Formation Of Toxic Intermediates Research Articles

Bacterial microcompartments as a next-generation metabolic engineering tool: utilizing nature's solution for confining challenging catabolic pathways.

Advancements in synthetic biology have facilitated the incorporation of heterologous metabolic pathways into various bacterial chassis, leading to the synthesis of targeted bioproducts. However, total output from heterologous production pathways can suffer from low flux, enzyme promiscuity, formation of toxic intermediates, or intermediate loss to competing reactions, which ultimately hinder their full potential. The self-assembling, easy-to-modify, protein-based bacterial microcompartments (BMCs) offer a sophisticated way to overcome these obstacles by acting as an autonomous catalytic module decoupled from the cell's regulatory and metabolic networks. More than a decade of fundamental research on various types of BMCs, particularly structural studies of shells and their self-assembly, the recruitment of enzymes to BMC shell scaffolds, and the involvement of ancillary proteins such as transporters, regulators, and activating enzymes in the integration of BMCs into the cell's metabolism, has significantly moved the field forward. These advances have enabled bioengineers to design synthetic multi-enzyme BMCs to promote ethanol or hydrogen production, increase cellular polyphosphate levels, and convert glycerol to propanediol or formate to pyruvate. These pioneering efforts demonstrate the enormous potential of synthetic BMCs to encapsulate non-native multi-enzyme biochemical pathways for the synthesis of high-value products.

Photoresponsive Activity of the Zn0.94Er0.02Cr0.04O Compound with Hemisphere-like Structure Obtained by Co-Precipitation

In this work, a ZnO hemisphere-like structure co-doped with Er and Cr was obtained by the co-precipitation method for photocatalytic applications. The dopant's effect on the ZnO lattice was investigated using X-ray diffraction, Raman, photoluminescence, UV-Vis and scanning electron microscopy/energy dispersive spectroscopy techniques. The photocatalytic response of the material was analyzed using methylene blue (MB) as the model pollutant under UV irradiation. The wurtzite structure of the Zn0.94Er0.02Cr0.04O compound presented distortions in the lattice due to the difference between the ionic radii of the Cr3+, Er3+ and Zn2+ cations. Oxygen vacancy defects were predominant, and the energy competition of the dopants interfered in the band gap energy of the material. In the photocatalytic test, the MB degradation rate was 42.3%. However, using optimized H2O2 concentration, the dye removal capacity reached 90.1%. Inhibitor tests showed that •OH radicals were the main species involved in MB degradation that occurred without the formation of toxic intermediates, as demonstrated in the ecotoxicity assays in Artemia salina. In short, the co-doping with Er and Cr proved to be an efficient strategy to obtain new materials for environmental remediation.

Experimental investigation for treating ibuprofen and triclosan by biosurfactant from domestic wastewater

The presence of emerging pollutants of pharmaceutical products and personal care products (PPCPs) in the aquatic environment overspreads the threat on living beings. Bioremediation is a promising option for treating wastewater. In the present study, an experimental investigation was carried out to produce a biosurfactant by Pseudomonas aeruginosa (MTCC 1688) for the removal of Ibuprofen (IBU) and Triclosan (TCS) from domestic wastewater. It was performed in three stages. Firstly, the production and optimization of biosurfactant was carried out to arrive at the best combination of crude sunflower oil, sucrose and ammonium bicarbonate (10%: 5.5 g/L: 1 g/L) to yield effective biosurfactant production (crude biosurfactant) and further extended to achieve critical micelle concentration (CMC) formation by dilution (biosurfactant at 10.5%). The stability of the biosurfactant was also confirmed. Biosurfactant showed a reduction in the surface tension to 41 mN/m with a yield concentration of 11.2 g/L. Secondly, its effectiveness was evaluated for the removal of IBU and TCS from the domestic wastewater collected during the dry and rainy seasons. Complete removal of IBU was achieved at 36 h & 6 h and TCS at 6 h & 1 h by crude biosurfactant and biosurfactant at CMC formation for the dry season sample. IBU removal was achieved in 2 h by both crude and biosurfactant at CMC and no TCS was detected in the rainy season sample. Thirdly, biotransformation intermediates of IBU and TCS formed during the application of the biosurfactant and degradation pathways are proposed based on the Liquid Chromatography-Mass Spectrometry (LC-MS) and it indicates that there is no formation of toxic by-products. Based on the results, it is evident that biosurfactant at CMC has performed better for the removal of IBU and TCS than crude biosurfactants without any formation of toxic intermediates. Hence, this study proved to be an eco-friendly, cost-effective and sustainable treatment option for domestic wastewater treatment.

