Synthesis and Antifungal Activity of Novel L-Menthol Hydrazide Derivatives as Potential Laccase Inhibitors

To discover novel laccase inhibitors as potential fungicides, twenty-six novel L-menthol hydrazide derivatives were designed and synthesized. In the in vitro antifungal assay, most of the target compounds displayed pronounced antifungal activity against Sclerotinia sclerotiorum, Fusarium graminearum, and Botryosphaeria dothidea. Especially, the EC₅₀ of compounds 3 b and 3 q against B. dothidea was 0.465 and 0.622 mg/L, which was close to the positive compound fluxapyroxad (EC₅₀=0.322 mg/L). Scanning electron microscopy (SEM) analysis showed that compound 3 b could significantly damage the mycelial morphology of B. dothidea. In vivo antifungal experiments on apple fruits showed that 3 b exhibited excellent protective and curative effects. Furthermore, in the in vitro laccase inhibition assay, 3 b showed outstanding inhibitory activity with the IC₅₀ value of 2.08 μM, which is much stronger than positive control cysteine and PMDD-5Y. These results indicated that this class of L-menthol derivatives could be promising leads for the discovery of laccase-targeting fungicides.     

Synthesis and anthelmintic activity of arctigenin derivatives against Dactylogyrus intermedius in goldfish

To control the parasitic disease of Dactylogyrus intermedius, a series of new arctigenin derivatives were designed, synthesized and tested in our study. The anthelmintic activity of most of the derivatives ranged from 1 to 10 mg/L. Compared to traditional drug praziquantel (EC₅₀ = 2.69 mg/L), ether derivatives 2g and 2h exhibited slightly higher anti-parasitic activity, with the EC₅₀ values of 2.48 and 1.52 mg/L, respectively. Furthermore, the arctigenin-imidazole hybrids 4a and 4b also removed D. intermedius effectively, with the EC₅₀ values of 2.13 and 2.07 mg/L, respectively. The structure-activity relationship analysis indicated that four carbon atoms length of linker and imidazole substitute group could significantly increase the anthelmintic activity, and reduced the toxicity. Through the scanning electron microscope observation, compounds 4a and 4b caused the D. intermedius tegumental damage such as intensive wrinkles, holes and nodular structures. Overall, the structural optimization analysis of arctigenin suggested that 4a and 4b can be used for preventing and controlling Dactylogyrus infections and considered as promising lead compounds for the development of commercial drugs.     

Design,Synthesis, and Antifungal Activities of Novel Carboxamides Derivatives Bearing a Chalcone Scaffold as Potential SDHIs

In search for SDHIs fungicides, twenty-five novel carboxamides containing a chalcone scaffold were designed, synthesized, and evaluated for antifungal activities against five pathogenic fungi. The results showed that compound 5 k exhibited outstanding antifungal activity against R. solani with an EC₅₀ value of 0.20 μg/mL, which was much better than that of commercial SDHIs Boscalid (EC₅₀=0.74 μg/mL). Moreover, compound 5 k also displayed promising antifungal activities against S. sclerotiorum, B. cinerea, and A. alternate (IC₅₀=2.53–4.06 μg/mL), indicating that 5 k had broad-spectrum antifungal activity. Additionally, in vivo antifungal activities results showed that 5 k could significantly inhibit the growth of R. solani in rice leaves with good protective efficacy (57.78 %) and curative efficacy (58.45 %) at 100 μg/mL, both of which were much better than those of Boscalid, indicating a promising application prospect. Moreover, SEM analysis showed that compound 5 k could remarkably disrupt the typical structure and morphology of R. solani hyphae. Further SDH enzyme inhibition assay and molecular docking study revealed that lead compound 5 k had a similar mechanism of action as commercial SDHI Boscalid. These results indicated that compound 5 k showed potential as a SDHIs fungicide and deserved further investigation.     

