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1.
Growth of Penicillium digitatum was inhibited after a 40-min incubation in a culture medium containing 0.5 mM sec-butylamine, and the dry weight of the hyphae was 50% of the control value after 180 min. Respiration of the hyphae was reduced 13% after a 20-min contact with 0.5 mM sec-butylamine but this treatment did not influence the uptake of amino acids, glucose, or phosphate nor intensify the efflux of 33P- or 14C-labeled metabolites from the cells. The syntheses of cell walls and total lipids were inhibited 20–30% after a 90-min incubation with sec-butylamine, and nucleic acid synthesis was reduced to about 50% of the control value at this time. sec-Butylamine inhibited the incorporation of labeled carbon from [14C]glucose into the protein fraction of the hyphae to a greater degree than 14C derived from labeled proline, lysine, or leucine. These observations suggested that sec-butylamine interfered primarily with the intermediary metabolism of glucose rather than inhibiting a later stage of macromolecule synthesis. Hyphae incubated with [14C]glucose and sec-butylamine accumulated pyruvic acid to a level seven times greater than in control hyphae. Furthermore, sec-butylamine strongly inhibited 14CO2 evolution from hyphae metabolizing [14C]pyruvate whereas CO2 derived from acetate or glucose after a 45-min incubation was only slightly reduced by sec-butylamine. These observations implicate pyruvate oxidation as the primary site of sec-butylamine action in young hyphae of P. digitatum.  相似文献   

2.
A comparison of the uptake of simple (C1–C4) aliphatic amines byPenicillium digitatum revealed that the fungitoxicity of (?)sec-butylamine [(?)SBA] was not due to its accumulation by hyphae since (+)SBA was accumulated to the same extent and methylamine, which showed negligible antifungal activity, accumulated to twice the level of SBA. Amines with a secondary alkyl structure were resistant to fungal metabolism, whereas primary amines were degraded to a significant extent byP. digitatum during a 4 h incubation period.(?)SBA accumulated in the fresh hyphae ofP. digitatum to a level 24 times higher than that in the culture medium containing 1 μmol ml?1; most of the SBA effluxed from the hyphae when transferred to fresh culture medium minus SBA. SBA did not accumulate when hyphae were incubated in N2 or in the presence of respiratory inhibitors. The absorption and accumulation of SBA is characteristic of active transport.Penicillium species and biotypes that are sensitive to SBA did not accumulate more (?)SBA than resistant fungi. SBA-resistant biotypes ofP. digitatum accumulated SBA to twice the level of SBA-sensitive biotypes, but did not accumulate pyruvate in the hyphae, which is characteristic of SBA-altered metabolism.No evidence was found to implicate exclusion or metabolic detoxification as mechanisms of SBA resistance. More probably, resistance involves cytoplasmic sequestration of SBA or low affinity of the biochemical target, pyruvic dehydrogenase.  相似文献   

3.
Fluorescamine, 4-phenylspiro[furan-2(3H), 1′(3′H)-isobenzofuran]-3,3′-dione, derivatives of primary aliphatic amines were separated by high-pressure liquid chromatography using a reverse-phase system with fluorescence detection. This technique was applied to the determination of residues of the fungicide sec-butylamine in potato tubers; the limit of detection was 0.36 pmol, equivalent to a residue of 0.1 mg kg?1 in potato samples. A second amine, phenethylamine, was identified in extracts from artificially rotted potato flesh but this did not interfere with the analysis of sec-butylamine residues.  相似文献   

4.
At concentrations near 2 × 10?4M, barban, chlorpropham, and phenmedipham are inhibitors of the electron transfer in potato and mung bean mitochondria. The inhibition seems to be localized in the flavoprotein region. It affects preferentially the exogenous NADH dehydrogenation, in potato mitochondria (I50, 10?4M). Succinate dehydrogenation is less inhibited. At noninhibiting concentrations, the studied carbamates cannot uncouple the oxidative phosphorylations. Photosynthesis is completely inhibited by 2.10?7M phenmedipham, 5 × 10?5M barban, and 2 × 10?4M chlorpropham. The inhibition takes place at the PS II level. Moreover, barban and chlorpropham are uncouplers of the photophosphorylations for concentrations between 5 × 10?5 and 5 × 10?4M. The effects observed on mitochondrial respiration can also be found on respiration of Acer cultured cells. The effects on isolated chloroplast photosynthesis are also observed for slightly higher concentrations on cultured Chlorella and on pea and oat leaf fragments.  相似文献   

