Symmetrically substituted dichlorophenes inhibit N-acyl-phosphatidylethanolamine phospholipase D.

N-Acyl-phosphatidylethanolamine phospholipase D (NAPE-PLD) (EC 3.1.4.4) catalyzes the final step in the biosynthesis of N-acyl-ethanolamides. Reduced NAPE-PLD expression and activity may contribute to obesity and inflammation, but a lack of effective NAPE-PLD inhibitors has been a major obstacle to elucidating the role of NAPE-PLD and N-acyl-ethanolamide biosynthesis in these processes. The endogenous bile acid lithocholic acid (LCA) inhibits NAPE-PLD activity (with an IC50 of 68 µm), but LCA is also a highly potent ligand for TGR5 (EC50 0.52 µm). Recently, the first selective small-molecule inhibitor of NAPE-PLD, ARN19874, has been reported (having an IC50 of 34 µm). To identify more potent inhibitors of NAPE-PLD, here we used a quenched fluorescent NAPE analog, PED-A1, as a substrate for recombinant mouse Nape-pld to screen a panel of bile acids and a library of experimental compounds (the Spectrum Collection). Muricholic acids and several other bile acids inhibited Nape-pld with potency similar to that of LCA. We identified 14 potent Nape-pld inhibitors in the Spectrum Collection, with the two most potent (IC50 = ∼2 µm) being symmetrically substituted dichlorophenes, i.e. hexachlorophene and bithionol. Structure-activity relationship assays using additional substituted dichlorophenes identified key moieties needed for Nape-pld inhibition. Both hexachlorophene and bithionol exhibited significant selectivity for Nape-pld compared with nontarget lipase activities such as Streptomyces chromofuscus PLD or serum lipase. Both also effectively inhibited NAPE-PLD activity in cultured HEK293 cells. We conclude that symmetrically substituted dichlorophenes potently inhibit NAPE-PLD in cultured cells and have significant selectivity for NAPE-PLD versus other tissue-associated lipases.

Enhanced Degradation of Pesticide Dichlorophen by Laccase Immobilized on Nanoporous Materials: A Cytotoxic and Molecular Simulation Investigation.

Use of pesticides is usually related to overproduction of crops in order to overcome worldwide demand of food and alimentary safety. Nevertheless, pesticides are environmental persistent molecules, such as the organochlorine pesticides, which are often found in undesired places. In this work, we show that a hybrid nanomaterial (laccase-MSU-F) readily oxidizes the pesticide dichlorophen, reducing its acute genotoxicity and apoptotic effects. In order to predict chronic alterations related to endocrine disruption, we compared the calculated affinity of dichlorophen oxidized subproducts to steroid hormone nuclear receptors (NRs), using molecular simulation methods. We found a reduction in theoretical affinity of subproducts of oxidized dichlorophen for the ligand-binding pocket of NRs (∼5 kcal/mol), likewise of changes in binding modes, that suggests a reduction in binding events (RMSD values < 10 Å).

Identification, library synthesis and anti-vibriosis activity of 2-benzyl-4-chlorophenol from cultures of the marine bacterium Shewanella halifaxensis.

Summer Gut Syndrome (SGS) is caused by various Vibrio bacterial species and can have negative effects on aquaculture farms worldwide. In New Zealand, SGS is caused by Vibrio harveyii infecting King Salmon (Oncorhynchus tshawytscha). To find leads for the prevention of SGS, we screened the inhibitory effects of 16 strains of Shewanella upon V. harveyii growth in competitive solid phase cultures. The detailed investigation of Shewanella halifaxensis IRL548 revealed 2-benzyl-4-chlorophenol (1), a known, commercially available antibacterial agent, as the major bioactive component. Synthesis of a small library of congeners to confirm the natural product identity and to provide a structure-activity relationship for the observed activity was also completed. Compound 1 exhibits moderate activity against two pathogenic microorganisms.

Effects of new-generation inhibitors of the calcium-activated chloride channel anoctamin 1 on slow waves in the gastrointestinal tract.

