Design and Synthesis of Novel Diphenyl Ether Carboxamide Derivatives To Control the Phytopathogenic Fungus Sclerotinia sclerotiorum.

Sclerotinia stem rot (SSR) caused by the phytopathogenic fungus Sclerotinia sclerotiorum has led to serious losses in the yields of oilseed rape and other crops every year. Here, we designed and synthesized a series of carboxamide derivatives containing a diphenyl ether skeleton by adopting the scaffold splicing strategy. From the results of the mycelium growth inhibition experiment, inhibition rates of compounds 4j and 4i showed more than 80% to control S. sclerotiorum at a dose of 50 µg/mL, which is close to that of the positive control (flubeneteram, 95%). Then, the results of a structure-activity relationship study showed that the benzyl scaffold was very important for antifungal activity and that introducing a halogen atom on the benzyl ring would improve antifungal activity. Furthermore, the results of an in vitro activity test suggested that these novel compounds can inhibit the activity of succinate dehydrogenase (SDH), and the binding mode of 4j with SDH was basically similar to that of the flutolanil derivative. Morphological observation of mycelium revealed that compound 4j could cause a damage on the mycelial morphology and cell structure of S. sclerotiorum, resulting in inhibition of the growth of mycelia. Furthermore, in vivo antifungal activity assessment of 4j displayed a good control of S. sclerotiorum (>97%) with a result similar to that of the positive control at a concentration of 200 mg/L. Thus, the diphenyl ether carboxamide skeleton is a new starting point for the discovery of new SDH inhibitors and is worthy of further development.

Generation of Indoles with Agrochemical Significance through Biotransformation by Chaetomium globosum.

Six new (1-6) and two known (7 and 8) indole alkaloids were produced by the marine fish-derived fungus Chaetomium globosum 1C51 through biotransformation. The structures of these alkaloids were elucidated by a combination of MS, NMR, and X-ray crystallography analyses. Chaetoindolone A (1) was shown to inhibit the growth of the rice-pathogenic bacteria Xanthomonas oryzae pv. oryzae (xoo) both in vitro and in vivo. Chaetogline A (7) was found to be fungicidal against Sclerotinia sclerotiorum, a pathogen causing rape sclerotinia rot. Collectively, this work provides access to new indole alkaloids with potential agrochemical significance.

Design, synthesis, and antifungal activity of carboxamide derivatives possessing 1,2,3-triazole as potential succinate dehydrogenase inhibitors.

Succinate dehydrogenase (SDH) is demonstrably one of the most important molecular targets in development of new fungicide. In our continuous efforts to discover novel SDH inhibitors, forty-two carboxamide derivatives containing 1,2,3-triazole ring were designed and synthesized, which were precisely characterized by 1H NMR, ESI-MS, elemental analysis and X-ray single-crystal diffraction. The compounds were screened for antifungal activities against phytopathogenic fungi by mycelia growth inhibition assay in vitro. Compound A3-3 exhibited significant antifungal activity against Sclerotinia sclerotiorum, Botrytis cinerea, Rhizoctonia cerealis and Gaeumannomyces graminsis with EC50 values of 1.08, 8.75, 1.67 and 5.30 µg/mL, respectively, comparable to those of commercial SDHI boscalid. In vivo testing demonstrated that A3-3 was effective for suppressing rape sclerotinia rot, cucumber grey mould and wheat powdery mildew caused by S. sclerotiorum， B. cinerea and Blumeria graminis at a dosage of 200 µg/mL. Inhibition activities against SDH test proved the designed analogues were effective in the enzyme level. The molecular docking simulation revealed that A3-3 interacted with ARG43，TYR58 and TRP173 of the SDH through hydrogen bond and pi-pi interaction, which could explain the probable mechanism of action between the inhibitor and target protein.

New Metabolites from Endophytic Fungus Chaetomium globosum CDW7.

Five metabolites including two new ones, prochaetoviridin A (1) and chaetoindolin A (2), were isolated from the endophytic fungus Chaetomium globosum CDW7. Compounds 1 and 2 were characterized as an isocoumarin and an indole alkaloid derivative, respectively, with their structures elucidated by comprehensive spectroscopic analyses including high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), NMR, and circular dichroism (CD) comparison. Compounds 3⁻5 were identified as chaetoviridin A, chaetoglobosin R, and chaetoglobosin T, respectively. Chaetoviridin A (3) exhibited antifungal activity against Sclerotinia sclerotiorum with an EC50 value of 1.97 µg/mL. In vivo test showed that 3 displayed a protective efficacy of 64.3% against rape Sclerotinia rot at the dosage of 200 µg/mL, comparable to that of carbendazim (69.2%).