四霉素对茶藨子葡萄座腔菌Botryosphaeria ribis室内毒力测定及其田间防效试验

为确定四霉素对杨树溃疡病的防治效果,采用菌丝生长速率法测定四霉素水剂对茶藨子葡萄座腔菌Botryosphaeria ribis的室内毒力,采用刮治喷涂法对茶藨子葡萄座腔菌引起的杨树溃疡病进行田间应用效果研究。试验结果表明,四霉素对茶藨子葡萄座腔菌的EC_(50)和EC_(90)分别为114.82 a.i.mg/L、389.05 a.i.mg/L,对茶藨子葡萄座腔菌具有较好的抑菌效果,优于对照药剂多抗霉素。田间防效试验结果表明,刮治喷涂0.3%四霉素水剂可以有效防治真菌病原茶藨子葡萄座腔菌导致的杨树溃疡病,其有效成分浓度为60～150 a.i.mg/L时,田间防效达68.25%～82.41%,与室内毒力测定结果相符。     

Structure-based design, synthesis, and biological evaluation of 1,1-dioxoisothiazole and benzo[b]thiophene-1,1-dioxide derivatives as novel inhibitors of hepatitis C virus NS5B polymerase

A novel series of HCV NS5B polymerase inhibitors comprising 1,1-dioxoisothiazoles and benzo[b]thiophene-1,1-dioxides were designed, synthesized, and evaluated. SAR studies guided by structure-based design led to the identification of a number of potent NS5B inhibitors with nanomolar IC₅₀ values. The most potent compound exhibited IC₅₀ less than 10 nM against the genotype 1b HCV polymerase and EC₅₀ of 70 nM against a genotype 1b replicon in cell culture. The DMPK properties of selected compounds were also evaluated.     

Baseline sensitivity of Botrytis cinerea to fluazinam and cross‐resistance

Grey mould, caused by the fungus Botrytis cinerea Pers ex Fr., is a very destructive and important disease worldwide. Fluazinam is a phenylpyridinamine fungicide with broad‐spectrum activities. The baseline sensitivity of B. cinerea to fluazinam is yet to be established in Henan Province, China. In this study, a total of 117 field isolates of B. cinerea were collected from 49 commercial greenhouses in different locations of Henan Province, in 2016, and the sensitivities of these isolates to fluazinam were determined based on mycelial growth. The effective concentration for 50% (EC₅₀) values ranged from 0.0038 to 0.0441 μg/ml, and the mean EC₅₀ value was 0.0201 ± 0.0081 μg/ml for mycelial growth. The frequency distribution range presented a unimodal curve. To define the cross‐resistance relationships, the linear correlation coefficients of the EC₅₀ values between fluazinam and carbendazim, procymidone, pyrimethanil or boscalid were analysed. The results showed that no correlation was observed between fluazinam and the other tested fungicides. These results provide important information to growers for the prevention and control of grey mould.     

Synthesis and Fungicidal Activities of New 1,2,4‐Triazolo[1,5‐a]pyrimidines

A series of new acetohydrazone‐containing 1,2,4‐triazolo[1,5‐a]pyrimidine derivatives were designed and synthesized for the purpose of searching for novel agrochemicals with higher fungicidal activity. Their in vitro fungicidal activities against Rhizoctonia solani were evaluated, and the most promising compound, 2‐[(5,7‐dimethyl[1,2,4]triazolo[1,5‐a]pyrimidin‐2‐yl)sulfanyl]‐2′‐[(2‐hydroxyphenyl)methylidene]acetohydrazide ( 2‐17 ), showed a lower EC₅₀ value (5.34 μg ml^?1) than that of commercial carbendazim (EC₅₀=7.62 μg ml^?1). Additionally, compound 2‐17 was also found to display broad‐spectrum fungicidal activities, and its EC₅₀ value (4.56 μg ml^?1) against Botrytis cinereapers was very similar to that of carbendazim. Qualitative structure–activity relationships (QSARs) of the synthesized compounds were also discussed.     

New insight into the antifibrotic effects of praziquantel on mice in infection with Schistosoma japonicum

Background

Schistosomiasis is a parasitic disease infecting more than 200 million people in the world. Although chemotherapy targeting on killing schistosomes is one of the main strategies in the disease control, there are few effective ways of dealing with liver fibrosis caused by the parasite infection in the chronic and advanced stages of schistosomiasis. For this reason, new strategies and prospective drugs, which exert antifibrotic effects, are urgently required.