BACKGROUND AND PURPOSE: High-throughput screening of compound libraries using genetically encoded fluorescent biosensors has identified several second-generation. low MW inhibitors of the calcium-activated chloride channel anoctamin 1 (CaCC/Ano1). Here we have (i) examined the effects of these Ano1 inhibitors on gastric and intestinal pacemaker activity; (ii) compared the effects of these inhibitors with those of the more classical CaCC inhibitor, 5-nitro-2-(3-phenylpropylalanine) benzoate (NPPB); (ii) examined the mode of action of these compounds on the waveform of pacemaker activity; and (iii) compared differences in the sensitivity between gastric and intestinal pacemaker activity to the Ano1 inhibitors. EXPERIMENTAL APPROACH: Using intracellular microelectrode recordings of gastric and intestinal muscle preparations from C57BL/6 mice, the dose-dependent effects of Ano1 inhibitors were examined on spontaneous electrical slow waves. KEY RESULTS: The efficacy of second-generation Ano1 inhibitors on gastric and intestinal pacemaker activity differed significantly. Antral slow waves were more sensitive to these inhibitors than intestinal slow waves. CaCCinh -A01 and benzbromarone were the most potent at inhibiting slow waves in both muscle preparations and more potent than NPPB. Dichlorophene and hexachlorophene were equally potent at inhibiting slow waves. Surprisingly, slow waves were relatively insensitive to T16Ainh -A01 in both preparations. CONCLUSIONS AND IMPLICATIONS: We have identified several second-generation Ano1 inhibitors, blocking gastric and intestinal pacemaker activity. Different sensitivities to Ano1 inhibitors between stomach and intestine suggest the possibility of different splice variants in these two organs or the involvement of other conductances in the generation and propagation of pacemaker activity in these tissues.

Prediction model based on decision tree analysis for laccase mediators.

A Structure Activity Relationship (SAR) study for laccase mediator systems was performed in order to correctly classify different natural phenolic mediators. Decision tree (DT) classification models with a set of five quantum-chemical calculated molecular descriptors were used. These descriptors included redox potential (ɛ°), ionization energy (E(i)), pK(a), enthalpy of formation of radical (Δ(f)H), and OH bond dissociation energy (D(O-H)). The rationale for selecting these descriptors is derived from the laccase-mediator mechanism. To validate the DT predictions, the kinetic constants of different compounds as laccase substrates, their ability for pesticide transformation as laccase-mediators, and radical stability were experimentally determined using Coriolopsis gallica laccase and the pesticide dichlorophen. The prediction capability of the DT model based on three proposed descriptors showed a complete agreement with the obtained experimental results.

Synthesis and pharmacological evaluation of bis-3-(3,4-dichlorophenyl)acrylamide derivatives as glycogen phosphorylase inhibitors.

During our research using a high-throughput screening system for discovery of a new class of human liver glycogen phosphorylase a (hLGPa) inhibitors, a series of 3-(3,4-dichlorophenyl)acrylamide derivatives were synthesized, and their inhibitory activities toward hLGPa were evaluated. Among the derivatives, (2E,2'E)-N,N'-pentane-1,5-diylbis[3-(3,4-dichlorophenyl)acrylamide] (6c) inhibited hLGPa with an IC(50) value of 0.023 microM. An X-ray crystallographic study of the enzyme-6c complex showed that the inhibitor is bound at the dimer interface site, where the 3,4-dichlorophenyl moiety interacts hydrophobically with the enzyme.

[Study of combinations of antimicrobial agents. Development of the method]. / Etude des associations d'agents antimicrobiens. Mise au point de méthode.

We developed and validated a micromethod similar to the checkerboard method in antibiotherapy to study the efficiency of antiseptic and disinfectant molecules. The determination of the FBC index (Fractional Bactericidal Concentration) on 4 reference strains (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Enterococcus hirae) for 4 associations (2-benzyl-4 chlorophenol--iodophor; cetrimide-iodophor, 2 benzyl-4 chlorophenol--formaldehyde; hexamidine--formaldehyde) revealed 2 synergistic associations and 2 antagonistic associations for one or more strains.

Adaptation of Pseudomonas aeruginosa to 2,2'-methylenebis (4-chlorophenol).

A culture of Pseudomonas aeruginosa, isolated from a cooling water system, was grown in the presence of sub-inhibitory concentrations of 2,2'-methylenebis(4-chlorophenol) (MBC). It adapted to increasing concentrations from an initial minimum inhibitory concentration of 36 micrograms ml-1 to the highest, 80 micrograms ml-1. Resistant cultures exhibited a higher survival rate when exposed to 320 micrograms ml-1 than did the original strain. Lipopolysaccharide and outer membrane protein profiles were determined by SDS PAGE. No changes were detected in lipopolysaccharide profiles. The quantity of OprP, the phosphate uptake protein in the outer membrane, decreased to a low level correlating with decreased phosphate (P(i)) uptake during growth. It is proposed that OprP is the place of entry for MBC and that the cell can adapt by decreasing the level of OprP in the outer membrane.

