Bioavailability of flumequine and diclofenac in mice exposed to a metal-drug chemical cocktail. Evaluation of the protective role of selenium.

BACKGROUND AND PURPOSE: Organisms, including humans, are subjected to the simultaneous action of a wide variety of pollutants, the effects of which should not be considered in isolation, as many synergies and antagonisms have been found between many of them. Therefore, this work proposes an in vivo study to evaluate the effect of certain metal contaminants on the bioavailability and metabolism of pharmacologically active compounds. Because the most frequent entry vector is through ingestion, the influence of the gut microbiota and the possible protective effects of selenium has been additionally evaluated. EXPERIMENTAL APPROACH: A controlled exposure experiment in mammals (Mus musculus) to a "chemical cocktail" consisting of metals and pharmaceuticals (diclofenac and flumequine). The presence of selenium has also been evaluated as an antagonist. Mouse plasma samples were measured by UPLC-QTOF. A targeted search of 48 metabolites was also performed. KEY RESULTS: Metals significantly affected the FMQ plasma levels when the gut microbiota was depleted. Hydroxy FMQ decreased if metals were present. Selenium minimized this decrease. The 3-hydroxy DCF metabolite was not found in any case. Changes in some metabolic pathways are discussed. CONCLUSIONS AND IMPLICATIONS: The presence of metals in the mouse diet as well as the prior treatment of mice with an antibiotic mixture (Abxs), which deplete the gut microbiota, has a decisive effect on the bioavailability and metabolism of the tested pharmaceuticals and dietary selenium minimize some of their effects.

An In Vivo Whole-Transcriptomic Approach to Assess Developmental and Reproductive Impairments Caused by Flumequine in Daphnia magna.

Among veterinary antibiotics, flumequine (FLU) is still widely used in aquaculture due to its efficacy and cost-effectiveness. Although it was synthesized more than 50 years ago, a complete toxicological framework of possible side effects on non-target species is still far from being achieved. The aim of this research was to investigate the FLU molecular mechanisms in Daphnia magna, a planktonic crustacean recognized as a model species for ecotoxicological studies. Two different FLU concentrations (2.0 mg L-1 and 0.2 mg L-1) were assayed in general accordance with OECD Guideline 211, with some proper adaptations. Exposure to FLU (2.0 mg L-1) caused alteration of phenotypic traits, with a significant reduction in survival rate, body growth, and reproduction. The lower concentration (0.2 mg L-1) did not affect phenotypic traits but modulated gene expression, an effect which was even more evident under the higher exposure level. Indeed, in daphnids exposed to 2.0 mg L-1 FLU, several genes related with growth, development, structural components, and antioxidant response were significantly modulated. To the best of our knowledge, this is the first work showing the impact of FLU on the transcriptome of D. magna.

Flumequine-mediated fluorescent zeolitic imidazolate framework functionalized by Eu3+ for sensitive and selective detection of UO22+, Ni2+ and Cu2+ in nuclear wastewater.

Currently, antibiotics and heavy metal contaminants have posed a great threat for ecological security and human health. Herein, the lanthanide functionalized ZIF (named ZIF-90-PABA-Eu) is constructed by coordinating with Eu3+ via p-aminobenzoic acid intermediate. Due to the excellent fluorescence properties, the novel fluorescent probe can selectively monitor flumequine based on "turn on" mode. Furthermore, the obtained new material (named ZIF-90-PABA-Eu-Flu) can be used as "turn off" sensor for selective detection of both radioactive and nonradioactive heavy metal ions (UO22+, Ni2+ and Cu2+) which are the main component of nuclear industrial wastewater. ZIF-90-PABA-Eu-Flu shows ultra-short fluorescence response time (3 s) and ultra-low limit of detection (9.0 × 10-3, 1.3 × 10-2 and 6.1 × 10-4 ppm) for three metal ions, which may be attributed to its good affinity with UO22+, Ni2+ and Cu2+. Moreover, principal component analysis (PCA) is applied to distinguish the three metal ions. Additionally, the possible sensing mechanism is investigated by the UV-vis spectra, luminescence lifetimes and theoretical calculation analysis. Based on these results, ZIF-90-PABA-Eu possesses promising potential in practical application and provides insight for the design of novel probes to continuously monitor flumequine, radioactive and nonradioactive heavy metal ions.

