Design, synthesis and antifungal/insecticidal evaluation of novel nicotinamide derivatives

A series of nicotinamide derivatives based on Boscalid were prepared and tested for their activities against seven plant pathogenic fungi and two insects. The preliminary bioassays indicated that almost all of the synthesized target compounds displayed the antifungal activities and some of them also had certain insecticidal activities. And, compound 12 showed the strongest activity of all against Rhizoctonia solani (EC₅₀ = 0.010 mg L⁻¹) and Sphaceloma ampelimum (EC₅₀ = 0.040 mg L⁻¹), even stronger than Boscalid, a new nicotinamide fungicide. Additionally, both compounds 1 and 2 showed strong activities against Plutella xylostella (90% and 80%, respectively, at 1000 mg L⁻¹).     

Inhibition of plant pathogens in vitro and in vivo with essential oil and organic extracts of Cestrum nocturnum L.

The efficacy of the essential oil and various organic extracts from flowers of Cestrum nocturnum L. was evaluated for controlling the growth of some important phytopathogenic fungi. The oil (1000 ppm) and the organic extracts (1500 μg/disc) revealed antifungal effects against Botrytis cinerea, Colletotrichum capsici, Fusarium oxysporum, Fusarium solani, Phytophthora capsici, Rhizoctonia solani and Sclerotinia sclerotiorum in the growth inhibition range of 59.2-80.6% and 46.6-78.9%, respectively, and their MIC values were ranged from 62.5 to 500 and 125 to 1000 μg/mL. The essential oil had a remarkable effect on spore germination of all the plant pathogens with concentration and time-dependent kinetic inhibition of P. capsici. Further, the oil displayed remarkable in vivo antifungal effect up to 82.4-100% disease suppression efficacy on greenhouse-grown pepper plants. The results obtained from this study may contribute to the development of new antifungal agents to protect the crops from fungal diseases.     

6,7-亚甲二氧基硫代苯并二氢吡喃-4-酮衍生物的合成及抑菌活性

以亚甲二氧基苯为起始原料,经过溴化、硫化、环化等反应合成了19个未见文献报道的6,7-亚甲二氧基硫代苯并二氢吡喃-4-酮衍生物,其结构均经核磁共振氢谱和质谱确认。初步抑菌活性测试结果表明,在50 mg/L下,目标化合物对供试7种植物病原菌均有不同程度的抑制作用,其中化合物5b、5i、5k和5l对马铃薯干腐病菌 Fusarium solani的抑制率均达75%以上,5h和5i对番茄灰霉病菌Botrytis cinerea的抑制率分别是67.7%和70.4%。     

Comparative antifungal activities and biochemical effects of monoterpenes on plant pathogenic fungi

The antifungal activity of twelve monoterpenes, camphene, (R)-camphor, (R)-carvone, 1,8-cineole, cuminaldehyde, (S)-fenchone, geraniol, (S)-limonene, (R)-linalool, (1R,2S,5R)-menthol, myrcene and thymol was evaluated against four plant pathogenic fungi Rhizoctonia solani, Fusarium oxysporum, Penecillium digitatum and Asperigallus niger by using mycelial growth inhibitory technique. (S)-limonene and thymol were examined for their inhibitory effects on pectin methyl esterase (PME), cellulase and polyphenol oxidase (PPO) of tested fungi. Thymol was the most potent antifungal compound against the four test fungi with EC₅₀ values of 33.50, 50.35, 20.14 and 23.80 mg/L on R. solani, F. oxysporum, P. digitatum and A. niger, respectively. The antifungal activity of thymol was comparable to a reference fungicide, carbendazim. (S)-limonene and 1,8-cineole exhibited pronounced antifungal activity against the four tested fungi. The most effective antifungal compounds thymol and (S)-limonene showed strong inhibitory effect on the activity of PME and cellulase but revealed no inhibitory effect on PPO. The results showed that PME was more sensitive than cellulase to thymol and (S)-limonene. This is the first report on the inhibitory effects of monoterpenes thymol and (S)-limonene on PME, cellulase and PPO. The results indicated that monoterpenes may cause their antifungal activity by inhibiting PME and cellulase. The strong antifungal activity of thymol, (S)-limonene and 1,8-cineole reported in this study indicated that these compounds have a potential to be used as fungicides.     

