Toxicity toward freshwater and marine water organisms of the cytostatic drugs bleomycin and vincristine and their binary mixture.

Antineoplastic drugs are not effectively removed by wastewater treatment plants, ending up in surface waters. Since these drugs can interfere with the structure and functions of DNA, they pose a potential threat to aquatic biota. Unfortunately, many chemotherapeutic agents have not been studied in an environmental context. Additionally, there is a significant lack of information about the impact of anticancer drugs on marine organisms compared to freshwater species, and most studies only focus on the toxicity of single compounds rather than considering their occurrence as complex mixtures in the environment. Therefore, the aim of this study was to evaluate the ecotoxicity of two commonly used cytostatics, bleomycin and vincristine, toward six biomodels: Pseudokirchneriella subcapitata, Phaeodactylum tricornutum, Brachionus plicatilis, Brachionus calyciflorus, Thamnocephalus platyurus, and Artemia franciscana. These selected aquatic organisms are representatives of both freshwater and marine environments and belong to different trophic levels. The pharmaceuticals were investigated both individually and in combination. Binary mixture toxicity predictions were performed according to the Response Additivity and Independent Action models. Additionally, the toxicity data obtained from these experiments were utilized for risk assessment in the context of the drugs' environmental occurrence. The results indicated that freshwater species were generally more sensitive to both tested compounds than marine organisms, with T. platyurus being the most sensitive. Based on the tests performed on this biomodel, bleomycin was categorized as extremely toxic, while vincristine was considered moderately toxic. Neither of the applied models suitably predicted binary mixture toxicity, as the combination of drugs showed additive, synergistic, and antagonistic effects, suggesting that single compound toxicity data are insufficient for predicting the aquatic toxicities of cytostatics mixtures. The environmental risk of vincristine ranged from low to high, and for bleomycin varied from moderate to high, depending on the matrices examined. Therefore, further research on drug removal is recommended.

Bifunctional conducting polymer matrices with antibacterial and neuroprotective effects.

Current trends in the field of neural tissue engineering include the design of advanced biomaterials combining excellent electrochemical performance with versatile biological characteristics. The purpose of this work was to develop an antibacterial and neuroprotective coating based on a conducting polymer - poly(3,4-ethylenedioxypyrrole) (PEDOP), loaded with an antibiotic agent - tetracycline (Tc). Employing an electrochemical technique to immobilize Tc within a growing polymer matrix allowed to fabricate robust PEDOP/Tc coatings with a high charge storage capacity (63.65 ± 6.05 mC/cm2), drug release efficiency (629.4 µg/cm2 ± 62.7 µg/cm2), and low charge transfer resistance (2.4 ± 0.1 kΩ), able to deliver a stable electrical signal. PEDOP/Tc were found to exhibit strong antimicrobial effects against Gram-negative bacteria Escherichia coli, expressed through negligible adhesion, reduction in viability, and a characteristic elongation of bacterial cells. Cytocompatibility and neuroprotective effects were evaluated using a rat neuroblastoma B35 cell line, and were analyzed using MTT, cell cycle, and Annexin-V apoptosis assays. The presence of Tc was found to enhance neural cell viability and neurite outgrowth. The results confirmed that PEDOP/Tc can serve as an efficient neural electrode coating able to enhance charge transfer, as well as to exhibit bifunctional biological characteristics, different for eukaryotic and prokaryotic cells.

Covalent Immobilization of Organic Photosensitizers on the Glass Surface: Toward the Formation of the Light-Activated Antimicrobial Nanocoating.

