Difloxacin Research Articles

Tb3+ assisted dithioerythritol stabilized copper nanocluster with AIE behavior for ratiometric fluorescent determination of fluoroquinolones

BackgroundFluoroquinolones (FQs) are widely used in livestock and poultry industry because of their satisfactory effects in preventing and treating bacterial infection. However, due to irrational use and poor biodegradability, FQs can easily remain in food animals and further enter the human body through the food chain. Therefore, accurate and sensitive detection of FQs residues in animal-origin food is significant. The traditional methods commonly used for FQs detection have some limitations. Ratiometric fluorescence detection technology has the advantages of fast, sensitive, self-correcting, and easy visualization. However, the reports on the use of ratiometric fluorescence probes for FQs detection are limited. ResultsIn this work, a novel probe was proposed for ratiometric fluorescent analysis of FQs. In this probe, the fluorescence of dithioerythritol stabilized copper nanoclusters (DTE-Cu NCs) was significantly enhanced due to the Tb3+ triggered aggregation-induced emission effect. FQs bound Tb3+ in Tb3+/DTE-Cu NCs through carboxyl and carbonyl groups, so that Tb3+ was effectively sensitized to emit green fluorescence. However, the red fluorescence of DTE-Cu NCs was not interfered. The fluorescence of the probe transformed from red to green with the increase of FQs concentration. Using norfloxacin (NOR), difloxacin (DIF), and enrofloxacin (ENR) as FQs simulants, this probe showed a sensitive linear response ranged from 0.025 to 22.5 μM, with the limits of detection of 9.6 nM, 9.3 nM, and 7.7 nM. The application potential for FQs detection was verified via a standard addition assay of egg samples with the recovery rate of 90.4 %–114.7 %. SignificantThe fluorescence probe based on Tb3+/DTE-Cu NCs is expected to realize the ratiometric fluorescence sensitive detection of FQs. The establishment of this simple, effective, and rapid detection platform opens up a new way for the detection of FQs residues in animal-origin foods, and also provides a new idea for the design of rapid detection platforms for other hazard factors.

Fluorescence detection of difloxacin in milk using blue light-emitting carbon dots

Developing effective methods to detect contamination in milk during production, processing, and storage is quite important but challenging. Difloxacin (DIF), due to its great potential in veterinary medicine for the treatment of bacterial infections, is one of the most common organic contaminants in milk, posing a potential and significant threat to human health. However, there is a lack of high-performance means for the effective in-situ monitoring of DIF. Herein, we have successfully achieved high-sensitivity and high-selectivity fluorescence detection of DIF in milk using blue-luminescent carbon dot (b-CD) materials prepared. When a certain amount of DIF is present in the milk, titrating it into the reagent results in a significant quenching of the fluorescence. The calculated Ksv value is 1698.7 M−1, and the detection limit is 1.77 × 10-5 M, which is better than previously reported. Crucially, the preparation cost of these fluorescent carbon dots is low, making it hopeful to develop them into specific fluorescent detection products. They are potentially valuable for practical application in the detection of DIF in infant milk products and other emulsions.

An immunochromatographic strip sensor for marbofloxacin residues.

Marbofloxacin (MBF) was once widely used as a veterinary drug to control diseases in animals. MBF residues in animal food endanger human health. In the present study, an immunochromatographic strip assay (ICSA) utilizing a competitive principle was developed to rapidly detect MBF in beef samples. The 50% inhibitory concentration (IC50) and the limit of detection (LOD) of the ICSAs were 2.5 ng/mL and 0.5 ng/mL, respectively. The cross-reactivity (CR) of the MBF ICSAs to Ofloxacin (OFL), enrofloxacin (ENR), norfloxacin (NOR), and Ciprofloxacin (CIP) were 60.98%, 32.05%, 22.94%, and 23.58%, respectively. The CR for difloxacin (DIF) and sarafloxacin (SAR) was less than 0.1%. The recovery rates of MBF in spiked beef samples ranged from 82.0% to 90.4%. The intra-assay and interassay coefficients of variation (CVs) were below 10%. In addition, when the same authentic beef samples were detected in a side-by-side comparison between the ICSAs and HPLC‒MS, no statistically significant difference was observed. Therefore, the proposed ICSAs can be a useful tool for monitoring MBF residues in beef samples in a qualitative and quantitative manner.

