Recent developments in biodegradation of industrial pollutants by white rot fungi and their enzyme system

Increasing discharge and improper management of liquid and solid industrial wastes have created a great concern among industrialists and the scientific community over their economic treatment and safe disposal. White rot fungi (WRF) are versatile and robust organisms having enormous potential for oxidative bioremediation of a variety of toxic chemical pollutants due to high tolerance to toxic substances in the environment. WRF are capable of mineralizing a wide variety of toxic xenobiotics due to non-specific nature of their extracellular lignin mineralizing enzymes (LMEs). In recent years, a lot of work has been done on the development and optimization of bioremediation processes using WRF, with emphasis on the study of their enzyme systems involved in biodegradation of industrial pollutants. Many new strains have been identified and their LMEs isolated, purified and characterized. In this review, we have tried to cover the latest developments on enzyme systems of WRF, their low molecular mass mediators and their potential use for bioremediation of industrial pollutants.     

Dephenolization of industrial wastewaters catalyzed by polyphenol oxidase

A new enzymatic method for the removal of phenols from industrial aqueous effluents has been developed. The method uses the enzyme polyphenol oxidase which oxidizes phenols to the corresponding o-quinones; the latter then undergo a nonenzymatic polymerization to form water-insoluble aggregates. Therefore, the enzyme in effect precipitates phenols from water. Polyphenol oxidase has been found to nearly completely dephenolize solutions of phenol in the concentration range from 0.01 to 1.0 g/L. The enzymatic treatment is effective over a wide range of pH and temperature; a crude preparation of polyphenol oxidase (mushroom extract) is as effective as a purified, commercially obtained version. In addition to phenol itself, polyphenol oxidase is capable of precipitating from water a number of substituted phenols (cresols, chlorophenols, naphthol, etc.). Also, even pollutants which are unreactive towards polyphenol oxidase can be enzymatically coprecipitated with phenol. The polyphenol oxidase treatment has been successfully used to dephenolize two different real industrial waste-water samples, from a plant producing triarylphosphates and from a coke plant. The advantage of the polyphenol oxidase dephenolization over the peroxidase-catalyzed one previously elaborated by the authors is that the former enzyme uses molecular oxygen instead of costly hydrogen peroxide (used by peroxidase) as an oxidant.     

Microbial nitrogen cycles: physiology, genomics and applications

Genes and pathways involved in inorganic nitrogen cycles have been found in traditional as well as unusual microorganisms. These pathways or enzymes play a very important role in the adaptation or survival of these microorganisms under a variety of environmental conditions. Microbial nitrogen metabolism has industrial applications ranging from wastewater treatment to bioremediation and potential future use in biocatalysis for chemical production.     

Development of applications of industrial enzymes from Malaysian indigenous microbial sources

Malaysian enzyme industry is considered almost non-existence, although the import volume is large. Realizing the importance of enzymes, encompassing a wide range of applications in bioindustry, the development of home grown technologies for enzyme production and applications becomes one of the national priorities in industrial biotechnology. Enzyme production from indigenous microbial isolates was performed either by submerged or solid state fermentation processes. Based on its wide and unique spectrum of properties, enzymes have been developed for wide applications in various industrial processes. The development of the enzyme catalysed applications is based on the modification of the reaction systems to enhance their catalytic activities. Some of the applications of the industrial enzymes include the fine chemicals production, oleochemicals modification, detergent formulation, enzymatic drinking of waste papers, animal feed formulation and effluent treatment processes. Enzymes have also shown to be successfully used as analytical tool in the determination of compounds in body fluids. Although, most of these enzyme catalysed reactions were performed in aqueous phase, the use of enzymes in organic solvents was found to be significant for the production of new chemicals.     

