Tailor-made novel electrospun polycaprolactone/polyethyleneimine fiber membranes for laccase immobilization: An all-in-one material to biodegrade textile dyes and phenolic compounds.

In spite of many works on the biodegradation of textile dyes and phenolic compounds, we propose a new, inexpensive, environmentally friendly, and sustainable material based on electrospun fiber and immobilized laccase. The polycaprolactone (PCL)/polyethyleneimine (PEI) electrospun fibers were optimized and prepared by electrospinning technique according to the operational parameters like PCL concentration (12 wt%), PEI concentration (10 wt%), voltage (16 kV), needle tip-collector distance (20 cm), and injection speed (0.7 mL/h). Next, characterization studies were performed to investigate the morphology and structure of the electrospun fibers without and with laccase. The crude laccase was obtained by cultivating the white rot fungus T. trogii (TT), and T. versicolor (TV). The resulting electrospun fibers showed a smooth surface with a mean diameter of around 560 nm, and larger diameters were observed after laccase immobilization. According to the results, immobilization increased the stability properties of laccase such as storage, and operational. For instance, the residual activity of the PCL/PEI/TTL and PCL/PEI/TVL after 10 repeated cycles, was 33.2 ± 0.2% and 26.0 ± 0.9%, respectively. After 3 weeks of storage, they retained around 30% of their original activity. Moreover, the PCL/PEI/TTL and PCL/PEI/TVL were found to possess high decolorization yield to remove Orange II and Malachite Green textile dyes from solutions imitating polluted waters. Among them, the PCL/PEI/TTL exhibited the highest decolorization efficiencies of Orange II and Malachite Green after 8 continuous uses at pH 5 and a temperature of 50 °C, reaching over 86%, and 46%, respectively. Moreover, PCL/PEI/TTL and PCL/PEI/TVL effectively degraded the 2,6-dichlorophenol phenolic compound at an optimal pH and temperature range and exhibited maximum removal efficiency of 52.6 ± 0.1% and 64.5 ± 7.6%, respectively. Our approach combines the advantageous properties of electrospun fiber material and immobilization strategy for the efficient use of industrial scale important enzymes such as laccase in various enzymatic applications.

Design of laccase-metal-organic framework hybrid constructs for biocatalytic removal of textile dyes.

This study aims to present a simple and effective carrier matrix to immobilize laccase as opposed to complex and tedious immobilization processes and also to use it in the removal of textile dyes. For this purpose, Cobalt (Co) and Copper (Cu) based metal-organic frameworks (MOFs) were prepared and laccase was immobilized on two different MOFs via encapsulation. The characterization outcomes showed that laccase was well immobilized into MOF supports. Optimum pH and temperature were found for Lac/Co-MOF (pH 4.5 at 50 °C) and Lac/Cu-MOF (pH 5.0 at 50 °C). The Km (0.03 mM) and Vmax (97.4 µmol/min) values of Lac/Cu-MOF were lower than those of Lac/Co-MOF (Km = 0.13 mM, Vmax = 230.7 µmol/min). The immobilized laccases showed good reusability as well as improved resistance to temperature denaturation and high storage stability. For instance, the Lac/Co-MOF and Lac/Cu-MOF retained more than 58% activity after 4 weeks of storage at room temperature. Meanwhile, Lac/Co-MOF and Lac/Cu-MOF maintained 56.5% and 55.8% of their initial activity, respectively, after 12 reuse cycles. Moreover, thermal deactivation kinetic studies of immobilized laccases displayed lower k value, higher t1/2, and enhancement of thermodynamic parameters, which means better thermostability. Finally, the decolorization activities for the Lac/Co-MOF were 78% and 61% at the 5th cycle for Reactive Blue 171 and Reactive Blue 198, respectively. In conclusion, it can be inferred that the MOFs are more sustainable and beneficial support for laccase immobilization and they can be efficient for removing textile dyes from industrial wastes.

Synthesis of new 7-amino-3,4-dihydroquinolin-2(1H)-one-peptide derivatives and their carbonic anhydrase enzyme inhibition, antioxidant, and cytotoxic activities.