Comparing the photodegradation of typical antibiotics in ice and in water: Degradation kinetics, mechanisms, and effects of dissolved substances

New antibiotic contaminants have been detected in both surface waters and natural ice across cold regions. However, few studies have revealed distinctions between their ice and aqueous photochemistry. In this study, the photodegradation and effects of the main dissolved substances on the photolytic kinetics were investigated for sulfonamides (SAs) and fluoroquinolones (FQs) in ice/water under simulated sunlight. The results showed that the photolysis of sulfamethizole (SMT), sulfachloropyridazine (SCP), enrofloxacin (ENR) and difloxacin (DIF) in ice/water followed the pseudo-first-order kinetics with their quantum yields ranging from 4.93 × 10−3 to 11.15 × 10−2. The individual antibiotics experienced disparate photodegradation rates in ice and in water. This divergence was attributed to the concentration-enhancing effect and the solvent cage effect that occurred in the freezing process. Moreover, the main constituents (Cl−, HASS, NO3− and Fe(III)) exhibited varying degrees of promotion or inhibition on the photodegradation of SAs and FQs in the two phases (p < 0.05), and these effects were dependent on the individual antibiotics and the matrix. Extrapolation of the laboratory data to the field conditions provided a reasonable estimate of environmental photolytic half-lives (t1/2,E) during midsummer and midwinter in cold regions. The estimated t1/2,E values ranged from 0.02 h for ENR to 14 h for SCP, which depended on the reaction phases, latitudes and seasons. These results revealed the similarities and differences between the ice and aqueous photochemistry of antibiotics, which is important for the accurate assessment of the fate and risk of these new pollutants in cold environments.

Withdrawal time of danofloxacin and difloxacin and in vitro binding phenomenon to melanin in black-boned silky fowl.

Fluoroquinolones are commonly used in poultry breeding. Few studies have evaluated the causes of serious drug residues in black-boned silky fowl until enrofloxacin has been banned in black-boned silky fowl breeding in the Chinese Veterinary Commission of Chinese Veterinary Pharmacopoeia (2020). However, similarly structured fluoroquinolones have not been studied in black-boned silky fowl. In this study, the elimination of tissue residues of danofloxacin (DAN) and difloxacin (DIF) was studied in four tissues of black-boned silky fowl. The specific administration methods were 100mg/L of DIF aqueous solution for free drinking for 5 days and 50mg/L of DAN aqueous solution for free drinking for 3 days. Based on the experiment, the withdrawal times of 44 days for muscle, 95 days for skin+fat, 3 days for liver, and 44 days for kidney of DAN were acquired, of 43 days for muscle, 61 days for skin+fat, 0 days for liver, and 38 days for kidney of DIF were acquired, which showed that DIF and DAN should be used with caution for application in black-boned silky fowl. In vitro experiments showed that black-boned silky fowl tissues had stronger adsorption capacity to DAN and DIF than yellow chicken tissues (especially in skin+fat), and melanin has a strong adsorption effect on DAN and DIF, which is an important reason for the high residual concentrations of fluoroquinolone in black-boned silky fowl.

Identification and quantification of ciprofloxacin, enrofloxacin and difloxacin residues in milk using excitation-emission fluorescence and second-order standard addition methods

In the presence of non-modeled components and analyte-matrix interaction in real samples, a combination of second-order multivariate calibration and standard addition is desired to achieve accurate results via second-order advantage. Enrofloxacin (ENR), Difloxacin (DIF), and Ciprofloxacin (CIP) are Fluoroquinolones widely considered antibiotic residues in milk. Excitation-emission fluorescence spectra of these antibiotics at pH = 9.1 provide second-order data for identifying and quantifying them in a mixture using second-order calibration modeling.Rank annihilation factor analysis (RAFA) and multivariate curve resolution-alternating least squares (MCR-ALS) with trilinearity constraints are applied to determine the CIP, ENR, and DIF residues in synthetic mixtures and milk using second-order excitation-emission fluorescence spectroscopy. In both methods, the trilinearity constraint guarantees a unique true solution to the spectral profiles of analytes. The combination of second-order standard addition and higher-order calibration modeling with RAFA and MCR-ALS supports the second-order advantage in milk analysis.