Methods and Findings

The antifibrotic effects of praziquantel were assessed in the murine models of schistosomiasis japonica. Murine fibrosis models were established by cutaneous infection with 14±2 Schistosoma japonicum cercariae. Then, the mice of both chronic (8 weeks post-infection) and advanced (15 weeks post-infection) schistosomiasis were treated by gavage of praziquantel (250 mg/kg, once daily for 3 days) to eliminate worms, and followed by praziquantel anti-fibrosis treatment (300 mg/kg, twice daily for 30 days). The fibrosis-related parameters assessed were areas of collagen deposition, content of hydroxyproline and mRNA expressions of Col1α1, Col3α1, α-SMA, TGF-β, MMP9, TIMP1, IL-4, IL-10, IL-13 and IFN-γ of liver. Spleen weight index, alanine aminotransferase activity and liver portal venous pressure were also measured. The results showed that anti-fibrosis treatment improved liver fibrosis, splenomegaly, hepatic function, as well as liver portal hypertension. In order to confirm the anti-fibrotic properties of praziquantel, we established a CCL₄-induced model and revealed that CCL₄-induced liver fibrosis was inhibited by PZQ treatment for 30 days. Furthermore, we analyzed the effects of praziquantel on mouse primary hepatic stellate cells (HSCs). It is indicated that mRNA expressions of Col1α1, Col3α1, α-SMA, TGF-β, MMP9 and TIMP1 of HSCs were all inhibited after praziquantel anti-parasite treatments.

Conclusions

The significant amelioration of hepatic fibrosis by praziquantel treatment validates it as a promising drug of anti-fibrosis and offers potential of a new chemotherapy for hepatic fibrosis resulting from schistosomiasis.     

Synthesis and Antifungal Activity of Curcumol Derivatives

To discover novel and effective antifungal candidates, a series of new curcumol derivatives were designed, synthesized, and evaluated their antifungal activity against five phytopathogenic fungi by the mycelium growth rate method. Derivatives c4 , c22 and c23 exhibited excellent antifungal activity against Phomopsis sp. with EC₅₀ values of 3.06, 3.07, and 3.16 μM, respectively. Specifically, compound c4 exhibited the strongest antifungal activity against Phomopsis sp., which was 44 times that of pyrimethanil (EC₅₀=134.37 μM). The results of scanning electron microscopy (SEM) and transmission electron microscopy (TEM) indicated that compound c4 could cause cell senescence and death of Phomopsis sp. by changing the normal hyphal morphology and disrupting the normal metabolism of hyphal cells. Moreover, compound c4 showed excellent curative effect against Phomopsis sp. on kiwifruit. These findings confirmed that compound c4 has great potential as a potent antifungal agent.     

Structure optimization and bioactivity evaluation of ThDP analogs targeting cyanobacterial pyruvate dehydrogenase E1

Harmful cyanobacteria bloom (HCB) has occurred frequently in recent years and it is urgent to develop novel algicides to deal with this problem. In this paper, a series of novel thiamin diphosphate (ThDP) analogs 5a?5g were designed and synthesized targeting cyanobacterial pyruvate dehydrogenase complex E1 (Cy-PDHc E1). Our results showed that compounds 5a?5g have higher inhibitory activities against Cy-PDHc E1 (IC₅₀ 9.56–3.48 µM) and higher inhibitory activities against two model cyanobacteria strains Synechocystis sp PCC6803 (EC₅₀ 2.03–1.58 µM) and Microcystis aeruginosa FACHB905 (EC₅₀ 1.86–0.95 µM). Especially, compound 5b displayed highest inhibitory activities (IC₅₀ = 3.48 µM) against Cy-PDHc E1 and powerful inhibitory activities against cyanobacteria Synechocystis sp PCC6803 (EC₅₀ = 1.58 µM) and Microcystis aeruginosa FACHB905 (EC₅₀ = 1.04 µM). Moreover, the inhibitory activities of compound 5b were even higher than those of copper sulfate (EC₅₀ = 2.02 and 1.71 µM separately) which has been widely used as algicide against cyanobacteria PCC6803 and FACHB905. The more important was that compound 5b display much higher inhibitory selectivity between Cy-PDHc E1 (Inhibitory rate 97.4%) and porcine PDHc E1 (Inhibitory rate 11.8%) under the same concentration (100 μM). The inhibition kinetic experiment and molecular docking research showed that compound 5b can inhibit Cy-PDHc E1 by occupying the ThDP-binding pocket and then blocking Cy-PDHc E1 bound to ThDP as competitive inhibitor. The imagines of SEM and TEM showed that cellular microstructures were heavily destroyed under compound 5b stress. Our results demonstrated compound 5b could be taken as a potential lead compound targeting Cy-PDHc E1 to obtain environment-friendly algicide for harmful cyanobacterial blooms control.     

