insecticide, was entrapped into NOSCS micelles solution by reverse micelle method. And the highest rotenone concentration was up to 26.0 mg/mL, which was much higher than that in water (0.002 mg/mL). Therefore, NOSCS nano micelles may be useful as a prospective carrier for control released agrochemical.

Some compounds that contain glucose groups can be transported across the plasma membrane into the cells through hexose transporters. To test the hypothesis that glucose-conjugated insecticides also have similar uptake and translocation properties, a novel fluorescent conjugate (12) was prepared by conjugating glucose and 7-nitrobenz-2-oxa-1,3-diazole with 4-iodo-1-phenylpyrazoles. Its fluorescence spectra and uptake by suspension-cultured tobacco (Nicotiana tabacum L.cv.) cells were studied. The fluorescence spectra showed long wavelengths with maximum emission at 530 nm. After incubating tobacco cell suspensions in 10 μM conjugate for 0.5 h, green fluorescence of 12 was clearly visible in the cells under fluorescence microscopy. After 2 h of incubation, more than 70% of 12 was absorbed. Carbonyl cyanide m-chlorophenylhydrazone, phloridzin and glucose drastically inhibited uptake. In concentration-dependent uptakes, the uptake rate of 12 showed a saturable component and was in accordance with Michaelis–Menten kinetics. The results proved that the glucose moiety can guide 12 into tobacco cells and that hexose transporters mediated the uptake.

This study examined the photostabilization of rotenone when exposed to ultraviolet light (UV) in the presence of some photoprotectants. Rotenone with and without photoprotectants in methanol were applied onto the surface of glass slides. The remaining concentration of rotenone was analyzed by HPLC at particular intervals. The dissipation half-life values for the degradation of rotenone under ultraviolet radiation were obtained by first-order kinetic equation. It indicated that the addition of UV531, UV326, Photo-stabilizer 770, Photo-stabilizer 622, Vitamin C, Antioxidant 168, Antioxidant 1010 and Antioxidant 1098 provided moderate degree of photostabilization of rotenone and that addition of UV-P provided the best photostabilization, which due to the competitive energy absorption of UV photons by the photoprotectants. The dissipation half-life values of rotenone after irradiation under UV suggested that the addition of UV-P (in 1:1 mol ratio) could provide better photostabilization of rotenone molecule.

The synthesis of one pyrethroid insecticide [2-methyl-4-oxo-3-prop-2-yn-1-ylcyclopent-2-en-1-yl-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylate (Abbrev. JZ) (Fig. 1)] conjugated with a series of α-terthienyl derivatives (2–8) (Fig. 1) by palladium/copper-catalyzed cross-coupling reaction is presented here for evaluating the photoactivated cytotoxicity. The photoactivated cytotoxicity on Spodoptera litura (SL) cell line was detected by MTT assay. The inhibitory activity of all the conjugates was enhanced in the irradiation condition, compared with that of JZ. The IC50 values of the most effective compound 9 (Fig. 1) treated with irradiation were 11.60 μg mL−1 at 24 h and 8.93 μg mL−1 at 48 h, respectively. Generation of intracellular reactive oxygen species (ROS) and change of mitochondrial transmembrane potential (MMP) on SL cells treated with compound 9 were used for the further photoactivated study. A summary of these experiments on compound 9 demonstrated the notable ROS generation and dramatic MMP decrease when irradiated with UVA light. The results also represented statistically significant difference between dark and irradiation treatment of compound 9. However, in control and JZ groups, the effects were not statistically different. It was proved that our prepared compounds were ideal candidates for new photoactivated pyrethroid insecticides.

The photoactivatable insecticides have photoactive features and broad applications. The derivatives of the α-terthienyl analogues were synthesized for evaluating their photolarvicidal activities and 13 2,5-diarylethynylthiophenes were investigated to determine their effect on the second-instar larvae of Plutella xylostella L. Based on their photolarvicidal activities, the 2,5-Dithienylethynylthiophene, 2,5-Diphenylethynylthiophene, 2,5-Di-4-Methoxylphenylethynylthiophene and 2,5-Di-3,4-Methylenedioxyphenylethynylthiophene were found to be the most potent compounds, and their LC50 values were 34.1 mg l−1, 48.4 mg l−1, 60.8 mg l−1 and 42.7 mg l−1, respectively. The relationship analysis between structure and activity showed that the middle thiophene ring played an important role on the activities. The electron donor substituents increased the photolarvicidal activities and the length of the alkyl chain had negative influence on the activities.

Plenty of insect cells are sensitive to azadirachtin, an ideal biopesticide. However, the mode of action of azadirachtin on insect cells still remains unexplained. In this study, Trichoplusia ni BTI-Tn-5B1-4 cell line was used as the model system to evaluate induction of programmed death and cytoskeletal damage by azadirachtin. The result of colorimetric [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide] (MTT) assay indicated that the effect of azadirachtin on cells growth was time-dependent with different concentrations (0.75, 1.5 and 3 μg/mL). As autophagic vacuoles were observed obviously by transmission electron microscopy, azadirachtin could induce autophagy in cells. Furthermore, lysosomal function was affected after treatment with azadirachtin in time-dependent manner. Aberrant morphological figures appeared in treated cells, such as contraction and turning round, especially the appearance of apoptotic bodies and membrane blebbing, which was the evidence of azadirachtin-induced apoptosis, with apoptosis rate of 13.98% at 1.5 μg/mL for 120 h. As the stress fibers changed after treatment, the cytoskeletal function could be influenced. The effect on cell cycle was also analyzed by flow cytometry, with the result that azadirachtin enhanced cell cycle arrest at the G2/M phase gradually.

