Marine Natural Product for Pesticide Candidate: Pulmonarin Alkaloids as Novel Antiviral and Anti-Phytopathogenic-Fungus Agents.

Plant diseases are seriously endangering agricultural production. The emergence of drug resistance has brought great challenges to the prevention and control of plant diseases. There is an urgent need for the emergence of new drug candidates. In this work, we achieved the efficient synthesis of pulmonarins A and B in 64% and 59% overall yield, respectively. Pulmonarins A and B were found to have good antiviral activities against tobacco mosaic virus (TMV) for the first time. A series of pulmonarin derivatives were designed, synthesized, and evaluated for their antiviral and fungicidal activities systematically. Most compounds displayed higher anti-TMV activities than commercial ribavirin. Compounds 6a, 6c, and 6n with better inactivation effects than ningnanmycin emerged as new antiviral candidates. We selected 6c for further antiviral mechanism research, which revealed that it could inhibit virus assembly by interacting with TMV coat protein (CP). The molecular docking results further confirmed that these compounds could interact with CP through hydrogen bonding. These compounds also displayed broad spectrum fungicidal activities. Especially compound 6u with prominent antifungal activity emerged as a new fungicidal candidate for further research. The current work provides a reference for understanding the application of pulmonarin alkaloids in plant protection.

The first synthesis, carbonic anhydrase inhibition and anticholinergic activities of some bromophenol derivatives with S including natural products.

Starting from vanillin, known four benzyl bromides with Br were synthesized. The first synthesis of natural product 3,4-dibromo-5-((methylsulfonyl)methyl)benzene-1,2-diol (2) and 3,4,6-tribromo-5-((methylsulfonyl)methyl)benzene-1,2-diol (3) and derivatives were carried out by demethylation, acetylatilation, oxidation and hydrolysis reactions of the benzyl bromides. Also, these compounds were tested against some important enzymes like acetylcholinesterase and butyrylcholinesterase enzymes, carbonic anhydrase I, and II isoenzymes. The novel bromophenols showed Ki values of in range of 53.75 ±â€¯12.54-234.68 ±â€¯46.76 nM against hCA I, 42.84 ±â€¯9.36 and 200.54 ±â€¯57.25 nM against hCA II, 0.84 ±â€¯0.12-14.63 ±â€¯3.06 nM against AChE and 0.93 ±â€¯0.20-18.53 ±â€¯5.06 nM against BChE. Induced fit docking process performed on the compounds inhibiting hCA I, hCA II, AChE, and BChE receptors. Hydroxyl group should exist at the aromatic ring of the compounds for inhibition of the enzymes. The moieties reported in this study will be useful for design of more potent and selective inhibitors against the enzymes.

Synthesis of Transition-State Inhibitors of Chorismate Utilizing Enzymes from Bromobenzene cis-1,2-Dihydrodiol.

In order to survive in a mammalian host, Mycobacterium tuberculosis (Mtb) produces aryl-capped siderophores known as the mycobactins for iron acquisition. Salicylic acid is a key building block of the mycobactin core and is synthesized by the bifunctional enzyme MbtI, which converts chorismate into isochorismate via a SN2″ reaction followed by further transformation into salicylate through a [3,3]-sigmatropic rearrangement. MbtI belongs to a family of chorismate-utilizing enzymes (CUEs) that have conserved topology and active site residues. The transition-state inhibitor 1 described by Bartlett, Kozlowski, and co-workers is the most potent reported inhibitor to date of CUEs. Herein, we disclose a concise asymmetric synthesis and the accompanying biochemical characterization of 1 along with three closely related analogues beginning from bromobenzene cis-1S,2S-dihydrodiol produced through microbial oxidation that features a series of regio- and stereoselective transformations for introduction of the C-4 hydroxy and C-6 amino substituents. The flexible synthesis enables late-stage introduction of the carboxy group and other bioisosteres at the C-1 position as well as installation of the enol-pyruvate side chain at the C-5 position.

Synthesis of C2-Symmetric Benzimidazolium Salts and Their Application in Palladium-Catalyzed Enantioselective Intramolecular α-Arylation of Amides.