Two highly efficient commercial organic photosensitizers-azure A (AA) and 5-(4-aminophenyl)-10,15,20-(triphenyl)porphyrin (APTPP)-were covalently attached to the glass surface to form a photoactive monolayer. The proposed straightforward strategy consists of three steps, i.e., the initial chemical grafting of 3-aminopropyltriethoxysilane (APTES) followed by two chemical postmodification steps. The chemical structure of the resulting mixed monolayer (MIX_TC_APTES@glass) was widely characterized by X-ray photoelectron (XPS) and Raman spectroscopies, while its photoactive properties were investigated in situ by UV-Vis spectroscopy with α-terpinene as a chemical trap. It was shown that both photosensitizers retain their activity toward light-activated generation of reactive oxygen species (ROS) after immobilization on the glassy surface and that the resulting nanolayer shows high stability. Thanks to the complementarity of the spectral properties of AA and APTPP, the effectiveness of the ROS photogeneration under broadband illumination can be optimized. The reported light-activated nanocoating demonstrated promising antimicrobial activity toward Escherichia coli (E. coli), by reducing the number of adhered bacteria compared to the unmodified glass surface.

Removal of two cytostatic drugs: bleomycin and vincristine by white-rot fungi - a sorption study.

PURPOSE: Cytostatic drugs cannot be easily removed by conventional sewage treatment plants, resulting in their ultimate release into aquatic systems where they become a threat. Thus, new technologies which can be used to eliminate these drugs more effectively before they enter the environment are increasingly important. Fungal treatment of wastewaters is a promising and environmentally friendly technology for pharmaceutical remediation. The aim of this work is to examine the biosorption of two cytostatics, bleomycin and vincristine, in the aqueous solution by fungal biomass. METHODS: Five white-rot fungi were used in this study: Fomes fomentarius (CB13), Hypholoma fasciculare (CB15), Phyllotopsis nidulans (CB14), Pleurotus ostreatus (BWPH), and Trametes versicolor (CB8). Tests were conducted on different types of biomass (alive and dead - autoclaved) and in various physico-chemical conditions: varied drug concentrations (5, 10 and 15 mg/L), temperatures (from 15.4 to 29.6 °C), and pH (from 3.2 to 8.8). RESULTS: The results showed that among alive biomass, T. versicolor (CB8) had the greatest sorption ability for bleomycin and P. nidulans (CB14) worked best for vincristine. The tested sorption process could be described by a pseudo-second order kinetics model. Sorption equilibrium studies demonstrated that for bleomycin Redlich-Peterson, while for vincristine Langmuir model fitted best. The thermodynamic studies showed that the sorption process was endothermic chemisorption for bleomycin, and exothermic physisorption for vincristine. For both drugs the sorption ability increased with an increase of the pH value. CONCLUSION: The biosorption on fungal biomass is a favorable alternative to conventional wastewater treatment processes for anticancer drug removal.

Electrically-responsive antimicrobial coatings based on a tetracycline-loaded poly(3,4-ethylenedioxythiophene) matrix.

The growth of bacteria and the formation of complex bacterial structures on biomedical devices is a major challenge in modern medicine. The aim of this study was to develop a biocompatible, conducting and antibacterial polymer coating applicable in biomedical engineering. Since conjugated polymers have recently aroused strong interest as controlled delivery systems for biologically active compounds, we decided to employ a poly(3,4-ethylenedioxythiophene) (PEDOT) matrix to immobilize a powerful, first-line antibiotic: tetracycline (Tc). Drug immobilization was carried out simultaneously with the electrochemical polymerization process, allowing to obtain a polymer coating with good electrochemical behaviour (charge storage capacity of 19.15 ± 6.09 mC/cm2) and high drug loading capacity (194.7 ± 56.2 µg/cm2). Biological activity of PEDOT/Tc matrix was compared with PEDOT matrix and a bare Pt surface against a model Gram-negative bacteria strain of Escherichia coli with the use of LIVE/DEAD assay and SEM microscopy. Finally, PEDOT/Tc was shown to serve as a robust electroactive coating exhibiting antibacterial activity.

White-rot fungi-mediated biodegradation of cytostatic drugs - bleomycin and vincristine.

