| Fluoroquinolones(FQs)residues in animal food have attracted more and more attention due to the wide use of fluoroquinolones,abuse of fluoroquinolones and irrational use of FQs.In China’s national standard GB31650-2019,the residue limits of the three drugs(enrofloxacin is calculated by the sum of enrofloxacin and ciprofloxacin)are clearly stipulated,which is forbidden for laying hens.In order to ensure the accuracy of fluoroquinolone detection,fluoroquinolone detection technology is particularly important.Several national existing fluoroquinolone detection standards,high performance liquid chromatography mainly uses the traditional homogenate oscillation,the use of low speed centrifugation,the preparation of lipid removal liquid is complex,the complex way of lipid removal makes the processing time is long;The mobile phase used in elution has low acetonitrile content,the volume of elution liquid used is less,the elution is not sufficient,and the volume of purification liquid used when passing the column is too much,which is easy to block the column,resulting in low recovery rate.The amount of samples used in extraction and extraction liquid,the use of lipid removal liquid,resulting in reagent waste;The rotary evaporation method is used to purify,can not batch processing and other problemsIn this paper,the detection of fluoroquinolones residues in livestock,poultry,eggs,aquatic products and milk-HPLC pretreatment technology was studied in detail and improved,the optimization results are as follows:1.By improving the standard of Determination of fluoroquinolones residues in animal Food by HIGH Performance Liquid Chromatography(Announcement14-2008,Ministry of Agriculture,No.1025),homogenate oscillation was optimized to severe vortex,which had no effect on the recovery rate.It took 24 minutes for two homogenate oscillation extraction and only 6 minutes for one vortex extraction.When the ratio of acetonitrile in the eluent was changed,the recovery rate was different,and the elution effect was the best when acetonitrile was 30%.When the volume of purification liquid is reduced,the column passing will be less blocked,improve the column passing time,using 3m L column passing recovery rate is high and the column passing time is short;When the volume of eluted liquid was increased,the eluting time of 2.0m L mobile phase was reasonable and the recovery rate was higher.Column passing of other brands is used,and the recovery rate is not as high as Varian Bond Elut C18 column mentioned in the standard.2.By improving the standard of "Determination of fluoroquinolones residues in eggs-HIGH performance Liquid Chromatography"(Announcement no.781 of the Ministry of Agriculture-6-2006),homogenate oscillation was optimized to severe vortex,which had no effect on the recovery rate.It took 42 minutes for homogenate oscillation extraction and only 12 minutes for vortex extraction.The highest elution recovery was achieved when acetonitrile was 30% in the eluent.The recovery rate was higher and the time was shorter when 3m L was used.The elution time of 2.0m L mobile phase is reasonable and the recovery rate is good.It is more convenient and time saving to absorb the lower extract than to discard the upper fat.Saturated with water,ethyl acetate has a good degreasing effect but a low recovery.It is not suitable to use ethyl acetate in degreasing eggs.3.The standard of determination of Residues of Norfloxacin,ciprofloxacin hydrochloride and enrofloxacin in Aquatic Products(Announcement no.783-2-2006 of the Ministry of Agriculture)was improved,and the recovery rate remained unchanged after the original method was changed from stirring and oscillating to intense vortexing,which took 22 minutes for stirring and oscillating and 3 minutes for vortexing;With 2.00 g sample and 20 m L acidified acetonitrile,the recovery rate was almost unchanged,but the use of 40 m L acidified acetonitrile was saved.Without anhydrous sodium sulfate,the recovery rate was not affected,but 30 g of anhydrous sodium sulfate was reduced.The rotary evaporation is changed to 40 ℃ constant temperature water bath nitrogen blowing,has no effect on recovery,can greatly reduce the time cost,suitable for batch sample processing;The extraction solution was concentrated and dried with flow compatibility solution,and then the fat was removed.The use of n-hexane was reduced by 22 m L,and the recovery rate was not affected.The elution time of 2.0m L mobile phase is reasonable and the recovery rate is good.When centrifugation in this standard was optimized to 10000 r/min for 5min,the treatment of one sample could save 30 minutes and the recovery rate was not affected.4.By improving the standard of "Determination of fluoroquinolones residues in milk-HIGH performance Liquid Chromatography"(GB 29692-2013),the original standard was changed from 8m L acetonitrile extraction to 20 m L acetonitrile extraction,with the highest recovery rate;The rotary evaporation is changed to 40℃constant temperature water bath nitrogen blowing,which has no effect on the recovery rate and saves time,and is suitable for batch sample processing.The elution time of 2.0m L mobile phase was reasonable and the recovery was good. |
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