Antisettlement activity of synthetic analogues of polymeric 3-alkylpyridinium salts isolated from the sponge Reniera sarai

Based on the high, non-toxic and reversible antifouling activity of the polymeric 3-alkylpyridinium salts isolated from the sponge Reniera sarai, the anti-settlement activity and toxicity of a series of synthetic analogues has been studied. All the test compounds were less efficient than the natural polymers, suggesting that the high and reversible anti-macrofouling activity of the natural polymers could derive from their detergent-like properties. The values obtained for EC_50sett. of inhibition of cyprid settlement and EC_50imm. as naupliar toxicity for the synthetic compounds indicate that the presence of single or multiple charges in the structure is not relevant for the antifouling activity which, conversely, is favoured by increasing the length of the alkyl chain, or by the presence of uncharged pyridine units. The compound 1,8-di(3-pyridyl)octane was the most efficient (EC_50sett.= 0.44?μg?ml^–1), although with a higher toxicity on naupliar stage of B. amphitrite than the natural polymers.     

New 2‐Oxoimidazolidine Derivatives: Design,Synthesis and Evaluation of Anti‐BK Virus Activities in Vitro

A series of novel 2‐oxoimidazolidine derivatives were synthesized and their antiviral activities against BK human polyomavirus type 1 (BKPyV) were evaluated in vitro. Bioassays showed that the synthesized compounds 1‐{[(4E)‐5‐(dichloromethylidene)‐2‐oxoimidazolidin‐4‐ylidene]sulfamoyl}piperidine‐4‐carboxylic acid ( 5 ) and N‐Cyclobutyl‐N′‐[(4E)‐5‐(dichloromethylidene)‐2‐oxoimidazolidin‐4‐ylidene]sulfuric diamide ( 4 ) exhibited moderate activities against BKPyV (EC₅₀=5.4 and 5.5 μm , respectively) that are comparable to the standard drug Cidofovir. Compound 5 exhibited the same cytotoxicity in HFF cells and selectivity index (SI₅₀) as Cidofovir. The selectivity index of compound 4 is three times less than that of Cidofovir due to the higher toxicity of this compound. Hence, these compounds may be taken as lead compound for further development of novel ant‐BKPyV agents.     

Synthesis of novel fenfuram-diarylether hybrids as potent succinate dehydrogenase inhibitors

Twelve novel fenfuram-diarylether hybrids were designed, synthesized and characterized by ¹H NMR and MS. Their in vitro antifungal activities were evaluated against five phytopathogenic fungi by mycelial growth inhibition method. Most compounds showed significant antifungal effect on Rhizoctonia solani and Sclerotinia sclerotiorum. Compound 1c exhibited the most potent antifungal effect on R. solani with an EC₅₀ value of 0.242 mg/L, superior to the commercial fungicide boscalid (EC₅₀ = 1.758 mg/L) and the lead fungicide fenfuram (EC₅₀ = 7.691 mg/L). Molecular docking revealed that compound 1c featured a higher affinity for succinate dehydrogenase (SDH) than fenfuram. Furthermore, it was shown that the 2-chlorophenyl group of compound 1c formed a π-π stacking with D/Tyr-128 and a Cl-π interaction with B/His-249, which made compound 1c more active than fenfuram against SDH.     

