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1.
American cockroaches injected with sublethal doses of DDT (0.75 μg/roach) at 5-day intervals showed a 40% reduction in oligomycin-sensitive Mg2+ATPase from muscle homogenates, and a 23% reduction of Na+-K+ATPase from nerve cords. Thus, the maximum effect measured occurred with the same enzyme and tissue as determined from in vitro studies. The metabolite, DDE, used at 15 μg per roach, gave no significant change in activity of the ATPase system following injection. In contrast, high single doses of DDT (7.5 μg/roach) and 100 μg DDE and dicofol per roach caused over 30% increase in oligomycin-sensitive Mg2+ATPase of muscle and a 10–15% increase in Na+-K+ATPase of nerve cords measured 24 and 48 hr later. While a similar response was observed for Mg2+ATPase activities in cockroaches that were immobilized, the increase in enzyme activities were much greater than that caused by the pesticides.  相似文献   

2.
A study was conducted concerning the inhibition of calf thymus nuclear DNA synthesis by captan. Captan was shown to be toxic to the in vitro incorporation of [3H]dTTP into calf thymus DNA, with an ID50 value of 0.16 mM being measured. This inhibition was determined to be independent of Mg2+ concentration. Although intact nuclear activities were affected, the soluble DNA polymerizing activity isolated from calf thymus nuclei exhibited no inhibition when exposed to captan. Treatment of purified calf thymus DNA with 10?5 and 10?4M captan caused an elevation of the Tm by 2 and 6°C, respectively. The inhibitory characteristic of captan on DNA polymerizing activities and the influence of this compound on the thermostability of DNA indicate a mechanism of inhibition which is located in the nucleus and is possibly related to the template function of DNA and/or with the nuclear DNA polymerizing enzymes.  相似文献   

3.
Oligomycin-sensitive (O-S) Mg2+ ATPase from American cockroach muscle was more sensitive to DDT, TDE, methoxychlor, and DDE at cool temperatures than at warm temperature, thus showing a negative temperature effect. In contrast, inhibition by acaricides dicofol, chlorfenethol, and Plictran shows a positive temperature effect. Oxidative phosphorylation in a mitochondrial preparation from cockroach coxal muscle was reduced by DDT, but the reduction was greater at a higher temperature (32°C) than at a cooler temperature (22°C). In addition, Na+K+ ATPase from cockroach nerve cord showed a positive temperature effect with DDT. The inhibition by DDT was much less on Na+K+ ATPase than on O-S Mg2+ ATPase. The negative temperature effect by DDT and analogs on O-S Mg2+ ATPase parallels toxicity effects on insects and fish as reported by numerous researchers. The results provide further evidence for this energy-regulating enzyme being a critical component in the biological action of DDT.  相似文献   

4.
A Ca-ATPase highly sensitive to DDT has been found in peripheral nerves of lobster, Homarus americanus. The observed I50 for this Ca-ATPase toward DDT is on the order of 10?9M and has a low temperature quotien. The ATPase seems to work over a wide range of ATP concentrations. It is stimulated by Ca2+ (optimum 0.1 mM) and shows sensitivity to Na+ (optimum 20 mM) and K+ (optimum 20 mM) ions. The fact that it is highly sensitive to ruthenium red (I50 = 10 μM) suggests that the enzyme is a Ca-ATPase and not a Mg-ATPase. Furthermore the enzyme is not a CaMg-ATPase, since the presence of Mg2+ along with Ca2+ ion is not required for its activity. DDT is found to inhibit the process of Ca2+ binding in the axonic membrane only in the presence of ATP. The evidence suggests the important role of the Ca-ATPase in regulating Ca2+ concentrations in the membrane. The possible significance of DDT inhibition of the ATPase is discussed.  相似文献   

5.
Antibody molecules were produced by injection of BSA-Kelevan into chickens and rabbits. Pure antibody was obtained by a single pass of blood serum through an affinity column. The affinity gel was prepared by covalently binding BGG-Kelevan to activated Sepharose 4B-CN. Purity of the antibody was determined by ultracentrifugation and gel electrophoresis. Properties of the antibody included: sedimentation coefficient = 6.2, pI = 7.0, calculated MW = 150,000, and precipitin band formation using the microouchterlony test. The antibodies in free or immobilized form were able to prevent or reverse Kepone inhibition of ATPase activity from a variety of tissues from different sources. About 70 μg (approx 0.4 μM) of purified antibody was sufficient to restore the activity of mitochondrial (oligomycin-sensitive) Mg2+ ATPase activity which had been inhibited (in vitro) by 1 μM Kepone. The antibody was effective in preventing enzyme inhibition by other organochlorine pesticides with widely differing molecular structures. However, nonchlorinated inhibitors of mitochondrial oligomycin-sensitive Mg2+ ATPase activity were much less affected by the antibody. The available evidence suggests that the antibody binding site for the hapten may be specific for secondary or induced bonding forces due to the carbon-chlorine bonds rather than for a specific molecular structure.  相似文献   

