Chaotropic and hydrophobic stress mechanisms of antifungal substances

Growth and metabolism of fungi can be curtailed by chaotropic solutes and hydrophobic substances, both of which can weaken or inhibit non-covalent interactions within and between macromolecular systems. Here we explore the potential to utilize the fungistatic and fungicidal activities of such stressors as the basis for commercial formulations. A method was developed for the quantification of chaotropicity, which can be used for chemically diverse substances, in order elucidate roles of chaotropicity and hydrophobicity in microbial ecology (both of which are sufficiently potent to limit the Earth’s microbial biosphere). A large number of naturally occurring substances act as chaotropic or hydrophobic stressors including aliphatic alcohols, salts such as MgCl2, aromatics such as phenol, and hydrocarbons such as hexane and octene. We suggest that these stress parameters provide the (hitherto unidentified) modes-of-action for some extant antifungal products. The findings are discussed in relation to the development of a new generation of antifungals.

Physicochemical Investigation of Adiponitrile-Based Electrolytes for Electrical Double Layer Capacitor

Role of capsular modified heptose in the virulence of Campylobacter jejuni

The Campylobacter jejuni capsular polysaccharide is important for virulence and often contains a modified heptose. In strain ATCC 700819 (a.k.a. NCTC 11168), the modified heptose branches off from the capsular backbone and is directly exposed to the environment. We reported previously that the enzymes encoded by wcaG, mlghB and mlghC are involved in heptose modification. Here, we show that inactivation of any of these genes leads to production of capsule lacking modified heptose and alters the transcription of other capsule modification genes differentially. Inactivation of mlghB or mlghC, but not of wcaG, decreased susceptibility to bile salts and abrogated invasion of intestinal cells. All mutants showed increased sensitivity to serum killing, especially wcaG::cat, and had defects in colonization and persistence in chicken intestine, but did not show significant differences in adhesion, phagocytosis and intracellular survival in murine macrophages. Together, our findings suggest that the capsular heptose modification pathway contributes to bacterial resistance against gastrointestinal host defenses and supports bacterial persistence via its role in serum resistance and invasion of intestinal cells. Our data further suggest a dynamic regulation of expression of this pathway in the gastrointestinal tract.

Measurement of the Electrogenicity of Bile Salt/H+ Antiport in Escherichia coli

The transmembrane proton gradient (ΔpH) is the primary source of energy exploited by secondary active substrate/H+ antiporters to drive the electroneutral transport of substrates across the Escherichia coli (E. coli) inner membrane. Such electroneutral transport results in no net movement of charges across the membrane. The charge on the transported substrate and the stoichiometry of the exchange reaction, however, can result in an electrogenic reaction which is driven by both the ΔpH and the electrical (∆Ψ) components of the proton electrochemical gradient, resulting in a net movement of electrical charges across the membrane. We have shown that the major facilitator superfamily transporter MdtM - a multidrug efflux protein from E. coli that functions physiologically in protection of bacterial cells against bile salts - imparts bile salt resistance to the bacterial cell by coupling the exchange of external protons (H+) to the efflux of bile salts from the cell interior via an electrogenic antiport reaction (Paul et al., 2014). This protocol describes, using fluorometry, how to detect electrogenic antiport activity of MdtM in inverted membrane vesicles of an antiporter-deficient strain of E. coli TO114 cells by measuring transmembrane ∆Ψ. The method exploits changes that occur in the intensity of the fluorescence signal (quenching and dequenching) of the probe Oxonol V in response to changes in membrane potential due to the MdtM-catalysed sodium cholate/H+ exchange reaction. The protocol can be adapted to detect activity of any secondary active antiporter that couples the electrogenic translocation of H+ across a biological membrane to that of its counter-substrate, and may be used to unmask otherwise camouflaged transport activities and physiological roles.

