Indirect Photodegradation of Sulfamethoxazole and Trimethoprim by Hydroxyl Radicals in Aquatic Environment: Mechanisms, Transformation Products and Eco-Toxicity Evaluation.

The bacteriostatic antibiotics, sulfamethoxazole (SMX) and trimethoprim (TMP), have frequently been found in wastewater and surface water, which raises the concerns about their ecotoxicological effects. The indirect photochemical transformation has been proven to be an efficient way to degrade SMX and TMP. In this study, the reaction mechanisms of the degradation by SMX and TMF by OH radicals were investigated by theoretical calculations. Corresponding rate constants were determined and the eco-toxicity of SMX and TMP and its degradations products were predicted using theoretical models. The results indicate that the most favorable pathways for the transformation of SMX and TMP are both •OH-addition reaction of benzene ring site with lowest Gibbs free energy barriers (6.86 and 6.21 kcal mol-1). It was found that the overall reaction rate constants of •OH-initial reaction of SMX and TMP are 1.28 × 108 M-1 s-1 and 6.21 × 108 M-1 s-1 at 298 K, respectively. When comparing the eco-toxicity of transformation products with parent SMX and TMP, it can be concluded that the acute and chronic toxicities of the degraded products are reduced, but some products remain harmful for organisms, especially for daphnid (toxic or very toxic level). This study can give greater insight into the degradation of SMX and TMP by •OH through theoretical calculations in aquatic environment.

Sonochemical degradation of trimethoprim in water matrices: Effect of operating conditions, identification of transformation products and toxicity assessment.

The sonochemical degradation of trimethoprim (TMP), a widely used antibiotic, in various water matrices was investigated. The effect of several parameters, such as initial TMP concentration (0.5-3 mg/L), actual power density (20-60 W/L), initial solution pH (3-10), inorganic ions, humic acid and water matrix on degradation kinetics was examined. The pseudo-first order degradation rate of TMP was found to increase with increasing power density and decreasing pH, water complexity (ultrapure water > bottled water > secondary wastewater) and initial TMP concentration. TMP degradation is accompanied by the formation of several transformation products (TPs) as evidenced by LC-QToF-MS analysis. Nine such TPs were successfully identified and their time-trend profiles during degradation were followed. An in silico toxicity evaluation was performed showing that several TPs could potentially be more toxic than the parent compound towards Daphnia magna, Pimephales promelas and Pseudokirchneriella subcapitata.

Cranial neural tube defect after trimethoprim exposure.

OBJECTIVES: The Neural Tube Defects Research Group of University of Malaya was approached to analyze a tablet named TELSE, which may have resulted in a baby born with central nervous system malformation at the University of Malaya Medical Centre. In this animal experimental study, we investigated the content of TELSE and exposure of its contents that resulted in failure of primary neurulation. RESULTS: Liquid Chromatography Tandem Mass spectrophotometry analysis of the TELSE tablet confirmed the presence of trimethoprim as the active compound. The TELSE tablet-treated females produced significant numbers of embryos with exencephaly (n = 8, 36.4%, *P < 0.0001), in all litters. The TELSE tablet-treated females subsequently given folic acid did not result in pregnancies despite there being evidence of possible resorption. Furthermore, after multiple rounds of mating which did not yield viable pregnancies, eventually, 2 embryos with exencephaly were harvested in a litter of 6 at 0.05% w/v pure trimethoprim once. The use of trimethoprim, a folic acid antagonist, peri-conceptionally increased the risk of exencephaly in the mouse.

Dithiothreitol (DTT) rescues mitochondria from nitrofurantoin-induced mitotoxicity in rat.