Six new monopeptides, seven new dipeptides, and two deprotected monopeptide dihydroquinolinone conjugates were prepared by the benzothiazole-mediated method and their structures were confirmed by nuclear magnetic resonance, mass, infrared spectroscopy, and elemental analysis methods. The human carbonic anhydrase (hCA) I and hCA II enzyme inhibition activities of the compounds were determined using the stopped-flow instrument. The synthesized peptide-dihydroquinolinone conjugates 2, 3, 6, 10, 13, and 15 showed inhibition against the hCA II enzyme in the range of 15.7-65.7 µM. However, none of the compounds showed inhibition of hCA I at a concentration of 100 µM. The antioxidant activities of the compounds were also examined using the DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging method at concentrations of 12.5-125 µg/ml, but when compared with the standard antioxidant compounds α-tocopherol and butylated hydroxyanisole (BHA), weak antioxidant activities were detected. The cytotoxic effects of four compounds against the A549 and BEAS-2B cell lines were also investigated. Among the compounds studied, compound 7 was found to be most effective, with the IC50 values on the A549 cells for 48 and 72 h being 26.87 and 9.979 µg/ml, respectively, and the IC50 values on the BEAS-2B cells being >100 µg/ml. None of the tested compounds showed antimicrobial activity in the concentration range (800-1.56 µg/ml) studied.

Cytotoxic and antimicrobial potential of benzimidazole derivatives.

New benzimidazole derivatives were synthesized and their structures were characterized by spectroscopic and microanalysis techniques. The cytotoxic properties of ten benzimidazole derivatives, five of which were synthesized in our previous studies, were determined against the lung cancer cell line, A549, and the healthy lung epithelial cell line, BEAS-2B. Among the ten compounds tested, based on the 72-h incubation results, compound 12 was the most cytotoxic against the A549 cell line, whereas against the BEAS-2B cell line, it was as cytotoxic as cisplatin. The IC50 values of compound 12 were 3.98 and 2.94 µg/ml for A549 and BEAS-2B cells, respectively. The cisplatin values were 6.75 and 2.75 µg/ml for A549 and BEAS-2B cells, respectively. Compounds 10, 8, 7, and 13 showed toxic effects against A549 cells, but were less toxic against BEAS-2B cells than cisplatin. The antimicrobial activity of these compounds against pathogenic bacteria and yeasts was also evaluated based on their minimum inhibitory concentration (MIC) values. The compounds, except 12 and 13, generally showed higher antimicrobial activity against yeasts, compared with bacteria. Compound 12 showed better activity against Pseudomonas aeruginosa and Staphylococcus aureus than against Escherichia coli. Compounds 7, 8, and 11 were the most effective ones against the microorganisms, and yeasts were highly sensitive to these compounds with MIC values of 25-100 µg/ml.

Moringa oleifera leaves: could solvent and extraction method affect phenolic composition and bioactivities?

This study was carried out to evaluate the effect of extraction methods (direct maceration (DM) and successive maceration (SM)) and solvents (dichloromethane (DCM), ethyl acetate (EAc), butanol (But), and aqueous extraction (Aqu)) on the phenolic composition and biological activities of Moringa oleifera leaves cultivated for the first time in Tunisia. Total polyphenol content (TPC), total flavonoid content (TFC) and LC-MS analysis were performed for all extracts. Antioxidant potential by DPPH, metal chelating, and FRAP assays, antibacterial activity against Staphylococcus aureus (ATCC 29213) and Escherichia coli (ATCC 25922) by the minimum inhibitory concentration method (MIC) and cytotoxic properties against lung cancer cells (A549), were determined. Phenolic compounds and biological activities of M. oleifera leaves depend on the method and the solvent used for the extraction of these bioactive substances. The But extract prepared by SM method exhibited the highest amounts of TPC (103.06 ± 0.5 mg GAE/g DE) and showed the strongest potential antioxidant (CI50 = 0.26 mg/mL on DPPH test), antibacterial (MIC = 250 µg/mL) and cytotoxic activities (GI50 = 69.9 µg/mL). LC-MS analysis showed that 28 phenolic compounds of 33 tested standards were found in M. oleifera leaves. The But extract obtained by SM method exhibited the highest amounts of rutin, quercetin and syringic acid.

