Stephanie Gail Burton

Potential applications of laccase-mediated coupling and grafting reactions: A review

Many industries are currently pursuing enzymatic approaches for developing green chemistry technologies mainly due to shortcomings of physico-chemical methods, growing environmental concerns, legal restrictions, and increasing scientific knowledge. Laccase-assisted reactions, in particular, are being intensively investigated as they are generally eco-friendly and have wide application potential. Laccases only require oxygen as co-substrate, they release water as the only by-product and have a wide substrate range which can be further extended by use of laccase-mediator systems. Consequently, research covering various applications of laccase has been rapidly increasing in recent years, particularly in the areas of coupling and grafting reactions. This review summarizes the advances that have been made in developing technologies based on laccase-mediated coupling and grafting reactions for potential application in areas such as environmental pollution control, modification of lignocellulose materials, food industry, biosensors, textile industry, pharmaceutical industry, and in organic synthesis.

Developments in nitrile and amide biotransformation processes.

Nitrile and amide bioconversions have received attention through their ability to provide a range of commercially important chemicals. These bioconversions are mediated by distinct process strategies. Here, the processes performance is discussed, and the use of whole cells, cell extracts and enzymes as biocatalysts is compared. Additionally, the benefits of biocatalyst reuse through immobilization have been identified and immobilization matrices utilized for these bioconversions evaluated. Exploitation and commercial development will depend on optimization of the process performance and the capacity for scale-up in addition to the biocatalytic potential. High substrate concentrations and biocatalyst stability and reuse through immobilization strategies provide driving forces towards more efficient process kinetics. Membrane immobilization is specifically highlighted as a route to maximize process performance.

Microbial community structure stability, a key parameter in monitoring the development of constructed wetland mesocosms during start-up

Constructed wetlands (CWs) are known to be effective for treating waste streams, and pilot-scale CWs are useful for assessing the impact of pollutants and their remediation. However, little is known with respect to the establishment of these mesocosm systems or the parameters which should be monitored in assessing system equilibration, i.e. when they present stabilised physical and biological patterns. The aim of this study was to evaluate the temporal aspects of CW equilibration as a basis for future studies of system response to amendment. Microbial biomass and hydraulic conductivity values were monitored and microbial community fingerprints were obtained using denaturing gradient gel electrophoresis (DGGE). This study showed that microbial community fingerprinting provides a valuable tool for assessing the time scales of equilibration, as it was the last parameter which stabilised during the equilibration period. Hydraulic conductivity was also an important parameter in determining the time scale for initiation of the equilibration process during the study. For a CW of the dimensions used (173 cm long/106 cm large/30 cm depth), community equilibration times demonstrated on the basis of similar microbial community structures were found to be on the order of 100 days.

Actinobacterial Peroxidases: an Unexplored Resource for Biocatalysis

Peroxidases are redox enzymes that can be found in all forms of life where they play diverse roles. It is therefore not surprising that they can also be applied in a wide range of industrial applications. Peroxidases have been extensively studied with particular emphasis on those isolated from fungi and plants. In general, peroxidases can be grouped into haem-containing and non-haem-containing peroxidases, each containing protein families that share sequence similarity. The order Actinomycetales comprises a large group of bacteria that are often exploited for their diverse metabolic capabilities, and with recent increases in the number of sequenced genomes, it has become clear that this metabolically diverse group of organisms also represents a large resource for redox enzymes. It is therefore surprising that, to date, no review article has been written on the wide range of peroxidases found within the actinobacteria. In this review article, we focus on the different types of peroxidases found in actinobacteria, their natural role in these organisms and how they compare with the more well-described peroxidases. Finally, we also focus on work remaining to be done in this research field in order for peroxidases from actinobacteria to be applied in industrial processes.

Actinobacteria isolated from termite guts as a source of novel oxidative enzymes.

A multi-faceted screening programme was designed to search for the oxidases, laccase, peroxidase and tyrosinase. Actinobacteria were selectively isolated from the paunch and colon region of the hindguts of the higher termite, Amitermes hastatus. The isolates were subjected to solid media assays (dye decolourization, melanin production and the utilization of indulin AT as sole carbon source) and liquid media assays. Eleven of the 39 strains had the ability to decolourize the dye RBBR, an indicator for the production of peroxidases in actinobacteria. Melanin production on ISP6 and ISP7 agar plates served as a good indicator for laccase and/or tyrosinase production and the ability of the strains to grow in the presence of indulin AT as a sole carbon source served as a good indicator of lignin peroxidase and/or general peroxidase production. Enzyme-producing strains were cultivated in liquid media and extracellular enzyme activities measured. Strains with the ability to produce oxidative enzymes under the conditions tested were identified to genus level by 16S rRNA gene analysis and compared to known oxidase producers. A strong relationship was observed between the environment sampled (termite guts where lignocellulose degradation occurs) and the dominant type of oxidative enzyme activity detected (laccases and peroxidases), which suggests the possibility of future targeted screening protocols linking the physical properties of the target enzymes with specific operational conditions required, such as lignocellulosic degradation in the preparation of biofuel feedstocks.