Nitrofurantoin (N-(5-nitro-2-furfurylidine) 1-amino-hydantoine; NIT) is mainly used for the treatment of acute urinary tract infections. However, its administration can be associated with liver failure or cirrhosis. The aim of this study was to determine whether NIT is a mitochondrial toxicant, if so, what mechanism(s) is involved. The rat liver mitochondria were isolated and treated with different doses of NIT alone or in combination with a reagent of choice for protecting thiol groups, dithiothreitol (DTT). Several mitochondrial parameters, including succinate dehydrogenase activity (also called 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide assay), lipid peroxidation, superoxide dismutase activity, Reduced glutathione (GSH), and oxidized glutathione (GSSG), and GSSG (oxidized glutathione) levels were determined. The results from this study showed that simultaneous treatment of mitochondria with NIT and DTT significantly reduces the toxicity. Here, we provide evidence that mitochondrial dysfunction followed by depletion of reduced glutathione can be reversed by DTT administration.

Nitrofurantoin-induced pulmonary toxicity: A case report and review of the literature.

This paper describes a case of lung injury attributed to the use of Nitrofurantoin and a review of the relevant literature. An 88-year-old woman was admitted to the floor for the evaluation of recent symptoms of dyspnea, fatigue and productive cough. She was initiated on nitrofurantoin 300 mg per day for the treatment of a urinary tract infection 3 days earlier. Upon examination, chest auscultation revealed bilateral inspiratory crackles. Chest radiograph showed bilateral airspace and interstitial infiltrates. Laboratory studies revealed an elevated white blood cell count of 13,500/µL (reference range = 5200-12,400/µL) and blood eosinophilia (10%, reference range: 0-7%). Using clinical judgment and the algorithm of Naranjo, it was determined that nitrofurantoin use was the probable cause of the patient's lung injury. Symptomatic improvement was observed shortly after the drug was discontinued. A review of information from several European and North American pharmacovigilance databases (through June 2014) identified several reports of suspected nitrofurantoin-induced toxicity, including reports of acute toxicity reactions, which were related in many ways to the case we are reporting here.

Superiority of solar Fenton oxidation over TiO2 photocatalysis for the degradation of trimethoprim in secondary treated effluents.

The overall aim of this work was to examine the degradation of trimethoprim (TMP), which is an antibacterial agent, during the application of two advanced oxidation process (AOP) systems in secondary treated domestic effluents. The homogeneous solar Fenton process (hv/Fe(2+)/H2O2) and heterogeneous photocatalysis with titanium dioxide (TiO2) suspensions were tested. It was found that the degradation of TMP depends on several parameters such as the amount of iron salt and H2O2, concentration of TiO2, pH of solution, solar irradiation, temperature and initial substrate concentration. The optimum dosages of Fe(2+) and H2O2 for homogeneous ([Fe(2+)] = 5 mg L(-1), [H2O2] = 3.062 mmol L(-1)) and TiO2 ([TiO2] = 3 g L(-1)) for heterogeneous photocatalysis were established. The study indicated that the degradation of TMP during the solar Fenton process is described by a pseudo-first-order reaction and the substrate degradation during the heterogeneous photocatalysis by the Langmuir-Hinshelwood kinetics. The toxicity of the treated samples was evaluated using a Daphnia magna bioassay and was finally decreased by both processes. The results indicated that solar Fenton is more effective than the solar TiO2 process, yielding complete degradation of the examined substrate within 30 min of illumination and dissolved organic carbon (DOC) reduction of about 44% whereas the respective values for the TiO2 process were ∼70% degradation of TMP within 120 min of treatment and 13% DOC removal.

Evaluation of cytogenetic and DNA damage induced by the antibacterial drug, trimethoprim.

Trimethoprim, a widely used antimicrobial drug was tested for its effect on the level of nuclear DNA damage in cultured peripheral blood lymphocytes in terms of chromosome and DNA alterations. The extent of cytogenetic damage, expressed as chromosome breakage and chromosome loss, was evaluated employing the cytokinesis block micronucleus method (CBMN) in cultured peripheral blood lymphocytes coupled with fluorescence in situ hybridization (FISH) using a digoxigenin-labelled alphoid DNA probe specific for the centromere of all human chromosomes. The DNA breakage level was evaluated by the Comet assay. Cultures were set up by using blood samples from two healthy donors. A range of concentrations of the test agent (from 1 to 100 microg/ml) was used for the micronuclei (MN) frequency and cytogenetic origin of MN. For the Comet assay the range of doses used was from 0.5 to 150 microg/ml. From the results obtained it appears that this antifolic agent has a significant clastogenic potential, as detected by a dose-dependent increase of the incidence of C-MN and significantly greater than control levels at the highest concentrations tested (25,100 microg/ml). In addition, the results obtained in the Comet assay also show that trimethoprim induces a dose-dependent increase in the level of DNA breakage, this increase attaining statistical significance at the highest concentrations tested (25, 100, 150 microg/ml), which would confirm its genotoxicity.