Effects of plant essential oils and components from Oriental sweetgum (Liquidambar orientalis) on growth and morphogenesis of three phytopathogenic fungi

Commercial plant essential oils obtained from 40 plant species were tested for their antifungal activity against Phytophthora cactorum, Cryphonectria parasitica, and Fusarium circinatum. Strong antifungal activity against Phytophthora cactorum was achieved with the essential oil derived from Oriental sweetgum, Liquidambar orientalis at 28 × 10⁻³ mg/mL air concentration. In a test with C. parasitica, inhibition rate of patchouli was 51.0%, whereas the other essential oils showed weak activity. Essential oils of manuka (Leptospermum scoparium) and patchouli (Pagostemon patchouli) showed moderate activity against F. circinatum. Analysis by gas chromatography-mass spectrometry led to identification of 11 compounds in the oil of L. orientalis. The antifungal activity of identified compounds was tested singularly by using standard or synthesized compounds. Inhibition rates of cinnamyl aldehyde and benzaldehyde were 100% against P. cactorum at 28 × 10⁻³ mg/mL air concentration. There was a significant morphological alternation in three phytopathogenic fungi after oil or compound treatment.     

Antifungal bioactivity of 6-bromo-4-ethoxyethylthio quinazoline

In order to create novel potent antifungal agents, the antifungal effects of 6-bromo-4-ethoxyethylthio quinazoline on plant pathogenic fungi were evaluated by mycelial growth rate method. The bioassay results showed that title compound possesses high antifungal activities on fungi with EC₅₀ values ranging from 17.47 to 70.79 μg/mL. The mechanism of action of 6-bromo-4-ethoxyethylthio quinazoline against fungi was studied in Gibberella zeae model. After treated with title compound at 100 μg/mL for 12 h, the mycelial reducing sugar, chitosan, soluble protein and pyruvate content, chitinase activity showed declining tendency.     

Inhibitory effects of Schiff analogs of salicylidene aniline on phenoloxidase from Pieris rapae L. (Lepidoptera: Pieridae)

In order to gain insight into the development of insecticides with novel modes of action, the effects of salicylidene aniline (a), salicylidene-4-chloroaniline (b), salicylidene-4-bromoaniline (c), and salicylidene-4-nitroaniline (d) on partially purified phenoloxidase (PO) from Pieris rapae L. were investigated. The results showed that the 4 compounds could inhibit PO activity, and the inhibitor concentrations leading to a loss of 50% activity (IC₅₀) were estimated to be 0.025 mmol L⁻¹, 0.732 mmol L⁻¹, 0.471 mmol L⁻¹, and 0.675 mmol L⁻¹, respectively. Meanwhile, all the inhibitors showed reversible competitive inhibition, except (d), which showed reversible mixed inhibition. The K_I values were determined as 0.106 mmol L⁻¹, 10.059 mmol L⁻¹, 8.390 mmol L⁻¹, and 20.198 mmol L⁻¹ for the four compounds, respectively. The UV-vis spectra of (a) and (d) in the presence of copper ions and the enzyme showed that (a) could directly chelate the copper ions of PO; however, (d) could neither chelate the additional copper ions nor the copper ions of PO.     

Effect of waterborne benomyl on the hematological and antioxidant parameters of the Nile tilapia, Oreochromis niloticus