5.
Phenoloxidase from the mantle of the marine bivalve Modiolus demissus Dillwyn slowly catalyzes the oxidation of the rosewood ingredient obtusastyrene (Km 0.13 mM, Vmax 0.30 mM/min/mg). However, in the presence of another rosewood ingredient, obtusaquinone, the oxidation rate is increased to a limiting maximal velocity of 11 mM obtusastyrene/min/mg, without a concommitant change in the Km. The oxidation products of either the slow reaction or the obtusaquinone-enhanced reaction inhibit the catechol dehydrogenase function of phenoloxidase. The phenoloxidase-modifying properties of obtusaquinone and obtusastyrene may be related to the inhibition of shell formation in wood-boring bivalve larvae settling on rosewood.  相似文献   

6.
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   

7.
A “soluble” glutathione S-transferase that catalyzes the cleavage of the herbicide, 2,4′-dinitro-4-trifluoromethyl diphenylether (fluorodifen), was isolated and partially characterized from epicotyl tissues of pea seedlings. A 32-fold purification of the enzyme was achieved by differential centrifugation, ammonium sulfate precipitation, Sephadex gel filtration, and DEAE-cellulose ion exchange chromatography. The enzyme had a pH optimum of 9.3–9.5 and was specific for reduced glutathione, with an estimated apparent Km value of 7.4 × 10?4M. Limited specificity studies with four substituted 14C-labeled diphenylether compounds indicated that fluorodifen was the only effective substrate, with an estimated apparent Km value of 1.2 × 10?5M. Differences and similarities between the pea epicotyl enzyme and other plant and animal glutathione S-transferases were discussed from the standpoint of substrate specificity, pH optima, distribution, stability, and inhibitor studies.  相似文献   

8.
A rat hepatocyte suspension effectively epoxidized aldrin to dieldrin with a Vmax of 7.19 mol/mol P-450/min and a Km of 9.27 μM. Viability and metabolic activity were stable for 6 hr after isolation when cells were maintained at room temperature (20°C) with the gentle introduction of O2CO2 onto the surface of the suspension. The cytochrome P-450 content of the suspension was 303 pmol/106 cells. Primary maintenance culture of the cells also epoxidized aldrin. During culture for 3 days, metabolic activity decreased slowly day by day. Metabolic activity of microsomal fraction from rat liver was also examined. Microsomes epoxidized aldrin with a Vmax of 5.11 mol/mol P-450/min and a Km of 1.64 μM. Significant loss of some subspecies of cytochrome P-450 during fractionation of liver homogenate was indicated.  相似文献   

9.
The inhibition of eel acetylcholinesterase and bovine erythrocyte acetylcholinesterase by the 4-nitrophenyl esters of methyl-, ethyl-, and isopropyl(phenyl)phosphinic acid (MPP, EPP, and IPP, respectively) was investigated at pH 6.90 in 0.067 M phosphate buffer (25.0°C) using stopped-flow instrumentation and automated data processing. Our evaluation of the dissociation constant, Kd, the unimolecular bonding rate constant, k2, and the bimolecular reaction constant, ki, are the first reported values for these constants for a homologous series of this class of organophosphorus compounds. The largest k1 value (29,428 M?1 sec?1) was observed for the reaction of eel acetylcholinesterase with 4-nitrophenyl methyl(phenyl)phosphinate. The smallest ki value (9.6 M?1 sec?1) was observed for the reaction of bovine erythrocyte acetylcholinesterase with 4-nitrophenyl isopropyl(phenyl)phosphinate.  相似文献   

10.
Long-term experiments with dactyl cells of Nitella flexilis showed that the herbicide 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) at a concentration of 1 × 10?5M affected not only O2 evolution in the light but also O2 uptake in the dark. The inhibition of O2 production was transitory, but dark respiration did not recover. DCMU induced the formation of giant mitochondria which disappeared before cell death. It was concluded that the algicidic effect of 1 × 10?5M DCMU on N. flexilis, but not necessarily the elongation of mitochondria, was due to the inhibition of mitochondrial respiration and not of photosynthesis.  相似文献   