Advanced Photocatalytic Treatment of Wastewater Using Immobilized Titanium Dioxide as a Photocatalyst in a Pilot-Scale Reactor: Process Intensification.

In many nations, particularly those experiencing water scarcity, novel approaches are being applied to clean wastewater. Heterogeneous photocatalysis is the most widely used of these approaches because it entails the decomposition of organic molecules into water and carbon dioxide, which is a more ecologically benign process. In our study, we studied the photocatalytic degradation process on the effluent flumequine. This treatment is made through a solar pilot reactor in the presence of immobilized titanium dioxide with three light intensities and two types of water as solvents. A variety of factors that might influence the rate of deterioration, such as flow rate, light intensity, and initial concentration, have been investigated. The maximal degradation of flumequine was achieved at more than 90% after 2.5 h under optimal conditions (an initial concentration of 5 mg/L, three lamp light intensities, and a flow rate of 29 L/h). By combining the oxidized agent H2O2 with this process, the photocatalytic activity was improved further to 97% under the same conditions. The mineralization of this product has also been tested using total organic carbon (TOC) analysis. A high mineralization rate has been recorded at around 50% for a high initial concentration (20 mg/L) at a flow rate of 126 L/h. The results demonstrated the highly effective removal of flumequine and the efficacy of this photocatalytic system.

Water-sediment partitioning of flumequine and florfenicol, two antibiotics used in salmon aquaculture in Chile.

The water-sediment partitioning of flumequine and florfenicol, two antibiotics used in salmon aquaculture is a critical driver of their fate and environmental impact. Batch experiments, were carried out using pure water or seawater, with or without sediment, and at summer and winter temperatures of Chilean fjords. Log Kd (partition between water and sediment) of florfenicol in seawater varied from 0.62 ± 0.69 to 0.67 ± 0.13, and Log KOC (partition between water and organic fraction of sediment) from 2.15± 0.29 to 2.19 ± 0.13. Difference between KOC and the octanol-water partition constant (KOW) showed that for florfenicol, adsorption onto the surface of particles was more significant than the absorption driven by hydrophobicity whilst hydrophobic absorption was a major driver of flumequine sorption. Flumequine Log Kd (0.92 ± 0.25 to 1.36 ± 0.10) and Log KOC (from 2.44 ± 0.25 to 2.89 ± 0.10) demonstrated its greater affinity than florfenicol to particles and potential accumulation into marine sediments.

Antibacterial treatment of lumpfish (Cyclopterus lumpus) experimentally challenged with Vibrio anguillarum, atypical Aeromonas salmonicida and Pasteurella atlantica.

Lumpfish is a novel farmed species used as cleaner fish for the removal of lice from farmed salmon. As often with new, farmed species, there are challenges with bacterial infections. The frequency of prescription of antibiotic agents to lumpfish is increasing, despite the lack of knowledge about appropriate doses, duration of treatment and application protocols for the various antibacterial agents. In the current study, we have tested the effect of medicated feed with florfenicol (FFC), oxolinic acid (OA) and flumequine (FLU) on lumpfish experimentally challenged with Vibrio anguillarum, atypical Aeromonas salmonicida and Pasteurella atlantica. We found that all three antibacterial agents efficiently treated lumpfish with vibriosis using 10 and 20 mg kg-1  day-1 of FFC, 25 mg kg-1  day-1 of OA and 25 mg kg-1  day-1 FLU, whereas only FFC (20 mg kg-1  day-1 ) had good effect on lumpfish with pasteurellosis. None of the antibacterial agents were efficient to treat lumpfish with atypical furunculosis. FFC 20 mg kg-1  day-1 showed promising results in the beginning of the experiment, but the mortality increased rapidly 14 days post-medication. Efficient treatment is important for the welfare of lumpfish and for reducing the risk of development of antibiotic-resistant bacteria. To our knowledge, this is the first study to establish protocols for antibacterial treatment of lumpfish.

