LEVOFLOXACIN RESIDUES IN CHICKEN MEAT AND GIBLETS

Transcription

1 Bulgarian Journal of Veterinary Medicine (2013), 16, Suppl. 1, LEVOFLOXACIN RESIDUES IN CHICKEN MEAT AND GIBLETS R. KYUCHUKOVA 1, V. URUMOVA 2, M. LYUTSKANOV 2, V. PETROV 2 & A. PAVLOV 1 1 Department of Food Hygiene and Control, Veterinary Legislation and Management, 2 Department of Veterinary Microbiology, Infectious and Parasitic Diseases, Faculty of Veterinary Medicine, Trakia University, Stara Zagora, Bulgaria; Summary Kyuchukova, R., V. Urumova, M. Lyutskanov, V. Petrov & A. Pavlov, Studies on levofloxacin residues in chicken meat and giblets. Bulg. J. Vet. Med., 16, Suppl. 1, Levofloxacin is a third generation fluoroquinolone used in food-producing animals in some countries outside of EU. The aim of the study was to investigate the residues of this antibiotic in chicken tissues (meat, skin and giblets). Chicken (n=30) were treated with levofloxacin orally at a dose of 10 mg/kg BW for 5 days. Birds were divided into 5 groups and humanely killed as followed: on day 0 (the day after last administration of levofloxacin), 2, 4, 6 and 8 day. The results of the studies showed the highest residues levels of levofloxacin in liver (1051 µg/kg), followed by breast muscle, gizzard, heart and skin 428 µg/kg, 321 µg/kg, 303 µg/kg and 293 µg/kg, respectively. The rate of reduction of antimicrobial activity was different. Tissue concentrations in heart and gizzard decreased faster than these in muscles and skin. In the liver they decreased from the last day of the treatment to the 2nd day and the levels remained nearly equal up to the 8 th day after the end of drug administration. Key words: chicken, levofloxacin,residues INTRODUCTION Food-producing animals are treated with a variety of veterinary drugs, including a large number of different types of compounds which can be administered in the feed or in the drinking water. They are applied in animal husbandry for different reasons and may lead to residues in milk, eggs and in other edible tissues. These residues may include the non-altered parent compound as well as metabolites and/or conjugates, and have direct toxic effects on consumers, e.g. allergic reactions in hypersensitive individuals, or antibiotics may cause problems indirectly through selection of resistant strains of bacteria (Fàbrega et al., 2008). For controlling the residue problem, the EU has set maximum residue limits (MRLs) for a variety of veterinary drugs in milk, eggs and tissues (Reig & Toldra, 2008; Petersa et al., 2009). Antibiotics are widely used in all farm animals species and residues are often found in meat, and they should not exceed the maximal residue limits (Okerman et al., 2000). Fluoroquinolones are a group of synthetic antimicrobial agents widely used both in human and veterinary medicine. These agents exert their antibacterial effect through the inhibition of DNA gyrase, interfering with the supercoiling of bacterial chromosomal material. As a result, they have a broad spectrum of activity against Gram-negative and Gram-