Measurement of proton-driven antiport in Escherichia coli

Secondary active transport of substrates across the inner membrane is vital to the bacterial cell. Of the secondary active transporter families, the ubiquitous major facilitator superfamily (MFS) is the largest and most functionally diverse (Reddy et al., 2012). Recently, it was reported that the MFS multidrug efflux protein MdtM from Escherichia coli (E. coli) functions physiologically in protection of bacterial cells against bile salts (Paul et al., 2014). The MdtM transporter imparts bile salt resistance to the bacterial cell by coupling the exchange of external protons (H+) to the efflux of bile salts from the cell interior via an antiport reaction. This protocol describes, using fluorometry, how to detect the bile salt/H+ antiport activity of MdtM in inverted membrane vesicles of an antiporter-deficient strain of E. coli TO114 cells by measuring transmembrane ∆pH. This method exploits the changes that occur in the intensity of the fluorescence signal (quenching and dequenching) of the pH-sensitive dye acridine orange in response to changes in [H+] in the vesicular lumen. Due to low levels of endogenous transporter expression that would normally make the contribution of individual transporters such as MdtM to proton-driven antiport difficult to detect, the method typically necessitates that the transporter of interest be overexpressed from a multicopy plasmid. Although the first section of the protocol described here is very specific to the overexpression of MdtM from the pBAD/Myc-His A expression vector, the protocol describing the subsequent measurement of bile salt efflux by MdtM can be readily adapted for measurement of antiport of other substrates by any other antiporter that exchanges protons for countersubstrate.

EFFICIENT SYNTHESIS OF 3-MONO AND DISUBSTITUTED LACTAMS USING MEERWEIN-ESCHENMOSER [3,3] SIGMATROPIC REARRANGEMENTS

3-Allyl substituted five six and seven membered ring lactams, are readily available in good yields and reasonable selectivity by a formal Meerwein Eschenmoser [3,3] rearrangement, using readily available methoxymethyleniminium salts and lithium alkoxides derived from allyl alcohols.

THE SYNTHESIS OF BRIDGED-RING CARBOCYCLES AND HETEROCYCLES VIA PALLADIUM CATALYZED REGIOSPECIFIC CYCLIZATION REACTIONS

A catalyst system comprising 10 mol % (Pd(OAc) and 20 mol % PPh3 effects the cyclisation of aryl halides onto proximate alkenes via 5-, 6-, and 7-exo-trig, and 7-endo-trig processes giving a variety of bridged-ring carbo- and hetero-cycles in excellent yield. Double bond isomerisation in the product is rarely encountered and may be suppressed by the addition of Tl(1) salts. One example of diastereospecific bis-cyclisation is given and the crystal structure of 1-aza-2-sulphonyl-3,4-benzobicyclo[3.2.1]nona-6-ene is reported.

EFFICIENT SYNTHESIS OF 3-SUBSTITUTED LACTAMS USING MEERWEIN ESCHENMOSER CLAISEN [3,3] SIGMATROPIC REARRANGEMENTS

3-Allyl substituted five, six, and seven membered ring lactams are readily available in good yields and reasonable selectivity by a formal Meerwein Eschenmoser Claisen [3,3] rearrangement, using the readily available N,N-dialkylalkoxymethylene iminium salts and lithium alkoxides derived from allyl alcohols.

STEREOSELECTIVE ROUTE TO HIGHLY FUNCTIONALIZED AMINO-ALCOHOLS

Acyclic amino alcohols can be obtained in high stereoisomeric purity by Stevens [2,3] rearrangement of tetrahydropyridine salts followed by reduction of the ketones and ring opening of the disubstituted pyrrolidine.

3-Methylpiperidinium ionic liquids

A wide range of room temperature ionic liquids based on the 3-methylpiperdinium cation core were produced from 3-methylpiperidine, which is a derivative of DYTEK® A amine. First, reaction with 1-bromoalkanes or 1-bromoalkoxyalkanes generated the corresponding tertiary amines (Rmβpip, R = alkyl or alkoxyalkyl); further quaternisation reactions with the appropriate methylating agents yielded the quaternary [Rmmβpip]X salts (X(-) = I(-), [CF3CO2](-) or [OTf](-); Tf = -SO2CF3), and [Rmmβpip][NTf2] were prepared by anion metathesis from the corresponding iodides. All [NTf2](-) salts are liquids at room temperature. [Rmmβpip]X (X(-) = I(-), [CF3CO2](-) or [OTf](-)) are low-melting solids when R = alkyl, but room temperature liquids upon introduction of ether functionalities on R. Neither of the 3-methylpiperdinium ionic liquids showed any signs of crystallisation, even well below 0 °C. Some related non-C-substituted piperidinium and pyrrolidinium analogues were prepared and studied for comparison. Crystal structures of 1-hexyl-1,3-dimethylpiperidinium tetraphenylborate, 1-butyl-3-methylpiperidinium bromide, 1-(2-methoxyethyl)-1-methylpiperidinium chloride and 1-(2-methoxyethyl)-1-methylpyrrolidinium bromide are reported. Extensive structural and physical data are collected and compared to literature data, with special emphasis on the systematic study of the cation ring size and/or asymmetry effects on density, viscosity and ionic conductivity, allowing general trends to be outlined. Cyclic voltammetry shows that 3-methylpiperidinium ionic liquids, similarly to azepanium, piperidinium or pyrrolidinium counterparts, are extremely electrochemically stable; the portfolio of useful alternatives for safe and high-performing electrolytes is thus greatly extended.

