| Florfenicol,ciprofloxacin,enrofloxacin,and sarafloxacin are the most frequently identified veterinary drug residues in eggs and poultry muscles.In order to guarantee the safety and quality of poultry products available on the market and avert the introduction of veterinary drug residues into the food chain,which could jeopardize consumer health,the advancement of technology for the detection of veterinary drug residues in poultry products is of paramount importance.Presently,there are no documented instances of the employment of high-performance liquid chromatography with fluorescence detection(HPLC-FLD)methodology for the concurrent quantification of florfenicol,its metabolite florfenicol amine,and three fluoroquinolones(ciprofloxacin,enrofloxacin and sarafloxacin)residues in animal-derived comestibles on both a domestic and international scale.Within this study,liquid-liquid extraction(LLE)was employed to extract five targeted compounds from the muscle tissue of Shaobo chickens,Gaoyou ducks,and Yangzhou geese.A HPLC-FLD method was developed and refined for the concurrent quantification of florfenicol,its metabolite florfenicol amine,and three fluoroquinolones(ciprofloxacin,enrofloxacin,and sarafloxacin)residues in various poultry products,including chicken eggs(whole egg,egg yolk,and egg albumen)and poultry muscles(chicken muscle,duck muscle,and goose muscle).The primary outcomes of this research are as follows:1.A technique involving liquid-liquid extraction(LLE)was established and fine-tuned for extracting florfenicol,its metabolite florfenicol amine,and three fluoroquinolones(ciprofloxacin,enrofloxacin,and sarafloxacin)from chicken eggs.Specifically,2(± 0.02)g chicken eggs were precisely weighed,followed by the addition of 0.4 mL of 0.1 mol/L EDTA solution and 8 mL of acetonitrile.The resulting mixture was vortexed and centrifuged to obtain the supernatant.Subsequently,the residual sample was subjected to the addition of 10 mL 80%acetonitrile solution,which was extracted twice,and the resulting supernatant was combined.Fat removal was carried out using n-hexane that was saturated with acetonitrile.The sample was then subjected to vacuum centrifugal concentration,re-dissolved in the mobile phase,and centrifugally filtered through a membrane before injection.The recovery rates of the five targeted compounds exceeded 71.95%and satisfied the relevant standards specified by the NY/T 1896-2010 regulation.2.A method has been devised and refined for the extraction of florfenicol,its metabolite florfenicol amine and three fluoroquinolones(ciprofloxacin,enrofloxacin,and sarafloxacin)from poultry muscles utilizing the LLE technique.Similarly,2(±0.02)g of poultry muscles were precisely weighed,and then mixed with 0.4 mL 0.1 mol/L EDTA solution and 1 mL 90%acetonitrile solution,which was vortexed.Then,9 mL of acetonitrile was added and vortexed again before being centrifuged to collect the supernatant.The leftover sample residue was blended with the supernatant after being extracted twice with 10 mL 90%acetonitrile solution.N-hexane that had been saturated with acetonitrile was used to extract the fat.Before injection,the sample was centrifugally vacuum-concentrated,redissolved in the mobile phase,and filtered membrane.The five target compounds had recovery rates that were higher than 70.34%.3.An HPLC-FLD procedure was formulated and refined to concurrently detect florfenicol and its corresponding metabolite,florfenicol amine,as well as three fluoroquinolones(ciprofloxacin,enrofloxacin,and sarafloxacin)residues present in chicken eggs and poultry muscles.The chromatographic column employed in this study was the XBridge BEH C18 with dimensions of 4.6 mm x 150 mm and particle size of 5 μm.Separation of the five targeted compounds was achieved through an isocratic elution technique utilizing a mobile phase containing a 0.01 mol/L sodium dihydrogen phosphate solution,with a pH adjustment to 4.8 using phosphoric acid.The eluent was composed of acetonitrile(65:35,v/v)and was run at a rate of 1 mL/min.A sample volume of 100 μL was injected into the system,and the column temperature was maintained at 30℃.The detection was accomplished via a dual-channel fluorescence detector,whereby the excitation/emission wavelengths of 228/279 nm were utilized to detect florfenicol and florfenicol amine(channel A),while 272/450 nm were employed to detect enrofloxacin,ciprofloxacin,and sarafloxacin(channel B).The outcomes divulged that the limits of detection(LOD)for florfenicol,florfenicol amine,ciprofloxacin,enrofloxacin,and sarafloxacin in chicken eggs and poultry muscles were 1.5,0.5,0.05,0.03,and 0.1 μg/kg,respectively.The limits of quantification(LOQ)were accordingly 5.0,2.0,0.1,0.1,and 0.2 μg/kg.The calibration curves of florfenicol,florfenicol amine,ciprofloxacin,enrofloxacin and sarafloxacin standard working solutions demonstrated excellent linearity across the concentration ranges of 10.0-320.0 μg/L(chicken eggs)or 10.0-800.0 μg/L(poultry muscles),4.0-160.0 μg/L(chicken eggs)or 4.0-400.0 μg/L(poultry muscles),0.2-80.0 μg/L,0.2-20.0 μg/L and 0.4-80.0 μg/L,respectively.The peak areas of the target compounds displayed a strong linear relationship with their concentrations,with determination coefficients(R2)were greater than 0.9997.Upon addition of standard concentrations of LOQ,0.5 MRL,1.0 MRL and 2.0 MRL to blank chicken eggs(whole egg,egg yolk and egg albumen),the recoveries of florfenicol,florfenicol amine,ciprofloxacin,enrofloxacin and sarafloxacin were found to be in the range of 83.88%-94.62%,71.95%-82.38%,72.55%-84.10%,86.17%-94.85%and 73.52%-88.27%,respectively.The intra-day relative standard deviations(RSD)were 2.63%-4.82%,2.90%-4.90%,2.42%-5.14%,2.01%-6.59%and 2.48%-6.92%,respectively.While the inter-day RSDs were found to be 2.62%-5.50%2.96%-5.50%,2.47%-6.11%,1.95%-6.46%and 2.29%-6.17%,respectively,the recoveries of florfenicol,florfenicol amine,ciprofloxacin,enrofloxacin and sarafloxacin in blank poultry muscles(chicken muscle,duck muscle,and goose muscle)were 87.82%-94.98%,70.56%-82.22%,71.72%-93.24%,74.62%-92.66%and 70.34%-91.02%,respectively.The intra-day RSDs were found to be 1.66%-3.89%,1.94%-4.39%,1.76%-6.24%,1.46%-4.07%and 1.66%-5.80%,respectively,whereas the inter-day RSDs were 1.60%-3.96%,2.75%-4.41%,1.98%-7.31%,1.96%-4.14%and 1.95%-6.62%,respectively.The methodologies employed in this study were validated to meet the requirements of NY/T 1896-2010 standard of the Ministry of Agriculture and Rural Affairs of China for the detection of veterinary drug residues.The method was successfully applied to actual samples,meeting the technical requirements of different laboratories. |
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