Indirect Competitive Enzyme-Linked Immunosorbent Assay Based on Broad-Spectrum Antibody for Simultaneous Determination of Thirteen Fluoroquinolone Antibiotics in Rana catesbeianus

Fluoroquinolone (FQ) is a type of widely used antibiotic in agriculture and aquaculture, and exposure to low doses of FQs may result in the transfer of resistance between animal and human pathogens. Based on the optimization of the operating parameters, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) standard curve was constructed for the simultaneous detection of 13 FQs, including enrofloxacin (ENR), ciprofloxacin (CIP), sarafloxacin (SAR), ofloxacin (OFL), norfloxacin (NOR), pefloxacin mesylate (PM), pefloxacin (PEF), enoxacin (ENX), marbofloxacin (MAR), fleroxacin (FLE), lomefloxacin (LOM), danofloxacin (DAN), and difloxacin (DIF). The limit of detection (LOD, computed as IC10) and sensitivity (IC50) of the ic-ELISA for ENR were 0.59 μg/L and 19.23 μg/L, respectively. The precision and dependability of the detection results of this ic-ELISA were properly verified by HPLC in Rana catesbeianus samples. This indicated that the established ic-ELISA approach could be utilized to determine the FQs in Rana catesbeianus. In addition, this ic-ELISA, based on a broad-spectrum antibody, provides a technical reference and potential strategy for an immunoassay of hazard factors with similar structure.

Distribution Coefficient of QNs in Urban Typical Water and Its Main Environmental Influencing Factors

Sediment is the main storage medium of antibiotics in a water environment, and a growing body of research has focused on the distribution behavior of antibiotics in water-sediment. However, most of the previous studies were based on laboratory simulation, and less attention was paid to the distribution behavior of antibiotics in a natural water environment and its correlation with environmental factors. Thus, the surface water and sediment in Shijiazhuang were taken as the research object for this study. The temporal and spatial distribution characteristics of quinolone (QNs) antibiotics in Shijiazhuang water were analyzed by using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS), calculating the distribution coefficients of quinolone (QNs) antibiotics in water and sediment, and confirming the main environmental factors influencing the distribution coefficient in natural water. The results showed that:① the content of ΣQNs in water and sediment ranged from 8.0 to 4.4×103 ng·L-1 and 16 to 2.2×103 ng·g-1 in Shijiazhuang water, whereas the primary QNs in water and sediment were enrofloxacin (ENR) and ofloxacin (OFL), respectively. ② The total concentrations of ΣQNs in Shijiazhuang water showed a tendency of being higher in December (1.0×104 ng·L-1) than in April (5.5×103 ng·L-1), and QNs in sediment were also higher in December (7.8×103 ng·g-1) than in April (6.2×103 ng·g-1). ③ The distribution coefficient of QNs in Shijiazhuang water varied from 34 to 2.9×105 L·kg-1 and showed a trend of being greater in December than in April. ④ The results of correlation analysis showed that total nitrogen (TN), nitrate nitrogen (NO3--N), nitrite nitrogen (NO2--N), and ammonia nitrogen (NH4+-N) were significantly correlated with most distribution coefficients of QNs[OFL, norfloxacin (NOR), ENR, difloxacin (DIF), and oxolinic acid (OXO)], whereas temperature (T), total organic carbon (TOC), and total dissolved solids (TDS) were significantly correlated with individual distribution coefficients of QNs[marbofloxacin (MAR) and DIF]. Therefore, the eutrophication level of water affects the distribution behavior of antibiotics in water-sediment.

Simultaneous Determination of Nine Quinolones in Pure Milk Using PFSPE-HPLC-MS/MS with PS-PAN Nanofibers as a Sorbent.

