STUDY ON SULPHONAMIDE RESIDUES IN HONEY Teresa Szczêsna 1 , Helena Rybak-Chmielewska 1 , Ewa Waœ 1 , Krystyna Pohorecka 1,2 1 Research Institute of Pomology and Floriculture, Apiculture Division Kazimierska 2, 24-100 Pu³awy, Poland 2 National Veterinary Research Institute, Aleja Partyzantów 57, 24-100 Pu³awy, Poland Received 22 September; accepted 27 November 2009 Summary The aim of the study was to optimise and validate the procedure for sulphonamide residue analyses in honey by HPLC with fluorescence detector. Separation, identification and quantity analysis of sulphonamides in honey were done after synthesis of these compound derivatives with fluorescamine. Procedure of the validation was done by calculation of: detection and quantification limits, repeatability, reproducibility and recovery for each analysed sulphonamide in honey. Hydrolysis parameters (temperature, mixing conditions) of sugar-sulfonamide bonded derivatives for improvement of repeatability and recovery were also determined. Phenomenex Synergi 4μ Fusion HPLC column and chromatographic conditions applied in these studies allowed for satisfactory separation of 4-aminobenzoic acid (PABA), which occurs in some kinds of honey, from sulfamethazine and simultaneous analysis of the following sulfonamides: sulfanilamide, sulfathiazole, sulfacetamide, sulfamethazine, sulfamerazine, sulfamethoxypyridazine, sulfachloropyridazine, sulfamethoxazole, sulfadimethoxine. Detection and quantification limits for all investigated sulphonamides were calculated on 0.2 and 0.3 μg/kg, respectively. In routine analysis quantification limit was settled at 5 μg/kg. Recovery ranged from 37 for sulfacetamide to 93% for sulfanilamide. The coefficient of variation of repeatability ranged from 1.4 for sulfacetamide to 8.1% for sulfadimethoxine and of within- laboratory reproducibility - from 5.1 for sulfanilamide to 16.8% for sulfamethoxypyridazine. Over 2-year period (2007-2008), 241 commercial honey samples were sent to the Bee Products Quality Testing Laboratory. Forty-one of them (17%) contained sulphonamides on a higher level than quantification limit established by the Polish National Monitoring Program for honey (50 μg/kg). Most of these samples were contaminated by three sulphonamides: sulfathiazole, sulfacetamide and sulfamethazine. These results show that Polish beekeepers still use Polisulfamid and other sulphonamide drugs in their beekeeping practices. Keywords: honey, sulphonamides, residues, method, HPLC. INTRODUCTION Honey as a foodstuff of animal origin, is subject for constant monitoring in the presence of forbidden substances. Commission Regulation (EEC) No. 2377/1990 (1990) lists the maximum residue limits (MRL) of veterinary medicinal products in foodstuffs of animal origin. This regulation does not allow antibacterial substances (antibiotics and sulphonamides) in honey. Therefore, the finding of any amount of these substances in honey disqualifies this product. It is true, even though the amounts of the detected contaminants are varied and the results are directly affected by the analytical methods used for this purpose. Generally, the results of honey tests, indicate that beekeepers from EU countries have not ceased to administer drugs Vol. 53 No. 2 2009 Journal of Apicultural Science 39