11.
During storage of potato tubers (Solanum tuberosum L., var. Bintje), important changes appear which affect respiratory control, ADPO, intensity of O2 consumption in the presence of different substrates, and NAD+ dependence, In mitochondria extracted under strictly similar conditions, from patato tubers stored at 4°C, the respiratory control (RC) maintains a value near 4 for 4 to 5 months. It then declines progressively to low values. At 20°C, a stable RC of 4 can be observed for several months, which then decreases at the end of dormancy. Then, the RC increases sharply; at this stage, ADPO are abnormally low, and, some time later, NAD+ dependence disappears. Mitochondria treated with 250 μM chlorpropham show a 50% inhibition of the electron transfer with exogenous NADH as substrate. After tuber treatment with 1% chlorpropham, sprouting is inhibited for several months. The activities of mitochondria extracted from such tubers remain unaffected by the treatment. The use of this phenylcarbamate for potato tuber treatment permits obtaining functional mitochondria from tubers after a slightly longer period of storage.  相似文献   

12.
A series of compounds containing the trifluoromethylketone group have been synthesized utilizing either a modified Grignard procedure or by reacting selected aliphatic bromides or tosylates with the Collman reagent [Na2Fe(CO)4]. When tested in vitro as inhibitors of crude juvenile hormone esterase from the hemolymph of the cabbage looper, Trichoplusia ni (Noctuidae), the most active compounds were trifluoromethylketones possessing either a juvenoid-like structure or a straight aliphatic chain. The logarithm of the inhibitory potency of the aliphatic compounds was proportional to their chain length, up to 1,1,1-trifluorotetradecan-2-one (I50 = 1 × 10?7M). This powerful inhibition was found to be highly selective for JHE, reversible, competitive by Lineweaver-Burk analysis, and was characterized by high affinity of the inhibitor for the esterase (Ki = 3.2 × 10?9M, Km JH III = 2 × 10?7M). Other trifluoromethylketones were shown to be inhibitors of T. ni α-naphthylacetate esterase and bovine trypsin. By analogy with the mechanism of trypsin action, trifluoromethylketones are probably potent inhibitors due to their resemblance to a tetrahedral transition state on the reaction coordinate to the acylated enzyme.  相似文献   

13.
Several aryl N-hydroxy- and N-methoxy-N-methylcarbamates were examined as inhibitors of bovine erythrocyte acetylcholinesterase (AChE). These carbamate derivatives were generally strong inhibitors of AChE, but, unlike the typical N-methyl- and N,N-dimethylcarbamates which are carbamylating agents, they proved to be reversible, competitive inhibitors of the enzyme. The values for the dissociation constant (Ka) for the enzyme-inhibitor complex to enzyme and inhibitor were in the range of 2 × 10?5?1 × 10?7M.  相似文献   

14.
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   

15.
Homogenates of three strains of Myzus persicae, A, R, and E, with an LD50 for topically applied parathion of 9, 93, and 263 ng per aphid, showed an in vitro hydrolytic degradation of paraoxon of 2.3, 4.7, and 8.6 pmol/mg aphid/h, respectively. These values represent Vmax; Km was <10?7M. The three strains showed a malaoxon degradation of 2.4, 11.9, and 18.8 pmol/mg/h at 10?6M substrate concentration. Vmax for R and E was 21 and 27 pmol, respectively and Km 7 and 4 × 10?7M. Activity in strain A was too low to estimate these entities. The breakdown product of paraoxon was mainly diethyl phosphoric acid, that of malaoxon mainly dimethyl phosphoric acid. No hydrolysis of the carboxylester groups of malaoxon was found. Hydrolysis of paraoxon and malaoxon was inhibited by isopropyl and n-propyl paraoxon and by the salioxon-analog K2. The two latter compounds were shown to act as synergists with parathion when added in amounts that caused little mortality when given alone. The hydrolytic enzyme is soluble and retains its activity during incubations of several hours. It is likely that it is responsible for at least part of the resistance. Resistance was maintained without selection over a period of three years. There was no correlation between degree of resistance and carboxylesterase activity of the strains.  相似文献   