Removal of phenols from wastewater by soluble and immobilized tyrosinase

An enzymatic method for removal of phenols from industrial wastewater was investigated. Phenols in an aqueous solution were removed after treatment with mushroom tyrosinase. The reduction order of substituted phenols is catechol > p-cresol > p-chlorophenol > phenol > p-methoxyphenol. In the treatment of tyrosinase alone, no precipitate was formed but a color change from colorless to dark-brown was observed. The colored products were removed by chitin and chitosan which are available abundantly as shellfish waste. In addition, the reduction rate of phenols was observed to be accelerated in the presence of chitosan. Tyrosinase, immobilized by using amino groups in the enzyme on cation exchange resins, can be used repeatedly. By treatment with immobilized tyrosinase, 100% of phenol was removed after 2 h, and the activity was reduced very little even after 10 repeat treatments. (c) 1993 John Wiley & Sons, Inc.     

Polyphenoloxidase from six mature grape varieties and their activities towards various phenols

Juices were prepared from three white and three red grape varieties harvested at full maturity and comparative studies on their oxygen-uptake, absorbance at 420 nm (degree of browning), polyphenoloxidase (EC 1.10.3.1; PPO) activity, and their phenol compositions were done. There was no correlation among the amounts of oxygen-uptake and oxidizable phenols in the juices and their degree of browning. However, there was similarity among the PPO from the six grape varieties in their general enzymatic properties and substrate specificity towards twenty-five phenols. A partially purified PPO fraction from Koshu juice, which did not contain free phenols, showed strong activity towards (+)-catechin, (−)-epicatechin, caffeic acid, catechol, pyrogallol, and protocatechuic acid (oxidizable phenols), but had no activity towards the other fifiteen phenols. The oxidizable substrates were not always the only limiting factor in the oxidation and browning of phenols by the PPO. Some unoxidizable phenols such as gallic acid, p-cresol, and tannic acid which were not substrates for PPO inhibited the oxidation of the oxidizable phenols except pyrogallol which was not inhibited by gallic acid. On the other hand, hydroquinone promoted the oxidation of the oxidizable phenols except protocatechuic acid. These showed that there were competitive reactions and synergism during the enzymatic oxidation of phenols.     

Potential applications of the oxidoreductive enzymes in the decolorization and detoxification of textile and other synthetic dyes from polluted water: a review

Recently, the enzymatic approach has attracted much interest in the decolorization/degradation of textile and other industrially important dyes from wastewater as an alternative strategy to conventional chemical, physical and biological treatments, which pose serious limitations. Enzymatic treatment is very useful due to the action of enzymes on pollutants even when they are present in very dilute solutions and recalcitrant to the action of various microbes participating in the degradation of dyes. The potential of the enzymes (peroxidases, manganese peroxidases, lignin peroxidases, laccases, microperoxidase-11, polyphenol oxidases, and azoreductases) has been exploited in the decolorization and degradation of dyes. Some of the recalcitrant dyes were not degraded/decolorized in the presence of such enzymes. The addition of certain redox mediators enhanced the range of substrates and efficiency of degradation of the recalcitrant compounds. Several redox mediators have been reported in the literature, but very few of them are frequently used (e.g., 1-hydroxybenzotriazole, veratryl alcohol, violuric acid, 2-methoxy-phenothiazone). Soluble enzymes cannot be exploited at the large scale due to limitations such as stability and reusability. Therefore, the use of immobilized enzymes has significant advantages over soluble enzymes. In the near future, technology based on the enzymatic treatment of dyes present in the industrial effluents/wastewater will play a vital role. Treatment of wastewater on a large scale will also be possible by using reactors containing immobilized enzymes.     