The contamination of the environment with anticancer drugs, which show recalcitrance to conventional wastewater treatment, has become a significant ecological threat. Fungi represent a promising non-conventional biological alternative for water conditioning. The aim of this work was to evaluate the efficacy of five white-rot fungi (Fomes fomentarius (CB13), Hypholoma fasciculare (CB15), Phyllotopsis nidulans (CB14), Pleurotus ostreatus (BWPH) and Trametes versicolor (CB8)) in the removal of bleomycin and vincristine. The removal capacity was measured at 0, 4, 9, and 14 days of incubation using SPE-UPLC-MS. The enzymatic profiles of laccase, manganese, and lignin peroxidases and wide range of eco- and cytotoxicity, assays of the post-process samples were also conducted. We observed >94% vincristine elimination by F. fomentarius, H. fasciculare and T. versicolor after only 4 days. Bleomycin removal occurred after a minimum of 9 days and only when the drug was incubated with T. versicolor (36%) and H. fasciculare (25%). The removal of both cytostatics was associated with laccase production, and the loss of eco- and cytotoxicity, especially in regard to viability of Lemna minor and Daphnia magna, as well as fibroblasts morphology.

Bacterial Surface Colonization of Sputter-Coated Platinum Films.

Due to its biocompatibility and advantageous electrochemical properties, platinum is commonly used in the design of biomedical devices, e.g., surgical instruments, as well as electro-medical or orthopedic implants. This article verifies the hypothesis that a thin layer of sputter-coated platinum may possess antibacterial effects. The purpose of this research was to investigate the adhesion and growth ability of a model strain of Gram-negative bacteria, Escherichia coli, on a surface of a platinum-coated glass slide. Although some previous literature reports suggests that a thin layer of platinum would inhibit the formation of bacterial biofilm, the results of this study suggest otherwise. The decrease in the number of bacterial cells attached to the platinum-coated glass, which was observed within first three hours of culturing, was found to be a short-time effect, vanishing after 24 h. Consequently, it was shown that a thin layer of sputter-coated platinum did not exhibit any antibacterial effect. For this reason, this study indicates an urgent need for the development of new methods of surface modification that could reduce bacterial surface colonization of platinum-based biomedical devices.

Ecotoxicity risk of presence of two cytostatic drugs: Bleomycin and vincristine and their binary mixture in aquatic environment.

Cytostatic drugs have become one of the greatest environmental threats. They occur in surface, ground and even drinking water. Their key emission sources are hospital effluents, municipal wastewater, as well as drug manufacturers and their effluents. These compounds are extremely stable in natural waters and they are not significantly removed during wastewater treatment, because they are resistant to biodegradation. The aim of this work was to establish possible negative effects of chosen cytostatics: bleomycin and vincristine on the three trophic levels of surface waters. A single agent acute toxicity test was conducted on representatives of the producer - an aquatic freshwater plant Lemna minor, the consumer - crustaceans Daphnia magna, and the decomposer - bacteria Pseudomonas putida. Binary mixture tests were performed according to the Concentration Addition, Response Additivity, and Independent Action models. Both substances had a different effect on the tested organisms; bleomycin could be classified as a very toxic, while vincristine as a toxic water pollutant. Half maximal effective concentration (EC50) values designed in the presented single agent acute toxicity studies are < 10 mg/L in all the tests with bleomycin as well as vincristine conducted on L. minor. In tests with vincristine performed on D. magna and P. putida EC50 > 100 mg/L. The highest toxicity is demonstrated by bleomycin towards the aquatic freshwater plant (EC50 = 0.2 mg/L). The binary mixture of the tested chemicals showed antagonistic effects of environmental concern.

Possibilities of Obtaining from Highly Polluted Environments: New Bacterial Strains with a Significant Decolorization Potential of Different Synthetic Dyes.

