嗜卷书虱和嗜虫书虱体内磷酸酯酶的生化与毒理学特性比较研究

采用药膜法对嗜卷书虱和嗜虫书虱进行生物测定,从5种杀虫剂对两种书虱的LC50可以看出,嗜卷书虱对两种拟除虫菊酯类药剂较嗜虫书虱敏感,嗜虫书虱对丁硫克百威较为敏感,而对有机磷类药剂的作用两种书虱的敏感性相似。两种书虱体内酸性磷酸酯酶（ACP）和碱性磷酸酯酶（ALP）的活力比较结果表明：嗜卷书虱和嗜虫书虱体内ACP的比活力为分别14 μmol/mg&#8226;30 min和2.2 μmol/mg&#8226;30 min,ALP的比活力分别为0.092 μmol/mg&#8226;30 min和0.046 μmol/mg&#8226;30 min。经方差分析,两种书虱ACP和ALP的比活力之间均存在显著性差异(p<0.05)。动力学参数比较结果显示,两种书虱ACP的V _max值之间的差异达到显著水平,而ALP的K _m值之间存在显著性差异。离体试验结果表明,5种药剂对磷酸酯酶的作用各不相同,个别药剂在低浓度下表现诱导作用,说明磷酸酯酶在不同外源毒物的解毒代谢过程中所起的作用有所不同。     

嗜卷书虱和嗜虫书虱体内磷酸酯酶的生化与毒理学特性比较

采用药膜法对嗜卷书虱和嗜虫书虱进行生物测定,从5种杀虫剂对两种书虱的LC50可以看出,嗜卷书虱对两种拟除虫菊酯类药剂较嗜虫书虱敏感,嗜虫书虱对丁硫克百威较为敏感,而对有机磷类药剂的作用两种书虱的敏感性相似.两种书虱体内酸性磷酸酯酶(ACP)和碱性磷酸酯酶(ALP)的活力比较结果表明:嗜卷书虱和嗜虫书虱体内ACP的比活力为分别14μmol/mg·30 min和2.2 μmol/mg·30 min,ALP的比活力分别为0.092μmol/mg·30 min和0.046μmol/mg·30 min.经方差分析,两种书虱ACP和ALP的比活力之间均存在显著性差异(p<0.05).动力学参数比较结果显示,两种书虱ACP的Vmax值之间的差异达到显著水平,而ALP的Km值之间存在显著性差异.离体试验结果表明,5种药剂对磷酸酯酶的作用各不相同,个别药剂在低浓度下表现诱导作用,说明磷酸酯酶在不同外源毒物的解毒代谢过程中所起的作用有所不同.     

Purification and biochemical characterization of glutathione S-transferases from three strains of Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae): Implication of insecticide resistance

Glutathione S-transferases (GSTs) catalyzing the conjugation of reduced glutathione (GSH) to a vast range of xenobiotics including insecticides were investigated in the psocid Liposcelis bostrychophila Badonnel. GSTs from susceptible and two resistant strains (DDVP-R for dichlorvos-resistant strain and PH₃-R for phosphine-resistant strain) of L. bostrychophila were purified by glutathione-agarose affinity chromatography and characterized by their Michaelis-Menten kinetics towards artificial substrates, i.e., 1-chloro-2,4-dinitrobenzene (CDNB), in a photometric microplate assay. The specific activities of GSTs purified from two resistant strains were significantly higher than their susceptible counterpart. For the resistant strains, GSTs both showed a significantly higher affinity to the substrate GSH while a declined affinity to CDNB than those of susceptible strain. The inhibitory potential of ethacrynic acid was very effective with highest I₅₀ value (the concentration required to inhibit 50% of GSTs activity) of 1.21 μM recorded in DDVP-R. Carbosulfan also exhibited excellent inhibitory effects on purified GSTs. The N-terminus of the purified enzyme was sequenced by Edman degradation, and the alignment of first 13 amino acids of the N-terminal sequence with other insect GSTs suggested the purified protein was similar to those of Sigma class GSTs.     