In this study, a packed-fiber solid-phase extraction (PFSPE)-based method was developed to simultaneously detect nine quinolones, including enrofloxacin (ENR), ciprofloxacin (CIP), ofloxacin (OFL), pefloxacin (PEF), lomefloxacin (LOM), norfloxacin (NOR), sarafloxacin (SAR), danofloxacin (DAN), and difloxacin (DIF), in pure milk, using high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). Polystyrene (PS) and polyacrylonitrile (PAN) were combined to form PS-PAN composite nanofibers through electrospinning. The nanofibers were used to prepare the home-made extraction columns, and the process was optimized and validated using blank pure milk. The analytical method showed high accuracy, and the recoveries were 88.68–97.63%. Intra-day and inter-day relative standard deviations were in the ranges of 1.11–6.77% and 2.26–7.17%, respectively. In addition, the developed method showed good linearity (R2 ≥ 0.995) and low method quantification limits for the nine quinolones (between 1.0–100 ng/mL) for all samples studied. The nine quinolones in the complex matrix were directly extracted using 4.0 mg of PS-PAN composite nanofibers as a sorbent and completely eluted in 100 μL elution solvent. Therefore, the developed PFSPE-HPLC-MS/MS is a sensitive and cost-effective technique that can effectively detect and control nine quinolones in dairy products.

Effects of two kinds of fishery drugs on the expressions of GAD and GABA-T mRNA in crucian carp (Carassius auratus gibelio)

The objective of this study was to investigate the effects of difloxacin (DIF) and avermectin (AVM) on glutamate decarboxylase (GAD) and GABA-transaminase (GABA-T) in different tissues of crucian carp (Carassius auratus gibelio). After the treatments of DIF and AVM, the mRNA expressions of GAD and GABA-T in different tissues were detected by quantitative real-time PCR (qPCR). The results showed that the mRNA expressions of GAD65, GAD67, and GABA-T in the telencephalon (Tel), mesencephalon (Mes), cerebella (Cer), and medulla oblongata (Med) were downregulated significantly with the safe dose (SD, 20 mg/kg) of DIF (P < 0.05 or P < 0.01). While the expressions of GAD65 and GAD67 in the kidney at 12 h had strikingly upregulated to 13.81 ± 1.06** and 150.67 ± 12.85** times. Treated with the lethal dose of 50% (LD50, 2840 mg/kg b. W.) of DIF, the mRNA expressions of GAD65, GAD67, and GABA-T in all tissues were increased significantly (P < 0.01). The results of AVM group showed that the mRNA expressions of GAD65, GAD67, and GABA-T both in the central and peripheral tissues were all remarkably downregulated at the safe concentration (SC, 0.0039 mg/L) and the lethal concentration of 50% (LC50, 0.039 mg/L), except for the mRNA inhibitions of GAD65, GAD67, and GABA-T in the muscle at 2 h which sharply downregulated to 0.20 ± 0.02ΔΔ × 10−2, 0.57 ± 0.06ΔΔ × 10−1 and 0.44 ± 0.02ΔΔ × 10−1, respectively (P < 0.01).

Effects of N-1 substituent on the phototoxicity of fluoroquinolone antibiotics: comparison of pefloxacin and difloxacin

The photophysics and photochemistry of pefloxacin (PEF), a 1-ethyl-substituted fluoroquinolone (FQ) antibiotic, were studied using transient, steady-state experimental methods and computational methods. The fundamental photoproperties of PEF and its phototoxicity toward lysozyme, a single-chain protein, were compared with those of a 1-fluorophenyl-substituted FQ antibiotic, difloxacin (DIF). The results showed that the phototoxicity was significantly decreased by the insertion of the bulky 1-fluorophenyl substituent (the phototoxicity of DIF was approximately one-quarter of that observed for PEF). This trend was attributed to the lowest lying singlet state with sizeable oscillator strength (f ≥ 0.1) being shifted from 319 nm in PEF to 266 nm in DIF upon the insertion of the bulky substituent at the 1-position, as investigated by using computational methods. In addition, 95% of the solar UV irradiation that reaches the earth’s surface has wavelength > 315 nm. Therefore, reducing the most effective excitation wavelength by optimizing the substituent at the 1-position may be a promising strategy to alleviate the phototoxicity of FQ antibiotics. These findings may be applied to other FQ antibiotics because a large number of phototoxicity studies on FQ antibiotics with different substituents at the 1-position can prove these finding’s effectiveness. Delafloxacin, an FQ antibiotic bearing a chlorine and bulky substituent at the 8- and 1-positions, respectively, exhibits no phototoxicity is the most recent example reported to date. To the best of our knowledge, this is the first transient and steady-state study of the effect of the N-1 substituent on the photochemistry and phototoxicity of FQ antibiotics. These findings will be beneficial to the development of novel FQ antibiotics without phototoxicity.