Huanong AVM is a novel nematicide, which was synthesized from avermectin by selective oxidization of the hydroxyl group at C-4, followed by the reaction with glacial acetic acid. Effects of Huanong AVM on Bursaphelechus xylophilus are reported in this paper. Huanong AVM had strong nematicidal activity against B. xylophilus, with LC50 values of 14.84 μg/mL, 12.49 μg/mL and 11.05 μg/mL, respectively, at 48 h, 72 h and 96 h after treatment. The proteins that responded to Huanong AVM were identified by proteomic analysis. 2-DE analysis revealed eighteen differentially accumulated protein spots. Five spots were successfully identified by MS analysis, and three Huanong AVM induced proteins were annotated in the database as LEVamisole resistant family member (lev-11), ACTin family member (act-4) and Hypothetical protein Y52B11A.10. Two Huanong AVM-up-regulated proteins were assigned as PIP Kinase family member (ppk-3) and Hypothetical protein B0432.6. These proteins are involved in signal transduction, maintaining cytoskeletal structure and LEVamisole resistance. These results point to pathways that are important for the mode of the action of Huanong AVM on B. xylophilus.

Glycinyl fipronil, a new fipronil derivative with 5-amine acylated by glycine, was synthesized. The phloem mobility of the synthesized derivative was higher that of the parent compound in intact soybean seedling and its insecticidal activity against the 3rd instar larvae of Plutella xylostella was comparable with that of fipronil.

A new isoryanodane diterpene, 13-deoxyitol A (1), together with a known isoryanodane diterpene, itol A (2), was isolated from the methanol extract of the seeds of Itoa orientalis. Its structure was determined by spectroscopic means. Compounds 1 and 2 showed antifeedant and contact toxic activities against various insect pests.Download full-size image

The present study revealed differentially expressed genes responsive to azadirachtin A (Aza) in Spodoptera litura cell line through suppression subtractive hybridization. In the Aza-responsive SSH library, approximately 270 sequences represent 53 different identified genes encoding proteins with various predicted functions, and the percentages of the gene clusters were 26.09% (genetic information processing), 11.41% (cell growth and death), 7.07% (metabolism), 6.52% (signal transduction/transport) and 2.72% (immunity), respectively. Eleven clones homologous to identified genes were selected to be confirmed through quantitative real time polymerase chain reaction. Among the eleven clones validated, all but one transcript of lipase showed an increase in SL cell line collected from ETA, whereas the transcripts of other genes were lower in the SL cell line collected from ETA compared with that of UETA. These genes were considered to be related to the response of SL cell line to Aza. These will provide a new clue to uncover the molecular mechanisms of Aza acting on SL cell line.Highlights► A cDNA library of azadirachtin A acting on Spodoptera litura Fabricius cell line was constructed through suppression subtractive hybridization technique. ► Fifty-three differentially expressed genes were found from the Aza responsive SSH library. ► Eleven genes were verified to be regulated by Aza in QRT-PCR. ► The median inhibition concentration of Aza after 48 h exposed to Spodoptera litura cell line was examined.

•Azadirachtin provoked potent growth inhibiting effects in larval Drosophila.•Twenty-eight genes were significantly regulated by 10 μM azadirachtin.•Cuticular protein, amylase and odorant-binding protein were subjected to azadirachtin regulation.•Azadirachtin dysregulated insulin/insulin growth factor signaling pathway.Azadirachtin is a botanical insecticide that affects various biological processes. The effects of azadirachtin on the digital gene expression profile and growth inhibition in Drosophila larvae have not been investigated. In this study, we applied high-throughput sequencing technology to detect the differentially expressed genes of Drosophila larvae regulated by azadirachtin. A total of 15,322 genes were detected, and 28 genes were found to be significantly regulated by azadirachtin. Biological process and pathway analysis showed that azadirachtin affected starch and sucrose metabolism, defense response, signal transduction, instar larval or pupal development, and chemosensory behavior processes. The genes regulated by azadirachtin were mainly enriched in starch and sucrose metabolism. This study provided a general digital gene expression profile of dysregulated genes in response to azadirachtin and showed that azadirachtin provoked potent growth inhibitory effects in Drosophila larvae by regulating the genes of cuticular protein, amylase, and odorant-binding protein. Finally, we propose a potential mechanism underlying the dysregulation of the insulin/insulin-like growth factor signaling pathway by azadirachtin.

Understanding and visualizing the biodistribution of agricultural chemicals inside cells and living plants is very important for enhancing targeting and changing the application approaches of chemicals. Here, a novel material was synthesized through 2,4-dichlorophenoxyacetic acid functionalized small gold nanoparticles (2,4-D-MP-Au NPs). The successful modification of Au NPs (4.46 ± 0.70 nm) was ascertained by UV-vis, TEM, FTIR and XPS. TGA data revealed about 1197 molecules of 2,4-D were coupled to the surface of one Au nanoparticle, which was sufficient for bioapplications. The optical imaging of 2,4-D-MP-Au NPs inside BY-2 cells was directly examined, revealing that 2,4-D-MP-Au NPs could be internalized in BY-2 cells by the two-photo microscopy and endocytosis, as the internalization mechanism was energy dependent for 2,4-D-MP-Au NPs. Furthermore, the biodistribution of 2,4-D-MP-Au NPs in Ricinus cotyledons was measured, revealing that 2,4-D-MP-Au NPs could enter into mesophyll cells of Ricinus cotyledons; the cell recognition was enhanced after 2,4-D conjugated Au NPs. These results indicate that the conjugates have great potential for applications on bioimaging and biolabeling for agricultural chemicals in plant physiology.