Electrochemical oxidation of Naproxen in aqueous matrices: Elucidating the intermediates’ eco-toxicity, by assessing its degradation pathways via experimental and density functional theory (DFT) approaches

The removal of the non-steroidal anti-inflammatory drug (NSAID) Naproxen (NAX) in water by hydroxyl radicals (•OH) was performed by electrochemical advanced oxidation processes either with Pt or BDD anodes and a 3D carbon felt cathode. The degradation of NAX by (•OH vs electrolysis time) was well fitted to a pseudo-first-order reaction rate kinetic. The detected reaction intermediates (aromatic compounds and carboxylic acids) were experimentally monitored during the process via LC, while density functional theory (DFT) was applied to uncover undetected intermediates, some for the first time in literature. The formation of toxic intermediates with higher toxicity than NAX were identified, such as IMS4b (6-Methoxy-1-[1-(6-methoxynaphthalen-2-yl) ethyl] naphthalen-2-ol), catechol, and glycolic acid. Based on these data, a detailed oxidation pathway of NAX by •OH was proposed. The evolution of solution toxicity indicated that the formed toxic intermediates were subsequently removed during the TOC removal process. Finally, almost complete mineralization of NAX was achieved in simulated urine or wastewater by the electro-Fenton treatment, with an optimized dose of iron as catalyst, showing the EAOPs’ potential to efficiently remove NAX even from challenging matrices. In extension, the strategies developed can be applied to the treatment of other NSAIDs.

Aggregation and the Intrinsic Structural Disorder of Dipeptide Repeat Peptides of C9orf72-Related Amyotrophic Lateral Sclerosis and Frontotemporal Dementia Characterized by NMR.

Dipeptide repeats (DPRs) are known to play important roles in C9ORF72-related amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Studies on DPRs have reported on the kinetics of aggregation, toxicity, and low-resolution morphology of the aggregates of these peptides. While the dipeptide hexa-repeats of Gly-Pro [(GP)6] have been shown to be nonaggregating, Gly-Ala [(GA)6] and Gly-Arg [(GR)6] exhibited the formation of neurotoxic aggregates. However, structural studies of these DPRs have been elusive. In this study, we explored the feasibility of a high-resolution monitoring of a real-time aggregation of these peptides in a solution by using NMR experiments. Although (GP)6 is disordered and nonaggregating, the existence of cis and trans conformations was observed from NMR spectra. It was remarkable that the (GR)6 exhibited the formation of multiple conformations, whereas the hydrophobic and low-soluble (GA)6 aggregated fast in a temperature-dependent manner. These results demonstrate the feasibility of monitoring the minor conformational changes from highly disordered peptides, aggregation kinetics, and the formation of small molecular weight aggregates by solution NMR experiments. The ability to detect cis and trans local isomerizations in (GP)6 is noteworthy and could be valuable to study intrinsically disordered proteins/peptides by NMR. The early detection of minor conformational changes could be valuable in better understanding the mechanistic insights into the formation of toxic intermediates and the development of approaches to inhibit them and, potentially, aid in the development of compounds to treat the devastating C9ORF72-related ALS and FTD diseases.

