Aqueous fate of furaltadone: Kinetics, high-resolution mass spectrometry - based elucidation and toxicity assessment of photoproducts.

Furaltadone (FTD) is an antibiotic belonging to the nitrofurans group. It has been broadly used in livestock and aquaculture for therapeutic purposes, as well as for stimulating promotion. Although the European Union has imposed restrictions on the use of FTD since 1995 due to concerns regarding its toxicity, in many cases FTD has been excessively and/or illegally applied in productive animals in developing countries, because of its high efficacy and low-cost. Unlike other nitrofuran compounds, the hydrolytic and photolytic behavior of FTD in natural aquatic systems has not been thoroughly investigated. To this end, hydrolysis in different pH values and photolysis in aquatic environment, including lake, river and sea water have been both examined. Hydrolysis was found to have an insignificant impact on degradation of FTD in the aquatic environment relevant pH values, whereas indirect photolysis proved to be the main route of its elimination. The identification of tentative photoproducts (PPs) was performed using ultra high performance liquid chromatography coupled to hybrid LTQ/Orbitrap high resolution mass spectrometry. A possible pathway for photolytic transformation of FTD was proposed. Additionally, in silico simulations were used to evaluate the toxicity such as the mutagenicity of FTD and PPs. Complementary to the low-cost and time-limited simulations, an in vitro method (Vibrio Fischeri bioluminescence) was also used to assess ecotoxicity.

Evaluation of animal feeds for the presence of three important antibiotic classes in Bhutan.

OBJECTIVES: An alarming rate of emergence of antimicrobial resistance (AMR) globally has urged Bhutan to take firm steps towards antibiotic stewardship through rational use in humans, animals and agriculture. This study aimed to investigate the use of three important antibiotic classes in animal feeds in Bhutan. METHODS: Thirteen samples were selected randomly from a pool of feed samples submitted by bidders for Annual Animal Feed Tender (fiscal year 2015-2016) of the Department of Livestock. The 13 samples were tested for nitrofurans and their metabolites, tetracyclines and sulphonamides. The objectives of the study were to evaluate the feed samples for the presence of three antibiotic classes and, if present, to understand the level of presence (mg/kg) in terms of AMR emergence and spread in a Bhutanese context. RESULTS: Two feed samples, belonging to Indian Feed Agents, tested positive for oxytetracycline at levels of 0.072mg/kg and 0.037mg/kg, respectively. Compared with the Feed Additive Compendium recommended dose (10-50g/ton) of oxytetracycline for poultry, the levels found in these two feeds were low. Samples testing positive for antibiotics were rejected in the bidding procedure as being medicated at prophylactic levels and could be the cause of emergence and spread of AMR in the country. CONCLUSION: Although the samples containing antibiotics were rejected by the Department of Livestock for use in government farms, some farmers may still use medicated feeds. Therefore, more screening needs to be conducted to further confirm and to prevent antibiotic abuse in animal feed in Bhutan.

Determination of nitrofuran and chloramphenicol residues by high resolution mass spectrometry versus tandem quadrupole mass spectrometry.

An ultra-high performance liquid chromatography based method, coupled to high resolution mass spectrometry (UHPLC-HRMS), was developed to permit the detection and quantification of various nitrofuran and chloramphenicol residues in a number of animal based food products. This method is based on the hydrolysis of covalently bound metabolites and derivatization with 2-nitrobenzaldehyde. Clean-up is achieved by a liquid/liquid and a reversed phase/solid phase extraction. Not only are the four conventional nitrofurans (nitrofurantoin, furazolidone, nitrofurazone and furaltadone) detected, but also nifursol, nitrovin and nifuroxazide. Furthermore, an underivatizable nitrofuran (nifurpirinol) and another banned drug (chloramphenicol) can be quantified as well. The compounds are detected in the form of their precursor ions, [M+H](+) and [M-H](-), respectively. The mass resolving power of 70,000 FWHM, and the applied mass window ensure sufficient selectivity and sensitivity. Confirmation is obtained by monitoring the HRMS resolved product ions which were derived from the unit-mass resolved precursor ions. The multiplexing capability of the utilized Orbitrap instrument provides not only highly selective, but also sensitive confirmatory signals. This method has been validated according to the CD 2002/657/EC for the following matrices: muscle, liver, kidney, fish, honey, eggs and milk.