Liver microsomal biotransformation of nitro-aryl drugs: mechanism for potential oxidative stress induction.

Toxic effects of several nitro-aryl drugs are attributed to the nitro-reduction that may be suffered in vivo, a reaction that may be catalysed by different reductases. One of these enzymes is NADPH-cytochrome P450 reductase, which belongs to the cytochrome P450 oxidative system mainly localized in the endoplasmic reticulum of the hepatic cell. This system is responsible for the biotransformation of oxidative lipophilic compounds, so that oxidative and reductive metabolic pathways of lipophilic nitro-aryl drugs can take place simultaneously. Because of the affinity of nitro-aryl drugs (xenobiotics) for the endoplasmic reticulum, we propose this subcellular organelle as a good biological system for investigating the toxicity induced by the biotransformation of these or another compounds. In this work we used rat liver microsomes to assess the oxidative stress induced by nitro-aryl drug biotransformation. Incubation of microsomes of rat liver with nifurtimox and nitrofurantoin in the presence of NADPH induced lipoperoxidation, UDP-glucuronyltransferase activation and an increase in the basal microsomal oxygen consumption. Nitro-aryl-1,4-dihydropyridines did not elicit these prooxidant effects; furthermore, they inhibited lipoperoxidation and oxygen consumption induced by Fe3+/ascorbate. Nifurtimox and nitrofurantoin modified the maximum absorption of cytochrome P450 oxidase and inhibited p-nitroanisole O-demethylation, an oxidative reaction catalysed by the cytochrome P450 system, signifying that oxidation may proceed in a similar way to that described for nitro-aryl-1,4-dihydropyridines. Thus the balance between lipophilic nitro-aryl drug oxidation and reduction may be involved in the potential oxidative stress induced by biotransformation.

In vitro studies on DNA-photosensitization by different drug stereoisomers.

The possible stereoselectivity in DNA-photosensitization by carprofen (a NSAID drug) and ofloxacin (a fluoroquinolone agent) was investigated. The different drug stereoisomers or racemic mixtures were UVA-irradiated and the relaxation of the supercoiled circular pBR322 quantified by electrophoresis. Formation of single strand breaks was compared for each group of compounds. Moreover a mechanistic study by means of repair enzymes: T4 endonuclease V (specific of cyclobutane pyrimidine dimers), E. coli endonuclease III (revealing oxidized pyrimidines) and E. coli Formamidopyrimidine-DNA glycosylase (revealing oxidized purines) provided further insights into a possible stereoselectivity of the different reaction pathways in drug photosensitized-DNA damage. Ofloxacin and levofloxacin (its S stereoisomer) were responsible of single strand breaks formation as well as oxidation of pyrimidine and purine bases. No pyrimidine dimers were observed. Racemic, R and S stereoisomers of carprofen were less efficient than ofloxacin in DNA single strand breaks formation and did not induce enzyme-sensitive sites. The photoproducts distribution of drug-photosensitized reactions of 2'-deoxyguanosine and thymidine were established by HPLC as fingerprints for assignment of the DNA-photosensitization mechanism. Both Type I and Type II mechanisms were assigned to nucleoside-photosensitization by ofloxacin and levofloxacin. In the case of carprofen, a weak nucleoside degradation was obtained. The data suggest that levofloxacin, the (S) stereoisomer, might be slightly more efficient than racemic ofloxacin. In the case of carprofen the (S) isomer appears to be somewhat less active than its (R) enantiomer. However, due to the small differences found, the possible stereoselectivity has to be confirmed by future studies.