Degradation of insulin amyloid by antibiotic minocycline and formation of toxic intermediates

Insulin balls, localized insulin amyloids formed at subcutaneous insulin-injection sites in patients with diabetes, cause poor glycemic control owing to impairments in insulin absorption. Our previous study has shown that some insulin balls are cytotoxic, but others are not, implying amyloid polymorphism. Interestingly, the patient with toxic insulin balls had been treated with antibiotic minocycline, suggesting a possible relationship between toxicity of insulin balls and minocycline. However, the direct effect of minocycline on the structure and cytotoxicity of the insulin amyloid is still unclear. Herein, we demonstrated that that minocycline at physiological concentrations induced degradation of insulin amyloids formed from human insulin and insulin drug preparations used for diabetes patients. Interestingly, the process involved the initial appearance of the toxic species, which subsequently changed into less-toxic species. It is also shown that the structure of the toxic species was similar to that of sonicated fragments of human insulin amyloids. Our study shed new light on the clarification of the revelation of insulin balls and the development of the insulin analogs for diabetes therapy.

A model assessment of the importance of direct photolysis in the photo-fate of cephalosporins in surface waters: Possible formation of toxic intermediates

The direct and indirect photodegradation of six cephalosporins was predicted using a photochemical model, on the basis of literature values of photochemical reactivity. Environmental photodegradation would be important in surface water bodies with depth ⩽2–3m, and/or in deeper waters with low values of the dissolved organic carbon (DOC⩽1mgCL−1). The half-life times would range from a few days to a couple of weeks in summertime. In deeper and higher-DOC waters and/or in different seasons, hydrolysis could prevail over photodegradation. The direct photolysis of cephalosporins is environmentally concerning because it is known to produce toxic intermediates. It would be a major pathway for cefazolin, an important one for amoxicillin and cefotaxime and, at pH<6.5, for cefapirin as well. In contrast, direct photolysis would be negligible for cefradine and cefalexin. The DOC values would influence the fraction of photodegradation accounted for by direct photolysis in shallow water, to a different extent depending on the role of sensitisation by the triplet states of chromophoric dissolved organic matter.

Modelling of 1,2-dichloroethane Biodegradation, Stimulated by Constant Electric Field

Biodegradation of 1,2-dichloroethane by the strain Xanthobacter autotrophicusGJ10 passes through the formation of toxic intermediates, like 2-chloroethanol and chloro-acetaldehyde. This adverse effect could be minimized by their deliberate anodic oxidation in constant electric field.The present work presents the mathematical modelling of these processes taking into account the electric field influence on biomass growth and the effect of substrate and product inhibition. The experiments comprises control one (i.e. without application of electric field) and ones at electric field application at different anode potentials within 0.753 and 0.903 V vs. the potential of the normal hydrogen electrode. The effect of the electric field on microbial specific growth rate and on the rate of microbial decay was estimated quantitatively in terms of the corresponding rate constants. Additionally, the accumulation and the decay of the intermediates were taken into account.The results of the mathematical modelling showed that the effect of the electric field is strongly positive on the microbial growth and best kinetic parameters were observed at certain anode potential, i.e. at 0.803 V/S.H.E. The maximum specific growth rate in this case is 0.90 h\textsuperscript{-1}, i.e. three times higher than for the reference experiment. The rate constant of decay of the intermediates is almost four times higher than for the reference experiment. [1] D.B. Janssen, A. Scheper, L. Dijkhuizen, B. Witholt, Degradation of halogenated aliphatic compounds by Xanthobacter autotrophicus GJ10, Appl. Environ. Microbiol. 1985, 49, 673--677.[2] A. Mileva, Ts. Sapundzhiev, V. Beschkov, Modeling 1,2-dichloroethane biodegradation by Klebsiella oxytoca VA 8391 immobilized on granulated activated carbon, Bioprocess Biosyst Eng. 2008, 31, 75--85.