6.
Resistance to many insecticides demonstrated by the beet armyworm,Spodoptera exigua (Hübner), can be caused by the action of carboxylamidases. A colorimetric method, based on the hydrolysis of 4-nitroacetanilide to 4-nitroaniline by carboxylamidases, was used for evaluating biochemical properties of these detoxifying enzymes in beet armyworm. The optimum pH and temperature were 7.5 and 38°C, respectively. Km (Michaelis constant) and Vmax (maximal velocity) at 28°C were 2.3 X 10-4 M and 2.06 nmol min-1 mg protein-1, respectively. The enzyme activity was evaluated in several body parts and located mostly (66.2%) in the midgut. The soluble fraction (supernatant of 105,000g) contained the highest enzyme activity relative to the total (69.3%), and exhibited the highest specific activity. Carboxylamidase activity was totally inhibitedin vitro at a concentration of 10-6 M methomyl. The analysis of thein vitro inhibition kinetics indicated the ability of methomyl and diflubenzuron to inhibit carboxylamidases noncompetitively. Over 95% inhibitionin vivo was obtained when the larvae were fed with castor bean leaves dipped in 250 mgl -1 of methomyl. Thein vivo enzyme activity could be reduced to half with a pretreatment of 15 mgl -1 diflubenzuron.  相似文献   

7.
The enzymatic hydrolysis of trans-permethrin by midgut homogenates of the soybean looper, Pseudoplusia includens (Walker), was characterized through gel electrophoresis, isoelectric focusing, gel filtration chromatography, and selective inhibition. All three separation techniques indicated that one enzyme (or closely related enzyme forms) was responsible for the observed hydrolytic activity. The molecular weight was approximately 80,000, the isoelectric point ranged from pH 4.6 to 4.8, and the apparent Km was 59.5 ± 3.2 μM. Enzyme activity was inhibited by organophosphates, carbamates, and sulfhydryl group inhibitors, as well as some chelators. The hydrolysis of trans-permethrin was distinct from the majority of “general esterase” activity when enzyme activity was separated by electrophoretic techniques.  相似文献   

8.
A continuous steady-state assay procedure was used to investigate the effects of DDT and several analogs on the in vitro Mg2+-stimulated adenosinetriphosphatase of a trout brain mitochondrial fraction. Pharmacological dissection of the enzyme with oligomycin, dicyclohexyl-carbodiimide, and azide failed to yield a fraction specifically sensitive to the organochlorines. At 25°C, low doses of DDT (≤1.35 μmol/mg of protein) stimulated enzyme activity, while methoxychlor was stimulatory at all doses. Higher doses of DDT and of several analogs caused only 45.5% or less inhibition at 25°C, but inhibition increased at lower temperatures. The physiological significance of these effects is discussed.  相似文献   

9.
Earlier communications from this laboratory have shown that DDT inhibited oligomycin-sensitive Mg2+-ATPase (EC 3.6.1.3) but that its active component, F1, was not affected. In the present investigation evidence has been obtained to determine the nature of the requirements for DDT sensitivity. The results showed that DDT sensitivity was conferred to F1 from pig heart mitochondrial preparations when it was bound to F0 from the same preparation. The F1 from house fly (Musca domestica L) thorax was able to bind to F0 from pig heart. This combination showed similar sensitivity to that of the original F1-F0 combination from pig heart mitochondria. However, when F1 from pig heart mitochondria was incorporated into F0 depleted in oligomycin sensitivity-conferring protein (OSCP) from the same source, the resulting ATPase activity was insensitive to DDT. Addition of crude (50–200 μg) or purified (5–20 μg) OSCP in the above preparation restored DDT sensitivity. Presence of dioleyl or dipalmitoyl phosphatidyl choline or Triton X-100 in the reaction medium antagonized the DDT inhibitions. Depletion of phospholipids from submitochondrial membrane preparations (SMP) decreased ATPase activity. Addition of dioleyl or soybean phosphatidyl choline to this lipid-depleted preparation restored DDT sensitivity. Evidence presented suggests that DDT acted on F1 in association with one or more membrane components and that OSCP and phospholipid were essential for DDT sensitivity.  相似文献   

