Decolorization of synthetic dyes by Irpex lacteus in liquid cultures and packed-bed bioreactor

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Abstract

White rot fungus Irpex lacteus was grown in both stationary and agitated submerged liquid cultures as well as in cultures immobilized on polyurethane foam (PUF) and pine wood (PW) to study the effect of growth conditions on degradation of commercial dyes. The respective values of decolorization of the anthraquinone-based Remazol Brilliant Blue R (RBBR) dye used at 150 μg ml−1 were 100, 95, 97 and 100% within 6–10 days. Stationary cultures exhibited higher levels of lignin peroxidase (LiP), manganese-dependent peroxidase (MnP) and laccase than submerged cultures and selective inhibition by NaN3 and n-propylgallate brought evidence that MnP played a major role in the decolorization of RBBR by I. lacteus. When comparing PUF- and PW-immobilized cultures decolorizing RBBR, five-fold MnP levels were detected in the former cultures whereas laccase activities were similar. No LiP was detected in either immobilized culture. Both immobilized cultures were able to rapidly decolorize not only RBBR but also various textile-industry, color bath effluents. Reusability and regenerative capacity of the immobilized cultures, important for application to water bioremediation, were documented.

Introduction

Dyes have found broad application in various industries for dyeing and printing. Product processing methods often cause a loss of large amounts of dyes to wastewaters representing 10–15% of the dyes applied [1]. Anthraquinone belongs to major chromophores found in commercial dyes [2]. Synthetic dyes are resistant to conventional wastewater treatments such as activated sludge, trickling filters, etc. [3], [4] and ultimately enter the environment.

White rot fungi, a group of lignin degrading basidiomycetes, have received considerable attention for their bioremediation potential. They are able to degrade lignin and other recalcitrant molecules using relatively nonspecific extracellular enzymes [5], [6]. Azo, anthraquinone, triphenylmethane, heterocyclic, and phthalocyanine dyes as well as colored wastewater effluents were reported to be decolorized [7], [8], [9], [10]. Fungal peroxidases, laccase and Remazol Brilliant Blue R oxygenase (RBBROx) have been involved in decolorization of synthetic dyes and olive mill wastewater effluents [11], [12], [13], [14].

Irpex lacteus, a cosmopolitan white rot fungus, has been studied in connection with proteinase and cellulase production [15], [16], lignin degradation [17] and degradation of PAHs in shallow stationary cultures [18]. Good capacity for decolorization of azo, anthraquinone, phthalocyanine and triphenylmethane dyes by I. lacteus in stationary liquid cultures has been demonstrated [19]. The fungus was also able to degrade an anthraquinone dye in contaminated soil [20].

Growth conditions represent an important factor influencing the efficiency of ligninolytic fungi to degrade synthetic dyes. Agitated cultures have often been found more efficient in decolorization of various dyes, compared to the static ones, presumably because of an increased mass and oxygen transfer [9], [21]. Also, less sorption of dyes to the biomass was observed in the former type of fungal cultures [9]. Cultures of white rot fungi immobilized on solid surfaces such as polyurethane foam (PUF), nylon web or metal screen have been shown to decolorize single dyes and pigment plant effluents, and to degrade pentachlorophenol [22], [23], [24].

The effect of culture conditions on the ability of I. lacteus to degrade dye compounds is not known and, therefore, our study was aimed at a comparison of dye degradation capacities of submerged and stationary liquid cultures and fungal cultures immobilized on PUF or pine wood (PW) cubes using Remazol Brilliant Blue R (RBBR) dye. The synthesis of ligninolytic enzymes was measured. Selective inhibition with NaN3 and n-propylgallate (n-PG) was used to elucidate roles of ligninolytic enzymes in decolorization of RBBR in vivo. The immobilized fungal cultures were efficient in decolorizing both RBBR and coloring bath effluents and were reusable with intermittent regeneration steps in the presence of a fresh medium.

Section snippets

Chemicals

The specification of samples removed from coloring baths in the textile factory was the following: Drimaren Blue sample containing Drimaren Violet and Levafix Royal Blue dyes (dye concentration: 806 μg ml−1; 550 nm); Drimaren Red sample containing Drimaren Blue, Drimaren Brilliant Orange and Levafix Brilliant Red dyes (dye concentration: 2300 μg ml−1; 508 nm); Acid Black sample containing Gelonyl Black LD140Z dye (dye concentration: 2300 μg ml−1; 569 nm); and Remazol Green sample containing Yoracron

Decolorization of RBBR by stationary and submerged cultures

In stationary cultures, I. lacteus formed mycelial mats floating on the surface of liquid medium starting Days 3–4 after inoculation, whereas, under shaken conditions the mycelium was present in the form of 2- to 4-mm, dense pellets. The decolorization of anthraquinone-based RBBR by stationary cultures was slightly more rapid than by the submerged ones; the respective dye removals after 10 days were 100 and 95% (Fig. 1). Our results, thus, did not confirm an increased dye decolorization in

Conclusions

I. lacteus was shown to be able to efficiently decolorize the anthraquinone RBBR dye in submerged stationary, submerged agitated and immobilized cultures. Time correlation of the presence of extracellular enzyme activities with the decolorization process and selective inhibition of MnP and laccase documented a major role of MnP in biodegradation of RBBR in stationary cultures. In contrast, other factors than a high MnP level seem to ensure the efficient decolorization of RBBR in submerged

Acknowledgements

We thank M. Macková (VŠCHT Prague) and J. Horák (Institute of Physiology, AS CR, Prague) for kindly providing us the enzyme inhibitors. The work was supported by the project No. 526/00/1303 of the Grant Agency of the Czech Republic and by the Institutional Research Concept No. AVOZ5020903.

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      Immobilized bioreactors have been found to exhibit good biological activities and abilities for long time operation (Singh, 2006). Irpex lacteus immobilized in a pine wood (PW) reactor decolorized Remazol Brilliant Blue R more rapidly than a polyurethane foam (PUF) reactor (Kasinath et al., 2003). However, contrary to the performance shown in aseptic batch tests, the application of white rot fungi in continuous bioreactors for dye wastewater treatment has been so far impeded by problems such as excessive growth of fungi causing reactor-clogging (Zhang et al., 1999), bacterial contamination inhibiting fungal decolouration (Hai et al., 2009; Libra et al., 2003), and loss of the extracellular enzymes and mediators essential for dye degradation with treated effluent (Hai et al., 2012).

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