Combining efficiently catalytic hydrodechlorination and wet peroxide oxidation (HDC–CWPO) for the abatement of organochlorinated water pollutants

The combination of catalytic hydrodechlorination (HDC) and wet peroxide oxidation (CWPO) has been investigated for the breakdown of chlorophenols (CPs). The process was conducted as a sequential two-step reaction involving 30min HDC followed by CWPO. Direct application of CWPO to CPs has shown important drawbacks derived from the formation of toxic intermediates under substoichiometric H2O2 doses as well as from the fact that the reaction rate decreases significantly as the number of chlorine atoms in the molecule increases (pseudo-first order rate constant values of 20.6×10−3, 4.6×10−3 and 1.4×10−3Lgcat−1min−1 were obtained for 4-CP, 2,4-DCP and 2,4,6-TCP disappearance, respectively). On the opposite, HDC removes chlorine from CPs under mild conditions at a frankly high rate which is almost unaffected by the chlorine content of the starting chlorophenol (values of 0.19, 0.17 and 0.15Lgcat−1min−1 for the pseudo-first order rate constants were obtained with 4-CP, 2,4-DCP and 2,4,6-TCP, respectively). Thus, HDC can be used as a short-time pretreatment (≈30min) associated to a subsequent CWPO step in order to achieve a high mineralization in significantly lower reaction times. The sequential combination of HDC and CWPO as a two-step reaction was tested with different CPs using monometallic catalysts for each step (Pd/γ-Al2O3 and Fe/γ-Al2O3, respectively). This combined process allowed improving TOC reduction from 30% (CWPO alone) to 70% after 120min reaction time. Moreover, it allowed reaching a substantial reduction of ecotoxicity (>95%) with the three chlorophenols tested.

Characterisation of acute toxicity, genotoxicity and oxidative stress posed by textile effluent on zebrafish

Textile industries are important sources of toxic discharges and contribute enormously to water deterioration, while little attention has been paid to the toxicity of textile effluents in discharge regulation. Bioassays with zebrafish were employed to evaluate the toxicity of wastewater samples collected from different stages at a textile factory and sewage treatment plants (STPs). Physico-chemical parameters, acute toxicity, genotoxicity and oxidative stress biomarkers were analyzed. The wastewater samples from bleaching, rinsing and soaping of the textile factory exhibited high acute toxicity and genotoxicity. The coexisting components of dye compounds, as assistants and oxidants, seemed to cause some effect on the toxic response. After treatment employing the anoxic-oxic (A/O) process in STPs, the color and the chemical oxygen demand (COD) were reduced by 40% and 84%, respectively, falling within the criteria of the Chinese Sewage Discharge Standard. In contrast, increases in acute toxicity and genotoxicity were observed in the anaerobic tank, indicating the formation of toxic intermediates. The genotoxicity of the effluent of the STP was not significantly different from that of the influent, suggesting the wastewater treatment processes were not effective in removing the genotoxicity of the dye wastewater. Results indicated that the effluent contains pro-oxidants since the activities of glutathione (GSH), malondialdehyde (MDA), and total anti-oxidation capacity (T-AOC) were all elevated. In addition, decreases in superoxide dismutase (SOD) and glutathione-S transferase (GST) activities observed can be interpreted as a cytotoxicity sign due to an over-production of reactive oxygen species (ROS). The results of the present study suggest that the STPs were not capable of reducing the toxicity of wastewater sufficiently. Further treatment is needed to remove the potential risks posed by textile effluent to ecosystems and human health, and employing a toxicity index is necessary for discharge regulation.