A series of C2-symmetric chiral benzimidazolium salts, the precursor of N-heterocyclic carbene ligands, were designed and synthesized from 1,2-dibromobenzene. In situ prepared corresponding carbenes were tested in the asymmetric palladium-catalyzed intramolecular α-arylation of amides, affording chiral diarylmethanols with high yields and moderate enantioselectivities.

Synthesis and cytotoxic activity of novel hexahydropyrrolo[2,3-b]indole imidazolium salts.

A series of novel hexahydropyrrolo[2,3-b]indole-1H-imidazolium salts were synthesized and evaluated in vitro against a panel of human tumor cell lines. The results suggest that the 5,6-dimethyl-benzimidazole ring, and substitution of the imidazolyl-3-position with a 2-bromobenzyl or 2-naphthylmethyl group, were important for the cytotoxic activity. Notably, Compound 43, bearing a 2-bromobenzyl substituent at position-3 of 5,6-dimethyl-benzimidazole, was found to possess the most potent derivative against five human tumor cell lines with IC50 values below 2.68µM and more selective towards SMMC-7721, A549 and SW480 cell lines. Compounds 25 and 39 were more selective to HL-60 and MCF-7 cell lines with IC50 values of 0.47 and 1.46µM.

Antifouling compounds from the sub-arctic ascidian Synoicum pulmonaria: synoxazolidinones A and C, pulmonarins A and B, and synthetic analogues.

The current study describes the antifouling properties of four members belonging to the recently discovered synoxazolidinone and pulmonarin families, isolated from the sub-Arctic sessile ascidian Synoicum pulmonaria collected off the Norwegian coast. Four simplified synthetic analogues were also prepared and included in the study. Several of the studied compounds displayed MIC values in the micro-nanomolar range against 16 relevant marine species involved in both the micro- and macrofouling process. Settlement studies on Balanus improvisus cyprids indicated a deterrent effect and a low toxicity for selected compounds. The two synoxazolidinones displayed broad activity and are shown to be among the most active natural antifouling bromotyrosine derivatives described. Synoxazolidinone C displayed selected antifouling properties comparable to the commercial antifouling product Sea-Nine-211. The pulmonarins prevented the growth of several bacterial strains at nanomolar concentrations but displayed a lower activity toward microalgae and no effect on barnacles. The linear and cyclic synthetic peptidic mimics also displayed potent antifouling activities mainly directed against bacterial adhesion and growth.

Stereocontrolled synthesis of adjacent acyclic quaternary-tertiary motifs: application to a concise total synthesis of (-)-filiformin.

Lithiation/borylation methodology has been developed for the synthesis of acyclic quaternary-tertiary motifs with full control of relative and absolute stereochemistry, thus leading to all four possible isomers of a stereodiad. A novel intramolecular Zweifel-type olefination enabled acyclic stereocontrol to be transformed into cyclic stereocontrol. These key steps have been applied to the shortest enantioselective synthesis of (-)-filiformin to date (9 steps) with full stereocontrol.

Palladium-catalyzed three-component reaction of 2-alkynylbromobenzene, 2-alkynylaniline, and electrophile: an efficient pathway for the synthesis of diverse 11H-indeno[1,2-c]quinolines.

Diverse 11H-indeno[1,2-c]quinolines are produced via a palladium-catalyzed three-component reaction of 2-alkynylbromobenzene, 2-alkynylaniline, and electrophile. This conversion tolerates a wide variety of functionality and substitution patterns on the 11H-indeno[1,2-c]quinoline ring.

Synthesis and carbonic anhydrase inhibitory properties of novel bromophenols including natural products.