Phenolic removal processes in biological sand filters, sand columns and microcosms

This study evaluated the removal processes involved in the removal of the phenolic component of winery wastewater in biological sand filters, sand columns and sand microcosms. It was found that at low influent phenolic concentrations, complete organic removal was accomplished, but at high concentrations, there was incomplete substrate removal and an accumulation of potentially toxic metabolites, including catechol. The sand provided a suitable substrate for the treatment of phenolic-laden waste, and both biotic (48%) and abiotic (52%) removal mechanisms effected the removal of model phenolics. Prior acclimation of microbial communities increased the biodegradation rate of phenolic acids significantly.

Assessment of temporal and spatial evolution of bacterial communities in a biological sand filter mesocosm treating winery wastewater

To assess the impact of winery wastewater (WW) on biological sand filter (BSF) bacterial community structures, and to evaluate whether BSFs can constitute alternative and valuable treatment‐ processes to remediate WW.

Treatment of high ethanol concentration wastewater by biological sand filters : Enhanced COD removal and bacterial community dynamics

Winery wastewater is characterized by its high chemical oxygen demand (COD), seasonal occurrence and variable composition, including periodic high ethanol concentrations. In addition, winery wastewater may contain insufficient inorganic nutrients for optimal biodegradation of organic constituents. Two pilot-scale biological sand filters (BSFs) were used to treat artificial wastewater: the first was amended with ethanol and the second with ethanol, inorganic nitrogen (N) and phosphorus (P). A number of biochemical parameters involved in the removal of pollutants through BSF systems were monitored, including effluent chemistry and bacterial community structures. The nutrient supplemented BSF showed efficient COD, N and P removal. Comparison of the COD removal efficiencies of the two BSFs showed that N and P addition enhanced COD removal efficiency by up to 16%. Molecular fingerprinting of BSF sediment samples using denaturing gradient gel electrophoresis (DGGE) showed that amendment with high concentrations of ethanol destabilized the microbial community structure, but that nutrient supplementation countered this effect.

Novel, Biocatalytically Produced Hydroxytyrosol Dimer Protects against Ultraviolet-Induced Cell Death in Human Immortalized Keratinocytes

Compounds derived from botanicals, such as olive trees, have been shown to possess various qualities that make them function as ideal antioxidants and, in doing so, protect them against the damaging effect of ultraviolet (UV)-derived oxidative stress. The aim of this study was to biocatalytically synthesize a dimeric product (compound II) from a known botanical, 3-hydroxytyrosol, and test it for its antioxidant ability using a human immortalized keratinocyte cell line (HaCaT). 2,2-Diphenyl-picryhydrazyl (DPPH) antioxidant assays showed 33 and 86.7% radical scavenging activity for 3-hydroxytyrosol and its dimer, respectively. The ferric-reducing antioxidant power (FRAP) assay corroborated this by showing a 3-fold higher antioxidant activity for the dimer than 3-hydroxytyrosol. Western blot analyses, showing cells exposed to 500 μM of the dimeric product when ultraviolet A (UVA)-irradiated, increased the anti-apoptotic protein Bcl-2 expression by 16% and reduced the pro-apoptotic protein Bax by 87.5%. Collectively, the data show that the dimeric product of 3-hydroxytyrosol is a more effective antioxidant and could be considered for use in skin-care products, health, and nutraceuticals.

Minor differences in sand physicochemistry lead to major differences in bacterial community structure and function after exposure to synthetic acid mine drainage

The formation of environmentally toxic acidic waste from mining activities is a world-wide problem. Neutralization of this waste can be accomplished by physicochemical and/or biological means. In this short-term study, synthetic acid mine drainage was added to sand-filled mesocosms containing silica-dominated (quartz) sand. Glucose was added as a carbon source for microbial iron and/or sulphate reduction. Replicates contained two separate batches of sand obtained from the same quarry site. The investigations used to assess and compare the chemical and biological functioning of the replicates included system hydraulic conductivity measurements, sand chemistry, effluent chemistry and bacterial community fingerprinting. Minor differences in composition of the sand, including the levels of available nutrients and micronutrients, resulted in major differences in measured parameters. Significant differences in effluent chemistry were found in systems containing different batches of sand. It was demonstrated that the characteristics of the sand and the presence of acid mine drainage (AMD) impacted the bacterial community structure and function. The importance of the physical substrate on the selection of functional microbial communities in systems remediating AMD should not be under-estimated. The physical substrate should be carefully selected and it may be prudent to include small-scale comparative studies in each particular setting prior to full-scale implementation.