Charles E. Turick

Gamma radiation interacts with melanin to alter its oxidation-reduction potential and results in electric current production.

The presence of melanin pigments in organisms is implicated in radioprotection and in some cases, enhanced growth in the presence of high levels of ionizing radiation. An understanding of this phenomenon will be useful in the design of radioprotective materials. However, the protective mechanism of microbial melanin in ionizing radiation fields has not yet been elucidated. Here we demonstrate through the electrochemical techniques of chronoamperometry, chronopotentiometry and cyclic voltammetry that microbial melanin is continuously oxidized in the presence of gamma radiation. Our findings establish that ionizing radiation interacts with melanin to alter its oxidation-reduction potential. Sustained oxidation resulted in electric current production and was most pronounced in the presence of a reductant, which extended the redox cycling capacity of melanin. This work is the first to establish that gamma radiation alters the oxidation-reduction behavior of melanin, resulting in electric current production. The significance of the work is that it provides the first step in understanding the initial interactions between melanin and ionizing radiation taking place and offers some insight for production of biomimetic radioprotective materials.

The role of 4‐hydroxyphenylpyruvate dioxygenase in enhancement of solid‐phase electron transfer by Shewanella oneidensis MR‐1

We hypothesized that Shewanella oneidensis MR-1, a model dissimilatory metal-reducing bacterium, could utilize environmentally relevant concentrations of tyrosine to produce pyomelanin for enhanced Fe(III) oxide reduction. Because homogentisate is an intermediate of the tyrosine degradation pathway, and a precursor of a redox-cycling metabolite, pyomelanin, we evaluated the process of homogentisate production by S. oneidensis MR-1, in order to identify the key steps involved in pyomelanin production. We determined that two enzymes involved in this pathway, 4-hydroxyphenylpyruvate dioxygenase and homogentisate 1,2-dioxygenase are responsible for homogentisate production and oxidation, respectively. We used genetic analysis and physiological characterization of MR-1 strains either deficient in or displaying substantially increased pyomelanin production. The relative significance imparted by pyomelanin on solid-phase electron transfer was also addressed using electrochemical techniques, which allowed us to extend the genetic and physiological findings to biogeochemical cycling of metals. Based on our findings, environmental production of pyomelanin from available organic precursors could contribute to the survival of S. oneidensis MR-1 when dissolved oxygen concentrations become low, by providing an increased capacity for solid-phase metal reduction. This study demonstrates the role of organic precursors and their concentrations in pyomelanin production, solid phase metal reduction and biogeochemical cycling of iron.

IN SITU URANIUM STABILIZATION BY MICROBIAL METABOLITES

Microbial melanin production by autochthonous bacteria was explored in this study as a means to increase U immobilization in U contaminated soil. This article demonstrates the application of bacterial physiology and soil ecology for enhanced U immobilization in order to develop an in situ, U bio-immobilization technology. We have demonstrated microbial production of a metal chelating biopolymer, pyomelanin, in U contaminated soil from the Tims Branch area of the Department of Energy (DOE), Savannah River Site (SRS), South Carolina, as a result of tyrosine amendments. Bacterial densities of pyomelanin producers were >10(6) cells per g wet soil. Pyomelanin demonstrated U complexing and mineral binding capacities at pH 4 and 7. In laboratory studies, in the presence of goethite or illite, pyomelanin enhanced U sequestration by these minerals. Tyrosine amended soils in a field test demonstrated increased U sequestration capacity following pyomelanin production up to 13 months after tyrosine treatments.

Review of concrete biodeterioration in relation to nuclear waste

Storage of radioactive waste in concrete structures is a means of containing wastes and related radionuclides generated from nuclear operations in many countries. Previous efforts related to microbial impacts on concrete structures that are used to contain radioactive waste showed that microbial activity can play a significant role in the process of concrete degradation and ultimately structural deterioration. This literature review examines the research in this field and is focused on specific parameters that are applicable to modeling and prediction of the fate of concrete structures used to store or dispose of radioactive waste. Rates of concrete biodegradation vary with the environmental conditions, illustrating a need to understand the bioavailability of key compounds involved in microbial activity. Specific parameters require pH and osmotic pressure to be within a certain range to allow for microbial growth as well as the availability and abundance of energy sources such as components involved in sulfur, iron and nitrogen oxidation. Carbon flow and availability are also factors to consider in predicting concrete biodegradation. The microbial contribution to degradation of the concrete structures containing radioactive waste is a constant possibility. The rate and degree of concrete biodegradation is dependent on numerous physical, chemical and biological parameters. Parameters to focus on for modeling activities and possible options for mitigation that would minimize concrete biodegradation are discussed and include key conditions that drive microbial activity on concrete surfaces.