Effect of o-benzyl-p-chlorophenol on drug-metabolizing enzymes in rats.

o-Benzyl-p-chlorophenol (BCP) is widely used as a broad spectrum disinfectant. Treatment of male Fischer 344 rats with BCP resulted in an increase in cytochrome P-450 content and an accompanying decrease in aryl hydrocarbon hydroxylase (AHH) activity in both liver and kidney microsomes. Several other drug-metabolizing enzymes were not affected by BCP treatment. However, in kidney, BCP induced NADPH-cytochrome c reductase and uridine diphosphate glucuronyl transferase activities and caused a small increase in total cytochrome P-450 content and glutathione concentration. The cytochrome P-450 isozymes induced by BCP were fractionated by high pressure liquid chromatography (HPLC). The HPLC profile following BCP treatment most closely resembled that seen after phenobarbital. Using an immunoblotting procedure and a radioimmunoassay, it was shown that the increase in cytochrome P-450 content in the liver after BCP treatment was, in part, due to an increase in the phenobarbital-inducible isozymes, P-450b + e. In the kidney, the increase in total cytochrome P-450 content after BCP exposure was not due to an increase in P-450b + e. The decrease in AHH activity appeared to be caused by noncompetitive inhibition of constitutive AHH activity by BCP. BCP also inhibited benzphetamine demethylation, although to a lesser extent. The failure to observe an increase in benzphetamine demethylase activity in vivo, despite the induction of P-450b, was probably due to the concomitant induction and inhibition of drug-metabolizing enzymes by BCP.

In vitro inhibition of Giardia lamblia and Trichomonas vaginalis growth by bithionol, dichlorophene, and hexachlorophene.

Bithionol, dichlorophene, and hexachlorophene, which are used in treating some helminthic infections, killed trophozoites of Giardia lamblia and Trichomonas vaginalis in modified BI-S-33 and Asami media, respectively. Virtually all G. lamblia and T. vaginalis cells were killed within 24 h with a 0.42 mM concentration of these compounds, except that 0.93 mM dichlorophene was required for sterilizing T. vaginalis in the same period. In modified BI-S-33 and Asami media from which bovine and human sera were omitted, respectively, the inhibitory actions of the compounds against in vitro growth of these protozoa were significantly enhanced. Trophozoites of G. lamblia and T. vaginalis could be killed in shorter than 10 min with 0.074 mM dichlorophene and 0.0025 mM hexachlorophene, respectively, in serum-free media. G. lamblia, which was incubated in the complete medium containing dichlorophene, showed a characteristic swelling of the ventral side which led to disruption of the parasite, whereas bithionol caused a thin crack in the cytoplasm of T. vaginalis incubated in Asami medium. The crack appeared to enlarge and result in vacuolization of T. vaginalis. These observations suggest that bithionol, dichlorophene, and hexachlorophene merit further evaluation to ascertain whether they are useful for treatment of giardiasis and trichomoniasis.

Entamoeba histolytica: inhibition in vitro by bithionol of respiratory activity and growth.

Endogenous and 2-propanol-supported respiration of intact trophozoites of of Entamoeba histolytica (stain HM-1:IMSS) were inhibited by bithionol, an effective chemotherapeutic agent for some trematode and cestode infections in humans. Dichlorophene and hexachlorophene also inhibited 2-propanol-supported respiration of the parasite. In contrast, ethanol formation by E. histolytica extract in the presence of N2 was scarcely inhibited by bithionol. The compound also inhibited in vitro growth of axenic (HM-1 strain) and polyxenic (strain HJ-1:KEIO) amoebae in culture. It took less than 24 hr to kill and disrupt virtually all amoebae of either strain with 0.28 mM bithionol. Omission of bovine serum from BI-S-33 medium resulted in considerably less disruption of HM-1 strain amoebae by the compound. However, organisms that looked undisrupted were strained with trypan blue. Moreover, the number of amoebae incubated for 10 min in the serum-free BI-S-33 medium containing 0.14 mM bithionol did not increase, even after incubation for 24 hr following replacement of the experimental culture fluid with fresh complete BI-S-33 medium free of the compound. These findings suggest that, although serum appears to diminish the antiamoebic action, some halogenated bisphenols (in particular bithionol) may be useful for treatment of amoebiasis.

A comparison of the antibacterial properties of some udder creams and ointments.

The antibacterial activity of three udder creams and two ointments was assayed using three different methods. Seven different bacterial genera were used as test organisms providing a total of 17 strains. Only a single strain of corynebacterium was inhibited by all the preparations. Certain strains of staphylococci and streptococci were inhibited by four of the preparations. The gram-negative organisms showed greatest resistance to the antibacterial agents tested. The in vitro assays showed only one of the five preparations to have even slight bactericidal activity. These preparations when tested on the teats of dry and lactating cows showed similar results to the in vitro experiments.

[Synthetic antimycotica (author's transl)]. / Synthetische antimycotica

In addition to the antimycotic antibiotics described in a previous paper (Tijdschr. Diergeneesk., 101, 900-904, (1976)), the drugs used in the treatment of dermatomycosis include a fairly large number of synthtic compounds showing a wide variety of chemical structures. These are partly old and occasionally even obsolete agents (often in use as early as prior to 1945); a number of these drugs are of more recent date. They will be discussed in the present paper. So far as is known, the physicochemical and pharmacotoxicological properties, antifungal range, mechanisms of action and preparations will be reviewed.