Nigrosporanenes C and D, two new cyclohexene derivatives from the enphytic fungus Nigrospora oryzae S4.

Two new cyclohexene derivatives, nigrosporanenes C and D (1 and 2), together with three known compounds (3-5), were isolated from the culture of an endophyte Nigrospora oryzae S4. Their structures were characterized by a combination of detailed spectroscopic analysis and comparison of their NMR data with those reported in the literature. All compounds were tested for anti-phytopathogenic activity, however, none of them showed activity at a concentration of 20 µM.

Chaetomium globosum CDW7, a potential biological control strain and its antifungal metabolites.

Screening for endophytic fungi with antifungal activity is an effective strategy for the discovery of novel biopesticides. Our previous work indicated that Chaetomium globosum CDW7, an endophyte from Ginkgo biloba, exhibited strong inhibitory activity against plant pathogenic fungi in vitro. In this study, we evaluated the CDW7 strain for its antifungal activities against nine phytopathogenic fungi and its biocontrol potential against rape sclerotinia rot caused by Sclerotinia sclerotiorum. The fermentation broth of CDW7 could successfully inhibit disease development in S. sclerotiorum-infected rape in vivo with 57.8% protective efficiency, which is comparable to that of carbendazim (59.8%) at 250 µg mL-1. The fermentation broth also expressed significant activity stability when exposed to 60°C and UV illumination, or when stored at 4°C. Furthermore, we found that 10% fermentation broth can promote the germination and growth of rape seedlings. Followed by the bioassay-guided approach, seven known metabolites were isolated and identified by spectroscopic analyses. Among them, chaetoglobosin A and D exhibited inhibitory activity against S. sclerotiorum with IC50 values of 0.35 and 0.62 µg mL-1, respectively, compared with carbendazim (0.17 µg mL-1). Therefore, our study demonstrated that CDW7 is a promising biocontrol fungus against S. sclerotiorum in agriculture.

Synthesis of 1,2,3-triazole hydrazide derivatives exhibiting anti-phytopathogenic activity.

A series of new 1,2,3-triazole derivatives have been prepared and screened for their antifungal activity against phytopathogenic fungi using the mycelium growth inhibition method in vitro. The results indicated that the 1,2,3-triazole hydrazide scaffold displayed significant antifungal activity. Compound 6ad exhibited the most potent anti-phytopathogenic activity, with EC50 values of 0.18, 0.35, 0.37 and 2.25 µg mL-1 against Rhizoctonia solani, Sclerotinia sclerotiorum, Fusarium graminearum, and Magnaporthe oryzae, respectively. In vivo testing demonstrated that 6ad was effective for the control of rice sheath blight, rape sclerotinia rot, fusarium head blight and rice blast caused by the aforementioned phytopathogens. This work suggests that the combination of 1,2,3-triazole and hydrazide moiety could be a promising fungicide scaffold in the future.

Synthesis and Biological Evaluation of Benzimidazole Phenylhydrazone Derivatives as Antifungal Agents against Phytopathogenic Fungi.

A series of benzimidazole phenylhydrazone derivatives (6a-6ai) were synthesized and characterized by ¹H-NMR, ESI-MS, and elemental analysis. The structure of 6b was further confirmed by single crystal X-ray diffraction as (E)-configuration. All the compounds were screened for antifungal activity against Rhizoctonia solani and Magnaporthe oryzae employing a mycelium growth rate method. Compound 6f exhibited significant inhibitory activity against R. solani and M. oryzae with the EC50 values of 1.20 and 1.85 µg/mL, respectively. In vivo testing demonstrated that 6f could effectively control the development of rice sheath blight (RSB) and rice blast (RB) caused by the above two phytopathogens. This work indicated that the compound 6f with a benzimidazole phenylhydrazone scaffold could be considered as a leading structure for the development of novel fungicides.

Anti-phytopathogenic activity of sporothriolide, a metabolite from endophyte Nodulisporium sp. A21 in Ginkgo biloba.