Purification and partial characterization of a glutathione S-transferase from the two-spotted spider mite, Tetranychus urticae

Glutathione S-transferases (GSTs) are known to catalyze conjugations by facilitating the nucleophilic attack of the sulfhydryl group of endogenous reduced glutathione on electrophilic centers of a vast range of xenobiotic compounds, including insecticides and acaricides. Elevated levels of GSTs in the two-spotted spider mite, Tetranychus urticae Koch, have recently been associated with resistance to acaricides such as abamectin [Pestic. Biochem. Physiol. 72 (2002) 111]. GSTs from acaricide susceptible and resistant strains of T. urticae were purified by glutathione-agarose affinity chromatography and characterized by their Michaelis-Menten kinetics towards artificial substrates, i.e., 1-chloro-2,4-dinitrobenzene and monochlorobimane. The inhibitory potential of azocyclotin, dicumarol, and plumbagin was low (IC₅₀ values > 100 μM), whereas ethacrynic acid was much more effective, exhibiting an IC₅₀ value of 4.5 μM. GST activity is highest in 2-4-day-old female adults and dropped considerably with progressing age. Furthermore, molecular characteristics were determined for the first time of a GST from T. urticae, such as molecular weight (SDS-PAGE) and N-terminal amino acid sequencing (Edman degradation). Glutathione-agarose affinity purified GST from T. urticae strain WI has a molecular weight of 22.1 kDa. N-terminal amino acid sequencing revealed a homogeneity of ≈50% to insect GSTs closely related to insect class I GSTs (similar to mammalian Delta class GSTs).     

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.     

Oxidative stress, steroid hormone concentrations and acetylcholinesterase activity in Oreochromis niloticus exposed to chlorpyrifos

We investigated the endocrine disrupting effects of chlorpyrifos-ethyl which is suspected to be originated from oxidative stress. Initially, the 96 h LC₅₀ values of chlorpyrifos in juvenile and adult of Oreochromis niloticus were determined to be 98.67 μg/L and 154.01 μg/L, respectively. Sub-lethal concentrations of chlorpyrifos-ethyl (5 ppb, 10 ppb, 15 ppb) were administrated to adult fish for 15 and 30 days. Fish were then left to depurate for 15 days in pesticide-free water. Gonadal somatic indices, serum sex steroids as indicators of reproductive function and cortisol level as indicator of stress condition were measured to observe the endocrine disruption effects of chlorpyrifos-ethyl. Gonadal glutathione S-transferase and antioxidant enzyme activities and lipid peroxidation as indicators of oxidative stress were also measured. Acetylcholinesterase activity was measured as a marker of chlorpyrifos toxicity. Results showed that serum estradiol, testosteron and cortisol levels in fish exposed to chlorpyrifos were lower than those of the control fish while gonad somatic indices did not change during the experiments. After 30 days, chlorpyrifos exposure decreased GST activity, and increased SOD enzyme activity by up to 215-446% compared with the control, suggesting there was a oxidative stress. No statistically significant differences between GPx and CAT specific activities, protein contents and lipid peroxidation were determined between control and treatment groups in all exposure concentrations and periods. Acetylcholinesterase activity decreased (45.83-77.28%) in gonad tissues. After recovery serum estradiol and testosteron levels were similar to those of the control levels. An increase in the GST and SOD enzyme activities were determined. Cortisol level and AChE activity in all exposure groups decreased after the depuration period, and fish were unable to overcome the stress of chlorpyrifos. Thus, this study revealed that after chlorpyrifos treatments there exists a protective function of antioxidant enzymes against lipid peroxidation in gonad tissue of O. niloticus. There also exist lower testosteron and estradiol levels in exposed fish than those of the control fish without any alterations in oxidative stress, which is attributed to the capability of chlorpyrifos to impair steroid hormone levels.     

