Thiocarbamate fungicides: reliable tools in resistance management and future outlook.

Among contact fungicides, dithiocarbamates have remained successful and are used worldwide. These organic sulfur fungicides, viz. mancozeb, maneb, zineb, ziram, thiram, metiram and propineb, have helped growers manage several economically important plant diseases. Their multi-site mode of action and broad-spectrum disease control make them some of the most common partners in mixtures of a number of single-site fungicides as part of resistance management strategies. Indeed, it was the part played by ethylene-bis-dithiocarbamates such as mancozeb in delaying the evolution of phenylamide resistance in several oomycete phytopathogens that laid the groundwork for mixture strategies to become a cornerstone of anti-resistance management in plant disease control. Dithiocarbamates, however, do not have systemic action, are only surface protectants and have to be applied prior to pathogen infection. Dithiocarbamates will likely continue play a key role as reliable resistance management tools to prolong the efficacy of single-site fungicides. The primary metabolite ethylene thiourea produced by some of these fungicides is considered a reproductive and endocrine disrupter in animals. Therefore, dithiocarbamates need to be used at reduced rates or in slow-release formulations. © 2017 Society of Chemical Industry.

A novel dithiocarbamate derivative induces cell apoptosis through p53-dependent intrinsic pathway and suppresses the expression of the E6 oncogene of human papillomavirus 18 in HeLa cells.

Dithiocarbamates (DTCs) exhibit a broad spectrum of antitumor activities, however, their molecular mechanisms of antitumor have not yet been elucidated. Previously, we have synthesized a series of novel dithiocarbamate derivatives. These DTCs were examined for cytotoxic activities against five human cancer cell lines. In this study, one of dithiocarbamate (DTC1) with higher potential for HeLa cells was chosen to investigate molecular mechanisms for its anti-tumor activities. DTC1 could inhibit proliferation, and highly induce apoptosis in HeLa cells by activating caspase-3, -6 and -9; moreover, activities of caspase-3, -6 and -9 were inhibited by pan-caspase inhibitor, Z-VAD-FMK. Furthermore, DTC1 decreased the levels of Bcl-2 and Bcl-xL, and increased expression of cytosol cytochrome c, Bak, Bax and p53 in a time-dependent manner but had no effect on the level of Rb. It was shown that DTC1 induced HeLa cells apoptosis through a p53-dependent pathway as tested by the wild type p53 inhibitor, pifithrin-α. Additionally, the relative expression of E6 and E7 were evaluated in HPV18-positive (HeLa cells) by real-time PCR and western blotting. The results firstly demonstrated that DTC1 suppressed both expression of E6 mRNA and E6 oncoprotein, but had no effect on the expression of E7 mRNA and protein in HPV18. Our results suggested that DTC1 may serve as novel chemotherapeutic agents in the treatment of cervical cancer and potential anti-HPV virus candidates that merit further studies.

Synthesis, molecular modeling and biological evaluation of dithiocarbamates as novel antitubulin agents.

A series of novel dithiocarbamate compounds with the chalcone scaffold have been designed and synthesized, and their biological activities were also evaluated as potential antiproliferation and antitubulin polymerization inhibitors. Compound 2n showed the most potent biological activity in vitro, which inhibited the growth of MCF-7 cells with IC(50) of 0.04+/-0.01 microM and the polymerization of tubulin with IC(50) of 6.8+/-0.6 microM. To understand the tubulin-inhibitor interaction and the selectivity of the most active compound towards tubulin, molecular modeling studies were performed to dock compound 2n into the colchicine binding site, which suggested probable inhibition mechanism.

[Synthesis and in vitro antitumor activity of 4(3H)-quinazolinone derivatives bearing dithiocarbamate chains].

A series of 4(3H)-quinazolinone derivatives bearing dithiocarbamate side chains have been synthesized through the reaction of 6-bromomethyl-2-methyl-4(3H)-quinazolinone with CS2 and various amines in the presence of anhydrous K3PO4, and their structures were confirmed with ESI-MS, H NMR, elemental analysis or HRMS. The target compounds 8a -8q were tested for their in vitro antitumor activity against human myelogenous leukaemia K562 and human Hela cell lines by means of colorimetric MTT assay. Among the tested compounds, 8q exhibited in vitro inhibitory activity against K562 and Hela cells with IC50 values of 0.5 and 12.0 micromol x L(-01), respectively. Therefore, compound 8q is worthy to be a lead compound for the design and synthesis of new antitumor agents.