Development and characterization of an infection inhibiting urinary catheter.

Catheter associated bacturia is common in hospitals and nursing homes. The objective of this study was to develop an infection inhibiting urinary catheter for prolonged use. Methods were established to add chlorhexidine digluconate (CHG) to a silicone elastomer and to compression mold the material to form a urinary catheter. CHG was randomly dispersed in the elastomer to be released through elution. Samples of the material, with CHG concentrations ranging from 1 to 4% by weight, were tested for in vitro release characteristics over a 28 day period and for in vivo toxicity over a 7 day period. Release profiles followed a common pattern for each concentration: an initial peak during the first 24 hours was followed by a subsequent decline. CHG amounts released into the saline medium were directly related to the CHG concentration of the samples; 4% samples released the largest amounts and 1% samples released the least amounts. Both 3% and 4% CHG by weight samples released measurable amounts of CHG throughout the entire observation period, whereas 1% CHG by weight samples were depleted after 9 days, and 2% CHG by weight samples were depleted after 19 days. No samples were found to be toxic during in vivo evaluations. These studies suggest that CHG bearing silicone rubber urinary catheters could resist surface colonization and infection for extended periods without toxicity.

Lack of effects of oxolinic acid on spermatogenesis in young adult and aged Wistar rats.

Prolonged treatment with oxolinic acid is known to elevate serum luteinizing hormone (LH) levels, resulting in induction of Leydig cell tumors in rats. In a carcinogenicity study of the compound, tubular atrophy of the testis was also increased, suggesting that oxolinic acid might affect spermatogenesis. The present study was therefore performed using rats of different ages with a particular focus on seminiferous tubule alteration and its relation to Leydig cell proliferation. Young adult (7 weeks of age) and aged (52 weeks of age) males of the Wistar strain were administered oxolinic acid at dietary concentrations of 0 (basal diet), 300, 1000 or 3000 ppm for 4 (all groups), 13 (0 and 3000 ppm groups), 26 (0 and 3000 ppm groups), or 52 weeks (0 and 3000 ppm groups of aged rats). Serum LH levels were elevated in both young adult and aged animals treated with 3000 ppm at most examined time points. While testosterone levels were also increased at the early time points in young adult, this was not the case in older animals. Elevation of the incidences of foci and/or focal hyperplasia of Leydig cells was noted but was only slight limited to aged rats treated with 3000 ppm after 26 weeks. Furthermore, it did not appear to be related to seminiferous tubular alteration. No treatment-related histopathological abnormalities could be detected in any treatment group, and morphometrical stage analysis of spermatogenesis conducted for the control and 3000 ppm-treated groups demonstrated no lesions. These results provide strong evidence that prolonged oxolinic treatment does not directly induce testicular toxicity or altered spermatogenesis in either young adult or aged rats, except for slight increase of Leydig cell proliferative lesions caused by elevated serum LH levels. Aged rats might have higher sensitivity than young adults to the effects of oxolinic acid on proliferative lesions of Leydig cells.

Polymer-coated microparticles for the sustained release of nitrofurantoin.

Suspensions of nitrofurantoin (NTF) microparticles for controlled release were investigated in this study. The microparticles were enteric coated with various combinations of the two polymers, cellulose acetate phthalate/cellulose acetate butyrate (CAP/CAB) by a modified solvent evaporation method. Ratios of NTF to the two polymers (NTF/CAP/CAB) ranged from 1.0:1.6:0.4,1.0:1.0:1.0,1.0: 0.4:1.6 to 1.0:0.0:2.0. The encapsulation efficiency, percentage yield, determined by comparing the final mass of the microparticles with the initial mass of the ingredients used, distribution of particle size and the in-vitro dissolution profiles of the microparticles were determined. Based on light photographs for the evaluation of the microparticle morphology, the drug crystals appeared to be encapsulated sufficiently by the enteric polymers. In our study, the microparticles enteric coated with CAP/CAB in the ratio of 0.4:1.6 displayed the most satisfactory in-vitro release profile (reduced release in the simulated gastric fluid and sustained release in the simulated intestinal fluid). Thus, microparticles with NTF/CAP/CAB in the ratio of 1.0:0.4:1.6 were formulated into a suspension for further bioavailability and ulcerogenicity studies in Sprague-Dawley rats, with the suspension of NTF crystals as a control. The bioavailability study was carried out in eight rats fed with either the free NTF or the corresponding microparticles in a cross-over design. The ulcerogenicity study was carried out in three groups of six rats each: one group received no drug treatment; the control group was treated with free NTF; and the third group was treated with enteric-coated NTF microparticles. The bioavailability of NTF from the microparticles was comparable with the control. More importantly, there was notably less ulceration of the gastric mucosa observed after dosing with the microparticle suspension compared with that after the administration of the control suspension.

