Synthesis and Biological Evaluation of Novel Benodanil-Heterocyclic Carboxamide Hybrids as a Potential Succinate Dehydrogenase Inhibitors.
ABSTRACT: In order to discover new antifungal agents, twenty novel benodanil-heterocyclic carboxamide hybrids were designed, synthesized, and characterized by 1H NMR and HRMS. In vitro, their antifungal activities against four phytopathogenic fungi were evaluated, as well as some of the target compounds at 50 mg/L demonstrated significant antifungal activities against Rhizoctonia solani. Especially, compounds 17 (EC50 = 6.32 mg/L) and 18 (EC50 = 6.06 mg/L) exhibited good antifungal activities against R. solani and were superior to the lead fungicide benodanil (a succinate dehydrogenase inhibitor, SDHI) (EC50 = 6.38 mg/L). Furthermore, scanning electron microscopy images showed that the mycelia on treated media with the addition of compound 17 grew abnormally as compared with the negative control with tenuous, wizened, and overlapping colonies, and compounds 17 (IC50 = 52.58 mg/L) and 18 (IC50 = 56.86 mg/L) showed better inhibition abilities against succinate dehydrogenase (SDH) than benodanil (IC50 = 62.02 mg/L). Molecular docking revealed that compound 17 fit in the gap composed of subunit B, C, and D of SDH. Furthermore, it was shown that the main interaction, one hydrogen bond interaction, was observed between compound 17 and the residue C/Trp-73. These studies suggested that compound 17 could act as a potential fungicide to be used for further optimization.
Project description:A series of pyrazole carboxamide and isoxazolol pyrazole carboxylate derivatives were designed and synthesized in this study. The structures of the compounds were elucidated based on spectral data (infrared, proton nuclear magnetic resonance and mass spectroscopy). Then, all of the compounds were bioassayed in vitro against four types of phytopathogenic fungi (Alternaria porri, Marssonina coronaria, Cercospora petroselini and Rhizoctonia solani) using the mycelium growth inhibition method. The results showed that some of the synthesized pyrazole carboxamides displayed notable antifungal activity. The isoxazole pyrazole carboxylate 7ai exhibited significant antifungal activity against R. solani, with an EC50 value of 0.37 ?g/mL. Nonetheless, this value was lower than that of the commercial fungicide, carbendazol.
Project description:43 Actinobacteria were isolated from the nest of Odontotermes formosanus. A phylogenetic analysis of 23 Actinobacteria isolates with different morphotypes showed that they did not form a monophyletic group. Antifungal bioassays exhibited that many strains inhibit both the termite cultivar Termitomyces and the competitor Xylaria. However, Actinobacteria inhibited the competitor Xylaria more severely than the termite cultural fungus Termitomyces. Furthermore, two Actinobacteria (Streptomyces sp. T33 and S. bellus T37) had a selective antifungal effect on Xylaria, with the inhibition zone of 25.5 and 8.9 mm, respectively. An actinomycin D was isolated from the strain T33 and had potent antifungal activity against Xylaria with IC50 value of less than 3.1 µg/mL. In addition, further bioassays showed that actinomycin D possessed potent antifungal activities against Magnaporthe grisea (IC50?=?0.9 µg/mL), Fusarium oxysporum f. sp. cucumerinum (IC50?=?2.2 µg/mL), Valsa mali (IC50?=?1.7 µg/mL), Rhizoctonia solani (IC50?=?10.3 µg/mL), Dothiorella gregaria (IC50?=?12.5 µg/mL) and F. oxysporum f. sp. mornordicae (IC50?=?14.3 µg/mL), which were comparable to those of referenced cycloheximide. The findings of the present study suggest that the termite-associated Actinobacteria have a potential to be used as microbial fungicide.
Project description:Curcuma longa possesses powerful antifungal activity, as demonstrated in many studies. In this study, the antifungal spectrum of Curcuma longa alcohol extract was determined, and the resulting EC50 values (mg/mL) of its extract on eleven fungi, including Fusarium graminearum, Fusarium chlamydosporum, Alternaria alternate, Fusarium tricinctum, Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium culmorum, Rhizopus oryzae, Cladosporium cladosporioides, Fusarium oxysporum and Colletotrichum higginsianum, were 0.1088, 0.1742, 0.1888, 0.2547, 0.3135, 0.3825, 0.4229, 1.2086, 4.5176, 3.8833 and 5.0183, respectively. Among them, F. graminearum was selected to determine the inhibitory effects of the compounds (including curdione, isocurcumenol, curcumenol, curzerene, ?-elemene, curcumin, germacrone and curcumol) derived from Curcuma longa. In addition, the antifungal activities of curdione, curcumenol, curzerene, curcumol and isocurcumenol and the synergies of the complexes of curdione and seven other chemicals were investigated. Differential proteomics of F. graminearum was also compared, and at least 2021 reproducible protein spots were identified. Among these spots, 46 were classified as differentially expressed proteins, and these proteins are involved in energy metabolism, tRNA synthesis and glucose metabolism. Furthermore, several fungal physiological differences were also analysed. The antifungal effect included fungal cell membrane disruption and inhibition of ergosterol synthesis, respiration, succinate dehydrogenase (SDH) and NADH oxidase.
