【摘要】 该研究利用超高效液相色谱-串联四极杆静电场轨道阱高分辨质谱(UHPLC/ESI-Q-Orbitrap)技术,对硒代蛋氨酸(Se Met)的色谱信息、分子离子质荷比和碎裂片段的质荷比进行采集,并对特征离子碎裂途径进行解析,建立了豆类中硒代蛋氨酸的检测方法。样品用三羟甲基氨基甲烷-盐酸(Tris-HCl)缓冲溶液溶解后,涡旋混匀,超声提取,在恒温水浴条件下酶解,离心后取上清液过0.22μm滤膜后上机检测。采用Hypersil GOLD HILIC(50 mm×2.1 mm,1.9μm)色谱柱进行分离,以0.2%(体积分数,下同)甲酸6 mmol/L甲酸铵水溶液和0.2%甲酸6 mmol/L甲酸铵乙腈溶液为流动相梯度洗脱,采用电喷雾正离子模式电离,在全扫描/数据依赖扫描模式(Full MS/dd-MS2)下进行检测,基质匹配标准校正法定量。结果表明,硒代蛋氨酸的基质效应为15.75%,在0.05～0.5 mg/L范围内线性关系良好,相关系数(r~2)为0.997 6,方法检出限(LOD)为0.015 mg/kg,定量下限(LOQ)为0.05 mg/kg;空白样品在0.1、0.2、0.4 mg/kg 3个加标水平下的平均回收率为77.6%～83.2%,日内相对标准偏差(RSDr)为2.8%～4.8%,日间相对标准偏差(RSD_R)为4.1%～6.5%。将方法应用于实际样品的检测,得富硒黑豆、富硒红豆、富硒绿豆中硒代蛋氨酸的含量分别为0.252、0.163、0.184 mg/kg。该方法具有前处理操作简单、结果准确、重复性好等优点,适用于豆类中硒代蛋氨酸的检测。更多还原

【Abstract】 An ultrahigh performance liquid chromatography coupled with quadrupole orbitrap mass spectrometry(UHPLC/ESI-Q-Orbitrap)was established for the detection of selenomethionine in beans,by collecting the chromatographic information,mass charge ratio of molecular ion and mass charge ratio of fragmentation fragments of selenomethionine(Se Met)and analyzing the fragmentation pathway.The samples were firstly dissolved with Tris-HCl buffer solution,then blended well with a vortex mixer,followed by ultrasonic extraction and enzymatic hydrolysis under constant temperature bath with trypsin and protease K,and finally centrifuged.The supernatant was filtered through a 0.22μm microporous membrane,and then determined using the UHPLC/ESI-Q-Orbitrap system.The extract was separated on an Hypersil GOLD HILIC column(50 mm×2.1 mm,1.9μm)by gradient elution,using 0.2%(volume fraction)formic acid-6 mmol/L ammonium formate aqueous solution and 0.2%formic acid-6 mmol/L ammonium formate acetonitrile solution as the mobile phases.The Se Met was determined in positive electrospray ionization mode(ESI) under Full MS/dd-MS~2 acquisition mode,and quantified by matrix-matched standard calibration method due to the existence of matrix effect.The matrix effect for Se Met was 15.75%,evaluated by the ratio of solvent standard curve and matrix-matched standard curve.Under the optimal conditions,there was a good linear relationship for Se Met in the concentration range of 0.05-0.5 mg/L with a correlation coefficient(r~2)of 0.997 6,while the limit of detection(LOD)and the limit of quantification(LOQ)were 0.015 mg/kg and 0.05 mg/kg,respectively.At three spiked levels of 0.1,0.2 and 0.4 mg/kg,the average recoveries for Se Met in mung bean ranged from 77.6%-83.2%,with the intra-day relative standard deviations(RSDr)and the inter-day relative standard deviations(RSDR)of 2.8%-4.8%and 4.1%-6.5%,respectively.This method was applied to the detection on actual samples collected in laboratory.The results showed that the contents of selenomethionine in selenium-enriched black bean,selenium-enriched red bean and selenium-enriched mung bean were 0.252,0.163 and 0.184 mg/kg,respectively.With the advantages of simple pretreatment,accurate result and good repeatability,this method is suitable for the detection of Se Met in beans.更多还原