Biological activity of the essential oil from the leaves of Piper sarmentosum Roxb. (Piperaceae) and its chemical constituents on Brontispa longissima (Gestro) (Coleoptera: Hispidae)

The results showed that the essential oil from Piper sarmentosum has strong antifeedant and toxicity effects on Brontispa longissima. The best antifeedant and contact toxicity effects were observed in the 1st-2nd instar larvae. The essential oil also displayed a notable fumigation effect on the eggs and pupae of B. longissima. It took the control-treated insects 43.34 d to complete one generation, while the insects treated with 2000 mg/L essential oil needed 73.58 d. The essential oil was analyzed by gas chromatography/mass spectrometry and 41 components were identified. Myristicin (65.22%) and trans-caryophyllene (13.89%) were the major components. Myristicin exhibited strong antifeedant and contact toxicity effects on both the 3rd instar larvae and the imagoes of B. longissima, when it showed a significant fumigation effect on the eggs and pupae of B. longissima. Another P. sarmentosum essential oil and myristicin all showed a strong inhibiting effect on the growth and development of B. longissima along with the activity of AChE, CarE, GSTs and Na⁺, K⁺-ATPase in B. longissima larvae.     

Subacute chlorpyrifos-induced oxidative stress in rat erythrocytes and the protective effects of catechin and quercetin

This study examined the effects of chlorpyrifos in the rat erythrocyte antioxidant system and evaluated the ameliorating effects of catechin and quercetin on the oxidative damage induced by chlorpyrifos. Sexually mature male Wistar rats were given chlorpyrifos (5.4 mg/kg, 1/25 of the oral LD₅₀), catechin (20 mg/kg), quercetin (20 mg/kg), catechin plus chlorpyrifos, and quercetin plus chlorpyrifos daily via gavage for four weeks. No statistical differences were found in the catechin-only and quercetin-only groups compared with the control group. By the end of the fourth week, chlorpyrifos alone increased the levels of malondialdehyde (MDA) and decreased superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities compared with the control group in rat erythrocytes. In the catechin-plus-chlorpyrifos and quercetin-plus-chlorpyrifos groups, there were statistically significantly decreased MDA levels and increased SOD, CAT, and GPx activities compared with the chlorpyrifos-only group. Thus, it appears that catechin and quercetin ameliorate chlorpyrifos-induced oxidative stress in rat erythrocytes in vivo.     

Inhibition of Na-K-ATPase in different tissues of freshwater fish Channa punctatus (Bloch) exposed to monocrotophos

Freshwater fish, Channa punctatus, commonly known as the snakehead fish, was exposed to two sublethal concentrations (0.96 and 1.86 mg/L) (selected on the basis of 1/20 and 1/10 of 96 h LC₅₀ value) of monocrotophos for two exposure periods (15 and 60 days). Effects of monocrotophos on Na⁺, K⁺-ATPase in liver, kidney, muscle, intestine, brain, heart and gills were determined. Results indicate that Na⁺, K⁺-ATPase activity in tissues decreased as concentration of monocrotophos and exposure period increased. Monocrotophos induced significant inhibitory effects on the Na⁺, K⁺-ATPase activity of C. punctatus, ranging from gills (70%) > Kidney (63%) > brain (57%) > intestine (52%) > liver (50%) > muscle (47%) > heart (44%) inhibition at a sublethal concentration of 0.96 mg/L. Significant inhibition was detected in Na⁺, K⁺-ATPase activity, ranging from gills (90%) > heart (78%) > kidney (78%) > muscle (74%) > intestine (71%) > brain (67%) > liver (63%) at sublethal concentration of 1.86 mg/L. After subacute exposure (15 days) only gills and brain showed significant inhibition after higher concentration (1.86 mg/L). However, it is evident that exposure duration is more important than dose in the inhibition of the activity of enzyme. At lower concentration initial stimulation of the activity of Na⁺, K⁺-ATPase activity was also noticed. It is suggested that the inhibition of the ATPase by monocrotophos blocked the active transport system of the gill epithelial as well as chloride cells, glomerular and epithelial cells of the tubules and thus altered the osmoregulatory mechanism of the fish. In fact, the impairment of the activity of enzymes which carry out key physiological roles could cause alterations of the physiology of the whole organism.     