Potential Applications of the Oxidoreductive Enzymes in the Decolorization and Detoxification of Textile and Other Synthetic Dyes from Polluted Water: A Review

ABSTRACT

Recently, the enzymatic approach has attracted much interest in the decolorization/degradation of textile and other industrially important dyes from wastewater as an alternative strategy to conventional chemical, physical and biological treatments, which pose serious limitations. Enzymatic treatment is very useful due to the action of enzymes on pollutants even when they are present in very dilute solutions and recalcitrant to the action of various microbes participating in the degradation of dyes. The potential of the enzymes (peroxidases, manganese peroxidases, lignin peroxidases, laccases, microperoxidase-11, polyphenol oxidases, and azoreductases) has been exploited in the decolorization and degradation of dyes. Some of the recalcitrant dyes were not degraded/decolorized in the presence of such enzymes. The addition of certain redox mediators enhanced the range of substrates and efficiency of degradation of the recalcitrant compounds. Several redox mediators have been reported in the literature, but very few of them are frequently used (e.g., 1-hydroxybenzotriazole, veratryl alcohol, violuric acid, 2-methoxy-phenothiazone). Soluble enzymes cannot be exploited at the large scale due to limitations such as stability and reusability. Therefore, the use of immobilized enzymes has significant advantages over soluble enzymes. In the near future, technology based on the enzymatic treatment of dyes present in the industrial effluents/wastewater will play a vital role. Treatment of wastewater on a large scale will also be possible by using reactors containing immobilized enzymes.     

Development of functionalised polyelectrolyte capsules using filamentous Escherichia coli cells

Background

Chloronitrophenols (CNPs) are widely used in the synthesis of dyes, drugs and pesticides, and constitute a major group of environmental pollutants. 4-Chloro-2-nitrophenol (4C2NP) is an isomer of CNPs that has been detected in various industrial effluents. A number of physicochemical methods have been used for treatment of wastewater containing 4C2NP. These methods are not as effective as microbial degradation, however.

Results

A 4C2NP-degrading bacterium, Exiguobacterium sp. PMA, which uses 4C2NP as the sole carbon and energy source was isolated from a chemically-contaminated site in India. Exiguobacterium sp. PMA degraded 4C2NP with the release of stoichiometeric amounts of chloride and ammonium ions. The effects of different substrate concentrations and various inoculum sizes on degradation of 4C2NP were investigated. Exiguobacterium sp. PMA degraded 4C2NP up to a concentration of 0.6 mM. High performance liquid chromatography and gas chromatography–mass spectrometry identified 4-chloro-2-aminophenol (4C2AP) and 2-aminophenol (2AP) as possible metabolites of the 4C2NP degradation pathway. The crude extract of 4C2NP-induced PMA cells contained enzymatic activity for 4C2NP reductase and 4C2AP dehalogenase, suggesting the involvement of these enzymes in the degradation of 4C2NP. Microcosm studies using sterile and non-sterile soils spiked with 4C2NP were carried out to monitor the bioremediation potential of Exiguobacterium sp. PMA. The bioremediation of 4C2NP by Exiguobacterium sp. PMA was faster in non-sterilized soil than sterilized soil.

Conclusions

Our studies indicate that Exiguobacterium sp. PMA may be useful for the bioremediation of 4C2NP-contaminated sites. This is the first report of (i) the formation of 2AP in the 4C2NP degradation pathway by any bacterium and (iii) the bioremediation of 4C2NP by any bacterium.

     

A comprehensive review on application of bioreactor for industrial wastewater treatment

In the recent past, wastewater treatment processes performed a pivotal role in accordance with maintaining the sustainable environment and health of mankind at a proper hygiene level. It has been proved indispensable by government regulations throughout the world on account of the importance of preserving freshwater bodies. Human activities, predominantly from industrial sectors, generate an immeasurable amount of industrial wastewater loaded with toxic chemicals, which not only cause dreadful environmental problems, but also leave harmful impacts on public health. Hence, industrial wastewater effluent must be treated before being released into the environment to restrain the problems related to industrial wastewater discharged to the environment. Nowadays, biological wastewater treatment methods have been considered an excellent approach for industrial wastewater treatment process because of their cost-effectiveness in the treatment, high efficiency and their potential to counteract the drawbacks of conventional wastewater treatment methods. Recently, the treatment of industrial effluent through bioreactor has been proved as one of the best methods from the presently available methods. Reactors are the principal part of any biotechnology-based method for microbial or enzymatic biodegradation, biotransformation and bioremediation. This review aims to explore and compile the assessment of the most appropriate reactors such as packed bed reactor, membrane bioreactor, rotating biological contactor, up-flow anaerobic sludge blanket reactor, photobioreactor, biological fluidized bed reactor and continuous stirred tank bioreactor that are extensively used for distinct industrial wastewater treatment.     