Project description:Alternaria blight is an important foliage disease caused by Alternaria solani. The enzyme Succinate dehydrogenase (SDH) is a potential drug target because of its role in tricarboxylic acid cycle. Hence targeting Alternaria solani SDH enzyme could be efficient tool to design novel fungicides against A. solani. We employed computational methodologies to design new SDH inhibitors using homology modeling; pharmacophore modeling and structure based virtual screening. The three dimensional SDH model showed good stereo-chemical and structural properties. Based on virtual screening results twelve commercially available compounds were purchased and tested in vitro and in vivo. The compounds were found to inhibit mycelial growth of A. solani. Moreover in vitro trials showed that inhibitory effects were enhanced with increase in concentrations. Similarly increased disease control was observed in pre-treated potato tubers. Hence the applied in silico strategy led us to identify novel fungicides.
Project description:The title compounds can be considered as simple analogues of quaternary benzo[c]phenanthridine alkaloids (QBAs). In order to develop potent QBA-like antifungal agents, as our continuing study, a series of new title compounds were synthesized and evaluated for bioactivity against five plant pathogenic fungi by the mycelium growth rate method in this study. The SAR were also derived. The majority of the compounds showed good to excellent inhibition activity with average EC50 values of 7.87-20.0??M for the fungi, superior to sanguinarine and cherythrine (two QBAs) and the commercial fungicide azoxystrobin. Part of the compounds were more active than commercial fungicides thiabendazole or carbendazim against F. solani, F. graminearum and C. gloeosporioides. Six compounds with average EC50 of 3.5-5.1??g/mL possessed very great potential for development of new antifungal agents. SAR found that substitution patterns of the two aryl-rings significantly affect the activity. There exists a complex interaction effect between substituents of the two aryl-rings on the activity. Generally, the presence of electron-withdrawing groups on the C-ring can significantly increase the activity. These findings will be of great importance for the design of more potent antifungal isoquinoline agents.
Project description:The succinate dehydrogenase inhibitor (SDHI) fungicide, fluopyram, is used as a soybean seed treatment to manage Fusarium virguliforme, the casual agent of sudden death syndrome (SDS). More recently, other species within clade 2 of the Fusarium solani species, F. tucumaniae in South America and F. brasiliense in America and Africa, have been recognized as additional agents capable of causing SDS. To determine if fluopyram could be used for management of SDS caused by these species, in vitro sensitivity tests of the three Fusarium species to fluopyram were conducted. The mean EC50 values of F. brasiliense and F. virguliforme strains to fluopyram were 1.96 and 2.21 ?g ml-1, respectively, but interestingly F. tucumaniae strains were highly sensitive (mean EC50 = 0.25 ?g ml-1) to fluopyram compared to strains of the other two species. A sequence analysis of Sdh genes of Fusarium strains revealed that the F. tucumaniae strains contain an arginine at codon 277 in the SdhB gene instead of a glycine as in other Fusarium species. Replacement of glycine to arginine in SdhB-277 in a F. virguliforme wild-type strain Mont-1 through genetic transformation resulted in increased sensitivity to two SDHI fungicides, fluopyram and boscalid. Similar to a F. tucumaniae strain, the Mont-1 (SdhBG277R) mutant caused less SDS and root rot disease than Mont-1 on soybean seedlings with the fluopyram seed treatment. Our study suggests the amino acid difference in the SdhB in F. tucumaniae results in fluopyram being efficacious if used as a seed treatment for management of F. tucumaniae, which is the most abundant SDS causing species in South America. The establishment of baseline sensitivity of Fusarium species to fluopyram will contribute to effective strategies for managing Fusarium diseases in soybean and other pathosystems such as dry bean.
Project description:Fungicide resistance is a constant threat to agricultural production worldwide. Molecular mechanisms of fungicide resistance have been studied extensively in the wheat pathogen Zymoseptoria tritici. However, less is known about the evolutionary processes driving resistance development. In vitro evolutionary studies give the opportunity to investigate this. Here, we examine the adaptation of Z. tritici to fluxapyroxad, a succinate dehydrogenase (Sdh) inhibitor. Replicate populations of Z. tritici derived from the sensitive isolate IPO323 were exposed to increasing concentrations of fluxapyroxad with or without UV mutagenesis. After ten increases in fungicide concentration, sensitivity had decreased dramatically, with replicate populations showing similar phenotypic trajectories. Sequencing the Sdh subunit B, C, and D encoding genes identified seven mutations associated with resistance to fluxapyroxad. Mutation frequency over time was measured with a pyrosequencing assay, revealing sequential lineage replacement in the UV-mutagenized populations but not in the untreated populations. Repeating selection from set time-points with different fungicide concentrations revealed that haplotype replacement of Sdh variants was driven by dose-dependent selection as fungicide concentration changed, and was not mutation-limited. These findings suggest that fungicide field applications may select for highly insensitive Sdh variants with higher resistance factors if the fungicide concentration is increased to achieve a better disease control. However, in the absence or presence of lower fungicide concentrations, the spread of these strains might be restricted if the underlying Sdh mutations carry fitness penalties.