The present study was conducted to determine the 96 h-LC₅₀ of benomyl to the Nile tilapia, Oreochromis niloticus and to investigate the biochemical or hematological indices of blood and the alterations in the antioxidant enzymes of this fish in response to sublethal concentrations of benomyl. Fish weighing 71.61 ± 12.05 g were used in this study; they were subjected to fasting for 4 weeks before treatment. An aqueous solution of benomyl (0, 0.5, 1, 2, 4, 8, and 16 mg L⁻¹) was administered for 96 h to determine the LC₅₀. The 96 h-LC₅₀ value of benomyl was 4.39 (3.23-5.60) mg L⁻¹ in the present study. For 5 weeks, the aqueous solution of benomyl (0, 100, 200, and 400 μg L⁻¹) was administered to investigate its effect on the hematological parameters and antioxidant enzymes. The predominant hematological findings in fish exposed to benomyl were as follows: no significant change in the Hb (g dL⁻¹) level, MCV (μm³), MCH (pg) and MCHC (%) as compared to the control. Benomyl exposure led to greater increases in the GPT, GOT (Karmen-unit), LDH (Wroblewski unit), total cholesterol, Fe, and Ca (mg dL⁻¹) values, whereas the levels of ALP (KA unit), total protein, triglyceride, albumin, and Mg (mg dL⁻¹) did not increase. Benomyl increased the in vivo HSI (%), GST (nmol min⁻¹ mg protein⁻¹), and SOD (U mg protein⁻¹) values in the fish livers in the test group, unlike those in the control group for 5 weeks. At concentrations higher than 100 μg L⁻¹, benomyl affected the GST and SOD levels of Nile tilapia in a dose- and time-dependent manner. The present findings suggest that the in vivo hepatotoxicity associated with benomyl may, in part, result from the hematological index, and antioxidants may provide limited protection against benomyl toxicity.     

Plumbagin as a new natural herbicide candidate for Sicyon angulatus control agent with the target 8-amino-7-oxononanoate synthase

A natural compound plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone) was isolated from the leaves of Plumbago auriculata and found to inhibit the enzyme, 8-amino-7-oxononanoate synthase (AONS, also known as 7-keto-8-aminopelargonate synthase, KAPAS) an IC₅₀ of 2.1 μM in vitro. Biotin supplement significantly rescued the plant injury caused by the plumbagin treatment, and this result confirmed the target site, AONS. Foliar application of 1000 ∼ 2000 μg/mL plumbagin in a greenhouse condition showed lethal activity against eight species of weeds, containing three grass species of Sorghum bicolor, Echinochloa crus-galli, Digitaria sanguinalis and five broad leaf species of Solanum nigrum, Aeschynomene indica, Abutilon avicennae, Xanthium strumarium, Calystegia japonica. Field trial of foliar application with plumbagin 2000 μg/mL have successfully controlled 10 ∼ 15 leaf-stages and 2 ∼ 3 m vine lengths of Sicyos angulatus at the natural habitats around riparian zone in the Nam-Han River in Korea. Visual symptom of desiccation might be induced by the physiological cellular leakage which was significantly dose dependent on the plumbagin treatment regardless of light.     

Synthesis and insecticidal activities of pyridine ring derivatives of podophyllotoxin

In an attempt to find the biorational pesticides, we synthesized 12 pyridinyl derivatives of podophyllotoxin (PPT) and 4′-demthylepipodophylltoxin (4′-DMEP) in this study. Their structures and the α/β substitution at C-4 were confirmed by ¹H NMR, IR, MS spectral analyses and elemental analysis. The insecticidal activities were tested against fifth-instar larvae of Pieris rapae and the third-instar larvae of Cullex pipiens pallens at concentrations of 250 and 10 μg ml⁻¹. Four derivatives of PPT, 4.1, 4.2, 4.3 and 4.5, showed higher insecticidal activities against P. rapae than PPT, while three derivatives of PPT, 4.4, 4.5 and 4.6, displayed higher mosquito larvicidal activity than PPT, with LC₅₀ values of 1.66, 3.96 and 1.54 mg l⁻¹, respectively. Interestingly, we also found that the pyridine ring derivatives of PPT showed delayed insecticidal activity, which is different from traditional neurotic insecticides. The results suggest that 4′-OCH₃ in the PPT derivatives is essential to keep the insecticidal activity and the insecticidal activities of pyridine ring derivatives of PPT are higher than that of the derivatives of 4′-DMEP, supporting PPT has the potential to be a lead structure of semi-synthetic insecticides.     

