UHPLC-Q-TOF MS/MS测定稻米代谢物的方法研究

Study on Rice Metabolites Analysis by UHPLC - Q - TOF MS/MS Technique

稻米代谢物成分复杂,受多种环境因素和生长条件的影响,本文建立了一种超高效液相色谱-四极杆飞行时间串联质谱联用技术(UHPLC -Q-TOF MS/MS)测定稻米代谢物的方法,研究了不同前处理条件(包括提取溶剂、提取料液比、提取方式及提取时间)及液相色谱分离条件(包括梯度洗脱条件和色谱柱温)对大米样品中代谢物提取效率和分离效果的影响。在料液比1:5的情况下,50%甲醇经超声提取30min可获得较好的提取效果。确定的UHPLC - Q- TOF MS/MS的色谱条件为:流动相:乙腈-水-甲酸体系,梯度洗脱:23分钟,流速:0.3 mL/min,色谱柱温:30℃该方法具有良好的重复性、日内/日间精密度及线性范围,可以保证稻米代谢物被很好的分离和分析。采用该方法对两种不同稻米样品进行测定,共得到9146个特征峰,其中共有特征峰7843个。以上结果证明实验所建立的UHPLC -Q -TOF MS/MS技术分析稻米代谢物的方法准确、稳定,通过进一步鉴定和识别,可为后续稻米标志性成分的鉴定提供理论依据。

Rice metabolites are complex and affected by various environmental factors and growth conditions. In this manuscript, an ultra - high performance liquid chromatography - quadrupole time - of - flight tandem mass spectrometry ( UHPLC - Q - TOF MS/MS) method was developed for the determination of rice metabolites. The effects of different pretreatments conditions (including extraction solvent, extraction ratio, extraction method and extraction time) and liquid chromatography separation conditions ( including gradient elution and column temperature )on the extraction efficiency and separation efficiency of metabolites in rice samples were studied. In the case of a ratio of material to liquid of 1:5 , 50% methanol was extracted by ultrasonication for 30 min to obtain a better extraction effect. The determined UHPLC - Q - TOF MS/MS conditions were: mobile phase: acetonitrile -water - formic acid system, gradient elution: 23 min, flow rate: 0. 3 mL/min, column temperature: 30T . This method was proven that rice metabolites could be well separated and analyzed with good repeatability, intra 一 day/inter 一 day precision and linear range. Two different rice samples were determined by this method, and a total of 9146 characteristic peaks were obtained, of which 7843 were shared characteristic peaks. The above results demonstrate that the UHPLC - Q - TOF MS/MS technique established by the experiment can accurately and stably analyze rice metabolites. Further identification and identification can provide a theoretical basis for the identification of subsequent rice marker components.