(2-Bromo-3,4-dimethoxyphenyl) (3,4-dimethoxyphenyl)methanone (10) and its derivatives with Br, one dibromide and isomeric three tribromides, were synthesized. Demethylation of these compounds afforded a series of new bromophenols. Inhibition of human cytosolic carbonic anhydrase II (hCA II) isozyme by these new bromophenols and naturally occurring 3,4,6-tribromo-5-(2,5-dibromo-3,4-dihydroxybenzyl)benzene-1,2-diol (3), and 5,5'-methylenebis(3,4,6-tribromo-benzene-1,2-diol) (4) was investigated. The synthesized compounds showed carbonic anhydrase inhibitory capacities with IC(50) values in the range of 0.7-372 µM against hCA II. Some bromophenols investigated here showed effective hCA II inhibitory activity and might be used as leads for generating novel carbonic anhydrase inhibitors which are valuable drug candidates for the treatment of glaucoma, epilepsy, gastric and duodenal ulcers, neurological disorders, or osteoporosis.

PET imaging of norepinephrine transporter-expressing tumors using 76Br-meta-bromobenzylguanidine.

UNLABELLED: Meta-iodobenzylguanidine (MIBG) labeled with (123)I or (131)I has been widely used for the diagnosis and radiotherapy of norepinephrine transporter (NET)-expressing tumors. However, (123)I/(131)I-MIBG has limitations for detecting small lesions because of its lower spatial resolution than PET tracers. In this study, meta-bromobenzylguanidine (MBBG) labeled with (76)Br (half-life, 16.1 h), an attractive positron emitter, was prepared and evaluated as a potential PET tracer for imaging NET-expressing tumors. METHODS: (76)Br-MBBG was prepared by a halogen-exchange reaction between the (76)Br and iodine of nonradioactive MIBG. The stability of MBBG was evaluated in vitro and in vivo by high-performance liquid chromatography analysis. Cellular uptake studies with or without NET inhibitors were performed in NET-positive PC-12 cell lines. Biodistribution studies were performed in PC-12 tumor-bearing nude mice by administration of a mixed solution of MBBG, MIBG, and (18)F-FDG. The tumor was imaged using (76)Br-MBBG and (18)F-FDG with a small-animal PET scanner. RESULTS: MBBG was stable in vitro, but some time-dependent dehalogenation was observed after administration in mice. MBBG showed high uptake in PC-12 tumor cells that was significantly decreased by the addition of NET inhibitors. In biodistribution studies, MBBG showed high tumor accumulation (32.0 +/- 18.6 percentage injected dose per gram at 3 h after administration), and the tumor-to-blood ratio reached as high as 54.4 +/- 31.9 at 3 h after administration. The tumor uptake of MBBG correlated well with that of MIBG (r = 0.997) but not with that of (18)F-FDG. (76)Br-MBBG PET showed a clear image of the transplanted tumor, with high sensitivity, which was different from the lesion shown by (18)F-FDG PET. CONCLUSION: (76)Br-MBBG showed high tumor accumulation, which correlated well with that of MIBG, and provided a clear PET image. These results indicated that (76)Br-MBBG would be a potential PET tracer for imaging NET-expressing neuroendocrine tumors and could provide useful information for determining the indications for (131)I-MIBG therapy.

Synthesis and biological analysis of a new curcumin analogue for enhanced anti-tumor activity in HepG 2 cells.

The aim of the present study was to investigate the apoptosis of human hepatocellular carcinoma cell line HepG 2 induced by a new curcumin analogue, GL63. HepG 2 cells were treated with increasing doses of GL63 and curcumin for 48 h. The proliferation of cells was detected with MTT. The apoptosis were examined by flow cytometry. The caspase-3 activity was detected by western blotting. ER calcium stores were assessed by the fluorescent calcium indicator fura-2/AM. The protein expression of ER stress pathway, GRP78, XBP-1, ATF-4 and CHOP were examined with western blotting. Growth inhibitory effect was observed for treatment with GL63 in a dose-dependent manner and with more potential than curcumin. GL63 at 20 microM induced significant apoptosis in HepG 2 cells. Furthermore, GL63 induced the ER stress response, up-regulation of CHOP, XBP-1, ATF-4 and GRP78 expression in a dose-dependent, while curcumin had no effect on ER stress. These results suggest that GL63 has more potent anti-tumor activity than curcumin, which is associated with activation of ER stress and induction of apoptosis in HepG 2 cells.

Synthesis of cyanoformamides from primary amines and carbon dioxide under mild conditions. Synthesis of ceratinamine.