Effects of different treatment methods of the fungicide jinggangmycin on reproduction and vitellogenin gene (Nlvg) expression in the brown planthopper Nilaparvata lugens Stål (Hemiptera:Delphacidae)

Jinggangmycin is an antibiotic fungicide against the rice sheath blight, Rhizoctonia solani, in China. Previous investigations have shown that jinggangmycin leaf spray stimulated the fecundity of Nilaparvata lugens Stål (Hemiptera:Delphacidae), but the effects of different application methods and concentrations of jinggangmycin on the reproduction of N. lugens have not been investigated. Here, we investigated three treatment methods (foliar and stem sprays and topical treatment) and four concentrations (100, 200, 400 and 800 ppm) of jinggangmycin on the reproduction of adult female N. lugens. The results showed that leaf spray significantly stimulated the fecundity of N. lugens and increased the vitellin content in female ovaries and the gene expression level of vitellogenin (Nlvg), but there was no significant effect on reproduction found for stem spray. In four concentrations of jinggangmycin, leaf sprays at concentrations of 100, 200 and 400 ppm and topical treatments of 100 and 200 ppm significantly increased the number of eggs laid, the vitellin content and the gene expression level of Nlvg compared to the control. Therefore, we suggest that stem spray at high concentration should be applied (if possible) when jinggangmycin is used against rice sheath blight to facilitate harmonious control of rice pests.     

Insecticidal,repellent and fungicidal properties of novel trifluoromethylphenyl amides

Twenty trifluoromethylphenyl amides were synthesized and evaluated as fungicides and as mosquito toxicants and repellents. Against Aedes aegypti larvae, N-(2,6-dichloro-4-(trifluoromethyl)phenyl)-3,5-dinitrobenzamide (1e) was the most toxic compound (24 h LC₅₀ 1940 nM), while against adults N-(2,6-dichloro-4-(trifluoromethyl)phenyl)-2,2,2-trifluoroacetamide (1c) was most active (24 h LD₅₀ 19.182 nM, 0.5 μL/insect). However, the 24 h LC₅₀ and LD₅₀ values of fipronil against Ae. aegypti larvae and adults were significantly lower: 13.55 nM and 0.787 × 10⁻⁴ nM, respectively. Compound 1c was also active against Drosophila melanogaster adults with 24 h LC₅₀ values of 5.6 and 4.9 μg/cm² for the Oregon-R and 1675 strains, respectively. Fipronil had LC₅₀ values of 0.004 and 0.017 μg/cm² against the two strains of D. melanogaster, respectively. In repellency bioassays against female Ae. aegypti, 2,2,2-trifluoro-N-(2-(trifluoromethyl)phenyl)acetamide (4c) had the highest repellent potency with a minimum effective dosage (MED) of 0.039 μmol/cm² compared to DEET (MED of 0.091 μmol/cm²). Compound N-(2-(trifluoromethyl)phenyl)hexanamide (4a) had an MED of 0.091 μmol/cm² which was comparable to DEET. Compound 4c was the most potent fungicide against Phomopsis obscurans. Several trends were discerned between the structural configuration of these molecules and the effect of structural changes on toxicity and repellency. Para- or meta- trifluoromethylphenyl amides with an aromatic ring attached to the carbonyl carbon showed higher toxicity against Ae. aegypti larvae, than ortho- trifluoromethylphenyl amides. Ortho- trifluoromethylphenyl amides with trifluoromethyl or alkyl group attached to the carbonyl carbon produced higher repellent activity against female Ae. aegypti and Anopheles albimanus than meta- or para- trifluoromethylphenyl amides. The presence of 2,6-dichloro- substitution on the phenyl ring of the amide had an influence on larvicidal and repellent activity of para- trifluoromethylphenyl amides.     

Acetylcholinesterase in selected plant-parasitic nematodes: Inhibition, kinetic and comparative studies

