Study On Determination Of Tetracyclines Residue By New HPLC-CL Method

Objective: It is relatively common phenomenon that abusing tetracycline antibiotic drug and unreasonable application in China which is regarded as a major producer of antibiotics.The resulting antimicrobial resistance,animals products with drug residues,,allergic reactions and other hazards become increasingly obvious and has a direct threat to human health and safety.The aim of this study was to establish a new HPLC-CL detection method for the simultaneous determination of tetracycline(TC),oxytetracycline(OTC),chlortetracycline(CTC)and methyl oxytetracycline(MTC)and to research the application of this method in the aspect of food safety.Methods: Tetracycline antibiotics were determined by HPLC-CL under optimal reaction conditions.Firstly,the tetracycline sample enters the high performance liquid system through a manual injector and then it was fully separated under isocratic elution on C18 column.Secondly,the post-column effluent which was needed to be qualitative by UV detector was first mixed with the MCLA solution and the mixture of both met with Ce(IV)solution to generate chemiluminescent in the flow cell.Finally,the photomultiplier amplified chemiluminescent signal and converted into electrical signal which could be recorded by computer.The corresponding parameters were set as follows: The mobile phase was a solution of 3.5 x 10-3 mol / L CH4O3 S in acetonitrile-water(20:80,v: v);flow rate at 0.85mL/min,column temperature at 30°C and detection wavelength at 270 nm.Results: The detection limits of tetracycline(TC),oxytetracycline(OTC),chlortetracycline(CTC)and methyl oxytetracycline(MTC)were 2.5 × 10-9mol/L,4.1 × 10-9mol/L,1.0 × 10-8mol/L,4.5 × 10-9 mol/L,and their relative standard deviations were 3.54%-8.76%(n=7).The recoveries were 84.85%-111.00% in pork samples and the relative standard deviation ranged from 3.85% to 10.87%.The recoveries were 76.49%-115.7% in milk samples and the relative standard deviation ranged from 4.17% to 10.70%.Conclusion: In this work,a new HPLC-CL method for the rapid determination of tetracycline antibiotics in food was established.Compared with other systems,the elution method is simple and the mobile phase is simplified.The selectivity has been improved and the interference problem of post-column effluent has been solved in comparison to HPLC-UV;the cost of detection is reduced in comparison to HPLC-MS.A cursory experiments have clarified the mechanism of this method,which provides a theoretical basis for the development of chemiluminescence technology.Through the actual sample detection,it is shown that the method has the advantages of simple operation,rapid analysis,good selectivity,high sensitivity and satisfactory recovery.So this method has a promising application prospect.