Microglial activation induced by neurodegeneration: a proteomic analysis.

Neuroinflammation mediated by microglial activation appears to play an essential role in the pathogenesis of Parkinson disease; however, the mechanisms by which microglia are activated are not fully understood. Thus, we first evaluated the effects of two parkinsonian toxicants, manganese ethylene bisdithiocarbamate (Mn-EBDC) and 1-methyl-4-phenylpyridine (MPP+), on microglial activation as well as associated dopaminergic (DAergic) neurotoxicity in primary cell culture systems. The results demonstrated that, when rat primary mesencephalic neuron-enriched or neuron-microglia mixed cultures were treated with Mn-EBDC at 2-8 microm or MPP+ at 0.25-5 microm, respectively, for 7 days, both toxicants were capable of inducing DAergic neurodegeneration as well as activating microglia via a mechanism secondary to DAergic neurodegeneration. Furthermore activated microglia subsequently enhanced DAergic neurotoxicity induced by Mn-EBDC or MPP+. Detailed scrutiny of neuron-microglia interactions identified a fraction of the conditioned media derived from a DAergic cell line treated with Mn-EBDC or MPP+ that potently activated microglia. To further define potential mediators leading to microglial activation secondary to neurodegeneration, we utilized a quantitative proteomic technique termed SILAC (for stable isotope labeling by amino acids in cell culture) to compare the protein profiles of MPP+-treated cellular fraction that mediated microglial activation as compared with controls. The search revealed numerous novel proteins that are potentially important in neurodegeneration-mediated microglial activation, a process believed to be critical in Parkinson disease progression.

Proteasomal inhibition induced by manganese ethylene-bis-dithiocarbamate: relevance to Parkinson's disease.

Maneb, a widely used fungicide, has been associated with Parkinsonism in humans. In experimental models, maneb and its major active element, manganese ethylene-bis-dithiocarbamate (Mn-EBDC) cause selective nigrostriatal neurodegeneration in mice and in rats, respectively. To investigate the mechanisms underlying this neurodegeneration, we studied the effects of Mn-EBDC on proteasomal function, which is decreased in patients with Parkinson's disease (PD), in a dopaminergic neuronal cell line (MES 23.5 or MES). The results demonstrated that exposure of MES cells to 6 microM Mn-EBDC for 7 days produced not only significant neurotoxicity but also inhibition of proteasomal chymotrypsin-like and postglutamyl peptidase activities. Proteasomal dysfunction was accompanied by formation of cytoplasmic inclusions that were positive for alpha-synuclein immunostaining and significantly increased sodium dodecyl sulfate-insoluble alpha-synuclein aggregation seen by Western blot analysis. In addition, there was a significant increase in oxidative stress, evidenced by elevated total protein carbonyl content, in cells treated with Mn-EBDC. Manipulation of intracellular reduced glutathione levels with N-acetyl-L-cysteine or L-buthionine sulfoximine pretreatment to modulate Mn-EBDC-mediated oxidative stress altered Mn-EBDC-mediated neurotoxicity, proteasomal dysfunction, and alpha-synuclein aggregation in these cells. These data suggest that neurotoxicity-induced by Mn-EBDC is at least partially attributable to Mn-EBDC-mediated proteasomal inhibition, and that the proteasome may be an important target by which environmental exposure modifies the risk for developing PD in vulnerable populations.

Increased synaptosomal dopamine content and brain concentration of paraquat produced by selective dithiocarbamates.