Project description:A series of novel 3-caren-5-one oxime esters were designed and synthesized by multi-step reactions in an attempt to develop potent antifungal agents. Two E-Z stereoisomers of the intermediate 3-caren-5-one oxime were separated by column chromatography for the first time. The structures of all the intermediates and target compounds were confirmed by UV-Vis, FTIR, NMR, ESI-MS, and elemental analysis. The antifungal activity of the target compounds was preliminarily evaluated by the in vitro method against Fusarium oxysporum f. sp. cucumerinum, Physalospora piricola, Alternaria solani, Cercospora arachidicola, Gibberella zeae,Rhizoeotnia solani, Bipolaris maydis, and Colleterichum orbicalare at 50 µg/mL. The target compounds exhibited best antifungal activity against P. piricola, in which compounds (Z)-4r (R = ?-pyridyl), (Z)-4q (R = ?-thienyl), (E)-4f' (R = p-F Ph), (Z)-4i (R = m-Me Ph), (Z)-4j (R = p-Me Ph), and (Z)-4p (R = ?-furyl) had inhibition rates of 97.1%, 87.4%, 87.4%, 85.0%, 81.9%, and 77.7%, respectively, showing better antifungal activity than that of the commercial fungicide chlorothanil. Also, compound (Z)-4r (R = ?-pyridyl) displayed remarkable antifungal activity against all the tested fungi, with inhibition rates of 76.7%, 82.7%, 97.1%, 66.3%, 74.7%, 93.9%, 76.7% and 93.3%, respectively, showing better or comparable antifungal activity than that of the commercial fungicide chlorothanil. Besides, the E-Z isomers of the target oxime esters were found to show obvious differences in antifungal activity. These results provide an encouraging framework that could lead to the development of potent novel antifungal agents.
Project description:BACKGROUND:The widespread occurrence of fungicide resistance in fungal plant pathogens requires the development of new compounds with different mode(s) of action (MOA) to avoid cross resistance. This will require a rapid method to identify MOAs. RESULTS:Here, gas chromatography-mass spectrometry (GC-MS) based metabolic fingerprinting was used to elucidate the MOAs of fungicides. Botrytis cinerea, an important pathogen of vegetables and flowers, can be inhibited by a wide range of chemical fungicides with different MOAs. A sensitive strain of B. cinerea was exposed to EC50 concentrations of 13 fungicides with different known MOAs and one with unknown MOA. The mycelial extracts were analyzed for their "metabolic fingerprint" using GC-MS. A comparison among the GC-MS vector' profiles of cultures treated with fungicides were performeded. A model based on hierarchical clustering was established which allowed these antifungal compounds to be distinguished and classified coinciding with their MOAs. Thus, metabolic fingerprinting represents a rapid, convenient, and information-rich method for classifying the MOAs of antifungal substances. The biomarkers of fungicide MOAs were also established by an analysis of variance and included succinate for succinate dehydrogenase inhibitors and cystathionine for methionine synthesis inhibitors. Using the metabolic model and the common perturbation of metabolites, the new fungicide SYP-14288 was identified as having the same MOA as fluazinam. CONCLUSION:This study provides a comprehensive database of the metabolic perturbations of B. cinerea induced by diverse MOA inhibitors and highlights the utility of metabolic fingerprinting for defining MOAs, which will assist in the development and optimization of new fungicides.
Project description:Six novel nicotinamide derivatives bearing a diarylamine-modified scaffold with flexible heterocyclic patterns were designed, synthesized, and characterized in detail via Hydrogen nuclear magnetic resonance (1H-NMR), Carbon nuclear magnetic resonance (13C-NMR), and Electrospray ionization mass spectrometry (ESI-MS). Their fungicidal activities and succinate dehydrogenase (SDH) enzymatic inhibitory abilities were evaluated. Preliminary fungicidal bioassay results showed that some of the target compounds exhibited moderate fungicidal activity. Among them, compound 4a showed 40.54% inhibition against Botrytis cinerea fungi. An SDH enzymatic inhibition assay revealed that the IC50 of compound 4b was 3.18?µM. This result indicated that the enzymatic inhibition level of 4b was similar to that of boscalid. Compound 4f exhibited superior comprehensive fungicidal and SDH enzymatic inhibitory activities. Molecular docking results suggested that 4f could bind well to the substrate cavity and the entrance cavity of SDH (1YQ3). In particular, 4f could react with the key catalytic site Arg 297. This phenomenon implied that 4f could act as the lead compound for further optimization.