Mutagenic, genotoxic and enzyme inhibitory effects of carbosulfan in rainbow trout Oncorhynchus mykiss

Rainbow trout (Oncorhynchus mykiss; 116.88 ± 21.69 g) were exposed to sublethal concentrations (25 μg/L) of carbosulfan for 60 days to test if the long term exposure of fish to carbosulfan affects red blood cells acetylcholinesterase (AChE), δ-aminolevulinic acid dehydratase (ALA-D) and paraoxonase (PON) enzyme activity and induces genotoxic and/or mutagenic effects. The exposure resulted in inhibition of AChE and ALA-D activity of rainbow trout when compared to control fish. The activity of PON was not affected by carbosulfan. Interestingly, carbosulfan was found to induce DNA damage in red blood cells (comet assay) and proved to be mutagenic as revealed by the Ames test. Results indicate that blood AChE and ALA-D of rainbow trout may be a sensitive biomarker for assessment of carbosulfan contaminated water bodies. Furthermore, because the Ames test and comet assay were proven successful to detect the genotoxicity of carbosulfan, we proposed that nonlethal techniques such as blood collection from caudal vein of fish should be used to determine potential toxic effects of other pesticides to surrounding environment.     

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.     

Biotransformation of clomazone in rice (Oryza sativa) and early watergrass (Echinochloa oryzoides)

Rice (Oryza sativa), a relatively tolerant species, and early watergrass (Echinochloa oryzoides; EWG), a relatively susceptible species, were exposed to ¹⁴C-labeled clomazone to determine accumulation, biotransformation, and mass balance. On a total mass basis, rice absorbed more clomazone than EWG (p < 0.05), but on a nmol/g basis, there was no significant difference between the two species (p > 0.05). Rice contained more extractable ¹⁴C residues (7.7 ± 0.5 vs. 4.8 ± 0.5 nmol in rice vs. EWG, respectively; p < 0.5), but the concentration in EWG was significantly higher (4.2 ± 0.5 vs. 1.8 ± 0.1 nmol/g in EWG vs. rice, respectively; p < 0.01). More metabolized residue was measured in EWG compared to rice (84.1% vs. 67.9%; p < 0.01). Both species produced hydroxylated forms, β-d-glucoside conjugates, and several other unidentified polar metabolites, but EWG generally produced higher metabolite concentrations. The concentration of the suspected active metabolite, 5-ketoclomazone, was significantly higher in EWG vs. rice (21 ± 2 vs. 5.7 ± 0.5 pmol/g, respectively; p < 0.01). Differences in sensitivity to clomazone between rice and EWG appear to be due to differential metabolism, but in this case the more susceptible EWG qualitatively and quantitatively metabolized more clomazone than the more tolerant rice. This is consistent with the action of a metabolically activated herbicide. This metabolic difference could be exploited to develop herbicide safeners for use with clomazone.     

The effects of Artemisia annua L. and Achillea millefolium L. crude leaf extracts on the toxicity, development, feeding efficiency and chemical activities of small cabbage Pieris rapae L. (Lepidoptera: Pieridae)

The biological effects of two important medicinal plants, Artemisia annua L. and Achillea millefolium (L.) (viz, mortality, growth, and feeding indices as well as enzyme and non-enzymatic activities) were studied on small white Pieris rapae L a deleterious pest of cruciferous plants under controlled conditions (16:8 h L:D at 25 ± 1 °C and 65 ± 5% RH). The LC₅₀ and LC₂₅ values were 9.387% and 3.645% for A. annua L. and 4.19% and 1.69% for A. millefolium (L.), respectively. At the lowest concentration (0.625%), the deterrency was 29.826% and 44.185% for A. annua L. and A. millefolium (L.), respectively. Feeding indices were variously affected with changes in a number of parameters and an increase in larval and pupal duration. The activity level of alkaline phosphatase increased sharply while alanin and aspartate aminotransferases showed a sharp decrease. For non-enzymatic compounds, the amount of glucose and uric acid increased, but total protein and cholesterol decreased. These results indicate that these two medicinal plants might possess potential secondary metabolites that may be useful for controlling potential insect pests.     