The in vitro inhibition potency of some organophosphates (OPs) and carbamates (CAs) which are widely used to control plant-parasitic nematodes on acetylcholinesterase (AChE) of Meloidogyne javanica, Heterodera avenae and Tylenchulus semipenetrans, the major pathogens responsible for the damage of a wide range of crops in Al-Qassim region, Saudi Arabia was examined. AChE of H. avenae activity was 1.58- and 1.51-fold greater than that of T. semipenetrans or M. javanica, respectively. The order of inhibition potency of the tested compounds against T. semipenetrans AChE was: carbofuran > paraoxon > oxamyl > fenamiphos > phorate-sulfoxide > aldicarb, where the corresponding concentrations that inhibited 50% of the nematode AChE activity (I₅₀) were 5 × 10⁻⁸, 7 × 10⁻⁷, 7.5 × 10⁻⁷, 2 × 10⁻⁶, 2 × 10⁻⁴ and 2 × 10⁻³ M, respectively. Paraoxon, fenamiphos and carbofuran exhibited high inhibition potency against M. javanica AChE where the I₅₀ values were below 1 nM. Phorate-sulfoxide and aldicarb were potent inhibitors of M. javanica AChE with I₅₀ values of 3.8 and 8 nM, respectively, while oxamyl exhibited low inhibition potency with I₅₀ of 15 nM. Fenamiphos and paraoxon showed the highest I₅₀ values of <100 μM against H. avenae followed by oxamyl (I₅₀ < 1 mM), whereas paraoxon, carbofuran and aldicarb showed low potency with I₅₀ values >1 mM. All the tested compounds exhibited high inhibition potency to AChE of M. javanica than T. semipenetrans or H. avenae. Except phorate-sulfoxide in M. javanica the inhibition pattern and implied mechanism for all the tested compounds for the three nematodes is suggested to be a linear mixed type (a combination of competitive and non-completive type).     

橙酮类似物的合成及抑菌活性

以芝麻酚和芳香醛为主要原料,经傅-克酰基化、关环和羟醛缩合反应,设计合成了14个未见文献报道的橙酮类似物,其结构均经1H NMR、MS和IR确证。初步抑菌活性测定结果表明:在100 mg/L下,所有目标化合物对7种供试病原菌均有不同程度的抑制作用,其中化合物 3a、3b、3c和3d 对番茄灰霉病菌Botrytis cirerea的抑制率达到100%, 3g 对小麦赤霉病菌Fusarium graminearum、玉米弯孢叶斑病菌Curvulavia lunata、棉花枯萎病菌Fusarium oxysporum vasinfectum、水稻稻瘟病菌Magnaporthe grisea的抑制率均大于80%, 3m 对马铃薯干腐病菌Fusarium solani和玉米弯孢叶斑病菌的抑制率分别为100%和83.8%。     

Influence of three diets on susceptibility of selected insecticides and activities of detoxification esterases of Helicoverpa assulta (Lepidoptera: Noctuidae)

The oriental tobacco worm, Helicoverpa assulta Guenée, is one of the most destructive pests of tobacco and peppers in China. We determined the susceptibility of H. assulta reared on an artificial diet, chili pepper and tobacco to four insecticides (fenvalerate, phoxim, methomyl, indoxacarb) under laboratory conditions associated with the activities of acetylcholinesterase (AChE), carboxylesterase (CarE) and glutathione S-transferase (GST) in its larvae. H. assulta larvae that were fed with chili pepper were more susceptible to fenvalerate, indoxacarb, and phoxim than those that were fed with tobacco and the artificial diet, but not to methomyl. The larvae that were fed with chili pepper were 3.65-, 2.49-, 1.92- and 2.44-fold more susceptible to fenvalerate, phoxim, methomyl, and indoxacarb than those fed with tobacco, respectively. The AChE activities of H. assulta larvae that were fed with chili pepper and tobacco were 2.12 and 1.07 μmol mg⁻¹ 15 min⁻¹, respectively, almost 2-fold difference. The CarE activity of H. assulta larvae that were fed with chili pepper, tobacco and the artificial diet was 4.12, 7.40 and 7.12 μmol mg⁻¹ 30 min⁻¹, respectively. Similarly, the GST activities of H. assulta larvae that were fed with chili pepper, tobacco and the artificial diet was 52.02, 79.37 and 80.02 μmol mg⁻¹ min⁻¹, respectively. H. assulta larvae that were fed with chili pepper were more resistance to the tested insecticides. The low activities of AChE and the high activities of CarE and GST lead to H. assulta become more susceptible to the tested insecticides.     

