Karl-Heinz Robra

Decolorization and Detoxification of Textile Dyes with a Laccase from Trametes hirsuta

ABSTRACT Trametes hirsuta and a purified laccase from this organism were able to degrade triarylmethane, indigoid, azo, and anthraquinonic dyes. Initial decolorization velocities depended on the substituents on the phenolic rings of the dyes. Immobilization of theT. hirsuta laccase on alumina enhanced the thermal stabilities of the enzyme and its tolerance against some enzyme inhibitors, such as halides, copper chelators, and dyeing additives. The laccase lost 50% of its activity at 50 mM NaCl while the 50% inhibitory concentration (IC50) of the immobilized enzyme was 85 mM. Treatment of dyes with the immobilized laccase reduced their toxicities (based on the oxygen consumption rate of Pseudomonas putida) by up to 80% (anthraquinonic dyes). Textile effluents decolorized with T. hirsuta or the laccase were used for dyeing. Metabolites and/or enzyme protein strongly interacted with the dyeing process indicated by lower staining levels (K/S) values than obtained with a blank using water. However, when the effluents were decolorized with immobilized laccase, they could be used for dyeing and acceptable color differences (ΔE*) below 1.1 were measured for most dyes.

Indigo degradation with purified laccases from Trametes hirsuta and Sclerotium rolfsii

The degradation of the textile dye indigo with purified laccases from the fungi Trametes hirsuta (THL1 and THL2) and Sclerotium rolfsii (SRL1) was studied. All laccases were able to oxidize indigo yielding isatin (indole-2,3-dione), which was further decomposed to anthranilic acid (2-aminobenzoic acid). Based on the oxygen consumption rate of the laccases during indigo degradation, a potential mechanism for the oxidation of indigo involving the step-wise abstraction of four electrons from indigo by the enzyme was suggested. Comparing the effect of the known redox-mediators acetosyringone, 1-hydroxybenzotriazole (HOBT) and 4-hydroxybenzenesulfonic acid (PHBS) on laccase-catalyzed degradation of indigo, we found a maximum of about 30% increase in the oxidation rate of indigo with SRL1 and acetosyringone. The particle size of indigo agglomerates after laccase treatment was influenced by the origin of the laccase preparation and by the incubation time. Diameter distributions were found to have one maximum and compared to the indigo particle size distribution of the control, for all laccases, the indigo agglomerates seemed to have shifted to smaller diameters. Bleaching of fabrics by the laccases (based on K/S values) correlated with the release of indigo degradation products.

Purification and properties of a feruloyl esterase involved in lignocellulose degradation by Aureobasidium pullulans

ABSTRACT The lignocellulolytic fungus Aureobasidium pullulans NRRL Y 2311-1 produces feruloyl esterase activity when grown on birchwood xylan. Feruloyl esterase was purified from culture supernatant by ultrafiltration and anion-exchange, hydrophobic interaction, and gel filtration chromatography. The pure enzyme is a monomer with an estimated molecular mass of 210 kDa in both native and denatured forms and has an apparent degree of glycosylation of 48%. The enzyme has a pI of 6.5, and maximum activity is observed at pH 6.7 and 60°C. Specific activities for methyl ferulate, methyl p-coumarate, methyl sinapate, and methyl caffeate are 21.6, 35.3, 12.9, and 30.4 μmol/min/mg, respectively. The pure feruloyl esterase transforms both 2-O and 5-O arabinofuranosidase-linked ferulate equally well and also shows high activity on the substrates 4-O-trans-feruloyl-xylopyranoside, O-{5-O-[(E)-feruloyl]-α-l-arabinofuranosyl}-(1,3)-O-β-d-xylopyranosyl-(1,4)-d-xylopyranose, and p-nitrophenyl-acetate but reveals only low activity on p-nitrophenyl-butyrate. The catalytic efficiency (kcat/Km) of the enzyme was highest on methyl p-coumarate of all the substrates tested. Sequencing revealed the following eight N-terminal amino acids: AVYTLDGD.