Molecular biology and application of plant peroxidase genes

Peroxidases are a family of isozymes found in all plants; they are heme-containing monomeric glycoproteins that utilize either H(2)O(2) or O(2) to oxidize a wide variety of molecules. These important enzymes are used in enzyme immunoassays, diagnostic assays and industrial enzymatic reactions. Peroxidase genes and their promoters can be used for molecular breeding of useful plants. Transgenic techniques have also been used to investigate the physiological and molecular functions of peroxidase genes in plants. Here, we review transgenic studies of peroxidase genes, including the functional analyses of the enzymes and their promoters. Regarding application of peroxidase genes, it has been reported that overexpression of the tomato TPX2 gene or the sweet potato swpa1 gene conferred increased salt-tolerance or oxidative-stress tolerance, respectively. The growth stimulation effect in transgenic tobacco and hybrid aspen upon overexpression of horseradish peroxidase gene is also discussed.     

Isolation and characterization of the polyphenoloxidase from senescent leaves of black poplar

The polyphenoloxidase (PPO) from black poplar senescent leaves has been purified to almost complete homogeneity by a combination of ammonium sulphate precipitation, Sephadex G75 filtration and DEAE-cellulose chromatography. The purified enzyme has a MW of 60 000 and is probably a Cu⁺ enzyme. Peroxidase (PO) activity co-purifies with PPO and has the same MW as it. The two enzymes differ in pH optimum and in response to the effect of ionic strength. Natural phenols are either substrates, inhibitors or activators of black poplar PPO. This enzyme is an o-diphenoloxidase which binds substrates with K_m in the millimolar range. With caffeic and chlorogenic acids inhibition by excess substrate is observed. Benzoic acid phenols and cinnamic acid phenols are either competitive or non-competitive inhibitors of PPO. Hydroquinone is a highly potent non-competitive inhibitor of the enzyme (K_i  90 μM). Ferulic acid is a potent activator of the PPO-catalysed oxidation of catechol (K_a  0.34 mM, ν_sat/ν_o  7.7).     

Location and catalytic characteristics of a multipotent bacterial polyphenol oxidase.

The melanogenic marine bacterium Marinomonas mediterranea contains a multipotent polyphenol oxidase (PPO) able to oxidize substrates characteristic for tyrosinase and laccase. Thus, this enzyme shows tyrosine hydroxylase activity and it catalyzes the oxidation of a wide variety of o-diphenol as well as o-methoxy-activated phenols. The study of its sensitivity to different inhibitors also revealed intermediate features between laccase and tyrosinase. It is similar to tyrosinases in its sensitivity to tropolone, but it resembles laccases in its resistance to cinnamic acid and phenylthiourea, and in its sensitivity to fluoride anion. This enzyme is mostly membrane-bound and can be solubilized either by non-ionic detergent or lipase treatments of the membrane. The expression of this enzymatic activity is growth-phase regulated, reaching a maximum in the stationary phase of bacterial growth, but L-tyrosine, Cu(II) ions, or 2,5-xylidine do not induce it. This enzyme can be separated from a second PPO form by gel permeation chromatography. The second PPO is located in the soluble fraction and shows a sodium dodecyl sulfate (SDS)-activated action on the characteristic substrates for tyrosinase, L-tyrosine, and L-dopa, but it does not show activity towards laccase-specific substrates. The involvement of the multipotent PPO in melanogenesis and its relationship with the SDS-activated form and with the alternative functions proposed for multicopper oxidases in other microorganisms are discussed.     

