Determination Of Tetracycline Antibiotics In Honey By High Performance Liquid Chromatography

In recent years,with the constant increase of honey consumption in the world,the safety of honey have been seriously concerned.Tetracycline antibiotics is a kind of broad spectrum antibiotic.In bee industry,TCs are commonly added to the feedstuff for the treatment of bee chalkbrood diseases and foulbrood diseases or for the growth promotion.Therefore,trace residues can be found in honey.The concentration of TCs residues in honey are extremely low and the honey sample is complex.In sense,how to establish a TCs analysis method with ideal specificity and high sensitivity is critical to evaluate the TCs residues.This study mainly includes:(1)The determination of TCs in honey by Matrix solid-phase dispersion-High performance liquid chromatography(MSPD-HPLC)A new method of detecting the contents of TCs residues in the honey by matrix solid phase dispersion-high performance liquid chromatography was established.The experimental conditions for the MSPD and HPLC were studied and optimized.In MSPD,silica was selected as dispersion absorbent.The best mass radio of honey to silica was 1:4,with 6mL acetic acid/methanol(5/95,v/v)as elution.Chromatographic conditions were as follows:Mobile phase was acetonitrile/methanol/0.01 Mcitric acid(20/10/70,v/v/v),detection wavelength was 355nm,the column was maintained at 35℃ and the flow rate was fixed at 0.8 ml/min.The result showed that the linearity was fine at 0.1～20μg/mL concentration range.The average recoveries of OTC,TC and CTC were 83.27%～92.61%,88.26%～95.32%,87.33%～96.98%,respectively.(2)The preparation of molecular imprinted polymer(MIP)In this research,a novel surface molecular imprinted polymer(MIP)was synthesized by using silica as carrier,Oxytetracycline as template molecule,Methyl methacrylate(MAA)as functional monomer,Ethylene dimethacrylate(EDMA)as cross-linker and Azodiisobutyronitrile(AIBN)as initiator.The dosages of reagents in the polymerization reaction were 0.1282g OTC,140mL acetonitrile,0.130mL MAA,1.413mL EDMA and 5mg AIBN.The MIP was proved to be successful by scanning electron microscope(SEM)and Fourier transform infrared(FT-IR)analysis.The specific adsorption of the MIP was tested by adsorption isotherms,kinetic curve and selective adsorption experiments.The results showed that the TCs absoptive MIP were successfully synthesized.(3)The determination of TCs in honey by MIPSE-HPLCA molecularly imprinted solid phase extraction(MIPSE)coupled with HPLC method was developed to detect TCs in honey.The experimental conditions for the MIPSE were confirmed as follows:The mass of MIPs was 300mg,5mL methanol/water(5/95,v/v)was used as washing solution and 6mL methanol/ethyl acetate(20/80,v/v)as elution.The proposed method has been successfully applied to preconcentrate and determine of TCs in honey due to its celerity,accuracy and precision.