The aim of this study was the isolation of bacterial strains which have the ability to decolorize synthetic dyes belonging to different chemical groups. The samples for bacterial isolation were collected from aqueous environments-two activated sludges and polluted local river. At the first stage of screening (performed on the solid media supplemented with two dyes-azo Evans blue or triphenylmethane brilliant green), 67 bacterial strains were isolated capable to decolorize the used dyes. In the further study, six dyes with different chemical structures were used: fluorone dyes (Bengal rose, erythrosine), triphenylmethane dyes (brilliant green, crystal violet), azo dyes (Evans blue, Congo red). Initial concentration of each of these chemicals in samples was 0.1 g/l. Obtained results showed that only 31 isolates were able to decolorize all six used dyes (with different efficiencies). Among them, 11 strains were isolated from the river (55% of isolates from this site) and 20 from activated sludges collected from two different treatment plants (15 from the first water treatment plant and 5 from the second which were 42 and 43% of isolated cultures respectively). The decolorizing microorganisms are mostly isolated from different industrial sewages (e.g., textile industry), but results of the study showed that water from polluted river as well as municipal wastewaters may be a precious source for isolation of bacterial strains with the wide spectrum and high decolorization potential. In general, there were no statistically significant differences between decolorization abilities of strains isolated from different sites. The group of dyes that was removed with the highest yield was triphenylmethanes (75.6%), followed by fluorones (70.0%) and azo group (60.9%). The analysis of decolorization efficiency of the individual dyes revealed the best removal results in case of triphenylmethane brilliant green (average removal 85.7%), followed by fluorone erythrosine (average removal 78.9%), triphenylmethane crystal violet (average removal 65.5%), azo Evans blue (average removal 64.4%), fluorone Bengal rose (average removal 61.0%), and azo Congo red (average removal 57.4%). Obtained results revealed that the dye susceptibility to decolorization depends on the characteristic chemical structure of given dye groups but more important is chemical structure of strictly given dye within the group.

Efficiency of decolorization of different dyes using fungal biomass immobilized on different solid supports.

Different technologies may be used for decolorization of wastewater containing dyes. Among them, biological processes are the most promising because they seem to be environmentally safe. The aim of this study was to determine the efficiency of decolorization of two dyes belonging to different classes (azo and triphenylmethane dyes) by immobilized biomass of strains of fungi (Pleurotus ostreatus - BWPH, Gleophyllum odoratum - DCa and Polyporus picipes - RWP17). Different solid supports were tested for biomass immobilization. The best growth of fungal strains was observed on the washer, brush, grid and sawdust supports. Based on the results of dye adsorption, the brush and the washer were selected for further study. These solid supports adsorbed dyes at a negligible level, while the sawdust adsorbed 82.5% of brilliant green and 19.1% of Evans blue. Immobilization of biomass improved dye removal. Almost complete decolorization of diazo dye Evans blue was reached after 24h in samples of all strains immobilized on the washer. The process was slower when the brush was used for biomass immobilization. Comparable results were reached for brilliant green in samples with biomass of strains BWPH and RWP17. High decolorization effectiveness was reached in samples with dead fungal biomass. Intensive removal of the dyes by biomass immobilized on the washer corresponded to a significant decrease in phytotoxicity and a slight decrease in zootoxicity of the dye solutions. The best decolorization results as well as reduction in toxicity were observed for the strain P. picipes (RWP17).

Efficacy of fungal decolorization of a mixture of dyes belonging to different classes.

Dyes are the most difficult constituents to remove by conventional biological wastewater treatment. Colored wastewater is mainly eliminated by physical and chemical procedures, which are very expensive and have drawbacks. Therefore, the advantage of using biological processes, such as the biotransformation of dyes, is that they may lead to complete mineralization or formation of less toxic products. To prove the possibility of using fungal processes for decolorization and other applications, the analysis of the toxicity of the processes' products is required. The decolorization of the mixture of two dyes from different classes - triphenylmethane brilliant green and azo Evans blue (GB - total concentration 0.08 g/L, proportion 1:1 w/w) - by Pleurotus ostreatus (BWPH and MB), Gloeophyllum odoratum (DCa), RWP17 (Polyporus picipes) and Fusarium oxysporum (G1) was studied. Zootoxicity (Daphnia magna) and phytotoxicity (Lemna minor) changes were estimated at the end of the experiment. The mixture of dyes was significantly removed by all the strains that were tested with 96 h of experimental time. However, differences among strains from the same species (P. ostreatus) were noted. Shaking improved the efficacy and rate of the dye removal. In static samples, the removal of the mixture reached more than 51.9% and in shaken samples, more than 79.2%. Tests using the dead biomass of the fungi only adsorbed up to 37% of the dye mixture (strain BWPH), which suggests that the process with the living biomass involves the biotransformation of the dyes. The best results were reached for the MB strain, which removed 90% of the tested mixture under shaking conditions. Regardless of the efficacy of the dye removal, toxicity decreased from class V to class III in tests with D. magna. Tests with L. minor control samples were classified as class IV, and samples with certain strains were non-toxic. The highest phytotoxicity decrease was noted in shaken samples where the elimination of dye mixture was the best.