Biodegradation and bioremediation of hydrocarbons in extreme environments

Many hydrocarbon-contaminated environments are characterized by low or elevated temperatures, acidic or alkaline pH, high salt concentrations, or high pressure, Hydrocarbon-degrading microorganisms, adapted to grow and thrive in these environments, play an important role in the biological treatment of polluted extreme habitats. The biodegradation (transformation or mineralization) of a wide range of hydrocarbons, including aliphatic, aromatic, halogenated and nitrated compounds, has been shown to occur in various extreme habitats. The biodegradation of many components of petroleum hydrocarbons has been reported in a variety of terrestrial and marine cold ecosystems. Cold-adapted hydrocarbon degraders are also useful for wastewater treatment. The use of thermophiles for biodegradation of hydrocarbons with low water solubility is of interest, as solubility and thus bioavailability, are enhanced at elevated temperatures. Thermophiles, predominantly bacilli, possess a substantial potential for the degradation of environmental pollutants, including all major classes. Indigenous thermophilic hydrocarbon degraders are of special significance for the bioremediation of oil-polluted desert soil. Some studies have investigated composting as a bioremediation process. Hydrocarbon biodegradation in the presence of high salt concentrations is of interest for the bioremediation of oil-polluted salt marshes and industrial wastewaters, contaminated with aromatic hydrocarbons or with chlorinated hydrocarbons. Our knowledge of the biodegradation potential of acidophilic, alkaliphilic, or barophilic microorganisms is limited.     

Enzyme systems for biodegradation of polychlorinated dibenzo-p-dioxins

The angular dioxygenase, cytochrome P450, lignin peroxidase, and dehalogenase are known as dioxin-metabolizing enzymes. All of these enzymes have metal ions in their active centers, and the enzyme systems except for peroxidase have each distinct electron transport chain. Although the enzymatic properties of the angular dioxygenase, lignin peroxidase, and cytochrome P450 have been studied well, the information about dehalogenase is much less than other enzyme systems due to its instability under the aerobic conditions. However, this enzyme system appears to be quite promising from the viewpoint of practical use for bioremediation, because dehalogenases are capable of degradation of polychlorinated dibenzo-p-dioxins (PCDDs) with more than four chlorine substituents, whereas the other three enzyme systems prefer low-chlorinated PCDDs. On the other hand, protein engineering of angular dioxygenase, lignin peroxidase, and cytochrome P450 based on their tertiary structures has great potential to generate highly efficient dioxin-metabolizing enzymes. Actually, we successfully generated 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-metabolizing enzyme by site-directed mutagenesis of cytochrome P450. We hope that recombinant microorganisms harboring genetically engineered dioxin-metabolizing enzymes will be used for bioremediation of soil contaminated with PCDDs and polychlorinated dibenzofurans in the near future.     

Use of plant material for the decontamination of water polluted with phenols

Plant materials were found useful in the decontamination water polluted with phenolic contained in the plant tissue. The enzymes mediated oxidative coupling of the pollutants, followed by precipitation of the formed polymers from the aqueous phase. An industrial wastewater contaminated with 2,4-dichlorophenol (up to 850 ppm) and other chlorinated phenols was successfully treated using minced horseradish, potato, or white radish (amended with H(2)O(2)). Horseradish-mediated removal of 2,4-dichlorophenol from model solutions was comparable with that achieved using purified horseradish peroxidase. In addition, horseradish could be reused up to 30 times. Due to the apparent ease of application, the use of plat material may present a breakthrough in the enzyme treatment of contaminated water. (c) 1994 John Wiley & Sons, Inc.     