Monitoring Infection and Antibiotic Treatment in the Skin Microbiota of Farmed European Seabass (Dicentrarchus Labrax) Fingerlings.

The microbiota of fish skin, the primary barrier against disease, is highly dynamic and modulated by several factors. In fish aquaculture, disease outbreaks occur mainly during early-life stages, with associated high economic losses. Antibiotic treatments sometimes remain the best option to control bacterial diseases, despite many reported negative impacts of its use on fish and associated microbiota. Notwithstanding, studies monitoring the effects of disease and antibiotic treatment on the microbiota of fingerlings are scarce. We sequenced the bacterial 16S rRNA V4 gene region using a metabarcoding approach to assess the impact of a mixed infection with Photobacterium damselae ssp. piscicida and Vibrio harveyi and subsequent antibiotic treatment with flumequine, on the skin microbiota of farmed seabass (Dicentrarchus labrax) fingerlings. Both infection and antibiotic treatment led to a significant increase in bacterial diversity and core microbial communities and impacted microbiome structure. Dysbiosis was confirmed by changes in the abundance of potential pathogenic and opportunistic bacterial taxa. Skin bacterial metabolic function was also significantly affected by flumequine administration, suggesting a detriment to fish skin health. Our results add to an increasing body of literature, showing how fish microbiome response to infection and antibiotics cannot be easily predicted.

Maximum levels of cross-contamination for 24 antimicrobial active substances in non-target feed.Part 10: Quinolones: flumequine and oxolinic acid.

The specific concentrations of flumequine and oxolinic acid in non-target feed for food-producing animals, below which there would not be an effect on the emergence of, and/or selection for, resistance in bacteria relevant for human and animal health, as well as the specific antimicrobial concentrations in feed which have an effect in terms of growth promotion/increased yield were assessed by EFSA in collaboration with EMA. Details of the methodology used for this assessment, associated data gaps and uncertainties, are presented in a separate document. To address antimicrobial resistance, the Feed Antimicrobial Resistance Selection Concentration (FARSC) model developed specifically for the assessment was applied. However, due to the lack of data on the parameters required to calculate the FARSC, it was not possible to conclude the assessment until further experimental data are available. To address growth promotion, data from scientific publications obtained from an extensive literature review were used. No suitable data for the assessment were available. It was recommended to carry out studies to generate the data that are required to fill the gaps which prevented the calculation of the FARSC for these antimicrobials.

Distribution of Flumequine in Intestinal Contents and Colon Tissue in Pigs after Its Therapeutic Use in the Drinking Water.

Flumequine concentrations in plasma, colon tissue and intestinal contents were evaluated in 12 healthy pigs after oral administration (12 mg/kg every 24 h for 5 consecutive days in drinking water). Plasma, colon tissue and intestinal content samples were collected from animals sacrificed on days 3, 6 and 7. Concentrations were measured by high performance liquid chromatography after having validated the method, following the European Medicines Agency (EMA) requirements. The drug was not detected in any plasma sample. In colon tissue, concentrations were higher on day 3 (0.230 ± 0.033 µg/g, descending colon; 0.156 ± 0.093 µg/g, ascending colon) than on day 6 (0.187 ± 0.123 µg/g, descending colon; 0.107 ± 0.007 µg/g, ascending colon). Concentrations were considerably higher in intestinal contents, again on day 3 (1.349 ± 1.401 µg/g, descending colon; 0.591 ± 0.209 µg/g, ascending colon) than on days 6 (0.979 ± 0.346 µg/g, descending colon; 0.595 ± 0.075 µg/g, ascending colon) and 7 (0.247 ± 0.172 µg/g, descending colon; 0.172 ± 0.086 µg/g, ascending colon). Measured concentrations were lower than those effective against the most common intestinal pathogenic microorganisms in swine and, more specifically, Brachyspira hyodysenteriae.

A novel route for catalytic activation of peroxymonosulfate by oxygen vacancies improved bismuth-doped titania for the removal of recalcitrant organic contaminant.