Treatment of primary amines with tetramethylphenylguanidine (PhTMG) and a cyanophosphonate at -10 degrees Celsius under an atmosphere of carbon dioxide provides cyanoformamides in very high to excellent yields. The reaction proceeds efficiently within a short time. By-products were not detected in most runs and epimerization was not found when optically pure alpha-aminoesters were used as substrates. As an example, the reaction was applied to the synthesis of the marine natural product ceratinamine.

Flexible synthesis of pyrimidines with chiral monofluorinated and difluoromethyl side chains.

Chiral pyrimidines with a fluorine atom in the benzylic position are easily accessible in high enantiomeric excesses from optically active propargylic intermediates by two complementary routes. Both the use of optically active propargylic fluorides and the fluorination of the chiral pyrimidine in the final stage give excellent results in terms of enantiocontrol. On the other hand, original pyrimidines with a difluoromethyl side chain are also obtained in a few steps from new propargylic ketones bearing a CHF(2) substituent on the triple bond.

Synthetic approaches to the microtubule-stabilizing sponge alkaloid ceratamine A and desbromo analogues.

Two synthetic approaches to the microtubule-stabilizing ceratamine alkaloids are described. The first approach involved attempts to graft an aminoimidazole moiety onto an azepine ring to form partially hydrogenated versions of the unprecedented aromatic imidazo[4,5-d]azepine core of the ceratamines. This route ultimately failed because it was not possible to aromatize the partially hydrogenated ceratamine intermediates. A second approach started with tribromoimidazole that was sequentially metalated and functionalized to efficiently generate a key imidazole intermediate containing vinyl bromide and amide functionalities. An intramolecular Buchwald vinyl amidation reaction converted this key intermediate into a bicyclic imidazo[4,5-d]azepine that was at the same oxidation state as the aromatic core of the ceratamines. The 2-amino functionality present on the imidazole ring of the ceratamines was installed using a Buchwald/Hartwig amination reaction on a 2-chloroimidazole precursor. Deprotection and aromatization resulted in the first synthesis of desbromoceratamine A (55) and desmethyldesbromoceratamine A (60). An unanticipated addition of atmospheric oxygen was encountered during deprotection of the imidazole ring in the last step of the synthesis leading to C-11 oxygenated ceratamine analogues as byproducts. Evaluation of the synthetic ceratamines in a TG3 cell-based assay for mitotic arrest revealed that the C-14 and C-16 bromine substituents in ceratamine A (1) play a major role in the antimitotic potency of the natural product. The synthetic route to ceratamine analogues has provided sufficient quantities of desbromoceratamine A (55) for testing in mouse models of cancer.

A novel synthesis of bromobenzenes using molecular bromine.

Certain substituted bromobenzenes have been synthesized in acceptable yields using a novel Sandmeyer type reaction. The reactions are relatively quick and possibly proceed via a radical mechanism.

Synthesis, spectral studies of salicylidine-pyridines: crystal and molecular structure of 2-[(1E)-2-aza-2-(5-methyl(2-pyridyl)ethenyl)]-4-bromobenzen-1-ol.

Schiff bases derived from different meta-substituted salicylaldehyde and 5-methylaminopyridine have been synthesized and characterized by elemental analysis, FT-IR, NMR and UV-vis techniques. NMR assignments were made using (1)H, (13)C NMR and aided by 2D HETCOR and HMBC heteronuclear correlation techniques. The UV-vis spectra of the compounds were found useful in understanding the existence of tautomeric equilibria [phenol-imine (O-H...N) and keto-amine (O...H-N) forms] in polar and non-polar solvents. In order to rationalize the stabilization of tautomer in solid state, X-ray structure of 2-[(1E)-2-aza-2-(5-methyl(2-pyridyl)ethenyl)]-4-bromobenzen-1-ol (6) was determined. According to our crystallographic result, it has enol-imine tautomeric form.

Synthesis and biological evaluation of purealin and analogues as cytoplasmic dynein heavy chain inhibitors.