Environmental risk assessment of antibiotics: comparison of mecillinam, trimethoprim and ciprofloxacin.

The effects of mecillinam, trimethoprim and ciprofloxacin, antibiotics used in the treatment of urinary tract infections, on the aquatic environment were assessed. Mecillinam and ciprofloxacin were both readily biodegradable (primary degradation) in activated sludge, whereas trimethoprim persisted. The toxicity of these antibiotics towards sludge bacteria, a green alga, a cyanobacterium, a crustacean and a fish were investigated; both mecillinam and ciprofloxacin were highly toxic to the cyanobacterium Microcystis aeruginosa (EC50 in the range 5-60 microg/L). Risk characterization for the aquatic environment was performed for the three compounds by calculating the predicted environmental concentration (PEC) and the predicted no-effects concentration (PNEC). A PEC/PNEC ratio of <1 indicates that, with the present pattern of use, no environmental risk is expected. PEC/PNEC ratios of <1 for present usage in Europe were found for mecillinam and trimethoprim whereas a PEC/PNEC ratio >1 was found for ciprofloxacin.

Genotoxic evaluation of the antimicrobial drug, trimethoprim, in cultured human lymphocytes.

The antimicrobial drug, trimethoprim, was evaluated for genotoxicity in human peripheral blood lymphocyte cultures set-up from two healthy donors. Sister-chromatid exchanges (SCE) and micronuclei (MN) were scored as genetic endpoints. The treatment was done using different trimethoprim concentrations ranging from 1 to 100 microg/ml. From our results, we can conclude that this drug is able to induce both cytotoxic and moderate genotoxic effects, as revealed by the increases seen in SCE and MN frequencies in cultures from the two donors and, at least, at one of the concentrations tested.

Evaluation of certain veterinary drug residues in food. Forty-eighth report of the Joint FAO/WHO Expert Committee on Food Additives.

This report presents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of residues of certain veterinary drugs in foods and to recommend maximum levels for such residues in food. The first part of the report considers standards for the performance of studies, residues at the injection site, and several initiatives to promote transparency of the process for setting Maximum Residue Limits (MRLs). A summary follows of the Committee's evaluations of toxicological and residue data on a variety of veterinary drugs: two anthelminthic agents (moxidectin and tiabendazole), eight antimicrobial agents (ceftiofur, danofloxacin, dihydrostreptomycin, streptomycin, enrofloxacin, flumequine, gentamicin and spiramycin), one glucocorticosteroid (dexamethasone), and two insecticides (cyfluthrin and fluazuron). Annexed to the report are a summary of the Committee's recommendations on these drugs, including Acceptable Daily Intakes and MRL's and further toxicological studies and other information required.

Furazolidone toxicosis in female Japanese quail (Coturnix coturnix japonica): pathomorphological changes in reproductive tract and reversibility of the induced changes.

