ELECTROCHEMICAL APPROACH TO ERADICATION OF BIOFILMS AND RENEWABLE ENERGY PRODUCTION

Md Monzurul Islam Anoy

doi:10.7273/000006910

This dissertation presents the application of the multi-disciplinary field of electrochemistry in wound healing and renewable energy production. This electrochemical research applied to 1) control biofilms and promote wound healing; especially eradication of wound-related biofilms such as Acinetobacter baumannii, & Methicillin-resistant Staphylococcus aureus (MRSA), and 2) improve the anodic current density in a well-mixed single-chamber bioelectrochemical system (BES). For the health application, the counter electrode reaction of a hydrogen peroxide (H2O2) generating electrochemical bandage (e-bandage) was characterized and its biocidal activity against A. baumannii BAA-1605 biofilm was investigated. e-Bandage is a three-electrode system with a working electrode placed on top of the biofilm, a reference electrode placed between the working and the counter electrode, and a counter electrode placed on the top. This setup allows continuous production of H2O2 in the vicinity of a biofilm when the working electrode was polarized to –0.6 VAg/AgCl. It was found that the counter electrode potential varied between 1.2 and 1.5 VAg/AgCl; interestingly, ~125 µM hypochlorous acid (HOCl) was generated in the e-bandage within 24 hours. In a biological test, the counter and working electrode of H2O2-generating e-bandage were separated and placed onto two laboratory-grown biofilms. The counter electrode of an H2O2 e-bandage showed ~6 log CFU/cm2 reduction (vs. control) while the working electrode showed ~1 log CFU/cm2 reduction (vs. control). Furthermore, HOCl was not produced on the counter electrode, and cell reduction of A. baumannii BAA-1605 was 1.08 ± 0.38 log CFU/cm2 after 24 h treatment, whereas when HOCl was produced, cell reduction was 3.87 ± 1.44 log CFU/cm2. Finally, it was found that HOCl inhibited the catalase activity, abrogating H2O2 decomposition. Additionally, scanning electron microscopy (SEM) was done on the hydrogel to investigate the morphology of the hydrogel. SEM images showed the porous structure of hydrogel which indicated the key mechanism of the H2O2-generating e-bandage which also generated HOCl at the counter electrode. Later, an intermittent treatment approach using electrochemically generated H2O2 and HOCl by e-bandage was evaluated against MRSA biofilms. By changing the working electrode potential, the e-bandage generated either HOCl (1.5 VAg/AgCl) or H2O2 (-0.6 VAg/AgCl). The degree of biocidal activity of intermittent treatment with HOCl and H2O2 correlated with HOCl treatment time; HOCl treatment durations of 0, 1.5, 3, 4.5, and 6 hours (with the rest of the 6-hour total treatment time devoted to H2O2 generation) resulted in mean biofilm reductions of 1.36±0.2, 2.22±0.16, 3.46±0.38, 4.63±0.74 and 7.66±0.5 log CFU/cm2, respectively vs. non-polarized controls, respectively. It was found that the immediate generation of H2O2 after HOCl was detrimental to viable cells' reduction in biofilm. For example, 3-hour HOCl treatment followed by 3-hours H2O2 resulted in a 1.90±0.84 log CFU/cm2 lower mean biofilm reduction than 3-hour HOCl treatment followed by 3-hour non-polarization. Moreover, microelectrode measurement on intermittent e-bandage (which generated HOCl first and followed by H2O2 generation) was conducted to investigate the HOCl concentration profile in e-bandage. Microelectrode measurement showed that the immediate generation of H2O2 after HOCl eradicated the generated HOCl due to the reaction between HOCl and H2O2. This result confirms the reduced biocidal activity of intermittent treatment using H2O2 immediately after HOCl by an e-bandage. Thus, later, incorporation of a non-polarized treatment period was included which allowed the electrochemically generated HOCl to be dissipated. HOCl generated over 3 hours exhibited biocidal activity for at least 7.5 hours after e-bandage operation ceased; 3 hours of HOCl generation followed by 7.5 hours of non-polarization resulted in a biofilm cell reduction of 7.92±0.12 log CFU/cm2 vs. non-polarized controls. Finally, intermittent treatment with HOCl (i.e., interspersed with periods of e-bandage non-polarization) for various intervals showed similar effects (approximately 6 log CFU/cm2 reduction vs. non-polarized control) to continuous treatment with HOCl for 3-hours, followed by 3-hours of non-polarization. The environmental application part of the research focused on the improvement of anodic current density in a well-mixed single-chamber BES. A directional electrode separator was developed for a 3D printed well-mixed single-chamber BES to minimize hydrogen reoxidation. Hydrogen reoxidation occurs in a single-chamber BES when hydrogen generated at the cathode migrates to the anode and the anodic biofilm uses the hydrogen as an electron donor. In a controlled abiotic environment, cyclic voltammetry and electrochemical impedance spectroscopy were performed to evaluate the optimum placement of the electrodes (anode and cathode) in the 3D printed well-mixed single-chamber BES. It was found that attaching the electrodes to the directional electrode separator was the optimum configuration based on high oxidation-reduction peak currents and decreased internal system resistance. Furthermore, electrochemical impedance spectra showed that a 3D printed directional electrode separator has seven times higher proton conductivity than a conventional proton exchange membrane. A directional electrode separator effectively reduced hydrogen migration (21 folds vs. control) to the anode which indicates it can reduce the reoxidation of hydrogen. Finally, a 3D printed well-mixed single-chamber BES equipped with a directional electrode separator inoculated with homogenized anaerobic granules and operated with fermented corn stover. This system showed 3.3 times higher current densities (~53 A/m2) than a conventional single-chamber BES without a separator (~16 A/m2).

ELECTROCHEMICAL APPROACH TO ERADICATION OF BIOFILMS AND RENEWABLE ENERGY PRODUCTION

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