Phytopathogenic fungi such as Rhizoctonia solani and Sclerotinia sclerotiorum caused multiple plant diseases resulting in severe loss of crop production. Increasing documents endorsed that endophytes are a striking resource pool for numerous metabolites with various bioactivities such as anti-fungal. Here we reported the characterization and anti-phytopathogenic activity of sporothriolide, a metabolite produced by Nodulisporium sp. A21-an endophytic fungus in the leaves of Ginkgo biloba. Among the total twenty-five endophytic fungi isolated from the healthy leaves of G. biloba, the fermentation broth (FB) of the strain A21 was found potently inhibitory activity against R. solani and S. sclerotiorum using mycelia growth inhibition method. A21 was then identified as Nodulisporium sp., the asexual stage of Hypoxylon sp., by microscopic examination and ITS rDNA sequence data comparison. Under the bioassay-guided fractionation, sporothriolide was isolated from the petroleum ether extract of the FB of A21, whose structure was established by integrated interpretation of HR-ESI-MS and (1)H- and (13)C-NMR. Furthermore, the crystal structure of sporothriolide was first reported. In addition, sporothriolide was validated to be potently antifungal against R. solani, S. sclerotiorum and inhibit conidium germination of Magnaporthe oryzae in vitro and in vivo, indicating that it could be used as a lead compound for new fungicide development.

[DNA Barcode Screening and Identifying for Anredera cordifolia and Its Related Species].

Objective: To identify Anredera cordifolia and its closely related species using the DNA barcode. Methods: 28 individuals of Anredera cordifolia and its close related species were collected from different habitats. ITS and ITS2 of ribosomal DNA,matK,rbcL and psb A-trn H of chloroplast DNA were amplified and sequenced. The amplification and sequencing success rate,barcoding gap,and NJ trees were used to evaluate the efficiency of species identification. Results: After amplified and sequenced, base deletion was occurred in psb A-trnH sequences of Basella alba. The sequencing success rates of mat K,rbc L,ITS and ITS2 were 100%,100%,78. 75% and64. 28%,respectively. Among the four DNA barcoding sequences,ITS and mat K had remarkable barcoding gap. The NJ tree showed that Anredera cordifolia could differed obviously from its closely related species by ITS and mat K. Conclusion: The sequences of ITS and matK provide an effective and fast tool for the identification and authentication of medicinal plant of Anredera cordifolia and its related species.

Construction of an immobilised acetylcholinesterase column and its application in screening insecticidal constituents from Magnolia officinalis.

BACKGROUND: Application of a matrix-immobilised target enzyme for screening inhibitors is widely used in drug development, but there are few studies in insecticide discovery. In this paper, an economical and effective immobilised acetylcholinesterase (AChE) column was prepared using the sol-gel embedment method, which was further combined with high-performance liquid chromatography for screening the AChE inhibitors and insecticidal compounds from complex natural products. RESULTS: AChE inhibitory constituents magnolol and honokiol were isolated from the ethanol extract of Magnolia officinalis, with IC50 values of 0.069 and 0.057 mM respectively. In an in vivo bioassay, magnolol and honokiol showed insecticidal activity against Nilaparvata lugens, with LC50 values of 0.324 and 0.137 mM, which are comparable with that of commonly used insecticide chlorpyrifos (0.233 mM). Moreover, molecular docking was carried out against a homology model of N. lugens AChE. The complexes showed that magnolol and honokiol placed themselves nicely into the active site of the enzyme and exhibited an interaction energy that was in accordance with our activity profile data. CONCLUSION: These results demonstrate that magnolol and honokiol have great applied potential to be developed as natural insecticides, and an immobilised AChE column is very useful as a rapid screening tool for target enzymes towards potent inhibitors.

Synthesis and antifungal activity of 1,2,3-triazole phenylhydrazone derivatives.

A series of 1,2,3-triazole phenylhydrazone derivatives were designed and synthesized as antifungal agents. Their structures were determined based on (1)H-NMR spectroscopy, MS, elemental analysis and X-ray single-crystal diffraction. The antifungal activities were evaluated against four phytopathogenic fungi including Rhizoctonia solani, Sclerotinia sclerotiorum, Fusarium graminearum and Phytophthora capsici, by the mycelium growth inhibition method in vitro. Compound 5p exhibited significant anti-phytopathogenic activity, with the EC50 values of 0.18, 2.28, 1.01, and 1.85 µg mL(-1), respectively. In vivo testing demonstrated that 5p was effective in the control of rice sheath blight, rape sclerotinia rot and fusarium head blight. A 3D-QSAR model was built for a systematic SAR profile to explore more potent 1,2,3-triazole phenylhydrazone analogs as novel fungicides.

Design, synthesis, antifungal, and antioxidant activities of (E)-6-((2-phenylhydrazono)methyl)quinoxaline derivatives.