Biotechnological applications of peroxidases

Peroxidases are widely distributed in nature. Reduction of peroxides at the expense of electron donating substrates, make peroxidases useful in a number of biotechnological applications. Enzymes such as lignin peroxidase and manganese peroxidase, both associated with lignin degradation, may be successfully used for biopulping and biobleaching in the paper industry, and can produce oxidative breakdown of synthetic azo dyes. Oxidative polymerization of phenols and aromatic amines conducted by horseradish peroxidase (HRP) in water and water-miscible organic solvents, may lead to new types of aromatic polymers. Site directed mutagenesis of HRP has been used to improve the enantioselectivity of arylmethylsulfide oxidations. Peroxidase has a potential for soil detoxification, while HRP as well as soybean and turnip peroxidases have been applied for the bioremediation of wastewater contaminated with phenols, cresols, and chlorinated phenols. Peroxidase based biosensors have found use in analytical systems for determination of hydrogen peroxide and organic hydroperoxides, while co-immobilized with a hydrogen peroxide producing enzyme, they can be used for determination of glucose, alcohols, glutamate and choline. Peroxidase has also been used for practical analytical applications in diagnostic kits, such as quantitation of uric acid, glucose, cholesterol, lactose, and so on. Enzyme linked immunorbent assay (ELISA) tests on which peroxidase is probably the most common enzyme used for labeling an antibody, are a simple and reliable way of detecting toxins, pathogens, cancer risk in bladder and prostate, and many other analytes. Directed evolution methods, appear to be a valuable alternative to engineer new catalyst forms of plant peroxidases from different sources to overcome problems of stability and to increase thermal resistance.     

Dual Purpose of ligninolytic- basidiomycetes: mycoremediation of bioethanol distillation vinasse coupled to sustainable bio-based compounds production

Agro-industrial wastes consist of various agriculture and food industry residues produced worldwide at an estimated rate of a thousand million tons per year. Vinasse is the main wastewater resulting from sugarcane bioethanol production. This agro-industrial effluent is highly polluting and, when discarded with no previous treatment, may cause nutrient imbalance and salt saturation in the soil as well as a reduction in photosynthesis in aquatic ecosystems. Environmentally safe vinasse disposal requires reliable mechanisms, preferably involving the production of goods or services. Various cleanup technologies have been implemented and different microorganisms are being studied as viable remediation agents. Ligninolytic basidiomycetes, which have the ability to store, release and mineralize a wide variety of toxic materials and compounds, are a source of valuable biochemicals for agricultural and industrial uses. This review discusses the use of filamentous fungi in the bioremediation of distillery vinasses, their main characteristics, the enzymes implicated and the transformation processes involved in the production of several high-value bio-based compounds.     

Location and Catalytic Characteristics of a Multipotent Bacterial Polyphenol Oxidase

The melanogenic marine bacterium Marinomonas mediterranea contains a multipotent polyphenol oxidase (PPO) able to oxidize substrates characteristic for tyrosinase and laccase. Thus, this enzyme shows tyrosine hydroxylase activity and it catalyzes the oxidation of a wide variety of o-diphenol as well as o-methoxy-activated phenols. The study of its sensitivity to different inhibitors also revealed intermediate features between laccase and tyrosinase. It is similar to tyrosinases in its sensitivity to tropolone, but it resembles laccases in its resistance to cinnamic acid and phenylthiourea, and in its sensitivity to fluoride anion. This enzyme is mostly membrane-bound and can be solubilized either by non-ionic detergent or lipase treatments of the membrane. The expression of this enzymatic activity is growth-phase regulated, reaching a maximum in the stationary phase of bacterial growth, but l -tyrosine, Cu(II) ions, or 2,5-xylidine do not induce it. This enzyme can be separated from a second PPO form by gel permeation chromatography. The second PPO is located in the soluble fraction and shows a sodium dodecyl sulfate (SDS)-activated action on the characteristic substrates for tyrosinase, l -tyrosine, and l -dopa, but it does not show activity towards laccase-specific substrates. The involvement of the multipotent PPO in melanogenesis and its relationship with the SDS-activated form and with the alternative functions proposed for multicopper oxidases in other microorganisms are discussed.