10.
In apterous adults of the spirea aphid, Aphis citricola van der Goot, the optimum conditions for determining acetylcholinesterase (AChE) activity consist of reaction mixture of 0.1 M phosphate buffer (pH 7.5), 10?3M acetylthiocholine (ASCh), and enzyme extract equivalent to 80 ± 3 μg protein incubated for 15 min at 30°C. The Km value for ASCh (6.7 × 10?5M) was much lower than that of butyrylthiocholine (BuSCh) (1.25 × 10?2M). The enzyme activity was almost completely inhibited by 10?6M paraoxon or 10?5M eserine and was 84% inhibited by 10?5M BW284C51 (a specific AChE inhibitor). DTNB was found to inhibit the enzyme activity and was therefore added at the end of the reaction. AChE activity of A. citricola was inhibited in vitro and in vivo by dimethoxon > dimethoate, and aldicarb sulfoxide > aldicarb > aldicarb sulfone. The in vivo effect correlates well with the toxicity level of the various toxicants. A neurotoxicity index which combines both mortality and in vivo inhibition of the aphid AChE activity is suggested as a measure for determining the toxicity of organophosphorus and carbamate compounds toward aphids.  相似文献   

11.
An insect water-soluble ATPase (IF1) was isolated from mitochondria of cockroach coxal muscle. It was homogenous as assessed by polyacrylamide gel electrophoresis and electron microscopy. It was found to be composed of five subunits with molecular weights of 70,000, 68,000, 53,000, 43,000, and 34,000. IF1 was cold labile and showed maximal activity at 47–50°C. Its ATPase activity was Mg+ dependent and was stimulated by DNP and p-hydroxy-mercuribenzoate. This activity was inhibited by sodium azide and guanidine-HCl, not inhibited by oligomycin, and only slightly inhibited at a relatively high level of DDT. The site of action of DDT is discussed in light of relative insensitivity of IF1 ATPase to DDT compared to the high sensitivity of particulate mitochondrial ATPase activity.  相似文献   

12.
The development, fecundity and survival ofStethorus gilvifrons Mulsant (Coleoptera: Coccinellidae) fed onTetranychus cinnabarinus Boisduval (Acari: Tetranychidae) were recorded at three constant temperatures (20, 25 and 30±1°C) and 50±10% relative humidity, under two photoperiods (16:8 L:D and 8:16 L:D) produced using artificial light (4000 lux). The development rate for the egg stage (r[Te]) increased linearly with increasing temperature (r[Te]=0.0132*T ? 0.0955; R2=0.95). The theoretical egg-development threshold was estimated to be 7.24°C; 75.75 degree-days (DD) were required for hatching. The total development time (r[Tt]) also decreased linearly with increasing temperature (r[Tt]=0.0039*T ? 0.0325; R2=0.98). The development threshold was estimated to be 8.33°C and full development from egg to adult required 256.41 DD. Higher temperatures resulted in a shorter generation time (T 0) and decreased net reproductive rate (R 0). The length of the previposition and postoviposition period, as well as longevity, decreased significantly with increasing temperature under both photoperiods. The oviposition and postoviposition periods, longevity, and total fecundity were not significantly affected by photoperiod. The values of both the intrinsic rate of increase (r m ) andR 0 were highest under the long-day photoperiod at 25°C. The mortality rate was lowest at 20°C under the short-day photoperiod. Of the conditions tested, the optimum temperature for rearingS. gilvifrons was 25°C and the optimum photoperiod was 16:8 L:D.  相似文献   

13.
Effects of DDT (1,1,1-trichloro-2,2-bis-(p-chlorophenyl) ethane) on various ATP utilizing enzymes in the lobster peripheral nerve were studied. On the basis of inhibition by ouabain and DDT, four classes of ATPase enzymes were recognized. They are: (1) ATPase activity that is sensitive to both ouabain and DDT inhibition, or Type A, (2) ATPase activity that is sensitive to DDT inhibition only, or Type B, (3) ATPase activity that is sensitive to ouabain only, and (4) ATPase activity that is not sensitive to either ouabain or DDT. The Type A ATPase is considered to be a part of the total (Na+K+) ATPase enzyme associated with the electrogenic pump. The Type B ATPase consisted of an uncharacterized Na+, K+, and Mg2+ stimulated ATPase and includes also a small portion of Mg2+ stimulated ATPase. Ca2+ stimulated ATPase activity was also detected but was not significantly affected by DDT. Proteins with actomyosin-like properties were also recognized to be present, though this superprecipitation process was only slightly affected by DDT.Other systems studied include the transfer of (γ-32P) ATP to endogenous proteins and added histone in the presence and absence of c-AMP. DDT generally stimulated the process of 32P incorporation, while it inhibited a portion of the specific c-AMP dependent protein kinase activity.It was concluded from these studies that DDT has a potential to inhibit or otherwise interfere with a variety of enzymatic reactions that utilize ATP as a substrate. Of these systems, the Type B ATPase bore overall resemblance to the possible target for DDT.  相似文献   