16.
The association equilibrium constant, 1Kd, and the carbamylation constant, k2, of 53 o-, m-, and p-substituted phenyl N-methylcarbamates with bovine erythrocyte acetylcholinesterase were determined. The 1Kd value varied 1000-fold, whereas the k2 value did not depend upon the nature and position of substituents. The variation in log(1Kd) was analyzed using free energy related substituent parameters and regression analyses. The effect of substituents at o-, m-, and p-positions was nicely separated into hydrophobic, electronic, hydrogen bonding, and proximity (steric and field electronic for o-substituents) factors. The physicochemical significance of these factors was established by comparison with those for model organic reactivities. The mechanism of the whole reaction process was elucidated in terms of physical organic chemistry.  相似文献   

17.
The effects of chlordimeform on rectus abdominis muscle of frog were investigated. Chlordimeform (10?3M) caused a slow contraction, and at lower concentration (10?5–10?3M) it inhibited the acetylcholine-induced contraction in noncompetitive manner. When chlordimeform was applied to the muscle of Rana catesbiana, K+-induced contraction was also inhibited in noncompetitive manner. Whereas it had no effect on caffeine-induced contraction.Chlordimeform-induced contraction was not affected by respective addition of d-tubocurarine (10?4M), procaine (10?3M), or eserine (0.3 mM), which results were same as that of K+-induced contraction. Chlordimeform, at lower concentration (10?5–10?3M), inhibits the acetylcholine- and K+-induced contractions probably owing to depression of not only the sensitivity of endplate but also the excitability of cell membrane.  相似文献   

18.
The hydrolysis of malation by rabbit liver oligomeric and monomeric carboxylesterases (CE's) (EC 3.1.1.1) results in the formation of a mixture of α- and β-monoacids. A new chromatographic procedure was utilized to investigate the formation of α- and β-monoacids. The oligomeric carboxylesterase (oCE) produced an αβ ratio of monoacids of 4.55, and the monomeric carboxylesterase (mCE) produced an αβ ratio of monoacids of 2.33. The ratios of α- and β-monoacids were independent of the initial concentration of malathion and remained constant over the time course of the reaction. Kinetic studies demonstrated that the Km values were the same for the corresponding reactions which produced either α-monoacid or β-monoacid with the same enzyme. Since both carboxylesterases are electrophoretically pure, the kinetic data strongly supports the theory that the reactions which produced α- and β-monoacids are catalyzed by the same active site. Comparison of the kcat and Km values governing the hydrolysis of malathion by the two esterases, together with their relative abundance in liver, indicated that the oCE would be responsible for about 80 to 98% of the hydrolytic detoxication of malathion by rabbit liver.  相似文献   

19.
The action of insecticides on the spontaneous electrical activity of neurohemal tissue in the stick insect, Carausius morosus, has been studied using extracellular electrodes. The pyrethroid, permethrin, causes a massive increase in the frequency of the spontaneously generated action potentials at concentrations between 5 × 10?5 and 5 × 10?8M. Concentrations as low as 5 × 10?11M are still effective in producing bursting activity.DDT, at concentrations between 5 × 10?5M and 5 × 10?6M, produces an overall increase in activity although the bursting activity is less violent than that shown with permethrin. DDT, 5 × 10?7M, is ineffective at altering the resting pattern.Carbaryl and coroxon cause a transitory increase in electrical activity at 1 × 10?4M, but are ineffective at 1 × 10?5M.It is concluded that insecticides could have a direct effect upon the neurohormonal balance in insects.  相似文献   

20.
A series of 27 substituted aryl N-methoxy-N-methylcarbamates were synthesized and their ability to reversibly inhibit house fly-head and bovine-erythrocyte acetylcholinesterase and horse-serum cholinesterase was determined. These compounds were all competitive, reversible inhibitors of bovine erythrocyte acetylcholinesterase but some of them showed mixed competitive inhibition against the house fly-head and horse-serum enzymes. Dissociation constants (Ki) as small as 9.9 × 10?9M and as large as 1.4 × 10?4M were observed. A highly satisfactory correlation between log Ki for the inhibition of fly-head acetylcholinesterase by the N-methoxy-N-methylcarbamates and ?log I50 for the inhibition of the same enzyme by the corresponding methylcarbamates was noted. Analysis of the anticholinesterase data by multiple regression showed -log Ki to be related to Hansch's π constant and ring position terms. The results indicate that reversible binding of these compounds to acetylcholinesterase occurs by hydrophobic bonding.  相似文献   

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