Efficacy of fungal decolorization of a mixture of dyes belonging to different classes

Dyes are the most difficult constituents to remove by conventional biological wastewater treatment. Colored wastewater is mainly eliminated by physical and chemical procedures, which are very expensive and have drawbacks. Therefore, the advantage of using biological processes, such as the biotransformation of dyes, is that they may lead to complete mineralization or formation of less toxic products. To prove the possibility of using fungal processes for decolorization and other applications, the analysis of the toxicity of the processes' products is required. The decolorization of the mixture of two dyes from different classes - triphenylmethane brilliant green and azo Evans blue (GB - total concentration 0.08 g/L, proportion 1:1 w/w) - by Pleurotus ostreatus (BWPH and MB), Gloeophyllum odoratum (DCa), RWP17 (Polyporus picipes) and Fusarium oxysporum (G1) was studied. Zootoxicity (Daphnia magna) and phytotoxicity (Lemna minor) changes were estimated at the end of the experiment. The mixture of dyes was significantly removed by all the strains that were tested with 96 h of experimental time. However, differences among strains from the same species (P. ostreatus) were noted. Shaking improved the efficacy and rate of the dye removal. In static samples, the removal of the mixture reached more than 51.9% and in shaken samples, more than 79.2%. Tests using the dead biomass of the fungi only adsorbed up to 37% of the dye mixture (strain BWPH), which suggests that the process with the living biomass involves the biotransformation of the dyes. The best results were reached for the MB strain, which removed 90% of the tested mixture under shaking conditions. Regardless of the efficacy of the dye removal, toxicity decreased from class V to class III in tests with D. magna. Tests with L. minor control samples were classified as class IV, and samples with certain strains were non-toxic. The highest phytotoxicity decrease was noted in shaken samples where the elimination of dye mixture was the best.

Dye Decolourisation Using Two Klebsiella Strains.

This study aimed to decolourise different dyes using two Klebsiella strains (Bz4 and Rz7) in different concentrations and incubation conditions. Azo (Evans blue (EB)) and triphenylmethane (brilliant green (BG)) dyes were used individually and in mixture. The toxicity of the biotransformation products was estimated. Both strains had a significant potential to decolourise the dyes in the fluorone, azo and triphenylmethane classes. The type and concentration of dye affects the decolourisation effectiveness. Differences in the dye removal potential were observed particularly in the main experiment. The best results were obtained for Bz4 in the samples with EB (up to 95.4 %) and dye mixture (up to 99 %) and for Rz7 with BG (100 %). The living and dead biomass of the strain Bz4 highly absorbs the dyes. Significant effect of the process conditions was noticed for both strains. The best results were obtained in static and semistatic samples (89-99 %) for the removal of EB and a mixture of dyes and in static samples (100 %) for BG. The decrease in zootoxicity (from class IV/V) was noticed in all samples with living biomass of the strain Bz4 (to class III/IV) and in samples with single dyes for Rz7 (to class III/IV). The decrease in phytotoxicity (from class III/IV) was noticed for Bz4 in the samples with BG and a mixture (to class III) and for Rz7 in the samples with BG (to class III). The process conditions did not affect the changes in toxicity after the process.

Decolourisation of Different Dyes by two Pseudomonas Strains Under Various Growth Conditions.

