Croubels, Siska

De Backer, Patrick

De Baere, Siegrid

Devreese, Mathias

English

Ghent University Academic Bibliography

5-6-20141INTESTINAL CONCENTRATIONS OF ENROFLOXACIN AFTER DIFFERENT DOSAGE REGIMENS IN BROILER CHICKENS Mathias Devreese*, Siegrid De Baere, Patrick De Backer and Siska CroubelsDepartment of Pharmacology, Toxicology and Biochemistry, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium* Corresponding author: e-mail: mathias.devreese@ugent.beThe posology of veterinary antimicrobial drugs is currently determined by dose titration and confirmation studies solely monitoring clinical efficacy. Thesestudies generally do not take into account the objective of limiting the emergence and spread of resistance. Nevertheless, resistance to antimicrobial drugsis one of the leading health concerns in human and veterinary medicine worldwide. As resistance development and selection depends on the extent ofexposure to antimicrobials, it is of importance to know to which amount of drug the intestinal microflora is exposed after treatment.The aims of present study are to: 1/ develop a LC-MS/MS method for quantitation of enrofloxacin in intestinal content2/ assess and compare the intestinal concentration of enrofloxacin after treatment of broiler chickens with theconventional dosage regimen, after dose escalation and with different administration routes (oral vs. intramuscular)Sample clean-up: To 1 g of intestinal content were added the internal standard (sarafloxacin), 3 mL PBS buffer and 5 mL ethylacetate. After extraction andcentrifugation and evaporation to dryness, samples were resuspended in 250 µL of H2O.Chromatography: Zorbax® Eclipse plus C18 (100 mm x 3.0 mm i.d., d.p.: 3.5 μm) (Agilent). Mobile phases: (A) 0.1% glacial acetic acid in H2O (B) acetonitrile.Mass spectrometry: TSQ® Quantum Ultra triple quadrupole MS (Thermo Scientific) operated in the + ESI mode.SRM transitions for identification and quantification (*): enrofloxacin: m/z 360.0 > 316.2*/245.1, sarafloxacin: 386.1 > 368.0/299.0*Animal experiment: Ninety-six animals were divided in 4 groups, each allocated to a different dosage regimen of enrofloxacin (Baytril®):Group 1: conventional treatment: 10 mg/kg BW, 5 days, oral (by oral gavage) Group 3: 10 mg/kg BW, 5 days, intramuscularGroup 2: 50 mg/kg BW, 5 days, oral (by oral gavage) Group 4: 50 mg/kg BW, 5 days, intramuscularAt 2 and 4 h after first administration, and 4 h after the last administration (100 h after first administration), intestinal content (ileum, cecum, colon andcloaca) was collected from eight animals/group. Samples of the eight chickens/sampling point were pooled and 1 g aliquots were extracted in triplicate forquantitative analysis.Results and DiscussionIntroduction and AimsMaterials and MethodsMethod validation (as described by [1], [2]): results fell within the specified ranges: Linearity: concentration range 0.1 – 20 µg/g, r ≥ 0.99, g ≤ 10% Accuracy: within -20% to +10% of theoretical concentration (0.1, 0.5 and 10 µg/g) Within-day precision: RSD < RSDmax of 15.1% (0.1 µg/g), 11.8% (0.5 µg/g), 7.5% (10µg/g) Between-day precision: < RSDmax of 17.8% (0.5 µg/g), 11.3% (10 µg/g) LOQ: 0.1 µg/g LOD: S/N ≥ 3 (0.07 ng/g) Specificity: no interferences Matrix effect: 73.2%Animal experiment Enrofloxacin concentrations in ileum, cecum, colon and cloaca are presented in Figure 2 High concentrations (µg/g range) were observed after oral as well as intramuscularadministration Intestinal concentrations are dose dependent After oral administration, a time dependent shift was observed from ileum (2 h) to moredistal intestinal segments (4 and 100 h) after oral administration Following intramuscular administration, the highest concentrations were detected in thecloaca, most probably due to mainly urinary excretion of enrofloxacin Future in vitro experiments can be performed to evaluate the effect of the differentenrofloxacin concentrations on resistance selection in the intestinal microbiotaFigure 1. LC-MS/MS chromatogram showing the SRM traces of enrofloxacin(ENRO) and sarafloxacin (SARA) in standard solution (0.5 µg/mL)[1] De Baere S. et al. (2011). Journal of Chromatography B, 879, 2403-2415. [2] 2002/657/EC. Official Journal of the European Union, L221.0100200300400500600ileum cecum colon cloacaEnrofloxacinconcentration (µg/g)Group 10100200300400500600ileum cecum colon cloacaGroup 20100200300400500600ileum cecum colon cloacaGroup 42 h4 h100 h0100200300400500600ileum cecum colon cloacaGroup 3Figure 2. Enrofloxacin concentrations  in different parts of the intestinal tract (ileum, cecum, colon and cloaca) after oral (Group 1 and 2) or intramuscular (Group 3 and 4) administration of 10 mg/kg BW (Group 1 and 3) or 50 mg/kg BW (Group 2 and 4) to broiler chickensReferences

Intestinal concentrations of enrofloxacin after different dosage regimens in broiler chickens

https://biblio.ugent.be/publication/4410075/file/4410076

Intestinal concentrations of enrofloxacin after different dosage regimens in broiler chickens

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