This effort will serve to develop predictive biodegradation models based on time-dependent quantitative biochemical and spectroscopic data for separating polymer coating biodegradation from bulk autocatalytic hydrolysis with single strains and defined co-cultures using fungi and bacteria isolated from Military cargo aircraft.. These data will serve as predictors for mitigating the biodegradation of polyurethane coating systems in a variety of exposure environments with the expectation that research activities will translate to improved coating design and technologies.

The approach will group and classify the complex bioenergetics and chemical interactions between fungi and bacteria separately and as consortia during the biodegradation of defined synthetic polyester and polyether polyurethane formulations. The researchers will use active polyester degrading mold and mold-like fungi (Aureobasidium pullulans and Cladosporium brunhei) and non-motile polyester degrading yeast strains (Papiliotrema laurentii or Naganisha alibida) with either bacterial (Bacillus megaterium or Rhodococcus sp.) or fungal (Fusarium oxysporum or Candida orthropsilosis) species with a published ability to oxidize alkanes. All microorganisms were isolated from the same location from inside each aircraft. They will separate direct polyurethane coating biodegradation from chemical autocatalytic degradation induced by biology by synthesizing defined chain-extended polyester and polyether polyurethanes with pH chromo-responsive molecules interspersed or cross-linked throughout the polymer coating.

The temperature (25-37°C) and humidity (70-95+% relative humidity) of these experiments will be controlled to assess the role of water adsorption, absorption, ingress, and/or egress in the active degradation of these coatings as well. A defined block co-polymer formulation will be used for all experiments to compare how each additive or microbial challenge affects polymer degradation, biofragmentation, depolymerization, and bioassimilation. In addition to these defined synthetic coatings, they will include polyether polyurethane coatings since these coating specifications are of interest to the U.S. Department of Defense (DoD) and are resistant to biodegradation based on experiments with individual fungal strains. They will perform surface roughness, polymer volume measurements, and X-ray fluorescence (XRF) measurements with pristine coatings and coatings with defined defects and roughness. They will also assess the impacts of polymer interface environments on the cells under degrading and non-degrading conditions, including those that promote continued cellular respiration versus quiescence, cell death, and new cellular growth from dying cells. They will develop quantitative polymerase chain reaction (qPCR) primer sets and methods based on the conserved regions of each organism’s genome to measure the change in microbial biofilm ecology as the degradation of a coating initiates and proceeds with reverse transcription (RT)-qPCR SYBR Green assays.

CO2 evolution and whole cell luminescent and fluorescent nicotinamide adenine dinucleotide + hydrogen (NADH)/NAD+, adenosine triphosphate (ATP) cycling, mitochondrial staining will be used to compare bioenergetic states during the degradation of the polymer in addition to fungal and bacterial viability stains in 96-well plate and slide formats. They will use electrochemical impedance spectroscopy (EIS), confocal fluorescence, Raman and Infrared (IR) microscopies and atomic force microscopy (AFM) and AFM-IR techniques to quantify common biochemical kinetic and thermodynamic variables and coating failure mechanism during the biodegradation process.

In order to create novel sustainable and biodegradation resistant coating formulations for DoD weapon systems, the complexity of the biodegradation process has to be separated into general categories and features to identify where active biodegradation stops and purely autocatalytic hydrolysis begins. Their specific focus on the essential role of water and secreted compounds in the movement and activity of these organisms will identify how degradation is associated to kinetic or thermodynamic mechanisms which diminish cellular propagation or potentially even stop it within coatings. Their data will result in new approaches to coating design for DoD weapon systems by making fouling and degradation, in general, more difficult since the coatings would be designed to use an organism’s natural biodegradation process (coupled with limiting the movement of water) against them. This will potentially eliminate the use of toxic additives and delay the spread of the organism so that the surfaces will be easier to remediate which will affect the overall environmental impact of the coating.