2 R. Kyuchukova, V. Urumova, M. Lyutskanov, V. Petrov & A. Pavlov positive bacteria, Mycoplasma spp. and Rickettsia, including those resistant to beta-lactam antibiotics and sulphonamides (Brown, 1996; Ramos et al., 2003). Since their discovery in the early 1960s, the quinolone group of antibacterials has generated considerable clinical and scientific interest. Nalidixic acid, the first used quinolone was obtained as an impurity during the manufacture of quinine. Since that time, many derivatives have been synthesised and evaluated for their antibacterial potency (Andersson & Mac- Gowan, 2003). A number of new fluoroquinolones have become available for use worldwide since the initial introduction of ciprofloxacin in the late 1980s (Appelbaum & Hunter, 2000). Levofloxacin is a third generation fluoroquinolone, an optical isomer of ofloxacin having two-fold higher antimicrobial activity than the parent compound. Currently, it is successfully used in human medicine in the treatment of infections of upper and lower respiratory tract, genitourinary system, skin and soft tissue. This compound has been applied in foodproducing animals (Patel et al., 2009). However, the data about residues after repeated oral administration of levofloxacin in chickens are lacking. Therefore, the present study was planned to investigate the residues of this antibiotic in chicken tissues (meat, skin and giblets). MATERIAL AND METHODS The study was conducted on thirty chickens aged two months. Chickens were treated orally with levofloxacin at a dose of 10 mg/kg BW via the drinking water for 5 days. Water was provided ad libitum. Birds were divided into 5 groups and humanely killed on the day after last administration of levofloxacin (day 0), and on post treatment days 2, 4, 6 and 8. Breast muscle, liver, gizzard, heart and skin (with fats) were separated from each carcass. Samples were weighed and homogenised with Maximum Recovery Diluent (MRD, HIMEDIA, India) in an amount equal to the mass of the sample, then were centrifuged for 15 min at 2500 min -1 (for liver samples 20 min). The supernatant was collected and dropped (100 µl) on a medium with the test microorganism Escherichia coli ATCC It was inoculated on plain agar (HIMEDIA, India), previously sterilized and cooled to 50 C, with concentration of cells 0.5 of McFarland standard. Sterile plates (90 mm) were filled with 14 ml E.coli ATCC infected agar as described by Okerman et al. (1998; 2007). After incubation for 24 h at 37 C, the widths of each inhibition zone were measured from the edge of the sample to the edge of the inhibition zone. Results were processed by GraphPad statistical software. RESULTS AND DISCUSSION The results of the studies are presented in Table 1. The data show that the highest residues levels of levofloxacin were in the liver (1051 µg/kg), followed by breast muscle, gizzard, heart and skin 428 µg/kg, 321 µg/kg, 303 µg/kg and 293 µg/kg, respectively. Decreasing concentrations of residues were found in all investigated tissues up to the 8 th day after treatment. On the second day the levels of gizzard and heart were below the MRL for fluoroquinolones. In breast muscle and skin this was observed on day 4 and in the liver levels remained high until the 8 th day after the end of the treatment (rates with no statistically significant difference vs previous days). BJVM, 16, Suppl

3 Studies on levofloxacin residues in chicken meat and giblets Table 1. Levofloxacin residue levels (µg/kg) in the meat and in the giblets (mean ± SD; n=6) Days after the end of the treatment Tissue samples Muscle 428 ± ± ± ± ± 15 Liver 1051 ± ± ± ± ± 31 Gizzard 321 ± ± ± ± ± 17 Heart 303 ± ± ± 8 30 ± 6 43 ± 10 Skin 293 ± ± ± ± 8 25 ± 7 The rate of reduction of antimicrobial activity was different for the various tissues. Concentrations of levofloxacin in the heart and in the gizzard decreased faster in comparison to the levels in the muscle and in the skin. After initial decrease in the liver up to the 2 nd day after the treatment they remained similar between days 2 and 8. These results were different from our previous research on tissue concentration of gatifloxacin (Kyuchukova & Pavlov, 2012) where the residue levels in muscles, heart and gizzard were below MRL on the second day. Microbiological assays for investigation of antimicrobial residues are considered as multi-residue screening tests for antibiotics in milk, meat or other animal tissues. Karraouan et al. (2009) used a microbiological method for the detection of antibacterial substances in poultry muscles. The method is based on the inhibition of Escherichia coli growth on agar. and can be used as a screening method for the detection of antibiotics in animal tissue. Devada et al. (2012) studied the safety of gatifloxacin after repeated oral administration in broiler chickens and determined tissue concentration of the drug following oral administration. The liver concentration of gatifloxacin was 0.75± 0.04 µg/g after the fourth dose and 0.22±0.07 µg/g after the tenth dose, respectively, whereas in skeletal muscles the concentration of gatifloxacin was below the limit of quantification after the fourth dose. Gatifloxacin was not detected after the tenth dose of. In conclusion it should be noted that the proposed withdrawal period of eight days for levofloxacin is applicable in all tissues with exception of the liver, where relatively high values persisted after the end of the study. Therefore we could state that chicken meat producers have to keep in mind withdrawal period of veterinary drugs used in their farms. REFERENCES Andersson, M. I. & A. P. MacGowan, Dvelopment of the quinolones. Journal of Antimicrobial Chemotherapy, 51, Suppl. S1, Appelbaum, P. C. & P. A. Hunter, The fluoroquinolone antibacterials: Past, present and future perspectives. International Journal of Antimicrobial Agents, 16, Brown, S. A., Fluoroquinolones in animal health. Journal of Veterinary Pharmacology and Therapeutics, 19, Devada, S. S., U. D. Walunj, A. J. Patil, J. H. Patel, S. K. Bhavsar & A. M. Thaker, Safety and tissue residue determination of gatifloxacin in broiler chicken. 218 BJVM, 16, Suppl. 1