Pharmacokinetics of difloxacin in Japanese quails (Coturnix japonica) after single intravenous and oral administration

Pharmacokinetics of difloxacin (DF), a fluoroquinolone antibiotic, were investigated in Japanese quails (Coturnix japonica) after a single intravenous (IV) and oral (PO) administration of 10 mg/kg bodyweight. Plasma concentration profiles of DF were analyzed by a compartmental pharmacokinetic method. Following IV injection, the plasma concentration vs time profile was best described by a two-compartment open model. Elimination half-life (t1/2β), total body clearance (Cltot), volume of distribution at steady state (Vdss) and mean residence time (MRT) of DF were 5.45 ± 0.14 h, 0.22 ± 0.01 L/kg/h, 1.54 ± 0.06 L/kg and 6.92 ± 0.19 h, respectively. Following PO administration, DF was rapidly absorbed, with peak plasma concentration (Cmax) of 3.67 μg/mL attained at 1.90 h (Tmax) after administration. Absorption half-life (t1/2ab), elimination half-life (t1/2el), mean absorption time (MAT) were 0.5 h, 5.26 h and 1.11 h, respectively. The bioavailability (F) following PO administration of DF was high (84.40%). For a successful clinical effect of DF in quails, a multiple dosage regimen of 10 mg/kg bodyweight, administered orally every 24 h is recommended to maintain effective plasma concentrations with bacterial infections, in which MIC90 is <0.2 μg/mL.

Performance and bacterial community dynamics of vertical flow constructed wetlands during the treatment of antibiotics-enriched swine wastewater

This study aimed to evaluate the nutrient removal and microbial response during the treatment of antibiotics-enriched swine wastewater by vertical flow constructed wetlands with different water flow directions and substrates. Better NH4-N and TN removal were achieved in down-flow and up-flow treatments, respectively. The concentrations of all the chemical properties including oxytetracycline (OTC), difloxacin (DIF), copper (Cu) and zinc (Zn), nitrite/nitrate, sulphate and total organic carbon (TOC) in down-flow treatment soils were significantly higher than those in up-flow treatments on day 30, 60 and 90 (p<0.05). Bacterial α-diversity indices in soils were negatively correlated with the concentrations of OTC, DIF, Cu and Zn significantly (p<0.05). Among experimental factors, substrate type significantly contributed to variation of bacterial communities in waters, while water flow direction and sampling time significantly contributed to those in soils (p<0.01). Among environmental factors, OTC+DIF, Cu+Zn, nitrate/nitrite and sulphate significantly contributed to variation of bacterial communities in soils (p<0.05). The taxonomy assignment indicated that antibiotics accumulation in soils promoted the growth of 19 dominant genera, including several antibiotic-resistant or degrading bacteria, which however were generally not dominant in effluents. In addition, although the TN and NH4-N removals were reduced when antibiotic concentrations in soils increased, the available community data cannot strongly support the inhibition effect of antibiotics on nitrogen transformation. Further assessment found that some dominant genera in water samples and those enriched under the stress of antibiotics in soils tended to increase their abundances in the effluents of oyster shell-based CWs.

Using In Vitro Dynamic Models To Evaluate Fluoroquinolone Activity against Emergence of Resistant Salmonella enterica Serovar Typhimurium.