Exposure to pesticides may be a risk factor for Parkinson's disease based on epidemiologic data in humans, animal models and in vitro studies. Different dithiocarbamate pesticides potentiate the toxicity of both 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and paraquat in mouse models of Parkinsonism by an unknown mechanism. This study examined the effects of commercially used dithiocarbamates on [3H]dopamine transport in striatal synaptosomal vesicles and on the concentration of [14C]paraquat in vivo in mice. Different ethylenebis-dithiocarbamates and diethyl-dithiocarbamate increased dopamine accumulation in synaptosomes, whereas dimethyl-dithiocarbamate and methyl-dithiocarbamate did not. Increased dopamine accumulation in synaptosomes was dose dependent and was related to the carbon backbone of these molecules. The dithiocarbamates that increased accumulation of dopamine did not alter the influx of dopamine, but rather delayed the efflux out of synaptosomes. These same dithiocarbamates also increased the tissue content of [14C]paraquat in vivo by a mechanism that appeared to be distinct from the dopamine transporter. There was a consistent relationship between the dithiocarbamates that increased synaptosomal accumulation of dopamine and tissue content of paraquat, with those previously demonstrated to enhance paraquat toxicity in vivo. These results suggest that selective dithiocarbamates may alter the kinetics of different endogenous and exogenous compounds to enhance their neurotoxicity.

Ethylenebisdithiocarbamate enhances MPTP-induced striatal dopamine depletion in mice.

Diethyldithiocarbamate (DDC) has been shown to enhance 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced striatal dopamine depletion in mice. Surprisingly, although DDC is a prototypic member of a class of compounds called dithiocarbamates (DTCs) that are widely used in industry and agriculture, only one study has investigated the interaction of dithiocarbamates other than DDC with MPTP. The purpose of the present study was to investigate whether two other widely used dithiocarbamates, ethylenebisdithiocarbamate (EBDC) and methyldithiocarbamate (MDC), would also enhance MPTP toxicity. The dithiocarbamates were administered to mice intraperitoneally at various doses with or without MPTP. Doses were chosen based on the LD50 values for each compound. DDC was also tested (using a previously reported dose) for comparison. Striata were obtained one week later for dopamine measurements. Consistent with previous reports, DDC produced statistically significant enhancement in MPTP-induced striatal dopamine depletion. EBDC also produced significant exacerbation of MPTP-induced dopamine depletion. In contrast to DDC and EBDC, MDC failed to enhance the effects of MPTP, even when administered at doses of high lethality. Further studies of the dithiocarbamate class of compounds may help to elucidate the mechanism of DDC and EBDC enhancement of MPTP toxicity. Given the widespread use of these compounds in the environment such studies may also provide clues to the process of nigrostriatal cell degeneration in Parkinson's disease.

Effects of minute doses of ethylenebisdithiocarbamate disodium salt (nabam) and its degradative products on connective tissue envelopes of the notochord in Xenopus: an ultrastructural study.

The fungicide ethylenebisdithiocarbamate disodium salt (nabam) at 40 or 100 micrograms/l has deleterious effects on the survival of Xenopus laevis embryos. The tissues of the notochordal enveloping layer show definite alterations at the ultrastructural level. There is a significant reduction or absence of the collagen fibres of the outer connective tissue cells. Embryos treated with 100 micrograms/l nabam stored for 3 weeks developed more severe malformations of the enveloping connective tissue layers of the notochord similar to the embryos immersed in combinations of ethylene thiourea and methyl isothiocyanate. In these animals the arrangement of collagen fibres of the notochordal sheath showed complete disarray. Furthermore, the elastica externa was reduced and fragmented. These fine structural observations clearly demonstrate the deleterious effects of this pesticide on cells and tissues of developing systems.

Studies on the mechanism of nabam- and zineb-induced inhibition of the hepatic microsomal monooxygenases of the male rat.