Nitrile Hydratase and Amidase from Rhodococcus rhodochrous Hydrolyze Acrylic Fibers and Granular Polyacrylonitriles

ABSTRACT Rhodococcus rhodochrous NCIMB 11216 produced nitrile hydratase (320 nkat mg of protein−1) and amidase activity (38.4 nkat mg of protein−1) when grown on a medium containing propionitrile. These enzymes were able to hydrolyze nitrile groups of both granular polyacrylonitriles (PAN) and acrylic fibers. Nitrile groups of PAN40 (molecular mass, 40 kDa) and PAN190 (molecular mass, 190 kDa) were converted into the corresponding carbonic acids to 1.8 and 1.0%, respectively. In contrast, surfacial nitrile groups of acrylic fibers were only converted to the corresponding amides. X-ray photoelectron spectroscopy analysis showed that 16% of the surfacial nitrile groups were hydrolyzed by the R. rhodochrousenzymes. Due to the enzymatic modification, the acrylic fibers became more hydrophilic and thus, adsorption of dyes was enhanced. This was indicated by a 15% increase in the staining level (K/Svalue) for C.I. Basic Blue 9.

Thermo-alkali-stable catalases from newly isolated Bacillus sp. for the treatment and recycling of textile bleaching effluents

Three thermoalkaliphilic bacteria, which were grown at pH 9.3-10 and 60-65 degrees C were isolated out of a textile wastewater drain. The unknown micro-organisms were identified as thermoalkaliphilic Bacillus sp. Growth conditions were studied and catalase activities and stabilities compared. Catalases from Bacillus SF showed high stabilities at 60 degrees C and pH 9 (t1/2=38 h) and thus this strain was chosen for further investigations, such as electron microscopy, immobilization of catalase and hydrogen peroxide degradation studies. Degradation of hydrogen peroxide with an immobilized catalase from Bacillus SF enabled the reuse of the water for the dyeing process. In contrast, application of the free enzyme for treatment of bleaching effluents, caused interaction between the denaturated protein and the dye, resulting in reduced dye uptake, and a higher color difference of 1.3DeltaE* of dyed fabrics compared to 0.9DeltaE* when using the immobilized enzyme.

Two-stage anaerobic fermentation of organic waste in CSTR and UFAF-reactors

The mechanically separated liquid fraction of organic waste from households was used as a substrate for anaerobic fermentation. A two-step system consisting of a 2001 continuously stirred tank reactor (CSTR) and a 501 upflow anaerobic filter filled with glass foam pearls was constructed. The CSTR was operated for 5 months with a loading rate of 9.8 kg CSB m(-3) day(-1). At a resulting hydraulic retention time (HRT) of 24 days, 68% COD was degraded and a gas productivity of 4.0 m3 m(-3) day(-1) was achieved. Further digestion of the CSTR output was separately optimised in a 20 l-UFAF and based on these results a 50 l-UFAF was connected to the CSTR. At a resulting hydraulic retention time (HRT) of 6 days 38% COD was degraded and a gas productivity of 1.8 m3 m(-3) day(-1) was achieved with the 50 l-UFAF. Thus, the overall degradation efficiency of the two-phase system was 80%. The methane content (61%) of the biogas produced in the 50 l-UF

Indigo Degradation with Laccases from Polyporus sp. and Sclerotium rolfsii

We have investigated the potential of fungal laccases from Polyporus sp. and Sclerotium rolfsii to degrade insoluble indigo. Evidence shows that both laccases are able to oxidize insoluble indigo to give isatin (indole-2,3-dione), which further degrades to anthranilic acid (2-aminobenzoic acid). Adsorption studies show that the laccase from Polyporus sp. has a higher affinity for indigo than the laccase of Sclerotium rolfsii. The particle size of indigo agglomerates is influenced by the origin of the laccase preparation and the incubation time. The potential of laccases to modify indigo stained fabrics is assessed. Treatment of indigo dyed fabrics with laccase prevents indigo backstaining, and Polyporus sp. appears to be more effective for reducing backstaining.