三重四级杆串联质谱法测定乳粉类产品中 5种核苷酸的含量
目的 建立超高效液相色谱-三重四级杆串联质谱法测定乳粉类产品中5种核苷酸, 包括胞嘧啶核苷酸(cytidine monophosphate, CMP)、尿嘧啶核苷酸(uridine monophosphate, UMP)、腺嘌呤核苷酸(adenosine monophosphate, AMP)、鸟嘌呤核苷酸(guanosine monophosphate, GMP)、次黄嘌呤核苷酸(inosine monophosphate, IMP)含量的分析方法。方法 样品经温水溶解后, 经1% (V:V)甲酸沉淀蛋白离心后, 分别通过固相萃取SPE法和超滤离心法有效净化其他干扰成分, 从而进行前处理方法评价; 采用Waters ACQUITY UPLC HSS T3(2.1 mm×50 mm, 1.8 μm)为分析柱, 0.1%(V:V)甲酸水溶液和乙腈作为流动相, 等度洗脱分离。质谱采用正离子模式, 多反应监测模式(MRM)进行检测, 对纯水配制标准溶液和阴性基质溶液配制标准溶液进行比较, 外标法定量分析。结果 5种核苷酸在4.5 min内达到基线分离, AMP、IMP和GMP在10~1000.0 ng/mL浓度范围内线性关系良好, CMP和UMP分别在25~1000 ng/mL和100~1000 ng/mL浓度范围内线性关系良好, 相关系数均在0.9990以上, 方法检出限分别为UMP 5 mg/kg、 CMP 1 mg/kg、AMP 0.5 mg/kg、IMP 0.5 mg/kg、GMP 0.1 mg/kg。在10.0、50.0、100.0 mg/kg加标水平下的超滤离心法的平均回收率在73.6%~110.9%之间, 相对标准偏差(RSD)为3.56%~15.89%; SPE法的平均回收率在86.4%~109.5%之间, 相对标准偏差(RSD)为2.32%~14.45%。结论 该方法灵敏度高, 可以快速准确鉴别并定量乳粉产品中的核苷酸。
Objective To establish a quantitative method for determination of 5 kinds of nucleotides, including cytidine monophosphate (CMP), uridine monophosphate (UMP), adenosine monophosphate (AMP), guanosine monophosphate (GMP), inosine monophosphate (IMP), in milk powder products by high-performance liquid chromatography-tandem triple quadrupole mass spectrometry (HPLC-MS/MS). Methods After hydrolysis with warm water, the milk sample was deproteinated with 1% (V:V) formic acid solution. Then the supernatant was cleaned up by PRiME HLB (Waters) solid phase extraction (SPE) column or ultrafiltration spin tube (cut off 3kDa, Pall). The analysis was performed on a Waters ACQUITY UPLC HSS T3 column(2.1 mm×50 mm,1.8 μm) using 0.1% (V:V) formic acid and acetonitrile as the mobile phase, isocratic elution. The samples were determined by mass-spectrography in the positive ion mode with the multiple reaction monitoring mode. Matrix effect was evaluated by comparing 2 sets of external standard curves which were diluted by water and negative matrix respectively, with quantitative analysis by the external standard method. Results Five kinds of nucleotides could be separated within 4.5 min, AMP, IMP and GMP were all linear in the concentration range of 10~1000.0 ng/mL, and CM and UMP were linear in the concentration range of 25~1000 ng/mL and 100~1000 ng/mL respectively (r2>0.9990). The limit of detection of UMP, CMP, AMP, IMP, and GMP were 5, 1, 0.5, 0.5, and 0.1 mg/kg, respectively. The recoveries of spiked samples at 3 levels (10.0, 50.0, 100.0 mg/kg) ranged from 73.6% to 110.9%, with RSD of 3.56%~15.89%, which samples were pretreated by ultrafiltration method. Alternatively, the recoveries ranged from 86.4%~109.5%, RSD 2.32%~14.45%, which samples were passed through SPE column. Conclusions The developed method is sensitive. Five kinds of nucleotides can be identified and quantified by this method effectively.
标题：三重四级杆串联质谱法测定乳粉类产品中 5种核苷酸的含量
英文标题：Determination of 5 kinds of nucleotides in milk powder by triple quadrupole mass spectrometry
作者：
孙姗姗 中国食品药品检定研究院
刘文婧 中国食品药品检定研究院
张隆龙 中国食品药品检定研究院
丁艺 中国食品药品检定研究院
王海燕 中国食品药品检定研究院
中文关键词:核苷酸,三重四级杆串联质谱,PRiME HLB,超滤离心,乳粉,
英文关键词:nucleotides,triple quadrupole mass spectrometry,PRiME HLB,ultrafiltration,milk powder,
发表日期：2015-11-12
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