Female Japanese quail (Coturnix coturnix Japonica) placed in 3 equal groups were given 600, 400 or 0 mg furazolidone (Fz)/kg feed for 4 w and then withdrawn for another 4 w. Another (pair-fed) group of same size was given basal feed as much as was consumed by the quail fed 600 mg Fz/kg feed. Fz feeding decreased body weight, feed intake and egg production. Ovaries of the Fz-fed quail decreased in weight and size and were studded with small follicles. Magnum, isthmus and uterus in Fz-fed groups had decreased area, height and number of mucosal folds compared with the control group. Microscopically, in Fz-fed groups, the mucosal glands in magnum and isthmus had decreased cell height with centrally located nuclei and foamy cytoplasm. In the 600 mg Fz-fed group, some birds had atrophy of the glandular tissue in the mucosa and infiltration of mononuclear cells and fibroblasts. Upon cessation of the Fz feeding, all parameters reversed gradually and became non-significantly different from control quail. These observations suggested that Fz-induced changes in mature female quail were reversible.

Roles of DT diaphorase in the genotoxicity of nitroaromatic compounds in human and fish cell lines.

The genotoxicity of nitroaromatic compounds was examined in two cultured cell lines, namely, a human hepatoma cell line, HepG2, and a brown bullhead fibroblast cell line, BB. Furthermore, the role of the quinone-reducing enzyme DT diaphorase [NAD(P)H:(quinone acceptor) oxidoreductase] was examined with respect to its influence on the genotoxic effects of model nitroaromatic pollutants. The nitroreductive characteristics of these two cell lines were examined using an acetylated cytochrome c reduction assay for enzymatic nitroreductase activity. Subsequently, the influence of DT diaphorase on the genotoxicity of two model nitroaromatics, 4-nitroquinoline 1-oxide (4NQ) and nitrofurantoin (NF), revealed that DT diaphorase was the predominant 4NQ reductase in cytosols of both cell lines, but played a lesser role in NF reduction in both species. Despite these interspecific similarities, results revealed marked qualitative differences between the two species in terms of the influence of DT diaphorase on quinone-mediated genotoxicity. When pretreated with the DT diaphorase inhibitor dicoumarol, HepG2 cells exhibited an exacerbation of genotoxicity in the presence of 4NQ, indicating a protective influence of the enzyme. In contrast, 4NQ genotoxicity in BB cells was reduced in the presence of dicoumarol, indicating a deleterious effect of DT diaphorase activity. Conversely, dicoumarol pretreatment was moderately protective against NF-mediated genotoxicity in HepG2 cells but exacerbated NF toxicity in BB cells. This study illustrates the manner in which functionally analogous enzymes may have markedly distinct influences on xenobiotic toxicity in different cellular systems.

Mitigation of nitrofurantoin-induced toxicity in the perfused rat lung.

1. Nitrofurantoin is an antimicrobial agent which produces pulmonary toxicity via the redox cycling of the nitro group and its radical anion. This futile cycling triggers a complex series of events known collectively as oxidative stress. 2. In the isolated perfused rat lung, nitrofurantoin induced a decrease in tissue levels of glutathione but not protein thiols by the end of the 180 min experiment. There was no decline in tissue levels of angiotensin converting enzyme (a marker of cell disruption). However, edema was extensive as monitored in real time by weight gain (2.71 +/- 0.56 g vs 0.63 +/- 0.53 g in control, P < 0.05, n = 4) and lung mechanical functioning. The edema was matched by an increase in lavage proteins (85 +/- 15 mg vs 16 +/- 9 mg in controls, P < 0.05, n = 4). Electron microscopic examination of tissue indicated that the endothelial cells were detached from the basement membrane which would account for the edema. 3. Co-infusion of penicillamine, N-acetylcysteine or N-(2-mercaptopropionyl)-glycine which can protect tissue from oxidative stress failed to mitigate NFT-induced edema. Allopurinol, an inhibitor of xanthine oxidase and a metal chelator, significantly decreased weight gain but did not prevent the loss of glutathione. These results suggested that allopurinol was not blocking metabolic activation of NFT by xanthine oxidase but scavenging metal cations which can initiate and/or propagate the oxidative stress cascade. 4. We concluded that, in the isolated perfused rat lung, the classic pathway of oxidative stress induced by NFT is interrupted at the stage of GSH loss. These experiments demonstrated that organ function was compromised more than the individual cells. They also suggested that allopurinol may prove beneficial in modulating NFT pulmonary toxicity.