The objectives of this study were to determine pharmacokinetic/pharmacodynamic (PK/PD) indices of fluoroquinolones that minimize the emergence of resistant Salmonella enterica serovar Typhimurium (S Typhimurium) using in vitro dynamic models and to establish mechanisms of resistance. Three fluoroquinolones, difloxacin (DIF), enrofloxacin (ENR), and marbofloxacin (MAR), at five dose levels and 3 days of treatment were simulated. Bacterial killing-regrowth kinetics and emergence of resistant bacteria after antibacterial drug exposure were quantified. PK/PD indices associated with different levels of antibacterial activity were computed. Mechanisms of fluoroquinolone resistance were determined by analyzing target mutations in the quinolone resistance-determining regions (QRDRs) and by analyzing overexpression of efflux pumps. Maximum losses in susceptibility of fluoroquinolone-exposed S Typhimurium occurred at a simulated AUC/MIC ratio (area under the concentration-time curve over 24 h in the steady state divided by the MIC) of 47 to 71. Target mutations in gyrA (S83F) and overexpression of acrAB-tolC contributed to decreased susceptibility in fluoroquinolone-exposed S Typhimurium. The current data suggest AUC/MIC (AUC/mutant prevention concentration [MPC])-dependent selection of resistant mutants of S Typhimurium, with AUC/MPC ratios of 69 (DIF), 62 (ENR), and 39 (MAR) being protective against selection of resistant mutants. These values could not be achieved in veterinary clinical areas under the current recommended therapeutic doses of the fluoroquinolones, suggesting the need to reassess the current dosing regimen to include both clinical efficacy and minimization of emergence of resistant bacteria.

Selective Immunodetection of Difloxacin in Animal Muscles and Sera: Role of Hapten Orientation

The influence of sarafloxacin (SAR) hapten orientation in immunogen on antibody specificity was examined. The spatial orientation of SAR linked through its carboxyl group to the carrier resulted to SAR-selective response (Anal Methods 2016, 8:5843–5850). To provide opposite orientation of SAR in immunogen, it was linked by its secondary amine to (1) succinic anhydride-modified BSA using carbodiimide-mediated conjugation and to (2) unmodified BSA using formaldehyde condensation method. Two versions of indirect competitive enzyme-linked immunosorbent assays (ELISA) based on generated antibodies were developed. A panel of homo- and heterologous conjugates was examined as potential coating antigens. No cross-reactions were registered in ELISA-1 and ELISA-2 except for fluorophenyl-containing fluoroquinolones difloxacin (DIF), SAR, and tosufloxacin as 138, 100 and 25% and 175, 100 and 8%, respectively. Among the recognized analytes, only DIF is authorized in meat products, so the developed tests were selective for DIF determination in bovine and porcine muscles and sera samples. The values of half-maximal inhibition concentration (IC50) for these assay versions were estimated to be 1.2 and 0.25 ng/ml. DIF detection limits (IC10) were, respectively, 0.15 and 0.015 ng/ml that allowed measuring of MRL and ten times lower level of DIF in the animal muscles. The developed ELISAs were also suitable for DIF determination in animal sera in the concentration range 1000–1 ng/ml.

Effect of various storage conditions on the stability of quinolones in raw milk

ABSTRACTResearch on the storage stability of antibiotic residues in milk is important for method development or validation, milk quality control and risk assessment during screening, confirmation, qualitative or quantitative analysis. This study was conducted using UPLC-MS/MS to determine the stability of six quinolones – ciprofloxacin (CIP), danofloxacin (DAN), enrofloxacin (ENR), sarafloxacin (SAR), difloxacin (DIF) and flumequine (FLU) – in raw milk stored under various conditions to investigate if quinolones degrade during storage of milk, and finally to determine optimal storage conditions for analysis and scientific risk assessment of quinolone residues in raw milk. The storage conditions included different temperatures and durations (4°C for 4, 8, 24 and 48 h; –20°C for 1, 7 and 30 days; –80°C for 1, 7 and 30 days), thawing temperatures (25, 40 and 60°C), freeze–thaw cycles (1–5), and the addition of different preservatives (sodium thiocyanate, sodium azide, potassium dichromate, bronopol and methanal). Most quinolones exhibited high stability at 4°C for up to 24 h, but began to degrade after 48 h. In addition, no degradation of quinolones was seen when milk samples were stored at –20°C for up to 7 days; however, 30 days of storage at –20°C resulted in a small amount of degradation (about 30%). Similar results were seen when samples were stored at –80°C. Moreover, no losses were observed when frozen milk samples were thawed at 25, 40 or 60°C. All the quinolones of interest, except sarafloxacin, were stable when milk samples were thawed at 40°C once and three times, but unstable after five freeze–thaw cycles. Preservatives affected the stability of quinolones, but the effects differed depending on the preservative and quinolone. The results of this study indicate optimum storage protocols for milk samples, so that residue levels reflect those at the time of initial sample analysis, and should improve surveillance programmes for quinolones in raw milk.