In this work, bismuth-doped titania (BixTiO2) with improved oxygen vacancies was synthesized by sol-gel protocol as a novel peroxymonosulfate (PMS, HSO5-) activator. HSO5- and adsorbed oxygen molecules could efficiently be transformed into their respective radicals through defect ionization to attain charge balance after their trapping on oxygen vacancies of the catalyst. XRD study of BixTiO2 with 5 wt% Bi (5BiT) revealed anatase, crystalline nature, and successful doping of Bi into TiO2 crystal lattice. The particle size obtained from BET data and SEM observations was in good agreement. PL spectra showed the formation rates of •OH by 3BiT, 7BiT, 5BiTC, and 5BiT as 0.720, 1.200, 1.489, and 2.153 µmol/h, respectively. 5BiT catalyst with high surface area (216.87 m2 g-1) and high porosity (29.81%) was observed the excellent HSO5- activator. The catalytic performance of 0BiT, 3BiT, 5BiT, and 7BiT when coupled with 2 mM HSO5- for recalcitrant flumequine (FLU) removal under dark was 10, 27, 55, and 37%, respectively. Only 5.4% decrease in catalytic efficiency was observed at the end of seventh cyclic run. Radical scavenging studies indicate that SO4•- is the dominant species that caused 62.0% degradation. Moreover, strong interaction between Bi and TiO2 through Bi-O-Ti bonds prevents Bi leaching (0.081 mg L-1) as shown by AAS. The kinetics, degradation pathways, ecotoxicity, and catalytic mechanism for recalcitrant FLU were also elucidated. Cost-efficient, environment-friendly, and high mineralization recommends this design strategy; BixTiO2/HSO5- system is a promising advanced oxidation process for the aquatic environment remediation.

Green synthesis of spongy Nano-ZnO productive of hydroxyl radicals for unconventional solar-driven photocatalytic remediation of antibiotic enriched wastewater.

Herein, novel green/facile approach to synthesize spongy defective zinc oxide nanoparticles (ZnONPs) is presented using for the first time pomegranate seeds molasses as a green capping fuel/reducing mediator during an aqueous solution combustion process. The developed ZnONPs is characterized by UV-Vis. Spectrophotometry and fluorimetry, XRD, Raman spectroscopy, SEM, TEM and BET. Interestingly, pomegranate seeds molasses within a viable content of bio-capping molecules reveal a defective nanoporous ZnO NPs of smaller particle size, greater pore size/volume, and higher surface area compared to the bulky non-biogenic ZnONPs. Moreover, the biosynthesized defective ZnONPs showed narrowed band gap and higher absorption of visible photons that breed higher density of hydroxyl radicals (•OH) under Solar-illumination. Even further, the bulk ZnO and the biosynthesized ZnO photocatalysts were examined in photodegrading flumequine (FL) antibiotic. The bulk ZnO gives 41.46% photodegradation efficiency compared to 97.6% for the biosynthesized ZnO. In highly acidic or highly alkaline media, FL photodegradability is greatly retarded. Scavenging experiment infers considerable contribution of holes over electrons in photodegradation reaction. The biosynthesized ZnO shows high durability in FL photodegradation after four reusing cycles. These promising findings highlight new insights for biogenic synthesis of tuned size/controlled morphology semiconductor NPs relevant to environmental remediation applications.

Red mud-activated peroxymonosulfate process for the removal of fluoroquinolones in hospital wastewater.

In this study, a novel peroxymonosulfate (PMS) activation method, which combines a solid waste (i.e., red mud, RM) and a reducing agent (i.e., hydroxylamine, HA), for the oxidative degradation of fluoroquinolones (FQs; i.e., flumequine (FLU) and ciprofloxacin (CIP)) in hospital wastewater (HW) was developed. The addition of HA into the PMS/RM suspension significantly enhanced FLU removal, owing to its ability to enhance the Fe(III)/Fe(II) cycle on the RM surface. The results of the quenching experiments suggested the predominance of SO4•- over •OH in the PMS/RM/HA system. Moreover, owing to the greater reactivity between CIP and SO4•-, CIP removal was more effective than FLU removal. Additionally, the liquid chromatography-mass spectroscopy (LC-MS) analysis revealed that the oxidation of CIP and FLU by PMS/RM/HA occurred via sequential and separate processes, involving ring cleavage, hydroxylation, decarbonylation, and defluorination. Surprisingly, the wastewater components exhibited contrasting effects on FLU removal in HW. Natural organic matter, nitrate and sulfate showed a slight impact on the removal performance of FLU, whereas chloride improved the oxidation extent. However, phosphate significantly inhibited the FLU removal because of its competitive binding at the RM surface and its scavenging effect towards SO4•-. This inhibitory effect was overcome by increasing the PMS concentration and its sequential addition, thus guaranteeing successful mineralization of FLU in HW. These results show that the RM/HA system can be utilized to activate PMS for the removal of antibiotics in wastewater.