Laccase-conjugated thiolated chitosan-Fe3O4 hybrid composite for biocatalytic degradation of organic dyes.

In this study, a novel immobilization support for laccase was developed to enhance enzyme stability, efficiency and reusability. Firstly, Fe3O4 magnetic particles were synthesized and modified by the co-precipitation route using thiolated chitosan (TCS). The support was characterized using several methods. Afterward, laccase was attached to the surface of functional support. The biochemical properties of the immobilized laccase were comprehensively investigated. It was observed that immobilized laccase achieved maximum activity at pH 4.0 and the optimum temperature was found to be 50 °C. After storage at +4 °C and ~25 °C for 4 weeks, the residual activity of the immobilized laccase was 87% and 80% of its initial activity, respectively. At 55 °C, the activity of immobilized laccase decreased to 73.4% in 180 min and after reused 20 times, the relative activity of immobilized laccase still was approximately 50% of its initial activity. Moreover, the textile dye (Reactive Blue 171 and Acid Blue 74) decolorization activity of immobilized laccase was also tested and it showed long-term textile dye decolorization activity. These results are promising for the use of laccase in industrial and biotechnological applications. Therefore, this functionalized magnetic hybrid composite might be used to immobilize laccase, an industrially important enzyme.

Comparison of indigo carmine decolorization by Pseudomonas aeruginosa and crude laccase enzyme from Funalia trogii.

The effects of incubation time, temperature, initial pH, and dye concentration on the indigo carmine decolorization activity of Pseudomonas aeruginosa ATCC 10145 and some factors on the decolorization potential of crude laccase enzyme obtained from Funalia trogii ATCC 200800 were comparatively investigated. This bacterium showed effective decolorization activity at all agitation and temperature values. Indigo carmine was greatly decolorized by P. aeruginosa at all pH values except pH 10. A decrease in decolorization activity occurred with increasing dye concentration, but this bacterium effectively decolorized the dye within 24 h. The decolorization process was through microbial metabolism, not biosorption. No decolorization or laccase activity could be obtained by the cell-free intracellular extract or culture filtrate of this bacterium. On the other hand, crude laccase effectively decolorized indigo carmine under highly acidic conditions, especially at pH 3.0 as 57% in 300 seconds. This activity decreased progressively due to the increase in pH values. In a short incubation period and at high temperature values, the crude laccase enzyme removed the color of the dye at 50 °C (56%), 60 °C (45%), and 70 °C (38%). These data are important for improving methods for decolorization of textile dyes used at high temperatures in various industrial applications.

A comparative study on decolorization of reactive azo and indigoid dyes by free/immobilized pellets of Trametes versicolor and Funalia trogii.

The objective of the present study was to investigate decolorization of Acid Blue 74 and Reactive Blue 198 dyes by free and immobilized white rot fungal pellets in order to confirm the possibility of practical application via repeated-batch cultivation. Decolorization studies were conducted using free pellets (FP), fungal cells immobilized on activated carbon (IFCAC) and pinewood (IFCP), and also fungal cells entrapped in alginate beads (FCEAB). No additional nitrogen and carbon source was used and high decolorization rates were achieved in only dye-contained media without pH adjustment. Acid Blue 74 was decolorized 96 and 94% within 2 hr by Trametes versicolor and Funalia trogii free pellets, respectively. These values were 87 and 84% for Reactive Blue 198, in this respect. Immobilization of fungal cells on pinewood increased the usability of pellets and the average decolorization efficiency of both dyes. The micro environment changed in the presence of pinewood and increased the stability of immobilized pellets. Decolorization was performed rapidly and efficiently. Laccase activity enhanced with availability of pinewood, and high laccase production with F. trogii was obtained. After separation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), the molecular weight of T versicolor and F. trogii laccase bands was determined 64 and 61 kDa approximately. Green bands were obtained by the activity staining process with laccase substrate (ABTS) after gel renaturation step.