In vitro effects of the ethylene bis-dithiocarbamate fungicides, nabam and zineb, on the hepatic microsomal monooxygenases of male rats were examined. Incubation of nabam and zineb with hepatic microsomes, without NADPH, leads to an inhibition of the metabolism of aminopyrine and aniline and to a denaturation of cytochrome P-450 into cytochrome P-420; in addition nabam causes the destruction of cytochrome P-450. Addition of NADPH into the incubation medium increases the inhibition of the monooxygenases, principally the inhibition of the metabolism of aniline induced by nabam. We studied the in vitro effects of three of the chief breakdown products of these fungicides: ethylene bis-isothiocyanate sulfide (EBIS), ethylene thiourea (ETU), and carbon disulfide (CS2). EBIS appears to be the only metabolite affecting directly (without NADPH) the hepatic monooxygenases activity. EBIS accounted partly for nabam-induced inhibition of the hepatic microsomal monooxygenases. The data suggest that the decrease of monooxygenases activity seen on incubation of nabam with hepatic microsomes may be due to the denaturation and destruction of cytochrome P-450 resulting from covalent binding of the compounds with cysteine sulfhydryl groups in cytochrome P-450. Inhibition of monooxygenase activity induced by zineb seems to be due to the reaction with the sulfhydryl groups of cytochrome P-450 and to another mechanism, probably related to its lipophilic character.

Comparative effects of N,N-disubstituted dithiocarbamates and dimercaptosuccinate on mobilization of methylmercury in mice.

Diethyldithiocarbamate (DDTC) and five of its N,N-disubstituted analogs were compared with dimercaptosuccinate (DMSA) for effectiveness in mobilizing and promoting excretion of methylmercury (MeHg) following administration of a sublethal dose of MeHg labeled with Me203Hg. The previously reported effectiveness of DMSA was confirmed. Of the dithiocarbamates assessed, only di(carboxymethyl)-dithiocarbamate (DCDC) was active in mobilizing MeHg, and its activity on a molar dose basis was less than that of DMSA. The disparate structural requirements of dithiocarbamates as antagonists of MeHg and of cadmium were discussed.

Control of bacterial speck of tomato by copper and ethylenebisdithiocarbamate fungicides: their efficacy and residues on leaves.

Field studies were conducted to determine the synergism, persistence, and efficacy of a tank-mix spray of cupric hydroxide and mancozeb for control of bacterial speck of tomato. The increased efficacy of the cupric hydroxide-mancozeb mixture could not be attributed to the formation of a bacteriostatic compound in the tank mixture nor to a greater persistence of the mixture on the tomato foliage.

[Hepatic microsomal monoxygenase inhibition by nabam in the rat (author's transl)]. / Inhibition par le nabame des monooxygénases microsomales hépatiques chez le rat.

Nabam (fungicide dithiocarbamate) has been incorporated with the diet of rats during six months (the doses were: 0, 10, 50, 100, 500, 1000 and 2000 ppm). It decreases significantly the hepatic microsomal enzymes activity (aniline hydroxylase and aminopyrine N-demethylase and the liver P 450 content, but the cytochrome b 5 concentration doesn't seem to be modified. The microsomal lipidic and proteic content is only modified with the highest Nabam dose. Several hypotheses may be proposed to explain the Nabam effects: one is the inhibition of the monooxygenases and their biosynthesis repression.

Changes in metabolic activities of Fusarium oxysporum f. fabae and Rhizoctonia solani in response to Dithan A-40 fungicide.

The effect of different concentrations of Dithan A-40 fungicide on the metabolic activities of the wilt fungus Fusarium oxysporum f. fabae and the root rot agent Rhizoctonia solani was studied. All toxicant concentrations reduced energy generation, total phosphorus and nitrogen content of both fungi. In addition, the toxicant caused a shift in free amino acids pool. As a result of these changes, the mycelium dry weight of both fungi was greatly reduced. R. solani was more sensitive to the toxic effect of Dithan A-40 than F. oxysporum.

[Experimental inflammation after subchronic treatment with Nabam in the rat]. / Inflammation expérimentale après un traitement subchronique au Nabame chez le rat.

Inflammatory response to turpentine-induced granuloma components was studied in rats receiving 100 ppm Nabam (N,N'--ethylene--bis)dithiocarbamate) disodium) with food for one month. Results of nutritional, clinical and histopathological studies showed no modification. However, administration of Nabam modified the amounts of various mucopolysaccharides, which are important components of the inflammatory response; the amounts of total mucopolysaccharides, glucuronic acid mucopolysaccharides and sialic acid mucopolysaccharides were obviously lowered in treated animals, a symptom of impairment of the inflammatory response. When most classical means of investigation provide only little information, this test could allow toxicologists and nutritionists to evaluate the organisms reactivity to insidious toxic aggression.