High-performance liquid chromatography with fluorescence detection for the determination of nitrofuran metabolites in pork muscle.

A simple and sensitive HPLC method with fluorescence detection (HPLC-FLD) is reported for the simultaneous determination of metabolites of four nitrofuran drugs (furazolidone, furaltadone, nitrofurantoin and nitrofurazone) in pork muscle. The method involves acid hydrolysis of the protein-bound drug metabolites and the conjugation of the released side-chains with a novel fluorescence agent 2-hydroxy-1-naphthaldehyde. After liquid-liquid extraction and effective separation of the derivatives on a YMC-Pack Polymer C18 column at 40°C under alkaline conditions, the high fluorescence intensity of these derivatives at emission wavelength λem = 463 nm enables their simultaneous determination in pork muscle at concentrations as low as 1 µg kg⁻¹. The method was validated using blank pork muscle fortified with all four metabolites at 0.5, 1.0 and 2.0 µg kg⁻¹. Recoveries were > 92.3% with RSDs < 8.5% for all four metabolites. The results obtained with HPLC-FLD and LC-MS/MS methods showed very good agreement for pork muscle samples.

Residue analyses and exposure assessment of the Irish population to nitrofuran metabolites from different food commodities in 2009-2010.

An exposure assessment to nitrofuran residues was performed for three human populations (adults, teenagers and children), based on residue analyses of foods of animal origin (liver, honey, eggs and aquaculture) covering the 2-year period 2009-2010. The occurrence of nitrofuran metabolites in food on the Irish market was determined for the selected period using the data from Ireland's National Food Residue Database (NFRD) and from results obtained from the analysis of retail samples (aquaculture and honey). Laboratory analyses of residues were performed by methods validated in accordance with Commission Decision 2002/657/EC regarding performance of the analytical method and interpretation of results. Semicarbazide (SEM) was the contaminant most frequently identified and its content ranged from 0.09 to 1.27 µg kg(-1). SEM is currently used as a marker of nitrofuran abuse, but it may also occur from other sources. The presence of nitrofuran metabolite 3-amino-2-oxazolidinone (AOZ) was detected in two aquaculture samples (prawns) at 1.63 and 1.14 µg kg(-1), but such a low number of positive cases did not present sufficient data for a full AOZ exposure assessment. Therefore, the evaluation of exposure was focused on SEM-containing food groups only. Exposure assessments were completed using a probabilistic approach that generated 10 iterations. The results of both the upper- and lower-bound exposure assessments demonstrate that SEM exposure for Irish adults, teenagers and children from selected food commodities are well below EFSA-estimated safe levels.

Separation and determination of nitrofuran antibiotics in turbot fish by microemulsion electrokinetic chromatography.

A microemulsion electrokinetic chromatography (MEEKC) method has been developed for the determination of four nitrofuran antibiotics (furazolidone (FZD), furaltadone (FTD), nitrofurazone (NFZ) and nitrofurantoin (NFT)) in turbot fish. The effect of buffer the pH, the concentration of SDS and the concentrations of octane and butan-1-ol were studied systematically. With the optimized experimental conditions (octane, 0.82% (w/w); SDS, 3.48% (w/w); butan-1-ol, 6.48% (w/w); and 10 mM sodium tetraborate buffer (pH 9.70), with 30 kV as the applied voltage) all four analytes were baseline-separated within 8 min. Regression equations revealed a good linear relationship between the peak area of each compound and its concentration. The correlation coefficients of the four analytes were from 0.9945 to 0.9999. The relative standard deviations of the migration times and the peak areas were <1.84 and 5.16% (intra-day). The obtained recovery ranged between 97 and 104%. Moreover, the method was successfully validated and applied to the determination of nitrofuran antibiotics in contaminated fish.