Pro-fragrant ionic liquids with stable hemiacetal motifs: water-triggered release of fragrances

Stable liquid and solid salts in the form of elusive hemiacetals, appended with fragrant alcohols, have been synthesised as pro-fragrances, and the controlled release of these fragrances, triggered by water, is demonstrated

Nucleoside Azide-Alkyne Cycloaddition Reactions Under Solvothermal Conditions or Using Copper Vials in a Ball Mill

Novel nucleoside analogues containing photoswitchable moieties were prepared using 'click' cycloaddition reactions between 5 '-azido-5 '-deoxythymidine and mono- or bis-N-propargylamide-substituted azobenzenes. In solution, high to quantitative yields were achieved using 5mol% Cu(I) in the presence of a stabilizing ligand. 'Click' reactions using the monopropargylamides were also effected in the absence of added cuprous salts by the application of liquid assisted grinding (LAG) in metallic copper reaction vials. Specifically, high speed vibration ball milling (HSVBM) using a 3/32('') (2.38mm) diameter copper ball (62mg) at 60Hz overnight in the presence of ethyl acetate lead to complete consumption of the 5 '-azido nucleoside with clean conversion to the corresponding 1,3-triazole.

Simple preparation of positively charged silver nanoparticles for detection of anions by surface-enhanced Raman spectroscopy

Modification of citrate and hydroxylamine reduced Ag colloids with thiocholine bromide, a thiol functionalized quaternary ammonium salt, creates particles where the zeta potential is switched from the normal values of ca. -50 mV to ca. + 50 mV. These colloids are stable but can be aggregated with metal salts in much the same way as the parent colloids. They are excellent SERS substrates for detection of anionic targets since their positive zeta potentials promote adsorption of negatively charged ions. This is important because the vast majority of published SERS studies involve cationic or neutral targets. Moreover, the fact that the modifier is a quaternary ammonium ion means that the negative surface charge is maintained even at alkaline pH. The modified colloids can be used to detect compounds which cannot be detected using conventional negatively-charged citrate or hydroxylamine reduced metal nanoparticles, for example the detection limit was 5.0 x 10(-5) M for perchlorate and

Olefin / Paraffin separation using Task-Specific Materials based on Ionic Liquids: Advances in Gas Processing

In this work, 1-hexene was extracted from its mixtures with n-hexane in varying ratios using a task specific ionic liquid. Herein, the ionic liquid (IL) 1-butyl-3-methylimidazolium nitrate, [BMIM][NO3], was used and examined with and without the addition of a metal salt. The impact of water on both selectivity and distribution coefficient was also tested. Four potential metal salts were investigated, the results of which demonstrate that the dissolution of transition-metal salts in the IL improves the separation of 1-hexene from n-hexane through metal-olefin complexation. Additionally, the presence of water in IL solutions containing metal salt enhances this selectivity. Finally, UNIFAC was used to correlate the experimental LLE data with good accuracy.

Process for the preparation of heterocyclyl chalcogenones via reaction of elemental sulfur, selenium, or tellurium with heterocyclic cationic species.

Heterocyclic chalcogenones were prepd. by reaction of S, Se, or Te with ionic liqs. or salts [I; Ra = (substituted) alkyl, cycloalkyl, aryl, aralkyl, alkylaryl; Q = (unsatd.) (substituted) linker to form a ring of 5-10 members; X- = anion selected from conjugate bases of HX having a pKa value of >2.5]. Thus, 1-butyl-3-methylimidazolium acetate was heated with stoichiometric S at 75° for 48 h to give 1-butyl-3-methylimidazole-2-thione. [on SciFinder(R)]