Challenges and recommendations for conducting epidemiological studies in the field of epilepsy pharmacogenetics

Epilepsy is one of the most prevalent neurological disorders, afflicting approximately 50 million Indians. Owing to affordability and easy availability, use of first-generation antiepileptic drugs (AEDs) is heavily encouraged for the treatment of epilepsy in resource-limited countries such as India. Although first-generation AEDs are at par with second-generation AEDs in terms of efficacy, adverse drug reactions (ADRs) are quite common with them. This could be attributed to the inferior pharmacokinetic parameters such as nonlinear metabolism, narrow therapeutic index and formation of toxic intermediates. In addition, epilepsy patients may differ in the pharmacokinetic and pharmacodynamic profiles, with about 1/3rd of the population failing to respond to treatment. A proportion of this interindividual variability in response may be explained by genetic heterogeneity in the activity and expression of the network of proteins such as metabolizing enzymes, transporters and targets of AEDs. Over the last two decades, a considerable effort has been made by the scientific community for unraveling this genetic basis of variable response to AEDs. However, there have been inconsistencies in such genetic association studies conducted across different territories of the world. There could be several reasons underlying the poor replicability of these studies, mainly nonuniform phenotypic definitions, poor sample size and interethnic variability. In the present review article, we provide an overview of heterogeneity in study designs for conducting pharmacogenetic studies. In addition, critical recommendations required for overcoming such challenges imposed by pharmacogenetic epidemiological studies have been briefly discussed.

Photo-induced transformation of hexaconazole and dimethomorph over TiO 2 suspension

The photo-induced transformation in aqueous solution of hexaconazole and dimethomorph over irradiated titanium dioxide was studied. The investigation involved monitoring pesticides decomposition, identifying intermediate compounds, assessing mineralization, and evaluating toxicity of pesticides derivatives. HPLC/UV and HPLC/MS were used to follow the disappearance of the initial pesticides and the formation of intermediate products, while the acute toxicity was evaluated by using the Vibrio fischeri luminescent bacteria assay. Hexaconazole photocatalytic transformation proceeds through the formation of highly persistent compounds. The formation of cyanuric acid, a non-toxic compound refractory to photocatalytic treatment, was recognized. Conversely, the toxicity assays prove that neither hexaconazole nor its intermediates exhibit acute toxicity. Dimethomorph under photocatalytic treatment is completely mineralized within 14 h of irradiation. However, its transformation proceeds through the formation of toxic intermediates. A correlation exists between the evolution of the intermediate compounds and the toxicity profile, as the highest toxicity is measured when the intermediates with lower EC50 (hydroquinone and 4-chlorophenol) are formed.

Cytotoxicity of the dental composite component TEGDMA and selected metabolic by-products in human pulmonary cells

Objectives The comonomer triethyleneglycoldimethacrylate (TEGDMA) is a commonly used constituent of resin-based dental materials. Upon placement, light-cured dental polymers may release a wide spectrum of residual compounds due to incomplete monomer-conversion during polymerization. Apart from liberating unreacted monomers, additional compound release might occur due to mechanical wear and enzymatic degradation on the salivary surface of resin fillings. Following delivery into the local bio phase, leached compounds may encounter a variety of different enzymes, which might be present in their oral or systemic environment. Metabolic by-products formerly associated with TEGDMA-degradation include triethylene glycol (TEG), methacrylic acid (MA), 2,3-epoxymethacrylic acid (2,3-EMA), and formaldehyde. Methods Cytotoxicitiy of TEGDMA-derived intermediates was measured as mitochondrial dehydrogenase activity assessed by colorimetric measurement of formazan formation as a cleavage-product from the tetrazolium salt XTT by metabolically active A549 cells. EC 50-values were calculated by using curve fitting software (GraphPad Prism). Results The following EC 50-values (mmol/L) (95% confidence interval) were obtained: 2,3-EMA 1.65 (1.28–2.13), TEGDMA 1.83 (1.46–2.30), MA 4.91 (4.22–5.71), and paraformaldehyde (PFA) 5.48 (4.56–6.58). For TEG no cytotoxic effects up to a concentration of 10 mM could be found. Significance The epoxy compound 2,3-EMA induced comparable toxic effects as the raw comonomer TEGDMA. It is therefore concluded that the formation of toxic intermediates might significantly contribute to TEGDMA-induced cytotoxicity in human pulmonary cells.

