Detection Technology Of Vancomycin And Norvancomycin In Eggs And Honey By High Performance Liquid Chromatography-Tandem Mass Spectrometry

Presently,antibiotic residues in poultry products are highly worried by consumers.Vancomycin and norvancomycin are classified as glycopeptide antibiotics,in fame of"last-line drugs"for more powerful than most of other antibiotics which is widely used to treating bacterial infections and promoting animal growth.The veterinary use of vancomycin is prohibited in China,and is listed as“non-edible substances that may be illegally added”.There are many reports on the detection methods of vancomycin in meat products,feed,milk and milk powder at home and abroad.The detection methods of vancomycin and norvancomycin in eggs and honey have not been reported.In this paper,the HPLC-MS\MS method of vancomycin and norvancomycin in eggs and honey was studied.This method has good specificity and high sensitivity.It is suitable for vancomycin and nail in poultry eggs and honey.Detection of vancomycin.High Performance Liquid Chromatography-Tandem Mass Spectrometry?HPLC-MS\MS?is a high-resolution separation of complex components of the matrix by a high-resolution liquid chromatography?HPLC?and a quasi-deterministic and high-sensitivity detection and analysis method for mass spectrometry?MS?.Suitable for the detection of vancomycin and norvancomycin in eggs and honey.In this experiment,we used high performance liquid chromatography-tandem mass spectrometry?HPLC-MS\MS?to detect vancomycin and norvancomycin in eggs and honey,on the one hand,to provide data reference for the study of relevant national standards,on the other hand,to provide technical support for food safety supervision,timely monitoring and early warning.?1?The main research contents and results of the experiment are as follows:Different extract ratio test.0.1%formic acid Water was used and acetonitrile were 2:8,3:7,4:6,5:5,7:3,9:1,10:0,according to HPLC-MS\MS initial conditions to detect vancomycin and norvancomycin.The results showed that the extraction effect of 5:5 and 10:0 ratio was optimal,and 0.1%formic acid water was determined as the extract of vancomycin and norvancomycin.?2?Purification process.Comparison of six different mixed cation exchange solid phase extraction columns for the purification of vancomycin and norvancomycin.Due to the complexity of the sample matrix,the turbidity of the sample solution caused the difficulty of the extraction fluid,and the retention agent was treated with PSA adsorbent when the egg sample was treated.The recovery rate of PCX mixed cationic exchange solid phase extraction column as purification column was better by screening.?3?Two different eluent tests were used to analyze the elution profile to determine the amount of eluent.It was confirmed that the mixed cation exchange solid phase extraction column was eluted with 3 mL of 10%ammonia-methanol?V/V?.?4?Two different manufacturers of C₁₈ columns were selected to verify the difference in separation columns of the same source and different sources,and a C₁₈ column was recommended to test separation effect two mobile phases of 0.1%formic acid with acetonitrile?V/V?and 0.1%formic acid with methanol?V/V?.The column selection X-C₁₈ column was determined based on the chromatograms of the two C18 columns.The parameters were 2.1mm×150mm,3.5?m.The chromatographic mobile phase was determined to be eluted with 0.1%aqueous formic acid and acetonitrile?V/V?.The mass spectrometric conditions were determined by mass spectrometer setting the electrospray ionization?ESI?ion source and selecting the positive ion multiple reaction monitoring?MRM?mode to detect the target ions,and Determine m/z 144.1 and m/z 144.0 as quantitation ions for vancomycin and norvancomycin.?5?The test confirmed the method by matrix effect,method specificity,linear range,limit of quantitation,recovery,precision and stability.It showed that the method was linear relation in the range of 5 to 100?g/L and the correlation coefficient was greater than 0.999 by the standard curve drawn from the test results.The limit of quantitation?LOQ?of the method was 5?g/kg,the average recoveries of vancomycin and norvancomycin in the sample were70.19%⁹4.02%and 70.89%⁹3.36%,respectively,and the RSD%were 2.25%-3.13%and3.34%-5.57%,respectively.It showed that the method was highly accurate.?6?Based on the HPLC-MS\MS method established in this study,random sampling of eggs and honey in Changchun City was carried out to find no residual vancomycin and norvancomycin.It was shown that Changchun was not a contaminated area of vancomycin and norvancomycin.In this experiment,the efficient extraction and purification methods on vancomycin and norvancomycin in eggs and honey were optimized for HPLC-MS\MS method detection,so that was applied in practical samples test and found non positive residues of vancomycin and norvancomycin in Changchun.