Nano-immobilized flumequine with preserved antibacterial efficacy.

Flumequine was nano-immobilized by self-assembly on iron oxide nanoparticles, called surface active maghemite nanoparticles (SAMNs). The binding process was studied and the resulting core-shell nanocarrier (SAMN@FLU) was structurally characterized evidencing a firmly immobilized organic canopy on which the fluorine atom of the antibiotic was exposed to the solvent. The antibiotic efficacy of the SAMN@FLU nanocarrier was tested on a fish pathogenic bacterium (Aeromonas veronii), a flumequine sensitive strain, in comparison to soluble flumequine and the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were assessed. Noteworthy, the MIC and MBC of soluble and nanoparticle bound drug were superimposable. Moreover, the interactions between SAMN@FLU nanocarrrier and microorganism were studied by transmission electron microscopy evidencing the ability of the complex to disrupt the bacterial wall. Finally, a preliminary in vivo test was provided using Daphnia magna as animal model. SAMN@FLU was able to protect the crustacean from the fatal consequences of a bacterial infection and showed no sign of toxicity. Thus, in contrast with the strength of the interaction, nano-immobilized FLU displayed a fully preserved antimicrobial activity suggesting the crucial role of fluorine in the drug mechanism of action. Besides the importance for potential applications in aquaculture, the present study contributes to the nascent field of nanoantibiotics.

Pharmacokinetic Data Show That Oxolinic Acid and Flumequine Are Absorbed and Excreted Rapidly From Plasma and Tissues of Lumpfish.

This study examined the uptake, tissue distribution and elimination of the antibacterial agents oxolinic acid and flumequine in lumpfish (Cyclopterus lumpus L.) by use of LC-MS/MS following a single oral administration of 25 mg/kg fish given in feed. Lumpfish are increasingly used as cleaner fish for removal of sea lice on commercially farmed salmon. The production of lumpfish is successful, but there are challenges with bacterial infections and the number of antibacterial treatments has increased in recent years. As the lumpfish is a novel species to farming, there is a need for pharmacokinetic data and establishment of protocols for efficient antibacterial treatment. The current study describes the pharmacokinetic properties of oxolinic acid and flumequine in lumpfish. Absorption of oxolinic acid was moderate and was characterized by a calculated peak plasma concentration (Cmax) of 2.12 µg/ml after 10.3 h (Tmax) and an elimination half-life (t1/2ß) of 21 h. Area under curve (AUC) and AUC from 0 to 24 h (AUC0-24h) were calculated to be 60.9 and 34.0 h µg/ml, respectively. For flumequine, plasma Cmax was found to be 2.77 µg/ml after 7.7 h (Tmax) with t1/2ß of 22 h. The area under the curve (AUC) and AUC from 0 to 24 h (AUC0-24) were calculated as 104.3 and 50.3 h µg/ml, respectively. Corresponding Cmax values in muscle, liver, and head-kidney for oxolinic acid were 4.01, 3.04, and, 4.68 µg/g, respectively and Tmax of 11.1, 9.2, and 10.0 h, respectively. For flumequine, Cmax values of 4.16, 4.01, and 7.48 µg/g were obtained in muscle, liver, and head kidney, respectively, with corresponding Tmax values of 10.2, 10.3, and 6.0 h. Antimicrobial susceptibility values as determined by minimum inhibitory concentration (MIC) analyses against 28 isolates of Aeromonas salmonicida isolated from diseased lumpfish ranged from 0.06 to 15 µg/ml for oxolinic acid and 0.024 to 6.25 µg/ml for flumequine. Bimodal distributions in susceptibility to both oxolinic acid and flumequine were observed. The combination of pharmacokinetic properties and MIC data make possible calculation of efficient treatment doses, which are needed to improve the welfare of lumpfish and minimize development of antibiotic resistant bacteria.

