Topographical Imaging as a Means of Monitoring Biodegradation of Poly(hydroxyalkanoate) Films

Authors: Ashby, Richard - Rick; Cooke, Peter; Solaiman, Daniel

Submitted to: Polymers and the Environment
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/21/2007
Publication Date: 7/30/2007
Citation: Ashby, R.D., Cooke, P.H., Solaiman, D. 2007. Topographical Imaging as a Means of Monitoring Biodegradation of Poly(hydroxyalkanoate) Films. Polymers and the Environment. 15(3):179-187.

Interpretive Summary: Poly(hydroxyalkanoates) (PHAs) are a class of "eco-friendly" bioplastics that are gaining more industrial acceptance due to their wide range of properties (rigid – rubber-like) and their ability to be naturally degraded to carbon dioxide and water by indigenous microorganisms. While much of the recent research on these polymers has focused on low-cost synthesis, property enhancement, and new applications, equally important is the degradation cycle which must be understood in order to perform accurate life-cycle assessments which are vital to predict PHAs' true environmental impact. In this study we used two different bacterial strains with the capability of producing the necessary enzymes to break down different types of PHA polymers and applied these bacteria to thin PHA polymer films. By relying on the relatively narrow specificities of the degrading enzymes, we developed a protocol to visually monitor the surface of each film and correlate the roughness of the film surface to those of known controls. Increases in the irregularity of the film surface demonstrate the ability to, and the pace at which degradation occurs. This newly developed method provides a simple but effective means to quantitate degradation and ultimately will aid in the understanding of carbon cycling in PHA degradation.

Technical Abstract: Poly(hydroxyalkanoates) (PHAs) are a class of bacterially-derived polymers that are naturally biodegradable through the action of extracellular depolymerase enzymes secreted by a number of different bacteria and fungi. In this paper we describe the development of topographical imaging protocols (by both scanning electron microscopy; SEM, and confocal microscopy; CM) as a means of monitoring the degradation of solution cast films of poly(3-hydroxybutanoate-co-3-hydroxyhexanoate) (P3HB/3HHx; NODAX, Procter & Gamble) and medium-chain-length (mcl-) PHA. Pseudomonas lemoignei and Comamonas P37C were used as sources for PHA depolymerase enzymes as these bacteria are known to degrade at least one of the polymers in question. Scanning electron microscopy (SEM) revealed the bacterial colonization of the film surfaces but little on the quantitative determination of degradation. In contrast, CM permitted the comparative measurement of the roughness of the film surface. By dividing the total surface area of the film (A') by the total area of the scan (A) it was possible to monitor degradation by observing differences in the topography of the film surface. Prior to inoculation, P3HB/3HHx films had an A'/A ratio of 1.06. A 24-hour incubation with P. lemoignei increased the A'/A ratio to 1.47 while a 48- and 120-hour incubation with Comamonas resulted in A'/A ratios of 1.16 and 1.33, respectively. These increases in the A'/A ratios over time demonstrated an increase in the irregularity of the film surface, indicative of PHA polymer breakdown.