Baseline sensitivity and efficacy of fluopyram against Botrytis cinerea from table grape in Italy

Succinate dehydrogenase inhibitor (SDHI) fungicides constitute a relatively recent fungicide class registered for the treatment of grey mould on grapevine in Italy. The sensitivity profile to a novel compound fluopyram was established for a set of 203 Botrytis cinerea isolates collected from Sicilian vineyards within 2009–2012 prior its introduction into market. In addition, its performances were compared in in vitro and in vivo assays with other registered SDHI fungicide boscalid, to evaluate their frequency distributions EC₅₀ values and cross‐resistance patterns. Results of the article showed that EC₅₀ values for fluopyram ranged from 0.05 to 1.98 µg mL^?1. Although EC₅₀ values of boscalid ranged from 0.01 to 89.52 µg mL^?1, no cross‐resistance relationship was observed between the two fungicides (r = 0.003; P = 0.964) within our B. cinerea population. On further confirming these data, boscalid failed in controlling grey mould infections when boscalid‐resistant isolates were inoculated on grape berries whereas fluopyram exhibited a good efficacy against the same isolates. This study represents the first report on the baseline sensitivity to fluopyram within B. cinerea population from Sicilian table grape vineyards in Italy, and it clearly shows the lack of cross‐resistance in vitro and in vivo between fluopyram and boscalid for the field pathogen isolates. These results provided useful information for managing of fungicide resistance suggesting that fluopyram could be a valid alternative to boscalid for the control of grey mould of table grape.     

In vitro evaluation of the effects of cysticidal drugs in the Taenia crassiceps cysticerci ORF strain using the fluorescent CellTracker CMFDA

Using a murine model of cysticercosis caused by the Taenia crassiceps ORF strain, we developed a fluorescent quantitative evaluation of the action of two well known anti-helminthic drugs: albendazole sulfoxide and praziquantel. The fluorescence emitted by a biotransformed CellTracker Probe known as CellTracker Green CMFDA in the vesicular fluids of cysticerci was estimated, and the results were compared with macroscopic observations of the parasites. The pharmacological EC₅₀ value of each drug and changes in the level of biotransformation of the fluorescent tracker caused by the drugs could be easily calculated. These drug-induced changes in biotransformation could be related to changes in the GSH/GSSG ratio of parasites. Both the cysticercosis murine model and the CMFDA biotransformation assay could be used as an in vitro screening method to evaluate potential or well known cysticidal drugs.     

No Evidence for Resistance to Fenbendazole in Trichostrongylus tenuis,a Nematode Parasite of the Red Grouse

ABSTRACT The parasitic nematode Trichostrongylus tenuis has a detrimental effect on red grouse (Lagopus lagopus scoticus) at the individual and population levels. Treatment using grit coated with the anthelmintic fenbendazole hydrochloride reduces parasite infection and increases grouse density. However, a frequent and low dose of anthelmintic increases selection pressure for parasite resistance, a serious practical and economic problem. We used an egg hatch assay to test resistance of T. tenuis from 12 moors in northern England, which differed in grit treatment intensity. The anthelmintic concentration that prevented 50% and 95% of T. tenuis eggs from hatching (ED₅₀ and ED₉₅, respectively) did not differ among moors and were not related to treatment. We suggest annual monitoring and responsible anthelmintic use to prevent resistance so that medicated grit continues to enhance red grouse management.     

Uncovering Biological Application of Brazilian Green Propolis: A Phenotypic Screening against Schistosoma mansoni

The chemotherapy of schistosomiasis remains centered in the use of praziquantel, however, there has been growing resistant parasites to this drug. Thus, the aim of this work was to evaluate in vitro schistosomicidal activity of the hexanes/dichloromethane 1 : 1 extract of Brazilian green propolis (Pex), as well as its major isolated compounds artepillin C, caffeic acid, coumaric acid and drupanin against Schistosoma mansoni. The Pex was active by displaying an IC₅₀ value of 36.60 (26.26–51.13) μg mL^?1 at 72 h against adult worms of S. mansoni. The major isolated compounds were inactive with IC₅₀ values >100 μM, however, the combination of the isolated compounds (CM) in the same range found in the extract was active with an IC₅₀ value of 41.17 (39.89–42.46) μg mL^?1 at 72 h. Pex and CM induced alteration in the tegument of S. mansoni, and caffeic acid caused alteration in egg's maturation. Pex displayed in vitro activity against adult worms’ and eggs’ viability of S. mansoni, which opens new perspectives to better understand the synergistic and/or additive effects promoted by both Pex extract and CM against schistosomiasis features.     