Ultra-structural changes in the integument induced by the use of three of six forms of allylisothiocyanate tested against Plutella xylostella and Pieris rapae larvae

The insecticidal activity of four forms of Hong Jing (HJ) allylisothiocyanate (AITC), AITC + cypermethrin (HJ_A, HJ_B, and HJ_C) with ratio of (1:1, 4:1, and 2:1), pure AITC (HJ_D), and two forms of Hong Du (HD) AITC, AITC + chlorpyrifos (HD_A and HD_B) with ratio of (2:1 and 2:1), respectively, were studied on the major cruciferous insect larvae Plutella xylostella (L.) and Pieris rapae (L.) by combining both spraying and dipping methods. The P. rapae was more susceptible than P. xylostella larvae. The LC₅₀ values 72 h after treatment of AITC forms (HJ_B, HJ_A, HJ_C, HJ_D, HD_B, and HD_A) on the P. rapae were; 0.07, 0.08, 0.16, 0.83, 0.26, 1.08 gL⁻¹, and 0.69, 0.26, 5.45, 0.93, 3.01, 5.98 gL⁻¹ on the P. xylostella, respectively. The toxicity of some of the AITC forms was very close to or better than that of the commercial contact insecticides such as chlorpyrifos (LC₅₀ = 0.03 and 0.04 gL⁻¹ on P. rapae and P. xylostella, respectively), and cypermethrin (0.65 and 0.78 gL⁻¹, respectively, against P. rapae and P. xylostella). The ultrastructural studies on the integument of the third larval instar of P. xylostella treated by sub-lethal concentration (LC₂₀) of HJ_B, HJ_D, and HD_B were carried out by using transmission electron microscope. The more pronounced alterations in the hypodermis and mitochondria cells. They exhibited changes in all treated samples. The hypodermis was almost completely destroyed, and the mitochondria exhibited morphological alterations, represented by enlargement, matrix rarefaction and vacuolization of the mitochondria matrix, quantity of cristae reduced, and density electron matrix lessened. These AITC forms have potential as contact insecticides, and the ultra structural observations confirm the insecticidal efficiency of different AITC forms on P. rapae and P. xylostella.     

Inhibitory effects of validamycin compounds on the termites trehalase

Trehalase, with the target to control insects, nematodes and fungi, is of increasing interest and has been investigated extensively in recent years. Validamycin compounds, as competitive trehalase inhibitors and lead compounds with broad applications have attracted substantial attention as well. In this study, the characterizations of termites trehalase were investigated and the inhibitory effects of validamycin compounds on the termites trehalase were studied as well. Results showed that the termites trehalase is presumably belonging to the acid trehalase with optimal pH of 3.3 and optimal temperature of 37 °C. It was investigated that the concentrations of validoxylamine A (VAA), validoxylamine B (VBB), validamycin A (VA) and validamycin B (VB) required for 50% inhibition IC₅₀ of termites trehalase were calculated to be 14.73 mg l⁻¹, 20.80 mg l⁻¹, 3.17 × 10³ mg l⁻¹and 2.24 × 10³ mg l⁻¹, respectively. The inhibition kinetic constant K_i values for the above validamycin compounds were 3.2 × 10⁻⁶ mol l⁻¹, 1.03 × 10⁻⁵ mol l⁻¹_, 4.02 × 10⁻⁴ mol l⁻¹and 2.69 × 10⁻⁴ mol l⁻¹, respectively. Validoxylamine A appeared to be the most potential termites trehalase inhibitor among the four compounds.     

Comparison of kinetic properties of a hydrophilic form of acetylcholinesterase purified from strains susceptible and resistant to carbamate and organophosphorus insecticides of green rice leafhopper (Nephotettix cincticeps Uhler)

