Determination of the nucleosides and nucleobases in Tuber samples by dispersive solid-phase extraction combined with liquid chromatography-mass spectrometry
Determination of the nucleosides and nucleobases in Tuber samples by dispersive solid-phase extraction combined with liquid chromatography-mass spectrometry
复制标题
分散固相萃取-液相色谱-质谱联用法测定块茎样品中的核苷和核碱基
DOI：
10.1016/j.aca.2010.12.025
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发表时间：
2011-02-21
影响因子：
6.2
通讯作者：
Tang, Ya-Jie
中科院分区：
文献类型：
--
作者：
Liu, Ping;Li, Yuan-Yuan;Tang, Ya-Jie
建立了分散固相萃取（DSPE）联合LC-MS同时测定块茎子实体和发酵菌丝中腺嘌呤、次黄嘌呤、尿嘌呤、腺苷、鸟嘌呤、鸟嘌呤、肌苷等7种核苷和核碱基的方法。首先引入DSPE去除样品溶液中的蛋白质干扰，选择D3520大孔树脂作为DSPE吸附剂，因为它对蛋白质干扰的去除能力高，但对分析物的吸附率低。并对DSPE工艺的关键参数（大孔树脂类型、大孔树脂用量、甲醇浓度、涡流时间）进行了优化，优化后的蛋白去除率可达到95%左右。通过方法验证试验，验证了DSPE-LC-MS方法的精密度、准确度和灵敏度，成功解决了色谱柱的盲化问题。利用所建立的方法，测定了发酵菌丝中核苷和核碱基的总量为4881.5 ~ 12592.9 mu g g(-1)，是子实体（498.1 ~ 2274.1 mu g g(-1)）的2 ~ 25倍。发酵菌丝体中核苷和核碱基的形成相对稳定，而块茎子实体中核苷和核碱基的形成则随菌种的不同而有显著差异。层次聚类分析（HCA）表明，块茎发酵菌丝体与籼稻块茎和喜马拉雅块茎子实体的核苷和核碱基组成相似。从核苷和核碱基的角度，证实了块茎发酵菌丝体作为其子实体替代资源的潜力。(C) 2010 Elsevier B.V.版权所有
A simple, fast and inexpensive method based on dispersive solid phase extraction (DSPE) combined with LC-MS was developed for simultaneous determination of 7 nucleosides and nucleobases (i.e., adenine, hypoxanthine, uridine, adenosine, guanine, guanosine, and inosine) in Tuber fruiting-bodies and fermentation mycelia. The DSPE procedure was firstly introduced to remove the protein interference from sample solutions, and D3520 macroporous resin was chosen as the DSPE sorbent because of its high removal capability on protein interferences, but low adsorption rate on analytes. Besides, key parameters on DSPE procedure (i.e., macroporous resin type, macroporous resin amount, methanol concentration, and vortex time) were optimized, and the protein removal efficacy could achieve about 95% after the process optimization. Though the method validation test, the DSPE-LC-MS method was confirmed to be precise, accurate and sensitive, and the column blinding problem was solved successfully. By using this established method, the total amount of nucleosides and nucleobases in the fermentation mycelia was determined to range from 4881.5 to 12,592.9 mu g g(-1), which was about 2-25 times higher than the fruiting-bodies (from 498.1 to 2274.1 mu g g(-1)). The formulation of nucleosides and nucleobases in the fermentation mycelia maintained relatively constant, while the formulation in Tuber fruiting-bodies varied significantly with their species. Hierarchical cluster analysis (HCA) showed the formulation similarity of nucleosides and nucleobases between Tuber fermentation mycelia and the fruiting-bodies of Tuber indicum and Tuber himalayense. From the viewpoint of nucleosides and nucleobases, this work confirms the potentiality of Tuber fermentation mycelia as the alternative resource for its fruiting-bodies. (C) 2010 Elsevier B.V. All rights reserved.