The aim of the present study was the decolourisation of mixture of two dyes belonging to different groups by two Pseudomonas fluorescens strains (Sz6 and SDz3). Influence of different incubation conditions on decolourisation effectiveness was evaluated. Dyes used in the experiment were diazo Evans blue (EB) and triphenylmethane brilliant green (BG). Another goal of the experiment was the estimation of toxicity of process by-products. Incubation conditions had a significant influence on the rate of decolourisation. The best results were reached in shaken and semistatic samples (exception Evans blue). After 24 h of experiment in semistatic conditions, BG removal reached up to 95.4 %, EB 72.8 % and dyes mixture 88.9 %. After 120 h, all tested dyes were completely removed. In most cases, dyes were removed faster and better by strain Sz6 than SDz3. At the end of the experiment, in majority of the samples, decrease of phyto- and zootoxicity was observed.

Effectiveness of Dyes Removal by Mixed Fungal Cultures and Toxicity of Their Metabolites.

Decolorization of brilliant green (0.06 g/L), Evans blue (0.15 g/L), and their mixture (total concentration 0.08 g/L, proportion 1:1 w/w) by fungi was studied. Fungal strains [Pleurotus ostreatus (BWPH), Gloeophyllum odoratum (DCa), and Fusarium oxysporum (G1)] were used separately and as a mixture of them. Zootoxicity (Daphnia magna) and phytotoxicity (Lemna minor) changes were estimated after the end of experiment. Mixtures of fungal strains were less effective in decolorization process than the same strains used separately (as a single strains). After 96 h of experiment, living biomass of strain BWPH removed up to 95.5 %; DCa, up to 84.6 %; G1, up to 79.2 % where mixtures BWPH + DCa removed up to 74.3 %; and BWPH + G1, only up to 32.2 % of used dyes. High effectiveness of dyes removal not always corresponded with decrease of toxicity. The highest decrease of zootoxicity and phytotoxicity (from V to III toxicity class or to even nontoxic) was noticed for single strains, while no changes or slight toxicity decrease was noticed in samples with strains mixtures.

Decolourization of Diazo Evans Blue by Two Strains of Pseudomonas fluorescens Isolated from Different Wastewater Treatment Plants.

The use of azo dyes is popular in different branches of industry. Discharge of colourants to surface water cause harmful environmental effects. The aim of the present study was evaluation of effectiveness of diazo Evans blue decolourization by two Pseudomonas strains and estimation of process byproducts toxicity. In static conditions, both tested strains removed more than 85 % of dye after 48 h and completely decolorized samples after 120 h. Agitation had negative impact on Evans blue removal (less than 70 % of dye removed after 120 h). Ecotoxicological effects were different for both studied strains beside comparable decolourization effectiveness. Increase of zootoxicity was noticed for strain Sz6 and decrease from IV to III class was noticed for strain SDz3. Optimization of process conditions for the most promising strain SDz3 should be deeply examined.

Biological Removal of Azo and Triphenylmethane Dyes and Toxicity of Process By-Products.

Increasing environmental pollution is connected with broad applications of dyes and imperfection of dyeing technology. Decolourization of triphenylmethane brilliant green and disazo Evans blue by bacterial and fungal strains and toxicity (phyto- and zootoxicity) of degradation by-products were investigated. Influence of incubation method on dyes removal was evaluated (static, semi-static, shaken). Dead biomass was used for sorption estimation. Toxicity of treated dyes was measured to estimate possible influence on aquatic ecosystems. The zootoxicity test was done with Daphnia magna and phytotoxicity with Lemna minor. Samples were classified according to ACE 89/BE 2/D3 Final Report Commission EC. The best results of removal for all tested strains were reached in shaken samples. In opposite to fungi, bacterial strains decolourized brilliant green more effectively than Evans blue. The most effective bacterial strain was Erwinia spp. (s12) and fungal strains were Polyporus picipes (RWP17) and Pleurotus ostreatus (BWPH and MB). Decolourization of brilliant green was connected with decrease of zootoxicity (D. magna) and phytotoxicity (L. minor). Removal of Evans blue was connected with no changes in zootoxicity and decrease of phytotoxicity in most of samples.