Flumequine-Mediated Upregulation of p38 MAPK and JNK Results in Melanogenesis in B16F10 Cells and Zebrafish Larvae.

Flumequine is a well-known second generation quinolone antibiotic that induces phototoxicity. However, the effect of flumequine on skin melanogenesis is unclear. Therefore, we, for the first time, investigated whether flumequine regulates melanogenesis. The present study showed that flumequine slightly inhibited in vitro mushroom tyrosinase activity but significantly increased extracellular and intracellular melanin content in B16F10 cells and promoted the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase. Additionally, flumequine remarkably increased melanin pigmentation in zebrafish larvae without any toxicity. We also found that flumequine stimulated p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) phosphorylation; inhibition of p38 MAPK and JNK resulted in significant downregulation of extracellular and intracellular melanin content in B16F10 cells and pigmentation of zebrafish larvae accompanied with suppression of MITF and tyrosinase expression, indicating that flumequine-mediated p38 and JNK promote melanogenesis in vitro and in vivo. According to the molecular docking prediction, flumequine targeted dual-specificity MAPK phosphatase 16 (DUSP16), which is a major negative regulator of p38 MAPK and JNK. Our findings demonstrate that flumequine induces an increase in melanin content in B16F10 cells and zebrafish larvae by activating p38 MAPK and JNK. These data show the potential of flumequine for use as an anti-vitiligo agent.

Enantioseparation and determination of flumequine enantiomers in multiple food matrices with chiral liquid chromatography coupled with tandem mass spectrometry.

The present work firstly described the enantioseparation and determination of flumequine enantiomers in milk, yogurt, chicken, beef, egg, and honey samples by chiral liquid chromatography-tandem mass spectrometry. The enantioseparation was performed under reversed-phase conditions on a Chiralpak IC column at 20°C. The effects of chiral stationary phase, mobile phase components, and column temperature on the separation of flumequine enantiomers have been studied in detail. Target compounds were extracted from six different matrices with individual extraction procedure followed by cleanup using Cleanert C18 solid phase extraction cartridge. Good linearity (R2 >0.9913) was obtained over the concentration range of 0.125 to 12.5 ng g-1 for each enantiomer in matrix-matched standard calibration curves. The limits of detection and limits of quantification of two flumequine enantiomers were 0.015-0.024 and 0.045-0.063 ng g-1 , respectively. The average recoveries of the targeted compounds varied from 82.3 to 110.5%, with relative standard deviation less than 11.7%. The method was successfully applied to the determination of flumequine enantiomers in multiple food matrices, providing a reliable method for evaluating the potential risk in animal productions.

Oxidation of flumequine in aqueous solution by UV-activated peroxymonosulfate: Kinetics, water matrix effects, degradation products and reaction pathways.

The degradation of flumequine (FLU) in aqueous solution by ultraviolet (UV)-activated peroxymonosulfate (PMS) was investigated in this work. Under the conditions of [PMS]0:[FLU]0 = 1:1, T = 25 ±â€¯2 °C, pH = 7.0 ±â€¯0.1, nearly complete removal of FLU was achieved after 60 min. The effects of various operating parameters, including oxidant doses, pH, the presence of typical ions (NH4+、Mg2+、Fe3+、Cl-、NO3-、HCO3-) and humic acid were evaluated. It was found that the pseudo-first-order rate constants of FLU degradation increased with increasing PMS dosage and decreasing solution pH. The presence of Mg2+ could accelerate FLU removal, while Fe3+, HCO3-, NO3- and HA inhibited the reaction. Moreover, the degradation of FLU in different water matrices were also explored, and the removal followed the order of Tap water > Ultrapure water > River water > Secondary clarifier effluent. According to the control and radical quenching experiment results, direct photolysis and reactive radicals (SO4- and HO) contributed mainly to FLU degradation in the UV/PMS system. Initial FLU molecule underwent reactions such as hydroxylation, hydroxyl substitution, demethylation, decarboxylation/decarbonylation and ring opening, leading to the formation of nineteen oxidation products. The effective degradation by UV/PMS suggests a feasible technology for treating FLU in waters and wastewaters.

