Hydrophilic Interaction Liquid Chromatography-tandem Mass Spectrometry For The Determination Of Glyphosate In Agricultural Products

The retention and separation of glyphosate and its major metabolite aminomethylphosphoric acidwith strong polarity and hydrophilic characteristics has beening one of challenges for chromatographers.And matrix effects caused by complicated samples as well as other issues also bring more difficultiesfor their analysis. To a certain extent, these methods like ion exchange chromatography, ion pairchromatography and using gas chromatography or liquid chromatography after derivatization caneffectively improve the separation effect, but they also have many problems, such as poor compatibilitywith mass spectrometry detector, tedious pretreatment operation and froward derivative process. So theeffective way to solve technical analysis problems of glyphosate and its metabolite is to choose asuitable separation mode to efficiently separate them from the sample matrix disruptors, and simplifythe sample pretreatment operations as far as possible. Here we established a HILIC-MS/MS methodwith Shodex Asahipak NH2P-504E chromatography column and polymer matrix amino solid phaseextraction cartridge to analysis glyphosate and aminomethylphosphoric acid residues in agriculturalproducts. Research results were as follows:1.A method of HILIC-MS/MS was studied and established for the determination of glyphosate andaminomethylphosphoric acid with hydrophilic interaction liquid chromatographic technology. The bestconditions of mass spectrum were established. The results showed that compounds should be monitoredin negative mode and obtained their characteristic ions. The influences of five chromatographic columns(Obelisic N, ZIC-HILIC, Agilent Zorbax NH2, Asahipak ODP-50and Shodex Asahipak NH2P-504E)on retention behavior of target analytes were investigated. And the result showed that Shodex AsahipakNH2P-504E was the best of all. Analytes on this column could get proper retention time and goodchromatographic peak shape. Mobile phase pH, buffer concentration and initial proportion of strongelution solvent were important factors affecting the retention behavior of target analytes. The finalchromatographic separation conditions were with the mobile phases of aqueous solution ofpH=11.2-acetonitrile in gradient elution, the flow rate of300μL/min and column temperature of25℃.2.A sample pretreatment method was established. The compounds should be extracted with purewater and purified with polymer matrix amino solid phase extraction cartridge. The results showed thatusing pure water as extraction solvent, grain samples crushed over1.70mm sieve mesh, the ratio ofsample quality and extraction solvent volume1:5with homogenate extraction for two times, while fruitand vegetable samples using the same extraction method but with only one extraction time, could obtainthe maximum sample extraction rate and recovery rate. When extraction solution was diluted withacetonitrile until the percentage of sample matrix volume taking up less than20%, polymer matrixamino solid phase extraction could achieve effective adsorption of the analytes. The eluent containing20%acetonitrile in aqueous solution was used to wash away impurities, and finally eluted with aqueoussolution containing5%ammonia. This method had the advantages of simple operation and goodpurifying effect.3.The consequences of matrix effects on the analytical results were evaluated. The results showedthat the existence of matrix effects had a significant impact on the sensitivity and accurate quantitation.It mainly depended on the marix medium characteristics and the spiked levels. The matrix inhibitioneffects in rice and corn were weak, in wheat and soybean were strong, while in cabbage, carrot, appleand other fruit and vegetable samples were extremely strong. In addition, the lower the concentrationwas, the stronger the matrix effect was. We could reduce the precence of coeluting substances in matrix by polymer matrix amino solid phace extraction which had a good separation selectivity. And thepurification recoveries were82%-110%.4.The analytes in real samples were detected by the established determination method, using isotopeinternal standard method for quantitative analysis. The mean spiked recoveries of Gly and AMPA wereranged from63%to110%at3spiked levels (0.050,0.100and0.500mg/kg), and the relative standarddeviations (RSDs, n=3) were lower than16%. Under the optimally analytical conditions, the linearitiesof Gly and AMPA were in the concentration range of2.5to500μg/L and0.5to500μg/L respectively,with the correlation coefficients both0.9995. The limits of detection(S/N≥3) of the method were2-10μg/kg for grain samples and1-5μg/kg for fruit and vegetable samples.HILIC-MS/MS analytical method were established in this study with polymer matrix amino solidphase extraction cartridge for purification, hydrophilic interaction chromatography for separation andisotope internal standard method for quantitation. The method had the characteristics of high sensitivity,high recovery, without derivatization, easy operation, and accuracy in qualitative analysis, which wasapplicable to quantitative of glyphosate and its metabolite aminomethylphosphoric acid in mostagricultural products and confirmatory analysis.