Simultaneous and differential determination of drugs and metabolites using the same antibody: difloxacin and sarafloxacin case

To differentiate the close structurally related fluoroquinolones (FQs) difloxacin (DIF) and its active metabolite/degradant sarafloxacin (SAR) in animal tissues, rabbit polyclonal antibodies against SAR congugated to ethylenediamine modified BSA were used in double format immunoassay.

Dissociation Constants and Octanol–Water Partition Equilibria for Several Fluoroquinolones

The octanol/water partition and acid–base dissociation equilibria for a set of six fluoroquinolones have been studied by the shake-flask method at 25 °C and at 0.15 M NaCl ionic strength, determining the partition coefficient (logD) in the pH range 3 to 9. The substances studied are ciprofloxacin (CIP), danofloxacin (DAN), difloxacin (DIF), enrofloxacin (ENR), marbofloxacin (MAR), and sarafloxacin (SAR). After the partition equilibrium, the remaining fluoroquinolone concentration is determined in aqueous phase by liquid chromatography with fluorescence detection. The pKa values obtained are in accordance with the previously published data, ranging between 5.59 and 6.50 for pKa1 and between 7.99 and 8.96 for pKa2. The lipophilicity of the substances is low, with logKOW ranging between −1.09 to 0.68. Additionally, we have determined the octanol/water partition coefficient for the cationic species in all cases.

Fluoroquinolones and Tetracycline Antibiotics in a Portuguese Aquaculture System and Aquatic Surroundings: Occurrence and Environmental Impact

The growth of aquaculture over the past few years is widely recognized as one of the main sources of antibiotics, mainly fluoroquinolones (FQ) and tetracyclines (TC), in the aquatic environment, consequently, increasing the risk of the emergence of antibiotic bacterial resistance and promoting the spread of resistant genes. This study aimed to (1) develop and validate a multiresidue method for determination and quantification of ciprofloxacin (CIP), difloxacin (DIFL), enrofloxacin (ENR), norfloxacin (NOR), sarafloxacin (SARA), and oxytetracycline (OXY) in aquaculture waters and surrounding water bodies and (2) provide the first Portuguese data to utilize in assessment of risk of adverse effects. In addition, the potential environmental impact posed by these antibiotics to aquatic organisms, belonging to different trophic levels, when exposed to the studied aquaculture waters was also assessed. The analytical strategy comprised of solid-phase extraction (SPE) through Oasis HLB cartridges, and detection and quantification by liquid chromatography with tandem mass spectrometry (LC/MSn). Method detection limits (MDL) and method quantification limits (MQL) were in the range of 0.7–3 ng/L and 2.4–10 ng/L, respectively. Recoveries varied between 57.4 and 122.8%. The method was applied to 31 water samples collected from an aquaculture and surrounding water bodies located in north of Portugal. Residues of all antibiotics, except SARA and DIFL, were detected at concentrations ranging from 3 to 75.1 ng/L. Norfloxacin was the antibiotic present at highest frequency and concentration. Regarding the environmental impact assessment (EIA), a risk quotient higher than 1 was observed for NOR.

Determination of Fluoroquinolones Residual in Freshwater Fish by HPLC

HPLC (High Performance Liquid Chromatography) method for danofloxacin(DAN), enrofloxacin(ENR), sarafloxacin(SAR) and difloxacin(DIF) residues in freshwater fish was developed. The fish sample was extracted with methanol-water-acetic acid(2/8/0.01, V/V/V), performed on a C18 solid-phase extraction column. With methanol+malonic acid-magnesium chloride aqueous solution as mobile phase, gradient elution at 30°C, inject volume was 20μL. The wavelength of UV detector was 280 nm. The linear rang of four drugs was 0.1~10μg/mL. In the level of 0.5μg/kg and 4.0μg/kg, the average recovery of the four kinds of drugs was 79.81%~92.03% , the RSD was 1.03%~4.07%. The limit of DAN, ENR, SAR, and DIF was 0.1μg/kg, 0.1μg/kg, 0.05μg/kg, and 0.05μg/kg individual. This method was employed to analyze freshwater fishes, such as catfish, chub and carp, because of its simplicity and high efficiency.