14.
The effects of phosphine on electron transport and on some partial reactions of oxidative phosphorylation of mitochondria from mouse liver, housefly flight muscles and granary weevils has been studied. Phosphine was a strong inhibitor of respiration of mitochondria in the “active” state (state 3), uncoupled state, and ion-pumping state on glutamate, pyruvate plus malate, succinate, α-glycerophosphate, and ascorbate-cytochrome c as substrates. Respiration of mitochondria in state 3 was completely inhibited by about 250 μM phosphine. By contrast, the respiration of mitochondria in state 4 was much less sensitive. This inhibition could not be released by uncouplers suggesting that it is due to a direct effect on electron transport. Only site III was inhibited to any significant extent. Kinetic studies show that the inhibition was noncompetitive with Ki ranging from 1.6×10?5 to 7.2×10?5 depending on the source and purity of cytochrome oxidase. The inhibition of site III was also more pronounced in sonicated particles than in intact mitochrondria. The significance of this is discussed in relation to membrane sideness and topology of the components of the respiratory chain.Phosphine was unable to activate the “latent” ATPase nor did it have any inhibition of the Mg2+-simulated ATPase and only high levels (1.1 mM) showed modest inhibition (41%) of uncoupler-stimulated ATPase. Phosphine had no effect on the ATP-Pi exchange and on the ATP-ADP exchange reaction at concentrations causing strong respiratory inhibition.  相似文献   

15.
Analogues of DDT (ethoxymethyl and methoxymethio derivatives) compared with DDT for their inhibitory action on the ATPase system from tissues of the cockroach, Periplaneta americana show similar, but less inhibitory effects. The mitochondrial (oligomycin-sensitive) Mg2+ ATPase activity from coxal muscle preparations was more sensitive to DDT than the two analogues; whereas, the muscle and nerve cord homogenates showed about equal sensitivity to the biodegradable analogues. The mitochondrial Mg2+ ATPase from nerve cord preparation was more sensitive to the three compounds than the Na+K+ ATPase activity. The significance of these results in relation to recent reports on the effect of DDT on Na+K+ ATPase is discussed.  相似文献   

16.
The phytotoxicities of nine pesticides (paraquat, fluazifop-p-butyl, haloxyfop, flusilazole, cuproxat, cyazofamid, imidacloprid, chlorpyrifos, and abamectin) at practical dosages on photosynthesis were investigated in cucumber (Cucumis sativus L. cv. Jinyan No. 4) by gas exchange and chlorophyll fluorescent measurements. Plants treated with paraquat showed the severest phytotoxic symptom with the highest reduction in net photosynthetic rate (Pn), while other pesticides except flusilazole inhibited Pn to various degrees. The inhibition of Pn by cuproxat was accompanied by declines both in stomatal conductance (Gs) and intercellular CO2 concentration (Ci), whereas decreased Pn for the cyazofamid was associated with increased Ci. For other 6 pesticides, however, inhibition of Pn was accompanied by decrease in Gs, while Ci was increased or unaffected. Paraquat almost completely inhibited the maximal quantum efficiency of PSII (Fv/Fm), while other pesticides had no significant effect on Fv/Fm. Quantum efficiency of PSII (ΦPSII) was significantly reduced by paraquat, fluazifop-p-butyl, and chlorpyrifos and the reduction was mostly attributed to decrease in photochemical quenching coefficient (qP). In comparison, ΦPSII was not significantly affected by haloxyfop, flusilazole, cyazofamid, imidacloprid, and abamectin. Non-photochemical quenching (NPQ) was suppressed by paraquat and haloxyfop, while apparent upregulation was evident after exposure to other pesticides. Interestedly, inhibitions of Pn were alleviated by 24-epibrassinolide (EBR) pretreatment, as for the pesticides examined in this study except paraquat and flusilazole. EBR pretreatment also increased ΦPSII and qP. It is likely that EBR enhanced the resistance of cucumber seedlings to pesticides by increasing CO2 assimilation capacity and activities of antioxidant enzymes.  相似文献   