In vitro and in vivo effects of some pesticides on carbonic anhydrase enzyme from rainbow trout (Oncorhynchus mykiss) gills

Here we investigated the in vitro and in vivo effects of the pesticides, deltamethrin, diazinon, propoxur and cypermethrin, on the activity of rainbow trout (rt) gill carbonic anhydrase (CA). The enzyme was purified from rainbow trout gills using Sepharose 4B-aniline-sulfanilamide affinity chromatography method. The overall purification was approx. 214-fold. SDS-polyacrylamide gel electrophoresis showed a single band corresponding to a molecular weight of approx. 29 kDa. The four pesticides dose-dependently inhibited in vitro CA activity. IC₅₀ values for deltamethrin, diazinon, propoxur and cypermethrin were 0.137, 0.267, 0.420 and 0.460 μM, respectively. In vitro results showed that pesticides inhibit rtCA activity with rank order of deltamethrin > diazinon > propoxur > cypermethrin. Besides, in vivo studies of deltamethrin were performed on CA activity of rainbow trout gill. rtCA was significantly inhibited at three concentrations (0.25, 1.0 and 2.5 μg/L) at 24 and 48 h.     

Oxygen consumption and ammonia excretion of the freshwater crab Trichodactylus borellianus exposed to chlorpyrifos and endosulfan insecticides

The effects of lethal and sublethal concentrations of chlorpyrifos and endosulfan on oxygen consumption and ammonia excretion rate of the crab Trichodactylus borellianus were evaluated. Oxygen consumption and energy expenditure had significant effect in relation to exposure times. Regarding endosulfan, a significant difference in consumption among times of exposure was registered in 625 μg L⁻¹. Moreover, at the highest concentration, energy expenditure rate was observed stabilized during 1-3 h. A significant increase in ammonia excretion was evidenced in 150 and 300 μg L⁻¹ of chlorpyrifos. The O:N ratio showed a decrease in chlorpyrifos and in 2500 μg L⁻¹ of endosulfan. This indicated a shift towards protein primary metabolism. An increment in the O:N ratio was observed in the lower endosulfan solutions. The relation oxygen:nitrogen showed a shift towards lipid and carbohydrate primary metabolism. This work indicated the complexity of the metabolism in the freshwater crab affected by xenobiotic elements.     

Biochemical effects of acute phoxim administration on antioxidant system and acetylcholinesterase in Oxya chinensis (Thunberg) (Orthoptera: Acrididae)

The study was undertaken to evaluate the effects of different concentrations of phoxim on acetylcholinesterase (AChE) and esterase (EST) activities, and antioxidant system after topical application to Oxya chinensis. The results showed that phoxim inhibited AChE activity, and did not cause significant changes in the EST activity and the levels of malondialdehyde (MDA) and reduced glutathione (GSH). After phoxim administration, superoxide (SOD) and catalase (CAT) activities showed a biphasic response with an initial increase followed by a decline in their activities. Glutathione reductase (GR) and glutathione peroxidase (GPx) activities were inhibited in comparison with the control. Glutathione S-transferase (GST) activity showed irregular changes. Its activity increased significantly at the concentrations of 0.06 and 0.12 μg/μL and decreased at the concentrations of 0.09 and 0.24 μg/μL compared with the control. Changes in SOD, CAT, GST, GPx, and GR activities indicated that phoxim caused oxidative damage in O. chinensis. However, no significant changes in MDA content suggested that these enzymes played important roles in scavenging the oxidative free radicals induced by phoxim in O. chinensis. The formation of oxygen free radicals might be a factor in the toxicity of phoxim.     