DNA modifications that do not cause gene mutations confer the potential for mutagenicity by combined treatment with food chemicals.

Cell proliferation plays a key role in fixing mutations induced by DNA damage. We clarified whether this phenomenon occurred after combined treatment with chemicals in food. The effects of antibiotic flumequine (FL), a residue of veterinary medicinal products in foodstuffs, on mutagenicity in the liver were examined in mice treated with estragole (ES), a natural food flavouring compound. Gpt delta mice were orally administered 10 or 100 mg/kg/day ES and simultaneously fed a diet containing 0.4% FL for 4 weeks. Proliferating cell nuclear antigen-positive cells and cell cycle-related genes were additively increased in the livers of combined treatment groups as compared with high-dose ES or FL groups. Mutant frequencies (MFs) in gpt after cotreatment with low-dose ES and FL were significantly increased, although treatment with ES alone increased MFs only in the high-dose group. Sult1a1 mRNA levels were unchanged after FL treatment. Liquid chromatography with tandem-mass spectrometry analysis showed that FL did not affect the amount of ES-specific DNA adducts in the livers, indicating that FL treatment did not influence metabolic pathways of ES. Thus, enhancement of the mutagenic potential of a chemical by chemical-induced cell proliferation may occur as a result of the combined effects of chemicals in food.

Novel coordination imprinted polymer monolithic column applied to the solid-phase extraction of flumequine from fish samples.

In this study, a coordination imprinted polymer (CIP) solid-phase extraction (SPE) method was developed to determine the residues of flumequine (FLU) in fish samples. Silanized graphene oxide-doped CIP (SGO-CIP) monolithic column was prepared using FLU-Zn2+ as template in the presence of SGO. The synthesis conditions of SGO-CIP column were optimized by the response surface methodology. Under the optimum conditions, this column showed high specificity to FLU, and the adsorption capacity reached 61.74 ng mg-1. The enrichment factor of the monolithic column was over 40-fold. Various factors affecting the extraction efficiency of SGO-CIP column during SPE were tested to achieve optimal enrichment and to reduce non-specific adsorption. FLU in fish was detected by using a high-performance liquid chromatography-fluorescence detection system. The detection limit was as low as 0.32 ng g-1 and the recovery was as high as 95.2%, with relative standard deviations of below 5.9%. This simple and sensitive method may be applicable to the determination of FLU residues in foods.

Transgenerational toxicity of flumequine over four generations of Daphnia magna.

In this study, the effects of both continuous and alternate exposure to 2 mg L-1 of flumequine (FLU) on survival, growth and reproduction of Daphnia magna were evaluated over four generations. Mortality was the most evident effect, with an average mortality rate of 23 ±â€¯14% across generations. Individuals destined to succumb were identifiable well in advance through their discolouration and lack of development, and limited or zero reproductive capacity. Inhibition of reproduction in surviving mothers varied across the four generations (14.3 ±â€¯17%) without an apparent correlation with the duration of exposure over generations. Significant reproductive inhibition was observed in the generation that followed three non-exposed generations (the fourth generation), pointing to a transgenerational toxicity of FLU. In another experiment, in vitro exposure of 72 D. magna embryos to 2 mg L-1 FLU caused 14% mortality (versus 7% in the control). Among the 62 individuals that hatched alive, six showed birth defects and only one was able to survive the next few days. The other, apparently healthy newborns were randomly assigned to two groups and submitted to a reproduction test, either in the absence or in the presence of 2 mg L-1 FLU. A high mortality rate and/or strongly significantly inhibited reproduction were detected in both groups. As with previously run analogous tests with enrofloxacin, the multigenerational and embryonic tests showed a clear disruption to this crustacean population which would not be evidenced by the standard official acute and chronic tests. This indicates the necessity of taking a different and more comprehensive approach to the evaluation of substances having an inherent ability to interact with genetic material.