Different substituted phenylhydrazone groups were linked to the quinoxaline scaffold to provide 26 compounds (6a-6z). Their structures were confirmed by (1)H and (13)C NMR, MS, elemental analysis, and X-ray single-crystal diffraction. The antifungal activities of these compounds against Rhizoctonia solani were evaluated in vitro. Compound 6p is the most promising one among all the tested compounds with an EC50 of 0.16 µg·mL(-1), more potent than the coassayed positive control fungicide carbendazim (EC50: 1.42 µg·mL(-1)). In addition, these compounds were subjected to antioxidant assay by employing diphenylpicrylhydrazyl (DPPH) and mice microsome lipid peroxidation (LPO) methods. Most of these compounds are potent antioxidants. The strongest compounds are 6e (EC50: 7.60 µg·mL(-1), DPPH) and 6a (EC50: 0.96 µg·mL(-1), LPO), comparative to or more potent than the positive control Trolox [EC50: 5.90 µg·mL(-1) (DPPH) and 18.23 µg·mL(-1) (LPO)]. The structure and activity relationships were also discussed.

Synthesis and antifungal activity of nicotinamide derivatives as succinate dehydrogenase inhibitors.

Thirty-eight nicotinamide derivatives were designed and synthesized as potential succinate dehydrogenase inhibitors (SDHI) and precisely characterized by (1)H NMR, ESI-MS, and elemental analysis. The compounds were evaluated against two phytopathogenic fungi, Rhizoctonia solani and Sclerotinia sclerotiorum, by mycelia growth inhibition assay in vitro. Most of the compounds displayed moderate activity, in which, 3a-17 exhibited the most potent antifungal activity against R. solani and S. sclerotiorum with IC50 values of 15.8 and 20.3 µM, respectively, comparable to those of the commonly used fungicides boscalid and carbendazim. The structure-activity relationship (SAR) of nicotinamide derivatives demonstrated that the meta-position of aniline was a key position contributing to the antifungal activity. Inhibition activities against two fungal SDHs were tested and achieved the same tendency with the data acquired from in vitro antifungal assay. Significantly, 3a-17 was demonstrated to successfully suppress disease development in S. sclerotiorum infected cole in vivo. In the molecular docking simulation, sulfur and chlorine of 3a-17 were bound with PHE291 and PRO150 of the SDH homology model, respectively, which could explain the probable mechanism of action between the inhibitory and target protein.

Cerebroside C increases tolerance to chilling injury and alters lipid composition in wheat roots.

Chilling tolerance was increased in seed germination and root growth of wheat seedlings grown in media containing 20 µg/mL cerebroside C (CC), isolated from the endophytic Phyllosticta sp. TG78. Seeds treated with 20 µg/mL CC at 4 °C expressed the higher germination rate (77.78%), potential (23.46%), index (3.44) and the shorter germination time (6.19 d); root growth was also significantly improved by 13.76% in length, 13.44% in fresh weight and 6.88% in dry mass compared to controls. During the cultivation process at 4 °C for three days and the followed 24 h at 25 °C, lipid peroxidation, expressed by malondialdehyde (MDA) content and relative membrane permeability (RMP) was significantly reduced in CC-treated roots; activities of lipoxygenase (LOX), phospholipid C (PLC) and phospholipid D (PLD) were inhibited by 13.62-62.26%, 13.54-63.93% and 13.90-61.17%, respectively; unsaturation degree of fatty acids was enhanced through detecting the contents of CC-induced linoleic acid, linolenic acid, palmitic acid and stearic acid using GC-MS; capacities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) were individually increased by 7.69-46.06%, 3.37-37.96%, and -7.00-178.07%. These results suggest that increased chilling tolerance may be due, in part, to the reduction of lipid peroxidation and alternation of lipid composition of roots in the presence of CC.

Synthesis and antioxidant activity of novel Mannich base of 1,3,4-oxadiazole derivatives possessing 1,4-benzodioxan.

A new series of Mannich base of 1,3,4-oxadiazole derivatives possessing 1,4-benzodioxan (6a-6ae) were synthesized and characterized by (1)H NMR, ESI-MS and elemental analysis. The structure of 6b was further confirmed by single crystal X-ray diffraction. All these novel compounds were screened for their in vitro antioxidant activity employing 2,2'-diphenyl-1-picrylhydrazyl radical (DPPH), 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonate) cationic radical (ABTS(+)) and ferric reducing antioxidant power (FRAP) scavenging assays. Due to the combination of 1,4-benzodioxan, 1,3,4-oxadiazoles and substituted phenyl ring, most of them exhibited nice antioxidant activities. In all of these three assays mentioned above, compounds 6f and 6e showed significant radical scavenging ability comparable to the commonly used antioxidants, BHT and Trolox. Seven compounds with representative substituents or activities were selected for further assays in chemical simulation biological systems-inhibition of microsomal lipid peroxidation (LPO) and protection against 2,2'-azobis (2-amidinopropane hydrochloride) (AAPH) induced DNA strand breakage, in which, 6f and 6e were demonstrated to be of the most potent antioxidant activities.