Design,synthesis and antifungal activity of novel fenfuram-diarylamine hybrids

Ten novel fenfuram-diarylamine hybrids were designed and synthesized. And their antifungal activities against four phytopathogenic fungi have been evaluated in vitro and most of the compounds demonstrated a significant antifungal activities against Rhizoctonia solani and Sclerotinia sclerotiorum. Compound 5e exhibited the most potent antifungal activity against R. solani with an EC₅₀ value of 0.037 mg/L, far superior to the commercially available fungicide boscalid (EC₅₀ = 1.71 mg/L) and lead fungicide fenfuram (EC₅₀ = 6.18 mg/L). Furthermore, scanning electron microscopy images showed that the mycelia on treated media grew abnormally with tenuous, wizened and overlapping colonies compared to the negative control. Molecular docking studies revealed that compound 5e featured a higher affinity for succinate dehydrogenase (SDH) than fenfuram. Furthermore, it was shown that the 3-chlorophenyl group in compound 5e formed a CH-π interaction with B/Trp-206 and a Cl-π interaction with D/Tyr-128, rendering compound 5e more active than fenfuram against SDH.     

Electrophysiological characterization of a schistosome transient receptor potential channel activated by praziquantel

Ion channels have proved to be productive targets for anthelmintic chemotherapy. One example is the recent discovery of a parasitic flatworm ion channel targeted by praziquantel (PZQ), the main clinical therapy used for treatment of schistosomiasis. The ion channel activated by PZQ – a transient receptor potential ion channel of the melastatin subfamily, named TRPM_PZQ – is a Ca²⁺-permeable ion channel expressed in all parasitic flatworms that are PZQ-sensitive. However, little is currently known about the electrophysiological properties of this target that mediates the deleterious action of PZQ on many trematodes and cestodes. Here, we provide a detailed biophysical characterization of the properties of Schistosoma mansoni TRPM_PZQ channel (Sm.TRPM_PZQ) in response to PZQ. Single channel electrophysiological analysis demonstrated that Sm.TRPM_PZQ when activated by PZQ is a non-selective, large conductance, voltage-insensitive cation channel that displays distinct properties from human TRPM paralogs. Sm.TRPM_PZQ is Ca²⁺-permeable but does not require Ca²⁺ for channel gating in response to PZQ. TRPM_PZQ from Schistosoma japonicum (Sj.TRPM_PZQ) and Schistosoma haematobium (Sh.TRPM_PZQ) displayed similar characteristics. Profiling Sm.TRPM_PZQ responsiveness to PZQ has established a biophysical signature for this channel that will aid future investigation of endogenous TRPM_PZQ activity, as well as analyses of endogenous and exogenous regulators of this novel, druggable antiparasitic target.     

Semisynthesis and Antifungal Activity of Novel Oxime Ester Derivatives of Carabrone Modified at C(4) against Botrytis cinerea

To continuously improve the potential utility of the natural lead compound of carabrone in agrochemistry, carabrone oxime and 36 novel oxime ester derivatives of carabrone modified at C(4) were synthesized, and evaluated for their antifungal activities against Botrytis cinerea in vitro and in vivo. Of these 36 oxime ester derivatives, some compounds exhibited antifungal activities in vitro or in vivo. It was found that compounds with a pyridinyl residue can either efficiently inhibit spore germination or efficiently inhibit hyphal growth of B. cinerea, and compound 9 exhibited the highest activity in vitro and in vivo with IC₅₀ and EC₅₀ values of 1.17 and 12.9 μg/ml, respectively. Further, the structure? activity relationships are also discussed.