Evaluation of molluscicidal activities of Arisaema tubers extracts on the snail Oncomelania hupensis

Four extracts of Arisaema erubescens tubers by acetic acetal (AAE), benzinum (BZE), n-butanol (NBE) and chloroform (CFE) were obtained to evaluate their molluscicidal activities against the snail Oncomlania hupensis. The responses of choline esterase (ChE), alkaline phosphatase (ALP), esterase (EST), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) to the extracts (NBE) were also investigated. In the four extracts (AAE, BZE, NBE and CFE), NBE showed the highest toxicity on the snails after 48 h exposure. NBE also showed the time- and concentration-dependent effect, for example, the LC₉₀ values of the NBE were decreased from 365.5 mg/L (24 h) to 36.4 mg/L (96 h). At the end of exposure to NBE (LC₅₀ concentration), the activities of ChE and ALP in snail tissues (cephalopodium and liver) decreased significantly. Isozyme electrophoresis profiles indicated that responses of isozymes (EST, SOD and GSH-Px) to NBE were more intense in liver than in cephalopodium. After 72 h exposure to NBE, the EST activity in snail liver decreased and some enzyme bands (EST1 and EST4) disappeared. But the activities of SOD 1 and GSH 2 in liver increased after 48 h exposure. The results indicated that NBE was the highest toxic component in the four extracts. The decline of the detoxification ability and the oxidative damage in snail tissues might be the main reason for the molluscicidal activities.     

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.     

Brusatol isolated from Brucea javanica (L.) Merr. induces apoptotic death of insect cell lines

Brucea javanica (L.) Merr. is a medicine plant distributed widely throughout Asia where its bitter fruits have been used traditionally in medicine for treating various ailments and controlling some pests. In recent years, concerns over the potential impact of synthetic pesticides on human health and environment have now become more pressing to develop environmentally friendly pesticides. In this paper, brusatol, a quassinoid, was isolated from the fruit of B. javanica, and identified using X-ray crystallographic analysis. Results showed that brusatol has potent contact toxicity (LD₅₀, 2.91 μg/larva, 72 h) and anfieedant activity (AFC₅₀, 17.4 mg/L, 48 h) against the third-instar larvae of Spodoptera exigua. Brusatol demonstrated cytotoxic effects to the tested insect cell lines, IOZCAS-Spex-II and Sf21, in a time- and dose-dependent manner. After brusatol treatment, apoptotic cell death with the DNA fragmentation, activation of caspase-3 and release of cytochrome c was preliminarily observed in both IOZCAS-Spex-II and Sf21. These results indicated the existence of apoptotic death with the mitochondrial-dependent pathway induced by brusatol in Sf21 and IOZCAS-Spex-II cell lines. Our studies will provide important knowledge to understand mechanisms of action of brusatol and to develop brusatol and its derivatives as insecticides.     

One-tenth dose of LC50 of 4-tert-butylphenol causes endocrine disruption and metabolic changes in Cyprinus carpio

Of the huge annual worldwide production (500,000 MT in 1997) of alkylphenol polyethoxylates (APEs) that are widely used as nonionic surfactants and anti-oxidants in variety of products, 60% ends up in water bodies. They undergo biodegradation to form octyl-, butyl-, and nonyl-phenols. This experiment evaluated effects of 4-tert-butyl phenol (4-TBP) in Cyprinus carpio, a projected candidate species in sewage fed fisheries. The 96th h LC₅₀ of 4-TBP was found to be 6.9 mg/L. Fishes were treated with 1/10th (0.69 mg/L), 1/5th (1.38 mg/L), and 1/3rd (2.3 mg/L) dose of LC₅₀. Whereas there was significant (P < 0.01) decrease in alkaline phosphatase [EC 3.1.3.1] and aspartate aminotransferase [EC 2.6.1.1] activity; alanine aminotranferase [EC 2.6.1.2] and acid phosphatase [3.1.3.2] (except decrease at 1/10th dose of LC₅₀) activity, vitellogenin production in muscle and hepatic- and reno-somatic indices were increased compared to control. With all the dose levels tested, testicular-somatic index (testis size) was reduced (P < 0.01) and histo-architectural changes in testicular and liver tissue were found even in group given 1/3rd dose of LC₅₀.