Antifungal screening of endophytic fungi from Ginkgo biloba for discovery of potent anti-phytopathogenic fungicides.

Many endophytic fungi have been found to synthesize bioactive compounds to defend host plants against pathogenic organisms. Here we performed anti-fungal bioassay of 80 endophytic fungi isolated from Ginkgo biloba. Fifteen endophytes were active against at least one of the selected fungi, Fusarium graminearum, Sclerotinia sclerotiorum and Phytophthora capsici, using the agar diffusion method. The most bioactive strain CDW7 was identified as Chaetomium globosum by microscopic examination and ITS rRNA gene sequence data. Culture broth of CDW7 diluted 3-fold completely inhibited the mycelial growth and conidia germination of F. graminearum in vitro. Therefore, Fusarium head blight, a common disease in wheat and barley associated with Fusarium spp., was used to test the anti-phytopathogenic activity in vivo. The fermentation broth of CDW7 resulted in a protective efficacy of 54.9% and curative efficacy of 48.8%. Followed by a bioassay-guided approach, 1,2-benzenedicarboxaldehyde-3,4,5-trihydroxy-6-methyl (flavipin) was isolated and demonstrated to significantly inhibit the growth of several plant-pathogenic fungi, especially F. graminearum with an EC(50) value of 0.73 µg mL(-1) comparable to the commonly used fungicide carbendazim, indicating that it could be used as a fungicide or as a lead compound of new fungicides.

The combination of 4-anilinoquinazoline and cinnamic acid: a novel mode of binding to the epidermal growth factor receptor tyrosine kinase.

A novel type of cinnamic acid quinazoline amide derivatives (20-42), which designed the combination between quinazoline as the backbone and various substituted cinnamic acid as the side chain, have been synthesized and their biological activities were evaluated within cytotoxicity assay firstly and then potent EGFR inhibitory activity. Compound 42 demonstrated the most potent inhibitory activity (IC(50)=0.94 µM for EGFR), which could be optimized as a potential EGFR inhibitor in the further study. Docking simulation was performed to position compound 42 into the EGFR active site to determine the probable binding model. Analysis of the binding conformation of 42 in active site displayed compound 42 was stabilized by hydrogen bonding interactions with Lys822, which was different from other derivatives. In the further study, Compounds 43 and 44 had been synthesized and their biological activities were also evaluated, which were the same as that we expected. Compound 43 has demonstrated significant EGFR (IC(50)=0.12 µM) and tumor growth inhibitory activity as a potential anticancer agent.

Benzophenones from Guignardia sp. IFB-E028, an endophyte on Hopea hainanensis.

The first natural S-containing benzophenone dimer, named guignasulfide (3), was isolated from the culture of Guignardia sp. IFB-E028, an endophytic fungus residing in healthy leaves of Hopea hainanensis. Its structure was determined through correlative analyses of its MS, 1D- and 2D-NMR spectroscopic data. Two other known benzophenone derivatives, monomethylsulochrin and rhizoctonic acid (1 and 2, resp.) were also isolated. Guignasulfide (3) was more active against the human liver cancer cell line HepG2 (IC(50) value: 5.2+/-0.4 microM) than metabolites 1 and 2 (IC(50) values: 63.5+/-0.6 and 60.2+/-0.5 microM); compounds 1-3 showed also moderately inhibitory effects on the human bacterial pathogen Helicobacter pylori with MIC values of 28.9+/-0.1, 60.2+/-0.4, and 42.9+/-0.5 microM, respectively.

[Study on chemical constituents isolated from Juncus effuses].

OBJECTIVE: To study the constituents isolated from Juncus effuses Linn.. METHODS: The compounds were isolated through chromatographic approaches, such as Silica gel. CC, Sephadex LH-20 and were identified by a combination of spectroscopic methods (UV, MS, 1H-NMR, 13C-NMR and NOE). RESULTS: 8 kinds of compounds were identified which included four flavonoids, luteolin (I), eriodictyol (II), 2', 5', 5, 7-tetrahydroxyflavone (III), glucoluteolin (IV); two aromatic compounds, methyl p-hydroxybenzoate (V), vanillic acid (VI); a fatty acid, hexadecanoic acid (I) and an alkane, n-Tetradecane (VIII). CONCLUSION: Compound II, III, V, VI, VII, VIII were firstly isolated from this plant.