Background/Objectives:The use of electric fields in combination with small doses of antibiotics for enhanced treatment of biofilms is termed the ‘bioelectric effect’ (BE). Different mechanisms of action for the AC and DC fields have been reported in the literature over the last two decades. In this work, we conduct the first study on the correlation between the electrical energy and the treatment efficacy of the bioelectric effect on Escherichia coli K-12 W3110 biofilms.Methods:A thorough study was performed through the application of alternating (AC), direct (DC) and superimposed (SP) potentials of different amplitudes on mature E. coli biofilms. The electric fields were applied in combination with the antibiotic gentamicin (10 μg/ml) over a course of 24 h, after the biofilms had matured for 24 h. The biofilms were analysed using the crystal violet assay, the colony-forming unit method and fluorescence microscopy.Results:Results show that there is no statistical difference in treatment efficacy between the DC-, AC- and SP-based BE treatment of equivalent energies (analysis of variance (ANOVA) P>0.05) for voltages <1 V. We also demonstrate that the efficacy of the BE treatment as measured by the crystal violet staining method and colony-forming unit assay is proportional to the electrical energy applied (ANOVA P<0.05). We further verify that the treatment efficacy varies linearly with the energy of the BE treatment (r2 =0.984). Our results thus suggest that the energy of the electrical signal is the primary factor in determining the efficacy of the BE treatment, at potentials less than the media electrolysis voltage.Conclusions:Our results demonstrate that the energy of the electrical signal, and not the type of electrical signal (AC or DC or SP), is the key to determine the efficacy of the BE treatment. We anticipate that this observation will pave the way for further understanding of the mechanism of action of the BE treatment method and may open new doors to the use of electric fields in the treatment of bacterial biofilms.

S-(+)-Methamphetamine is frequently found as the only isomer in urine specimens from methamphetamine abuseres. Enantiomerically pure S-(+)-methamphetamine can be synthesized from ephedrine or pseudoephedrine via chloroephedrine intermediates. These intermediates are unstable and capable of cyclizing to form cis- and trans-1,2-dimethyl-3-phenyl aziridine. Studies were done to determine if these intermediates could be detected when using a common gas chromatographic-mass spectrometric analytical method (derivatization with heptafluorobutyric anhydride, HFBA) for toxicological screening of methamphetamine. Analysis of (+)- or (-)-chloroephedrine after extraction into hexane and derivatization with HFBA indicated that both pseudoephedrine and ephedrine were the major compounds detected. Direct derivatization of a hexane solution of cis-1,2-dimethyl-3-phenyl aziridine yielded only the derivatives of ephedrine and pseudoephedrine, indicating that the aziridine intermediate is responsible for the formation of the ephedrine or pseudoephedrine. These studies indicate that the aziridine intermediates would not be detected in methamphetamine samples following HFBA derivatization.

The Polo-like kinase (Plk) family comprises four cell cycle serine/threonine kinases, Plk1-4. Among these, Plk1 has been most thoroughly characterized; it contains a conserved kinase domain and a C-terminal docking site for S/T-phosphorylated proteins (polo-box domain, PBD). Polo-like kinases are deregulated in oncogenesis and therefore constitute a therapeutic target for cancer. A high throughput screening campaign was carried out by the Pittsburgh Molecular Library Screening Center (PMLSC), using a fluorescence polarization assay with recombinant Plk1-PBD to monitor the inhibition of binding of an optimal phosphopeptide substrate motif with recombinant Plk1-PBD. Screening of 97,090 small molecule library samples provided by the NIH Small Molecule Repository distributed by DPI Galapagos led to 11 confirmed hits. The Pittsburgh MLSCN Chemistry Core selected one of the structurally most tractable hits, SID 861574, for chemical hit-to-probe development. A broad chemistry program was initiated that developed new strategies for 6-amino- and 6-hydroxy uracil synthesis as well as acylanilides, and generated a total of 70 analogs. Out of 46 analogues tested, none, nor the resynthesized hit, showed affinity to Plk1-PBD in the follow up assays. In contrast, re-assays of the original screening materials displayed activities similar to the original HTS assay. We ultimately concluded that an impurity in the commercial material led to the positive screening artifact. This case study highlights our development of a synthesis of 6-position functionalized uracil analogs, but also illustrates the importance of careful quality and compound stability monitoring of screening collections.

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