Antioxidative and metabolic responses to extended cold exposure in rats.

In this work, we investigated whether extended cold exposure increases oxidative damage and susceptibility to oxidants of rat liver, heart, kidney and lung which are metabolically active tissues. Moreover in this study the effect of cold stress on some of the lipid metabolic mediators were studied in rat experimental model. Male albino Sprague-Dawley rats were randomly divided into two groups: The control group (n=12) and the cold-stress group (n=12). Tissue superoxide dismutase (SOD), catalase (CAT), glutathion S-transferase (GST) and glutathion reductase (GR) activities and glutathion (GSH) were measured using standard protocols. The biochemical analyses for total lipid, cholesterol, trigliceride, HDL, VLDL and LDL were done on autoanalyzer. In cold-stress groups SOD activity was decreased in the lung whereas it increased in the heart and kidney. CAT activity was significantly decreased (except liver) in all the tissues in treated rats. GST activity of cold-induced rats increased in liver and heart while decreased in the lung. GR activity was significantly decreased (except in liver) in all the tissues in cold-stressed rats. GSH level was significantly increased in the heart but decreased in the lung of animals exposed to cold when compared to controls. It was found that among the groups trigliceride, total lipid, HDL and VLDL parameters varied significantly but cholesterol and LDL had no significant variance. In this study, we found that exposure of extended (48 h) cold (8 degrees C) caused changes both in the antioxidant defense system (as tissue and enzyme specific) and serum lipoprotein profiles in rats.

Adsorptive removal of textile dyes from aqueous solutions by dead fungal biomass.

Dead fungal biomass prepared from Phanerochaete chrysosporium and Funalia trogii was tested for their efficiency in removal of textile dyes. The effects of contact time, initial dye concentration, amount of dead biomass and agitation rate on dye removal have been determined. Removal of all dyes required a very short time (60 min). Experimental results show that, P. chrysosporium was more effective than F. trogii . An increase in the amount of dead biomass positively affected of the dye removal. The removal efficiency of different amount of biomass was in order 1 g > 0.5 g > 0.2 g > 0.1 g. The highest removal was obtained at 150-200 rpm. Slightly lower removing activities were found at lower agitation rates. This study showed that it was possible to remove textile dyes by dead biomass of P. chrysosporium .

Role of white rot fungus Funalia trogii in detoxification of textile dyes.

Toxic and genotoxic effects of the textile dyes on organisms suggest the need for remediation of dyes before discharging them into the environment. For this reason, the ability of Funalia trogii pellets to detoxify textile dyes was investigated and evaluated. Although, textile dyes are toxic substances for many microorganisms, the pellets were able to decolorize and detoxify the azo dyes used. Astrazon Blue and Red dyes inhibit growth of F. trogii and S. aureus on solid medium in a concentration dependent manner. The toxicity of these dyes on a fungus, F. trogii and a bacterium, S. aureus was significantly decreased after pretreatment with fungal pellets.

Astrazon Red dye decolorization by growing cells and pellets of Funalia trogii.

The dye decolorization activity of fungal pellets has been compared with another method based on the decolorization of dye by growing cells. The pellet method was more advantageous than the growing cell method. The growing cells of F. trogii decolorized 21% of the dye in distilled water medium and 16% in stock basal medium in 24 h. On the other hand, Funalia trogii pellets rapidly decolorized the Astrazon Red dye, mono-azo textile dye, in 24 h, without any visual sorption of any dye to the pellets. The effect of various supplements on longevity of decolorization by free pellets was also tested. Glucose and cheese whey supplementation improved dye decolorization performance of the pellets and remained high and stable for 10 days. We also tested the dye decolorization ability of pellets immobilized on activated carbon. These pellets showed the stable dye decolorization activity during the repeated batch experiments. The study revealed that dye decolorization by pellets is more effective method than the growing cell method.