Alaa-Eldin F. Nassar

Direct electron injection from electrodes to cytochromeP450cam in biomembrane-like films

Direct, reversible, electron transfer has been demonstrated between electrodes and native cytochrome P450 cam (cyt P450 cam ) in lipid films. Electron injection directly from electrodes into the haem Fe III of cyt P450 cam has been confirmed by the influence of the known reaction of cyt P450 cam Fe II with CO on the voltammetric midpoint potential. Analysis of square wave voltammograms suggested a distribution of enzyme formal potentials in the films. With oxygen present, more than one electron was injected into the enzyme in films, mimicking in vivo electron acceptance by cyt P450Fe III and cyt P450Fe II –O 2 during catalytic oxidations. Cyt P450 cam films also catalysed electrochemically driven reduction of trichloroacetic acid in anaerobic solutions. These stable enzyme–lipid films may find applications in fundamental biochemical and toxicity studies, biocatalysis and biosensors.

ELECTROCHEMICAL PROPERTIES OF MYOGLOBIN EMBEDDED IN LANGMUIR-BLODGETT AND CAST FILMS OF SYNTHETIC LIPIDS

Electrochemical parameters, formal potentials (E0′), electron-transfer rate constants (k0′), and diffusion coefficients (Dct) have been determined for myoglobin (Mb) embedded in the Langmuir–Blodgett (LB) and cast films of five synthetic lipids including two types of polymeric lipids bearing ammonium or poly(ethylene glycol) as a head group. The heterogeneous electron-transfer rate constants of MbFeIII/MbFeII in these films were ca. 102–103-fold larger than those for Mb in solution at an indium tin oxide electrode. Myoglobin-lipid LB films on basal-plane pyrolytic graphite (PG) electrodes gave 3–10-fold larger k0′ values compared with cast films of the same lipid. The Mb–lipid films showed gel to liquid-crystal phase transitions consistent with bilayer structures. The peak current of the square wave voltammograms for the cast Mb–lipid films showed breaks in the phase-transition temperature regions of the corresponding lipid films. Dependence of k0′ on lipid structure was not significant. A variety of synthetic lipids provided suitable microenvironments for the enhanced electron transfer of Mb. Immobilized Mb in the lipid films was stable for more than a month. The specific orientation of Mb in all Mb–lipid films was shown by linear dichroism. The orientation does not depend on the type of the lipids.

Salt and pH effects on electrochemistry of myoglobin in thick films of a bilayer-forming surfactant

Salt concentration and pH of external solutions were shown to control the electrochemistry of the heme protein myoglobin (MbFe(III)-H2O) in stable, ordered films of didodecyldimethylammonium bromide (DDAB). Protonation of aquometmyoglobin (MbFe(III)-H2O) in these films precedes electron transfer from electrodes, causing formal potentials to shift negative as pH increases from 5 to 8. At pH > 8, MbFe(III)-H2O dissociates to MbFe(III)-OH, which is reduced directly at the electrode at higher rates than MbFe(III)-H2O. Correlations of voltammetric data with FT-IR spectra suggested that at pH < 4.6, an unfolded form of Mb resides in the films and is reduced directly. The concentration of salt in solution influences electrochemical properties of Mb-DDAB films by its influence on Mb conformation and by effects on interfacial Donnan potentials. NMR indicated strong binding of anions to Mb within DDAB films. Bound anions may neutralize positive charge on Mbs surface so that it can reside in a partly hydrophobic environment, as postulated on the basis of previous ESR and linear dichroism studies.

Determination of O-Isopropyl Methylphosphonic Acid in Living Microorganism-Agar Matrixes using ION Chromatography/Conductivity Detection

ABSTRACT A direct anion exchange ion chromatography (IC)-based method for the detection of the chemical warfare (CW) agent degradation product, O-isopropyl methylphosphonic acid (IMPA) in agar medium has been developed. This is the first report of the development and validation of an IC-based method for the analysis of IMPA in a microorganism and agar matrix. In these experiments, IMPA served as a surrogate for the CW agent GB (Sarin) in a development program for living micro-organism-based CW agent destruction systems. Petri dishes containing the medium with 10, 100 and 1000 μg/mL IMPA were used. Samples were prepared by dilution of the agar medium with deionized water followed by sonication and then filtration before the analysis by IC/conductivity detection. Using this method, the amount of IMPA in the Petri dishes was measured. Excellent separation efficiency and freedom from interference due to common anions such as chloride and sulfate was obtained in the developed method. Retention time precision w...