TiO2-based solid phase extraction strategy for highly effective elimination of normal ribonucleosides before detection of 2'-deoxynucleosides/low-abundance 2'-O-modified ribonucleosides.
TiO2-based solid phase extraction strategy for highly effective elimination of normal ribonucleosides before detection of 2'-deoxynucleosides/low-abundance 2'-O-modified ribonucleosides.
复制标题
DOI：
10.1021/ac4025297
复制
发表时间：
2013-10
影响因子：
7.4
通讯作者：
Shaoting Wang;Wei Huang;W. Lu;Bifeng Yuan;Yuqi Feng
中科院分区：
文献类型：
--
作者：
Shaoting Wang;Wei Huang;W. Lu;Bifeng Yuan;Yuqi Feng
开发了一种新的基于TiO 2的固相萃取策略，用于在质谱（MS）分析2 '-脱氧核苷和2'-O-修饰的核糖核苷之前去除正常的核糖核苷。利用TiO 2分离柱对核糖核苷和2 '-脱氧核苷在TiO 2材料上的色谱保留行为进行了研究。结果表明，一个特定的亲和相互作用机制之间的二氧化钛和顺式二醇含核糖核苷，和相互作用被证明是有效的，即使在很宽的pH值条件和盐浓度。利用这些特点，我们开发了一种基于TiO 2的固相萃取（SPE）方法，用于高效去除基因组DNA中的RNA污染。与广泛使用的酶消化方法相比，本文提出的基于TiO 2的SPE方法对RNA的去除效率更高，并且对2 '-脱氧核苷的回收率也更高。此外，与传统的酶消化方法（~12 h）相比，使用基于TiO 2的SPE方法（~5 min）可显著缩短样品处理时间。最后，在HeLa细胞中成功实现了基于TiO 2的固相萃取法从RNA中纯化2 '-O-甲基化核糖核苷，为从大体积正常核糖核苷中纯化相关修饰核糖核苷提供了概念验证。总之，本研究中开发的这种策略提供了一种有前途的选择，通过消除生物样品中的正常核糖核苷来纯化2 '-脱氧核苷/2'-O-修饰的核糖核苷，以进行灵敏和准确的测定。
A novel TiO2-based SPE strategy was developed for eliminating normal ribonucleosides before mass spectrometry (MS) analysis of 2'-deoxynucleosides and 2'-O-modified ribonucleosides. The chromatographic research for the retention behavior of ribonucleosides and 2'-deoxynucleosides on TiO2 materials was investigated using TiO2 separation column. The results indicated a specific affinity interaction mechanism between TiO2 and cis-diol-containing ribonucleosides, and the interaction was proved effective even under a wide range of pH conditions and salt concentrations. Benefiting from these features, a TiO2-based solid phase extraction (SPE) method was developed for highly efficient elimination of RNA contamination from genomic DNA. Compared with the widely used enzymatic digestion method, the proposed TiO2-based SPE method showed much more efficiency for the removal of RNA as well as provided high recoveries for the 2'-deoxynucleosides. In addition, the sample processing time is dramatically shortened using the TiO2-based SPE method (~5 min) compared to the traditional enzymatic digestion method (~12 h). Finally, the purification of 2'-O-methylated ribonucleosides from RNA was successfully achieved in HeLa cells by the TiO2-based SPE method, which provided a proof-of-concept for the purification of relevant modified ribonucleosides from bulky normal ribonucleosides. Taken together, this strategy developed in the current study offers a promising option to purify 2'-deoxynucleosides/2'-O-modified ribonucleosides for their sensitive and accurate determination by eliminating normal ribonucleosides in biological samples.