Cytoplasmic dynein plays important roles in membrane transport, mitosis, and other cellular processes. A few small-molecule inhibitors of cytoplasmic dynein have been identified. We report here the first synthesis of purealin, a natural product isolated from the sea sponge Psammaplysilla purea, which is known to inhibit axonemal dynein. Also described are the first syntheses, by modular amide coupling reactions, of the natural product purealidin A (a component of purealin) and a small library of analogues. The library was examined for inhibition of cytoplasmic dynein heavy chain and cell growth. The compounds showed effective antiproliferative activity against a mouse leukemia cell line but selective activities against human carcinoma cell lines. Purealin and some of the analogues inhibited the microtubule-stimulated ATPase activity of recombinant cytoplasmic dynein heavy chain motor domain. The inhibitory effect of purealin was concentration dependent and uncompetitive, supporting the hypothesis that it does not compete with the binding of ATP.

An improved method for the bromination of metalated haloarenes via lithium, zinc transmetalation: a convenient synthesis of 1,2-dibromoarenes.

A facile protocol for the synthesis of 1,2-dibromoarenes is described. A standard ortho-lithiation/bromination procedure, when applied to bromoarenes, resulted in poor yields of the corresponding 1,2-dibromoarenes (13-62% yield). However, transmetalation of the transient aryllithium intermediate to an arylzinc species with ZnCl2, followed by bromination, resulted in dramatically improved yields of the synthetically useful 1,2-dibromoarenes (68-95% yield).

A small-molecule inhibitor of Mps1 blocks the spindle-checkpoint response to a lack of tension on mitotic chromosomes.

The spindle checkpoint prevents chromosome loss by preventing chromosome segregation in cells with improperly attached chromosomes [1, 2 and 3]. The checkpoint senses defects in the attachment of chromosomes to the mitotic spindle [4] and the tension exerted on chromosomes by spindle forces in mitosis [5, 6 and 7]. Because many cancers have defects in chromosome segregation, this checkpoint may be required for survival of tumor cells and may be a target for chemotherapy. We performed a phenotype-based chemical-genetic screen in budding yeast and identified an inhibitor of the spindle checkpoint, called cincreasin. We used a genome-wide collection of yeast gene-deletion strains and traditional genetic and biochemical analysis to show that the target of cincreasin is Mps1, a protein kinase required for checkpoint function [8]. Despite the requirement for Mps1 for sensing both the lack of microtubule attachment and tension at kinetochores, we find concentrations of cincreasin that selectively inhibit the tension-sensitive branch of the spindle checkpoint. At these concentrations, cincreasin causes lethal chromosome missegregation in mutants that display chromosomal instability. Our results demonstrate that Mps1 can be exploited as a target and that inhibiting the tension-sensitive branch of the spindle checkpoint may be a way of selectively killing cancer cells that display chromosomal instability.

Development of an ELISA for the detection of bromoxynil in water.

For development of an indirect competitive enzyme-linked immunosorbent assay (ELISA) for the nitrile herbicide bromoxynil, the polyclonal antibodies raised against 2,6-dibromo-4-cyano-phenoxyacetic acid (hapten) conjugated to bovine serum albumin (BSA) by the N-hydroxysuccinimide-activated ester method. Antiserum with a sufficiently high titer to provide the determinations of targeted compounds was obtained only 77 days after the primary immunization. Antiserum A2 was applied to the residual analysis of some water samples, under optimized ELISA condition, the quantitative working range was from 10 to 500 ppb with a limit of detection of 5 ppb. Cross-reactivity to structurally similar agrochemicals and related chemicals was determined. The antiserum showed little cross-reactivity with 2,6-dibromophenol and bromoxynil octanoate ester which have a dibromophenol group as common structure, but showed no cross-reactivity with other herbicides. Each water sample (river water, tap water, purified water, and bottled water) had a matrix effect and was investigated by adding Tween20 in the assay buffer. These four kinds of water samples were fortified with bromoxynil at several concentration levels and were directly analyzed with only dilution with an equal volume of antiserum solution, the mean recovery was 102.3%, and the mean coefficient of variation was 5.96%. The proposed ELISA turned out to be a powerful tool for monitoring of residual bromoxynil in water samples at trace level.