Formation of Toxic Intermediates Upon the Photocatalytic Degradation of the Pesticide Diuron

The photocatalyzed degradation of the pesticide N'-(3,4-Dichlorophenyl)-N,N-dimethyl-urea (Diuron) has been studied in aqueous suspensions of titanium dioxide by monotoring the change in substrate concentration and the depletion of the Total Organic Carbon (TOC) content, respectively, as a function of irradiation time. The degradation kinetics of the model compound were studied under several different conditions such as pH, catalyst concentration, different types of TiO 2 and in the presence of electron acceptors (H 2 O 2 , KBrO 3 and (NH 4 ) 2 S 2 O 8 ) besides molecular oxygen. The photonic efficiency for the depletion of Diuron itself as well as for the total mineralization is strongly influenced by the reaction pH. The choice of the TiO 2 photoctalyst, i.e., Degussa P25 or Sachtleben Hombikat UV 100, or the variation of the photocatalyst concentration between 0.2 and 2.0 g L -1 had no pronounced effect on the degradation kinetics of Diuron. All additives showed a beneficial effect on the degradation of the model compound. While the addition of hydrogen peroxide or persulfate results only in a slight increase, the addition of bromate ions improved the photonic efficiency by a factor about 3. However, for the total Diuron mineralization only the addition of bromate ions leads to an enhancement while in the presence of the other investigated additives even a slight decrease in the mineralization rate was observed. Toxicity tests (LUMIStox-Test) showed a dramatic increase of bacteria growth inhibition from an initial value of 30% to almost 100 % within the first two hours of irradiation indicating the generation of highly toxic intermediates. The latter are completely degraded upon prolonged irradiation as evidenced by a decrease to almost 0 % inhibition of the bacteria activity.

Whole-Cell Kinetics of Trichloroethylene Degradation by Phenol Hydroxylase in a Ralstonia eutropha JMP134 Derivative

The rate, progress, and limits of trichloroethylene (TCE) degradation by Ralstonia eutropha AEK301/pYK3021 whole cells were examined in the absence of aromatic induction. At TCE concentrations up to 800 &mgr;M, degradation rates were sustained until TCE was no longer detectable. The Ks and Vmax for TCE degradation by AEK301/pYK3021 whole cells were determined to be 630 &mgr;M and 22.6 nmol/min/mg of total protein, respectively. The sustained linear rates of TCE degradation by AEK301/pYK3021 up to a concentration of 800 &mgr;M TCE suggest that solvent effects are limited during the degradation of TCE and that this construct is little affected by the formation of toxic intermediates at the TCE levels and assay duration tested. TCE degradation by this strain is subject to carbon catabolite repression.

Dimethylsulfoxide as modifier of the organospecific mutagenicity of metronidazole in mice

Mutagenicity and carcinogenicity of metronidazole (MT) are imputable to the formation of toxic intermediates, which include radical forms derived from the nitroreductive process. Since dimethylsulfoxide (DMSO), the "universal" solvent, can quench free radicals in vitro, it was suggested that DMSO might protect by scavenging free radical generation in vivo. This study wanted to evaluate if DMSO (given concomitantly or prophylactically) protects against the organospecific mutagenicity of MT in vivo by means of the intrasanguineous host-mediated assay. DMSO used as solvent showed a 20%-30% reduction in the mutation frequencies by MT. Prophylactic administration of DMSO for 3 d caused a suppression of the organospecific mutagenicity. However, some increases in the spontaneous mutation frequency and enhancement of MT mutagenicity in kidney were observed. The protective effect was paralleled by a decrease in NADPH cytochrome c (P450) reductase in liver, kidney, and to a lesser extent in lung microsomes from pretreated mice. Inhibition of mutagenic activity might be related to scavenging of radical species as supported by the lack of tissue specificity and no appreciable changes in specific enzyme activity. However, changes in reductase content in prophylactically pretreated mice can affect the quantitative biotransformation of MT to the proximal mutagen contributing to the observed suppression in mutation frequencies.