4 R. Kyuchukova, V. Urumova, M. Lyutskanov, V. Petrov & A. Pavlov Journal of Advanced Veterinary Research, 2, Fàbrega, A., J. Sánchez-Céspedes, S. Soto & J. Vila, Quinolone resistance in the food chain. International Journal of Antimicrobial Agents, 31, Karraouan, B., B. Bouchrif, N. Ziyate, A. Talmi, K. I. S, Yahia, N. Cohen & A. Fassouane, Evaluation of multi-plate microbial assay for the screening of antibacterial residues in poultry muscle. European Journal of Scientific Research, 35, Kyuchukova, R. & A. Pavlov, Kinetics of the residue levels of gatifloxacin in poultry meat at storage. Days of Veterinary Medicine 2012, 3 rd International Scientific Meeting, Books of Proceedings, p.293. Okerman, L., H. Noppe, V. Cornet & L. Zutter, Microbiological detection of 10 quinolone antibiotic residues and its application to artificially contaminated poultry samples. Food Additives and Contaminants, 24, Okerman, L., K. Wasch & J. V. Hoof, Detection of antibiotics in muscle tissue with microbiological inhibition tests: Effects of the matrix. The Analyst, 123, Okerman, G., K. D. Wasch, & J. V. Hoof, An inhibition test intended to detect and to differentiate between penicillins, cephalosporins, tetracyclines and quinolones, for use in muscle tissue from different animal species. In: Proceedings of Euroresidue IV, Veldhoven 7 10/5, pp Patel, J. H., R. D. Varia, U. D. Patel, P. D. Vihol, S. K. Bhavsar & A. M. Thaker, Safety level of levofloxacin following repeated oral adminstration in White Leghorn layer birds. Veterinary World, 2, Petersa, R. J. B., Y. J. C. Bolcka, P. Rutgersa, A. A. M. Stolkera & M. W. F. Nielena, Multi-residue screening of veterinary drugs in egg, fish and meat using high-resolution liquid chromatography accurate mass time-of-flight mass spectrometry. Journal of Chromatography A, 1216, Ramos, M., A. Aranda, E. Garcia, T. Reuvers & H. Hooghuis, Simple and sensitive determination of five quinolones in food by liquid chromatography with fluorescence detection. Journal of Chromatography B, 789, Reig, M. & F. Toldra, Veterinary drug residues in meat: Concerns and rapid methods for detection. Meat Science, 78, Correspondence: Dr. Ralitsa Kyuchukova Department of Food Hygiene and Control, Veterinary Legislation and Management Faculty of Veterinary Medicine 6000 Stara Zagora, Bulgaria ralitsa.kjuchukova@abv.bg BJVM, 16, Suppl

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6 R. Kyuchukova, V. Urumova, M. Lyutskanov, V. Petrov & A. Pavlov BJVM, 16, Suppl