17.
An insect chitin synthetase (CS) is readily assayed using the microsomal fraction (～0.5 mg protein) from an homogenate of Tribolium castaneum larvae. This enzyme preparation is incubated at 22°C with uridine 5′-diphospho-N-acetyl[3H]glucosamine in 355 μl of 25 mM Tris-HCl buffer containing 10 mM MgCl2, 17 mM N-acetylglucosamine, and 1 mM dithiothreitol. Other divalent cations and amino sugars are less effective activators or are inhibitory. T. castaneum CS is strongly inhibited by polyoxin D and uridine 5′-diphosphate. These activation and inhibition properties of Tribolium castaneum gut CS are similar to those of fungal CS. The polymerization product formed by the Tribolium enzyme is stable in alkali but hydrolyzed by chitinase. Enzymes of Tribolium confusum, Tribolium brevicornis, Tenebrio molitor, and Galleria mellonella are also active under the same conditions. These enzymes are from the gut and probably from the peritrophic membrane. Integumental CS activity is not detected under the indicated assay conditions.  相似文献   

18.
Aspergillus nidulans is able to hydrolyze the herbicide propanil (3′,4′-dichloropropionanilide) with liberation of 3′,4′-dichloroaniline. When the fungus is grown with or without propanil, the hydrolytic activity is identical, but can be increased by starving the mycelium either for carbon, nitrogen, or both carbon and nitrogen. The enzyme which is responsible of this activity is of the aryl acylamidase type (EC 3.5.1 aryl acylamine amidohydrolase). It is also active on propionanilide and acetanilide, two structural analogs of propanil. A value of Km = 0.13 mM has been obtained for propanil. Temperature optimum is 40°C, when assayed with propanil as substrate. Although the pH optimum is 8, there is a relatively high enzyme activity over a wide range of pH values between 7.8 and 10.2. Carbaryl has been found to effectively inhibit the enzyme activity on propanil (Ki = 0.03 mM). The results indicated that the properties of this aryl acylamidase from A. nidulans are very similar to those of enzymes isolated from a variety of organisms such as rice, mammals and the fungus Fusarium solani.  相似文献   

19.
The inhibition of eel acetylcholinesterase by the 4-nitrophenyl esters of monochloromethyl(phenyl)-, dichloromethyl(phenyl)-, and phenyl(trichloromethyl)phosphinic acid (I, II, and III, respectively) was investigated at pH 6.90 in 0.067 M phosphate buffer (25.0°C) using stopped-flow instrumentation and automated data processing. The largest ki value (156,700 M−1 sec−1) was observed for the reaction of I with the enzyme. There is no direct relationship between the hydrolysis rates of the esters and their anticholinesterase activities on eel acetylcholinesterase. Spontaneous reactivation of the phosphinylated enzymes at pH 7.60 showed the following order of activity: I > III > II. The absence of aging is supported by oxime-induced reactivation studies.  相似文献   

20.
Pentachloromethylthiobenzene (PCTA) was synthesized in vitro from pentachloronitrobenzene (PCNB) at pH 7.9 by an enzyme system from onion root that required dithiothreitol, glutathione, and S-adenosylmethionine. The soluble enzyme system was isolated from onion root by ammonium sulfate fractionation and differential centrifugation. The system contained glutathione S-transferase activity with PCNB, C-S lyase activity with S-(pentachlorophenyl)cysteine, S-adenosylmethionine methyl transferase activity with pentachlorothiophenol (PCTP), and presumably several peptidase activities. All activities were stable when the crude enzyme system was stored at ?25°C. Evidence for the following sequence of reactions in PCTA synthesis was presented: PCNB→1S-(pentachlorophenyl)glutathione→2S-(pentachlorophenyl)-γ-glutamylcysteine→3S-(pentachlorophenyl)cysteine→4 PCTP→5 PCTA. The first reaction was studied with [14C]PCNB. Reactions 2–4 were studied with S-([14C]pentachlorophenyl)glutathione, S-([14C]pentachlorophenyl)cysteine, and peptide inhibitors. Reaction 5 was studied with [14C]PCTP, S-[14C]adenosylmethionine, and inhibitors. The possible use of the enzyme system in the characterization of other glutathione conjugates was discussed.  相似文献   

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