A hydrophilic form of acetylcholinesterase (AChE) was purified from N-methyl carbamate susceptible (SA) and highly N-methyl carbamate-resistant (N3D) strains of the green rice leafhopper (GRLH), Nephotettix cincticeps Uhler. Both of purified AChE from SA and N3D strains displayed the highest activities toward acetylthiocholine (ATCh) at pH 8.5. In the SA strain, the optimum concentrations for ATCh, propionylthiocholine (PTCh), and butyrylthiocholine (BTCh) were about 1 × 10⁻³, 2.5 × 10⁻³, and 1 × 10⁻³ M, respectively. However, in the N3D strain, substrate inhibition was not identified for ATCh, PTCh, and BTCh to 1 × 10⁻² M. The K_m value in the SA strain was 51.1, 39.1, and 41.6 μM and that in the N3D strain was 91.8, 88.1, and 85.2 μM for ATCh, PTCh, and BTCh, respectively. The K_m value in the N3D strain indicated about 1.80-, 2.25-, and 2.05-fold lower affinity than that of the SA strain for ATCh, PTCh, and BTCh, respectively. The V_max value in the SA strain was 70.2, 30.5, and 4.6 U/mg protein and that in the N3D strain was 123.0, 27.0, and 14.5 U/mg protein for ATCh, PTCh, and BTCh, respectively. The V_max value in the N3D strain was 1.75- and 3.15-fold higher for ATCh and BTCh than that in the N3D strain. However, it was 1.13-fold lower for PTCh. The increased activity of AChE in the N3D strain is due to the qualitatively modified enzyme with a higher catalytic efficiency. The bimolecular rate constant (k_i) for propoxur was 27.1 × 10⁴ and 0.51 × 10⁴ M⁻¹ min⁻¹ in the SA and N3D strain and that for monocrotophos was 0.031 × 10⁴ and 2.0 × 10⁴ M⁻¹ min⁻¹ in the SA and N3D strain. AChE from the N3D strain was 53-fold less sensitive than SA strain to inhibition by propoxur. In contrast, AChE from the N3D strain was 65-fold more sensitive to inhibition by monocrotophos than AChE from the SA strain. This indicated negatively correlated cross-insensitivity of AChE to propoxur and monocrotophos.     

The toxic effects of pyrethroid deltamethrin on the common carp (Cyprinus carpio L.) embryos and larvae

Deltamethrin, a synthetic pyrethroid pesticide contaminating aquatic ecosystems as a pollutant, was investigated in the present study for toxic effects on embryos and larvae of common carp, Cyprinus carpio as a model. The control and five test experiments were repeated five times. The water temperature in the experimental units was kept at 24 ± 1 °C. The number of dead embryos significantly increased in response to deltamethrin concentrations 0.005, 0.05, 0.5, 5, 25, and 50 μg L⁻¹ (p<0.05 for each cases). Dose-response decreases in hatching success were recorded as 75.2, 64.6, 47.4, 26.0, 14.4, and 9.0%, respectively. The lowest concentration of deltamethrin (0.005 μg L⁻¹) produced a significantly decrease in number of dead larvae compared to control group (p<0.05). With increasing deltamethrin concentrations, the larvae exposed duration 1-48 h significantly increased the number of dead larvae (p<0.05 for each cases). The 48 h LC₅₀ values (with 95% confidence limits) of deltamethrin for common carp embryos and larvae were estimated as 0.213 (0.103-0.404) and 0.074 (0.011-0.260) μg L⁻¹, respectively. The results provide evidence that deltamethrin pollution may have an adverse effect on the reproduction and development of carp, which should be considered when this chemical is used in agricultural areas near aquatic ecosystems.     

Functional expression of Helicoverpa armigera CYP9A12 and CYP9A14 in Saccharomyces cerevisiae

Elevated oxidative detoxification is a major mechanism responsible for pyrethroid resistance in Helicoverpa armigera from Asia. Constitutive overexpression of CYP9A12 and CYP9A14 was associated with pyrethroid resistance in the YGF strain of H. armigera. CYP9A12 and CYP9A14 were functionally expressed in the W(R) strain of yeast (Saccharomyces cerevisiae) transformed with a plasmid shuttle vector pYES2. The cell lysates prepared from yeast transformed with CYP9A12 and CYP9A14, respectively, exhibited considerable O-demethylation activities against two model substrates p-nitroanisole (0.59 and 0.42 nmol p-nitrophenol min⁻¹ mg protein⁻¹) and methoxyresorufin (2.98 and 5.41 pmol resorufin min⁻¹ mg protein⁻¹), and clearance activity against the pyrethroid esfenvalerate (8.18 and 4.29 pmol esfenvalerate min⁻¹ mg protein⁻¹). These results provide important evidence on the role of CYP9A12 and CYP9A14 in conferring pyrethroid resistance in H. armigera, and also demonstrate that the yeast expression system can provide necessary redox environment for insect P450s to metabolize xenobiotics.