Properties and Function of Pyomelanin

Melanin pigments are the most common pigments produced in nature and these complex biopolymers are found in species of all biological kingdoms. There are several categories of melanins which include eumelanins, pheomelanins and allomelanins. Eumelanins and pheomelanins are produced from oxidation of tyrosine or phenylalanine to odihydroxyphenylalanine (DOPA) and dopaquinone. Pheomelanin results from cysteinylation of DOPA. Allomelanins include a heterogeneous group of polymers that include pyomelanin. Melanin biochemistry and synthesis has been reviewed previously (Plonka and Grabacka 2006). This chapter will focus on the properties and function of pyomelanin and their potential utility in biotechnology. Pyomelanin originates from the catabolism of tyrosine or phenylalanine (Lehninger, 1975) (Fig. 1). Complete breakdown of tyrosine to acetoacetate and fumarate requires the enzymes 4-hydroxyphenylpyruvic acid dioxygenase (4-HPPD) and homogentisic acid oxidase (HGA-oxidase). In the absence of HGA-oxidase, or if homogentisic acid (HGA) production exceeds that of HGA-oxidase activity, HGA is over-produced and excreted from the cell (Yabuuchi and Ohyama 1972; Ruzafa et al. 1994; Katob et al. 1995). Autooxidation and selfpolymerization of HGA then results in pyomelanin. In addition, deletion of the gene that encodes for HGA-oxidase results in hyper production of pyomelanin while deletion of the gene that encodes for 4HPPD results in the inability to produce pyomelanin (Coon et al. 1994; Ruzafa et al. 1995). In humans with loss-of-function mutations in HGA-oxidase, pyomelanin (also known as alkapton or ochronotic pigment) forms in the urine due to the spontaneous auto oxidation of excess HGA (Beltran-Valero de Bernabe, et al. 1999). This condition is known as alkaptonuria in humans and can result in arthritis in adults. Pyomelanin production in microorganisms often is associated with numerous survival advantages and was first characterized in bacteria among numerous species of the genus Pseudomonas (Yabuuchi & Ohyama 1972). Since then several fungi and a number of bacteria, especially in the ┛ Proteobacteria have been shown to produce pyomelanin.

Life Span of Biopolymer Sequestering Agents for Contaminant Removal and Erosion Resistance

The objective of this paper is to report the development and life span of cross-linked biopolymers that remove contaminants, resist biodegradation over long periods of time, and resist erosion in dynamic aquatic environments. Biopolymers are polymeric compounds produced by living organisms (e.g., microorganisms, plants, crustaceans). They have repeated sequences that vary broadly in chemical composition including a variety of repeating functional groups (such as carboxyl, hydroxyl, amino, etc.). This makes them reactive and subject to cross-linking. Therefore, biopolymers, a great molecular weight compounds with repeated sequences, may have high opportunity for chemical interaction with other compounds. Depending on their functional groups, biopolymers can bind metals, organic contaminants, or soil particles and form interpenetrating cross-linking networks with other polymers. The ability of biopolymers (cross-linked or not) to bind a large variety of metals is supported by many studies (Chen et al., 1993; Etemadi et al., 2003; Knox et al., 2008 a, b). The capacity of alginate as a crosslinked product (calcium alginate) for Cr(VI) uptake was demonstrated by Fiol et al. (2004), who obtained an uptake of 86.42 mmol of Cr(VI) per L of wet sorbent volume using grape stalk wastes encapsulated into calcium alginate. The Cr(VI) removal ability of cross-linked calcium alginate was also shown by Araujo and Teixeira (1997), and its ability to bind Cu was shown by Chen et al. (1990 and 1993) and Wan et al. (2004). The removal of Cu, Cr, and As from treated wood onto the biopolymers, chitin and chitosan, was shown by Kartal and Iamamura (2004). The use of biopolymers based on elastine-like polypeptides for the selective removal of Hg was reported by Kostal et al. (2003), who also reported their potential for binding and removal of other metals such as As and Cr. Recently, the use of a similar elastin-like polypeptide composed of a polyhistidine tail was exploited as a metalbinding biopolymer with high affinity toward Cd by Prabhukumar et al. (2004). Knox et al. (2007 and 2008 a, b) showed that biopolymers (with and without cross-linking) have the ability to sequester a large variety of metals (e.g., Cu, Pb, Cd, As, Cr, Zn, and Ni) and organic contaminants (e.g., phenanthrene and pyrene).

In-situ electrochemical analysis of microbial activity

Microbes have a wide range of metabolic capabilities available that makes them industrially useful organisms. Monitoring these metabolic processes is a crucial component in efficient industrial application. Unfortunately, monitoring these metabolic processes can often be invasive and time consuming and expensive, especially within an anaerobic environment. Electrochemical techniques, such as cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) offer a non-invasive approach to monitor microbial activity and growth. EIS and CV were used to monitor Clostridium phytofermentans, an anaerobic and endospore-forming bacterium. C. phytofermentans ferments a wide range of sugars into hydrogen, acetate, and ethanol as fermentation by-products. For this study, both traditional microbiological and electrochemical techniques were used to monitor the growth of C. phytofermentans and the formation of fermentation products. An irreversible reduction peak was observed using CV beginning at mid-logarithmic phase of growth. This peak was associated with C. phytofermentans and not the spent medium and was indicative of a decrease in carbon and energy sources to the cells. Additionally, EIS analysis during growth provided information related to increased charge transfer resistance of the culture also as a function of carbon and energy source depletion. Results demonstrate that CV and EIS are useful tools in the monitoring the physiological status of bioprocesses.