Metal Oxide-Based Selective Enrichment Combined with Stable Isotope Labeling-Mass Spectrometry Analysis for Profiling of Ribose Conjugates
Metal Oxide-Based Selective Enrichment Combined with Stable Isotope Labeling-Mass Spectrometry Analysis for Profiling of Ribose Conjugates
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基于金属氧化物的选择性富集结合稳定同位素标记质谱分析核糖缀合物的分析
DOI：
10.1021/acs.analchem.5b01614
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发表时间：
2015-07-21
影响因子：
7.4
通讯作者：
Feng, Yu-Qi
中科院分区：
文献类型：
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作者：
Chu, Jie-Mei;Qi, Chu-Bo;Feng, Yu-Qi
生物体液中的一些修饰的核糖核苷已被评估为癌症相关代谢物。生物体液中内源性修饰核糖核苷的检测可作为一种非侵入性的癌症诊断方法。然而，由于修饰核糖核苷的丰度低且存在严重的基质干扰，因此其测定仍然具有挑战性。在这里，我们开发了一种新的策略，用于全面的，分析核糖共轭物从生物流体使用金属氧化物为基础的分散固相萃取（DSPE），然后与体外稳定同位素标记和双中性损失扫描质谱分析（DSPE-SIL-LC-DNLS-MS）。采用二氧化铈（CeO 2）在碱性环境下对复杂生物样品中的核糖结合物进行选择性识别和捕获。随后用一对同位素标记试剂（丙酮和丙酮-d（6））标记富集的核糖缀合物;丙酮标记的核糖缀合物的糖苷键容易断裂，并且所产生的携带同位素标签的核糖可以在碰撞诱导解离（CID）下作为中性片段丢失。由于轻（丙酮）和重（丙酮-d 6）标记的化合物具有相同的化学结构，并且可以产生不同的中性损失片段（NL 172和178 Da），因此在质谱分析中通过双中性损失扫描模式来分析核糖缀合物非常方便。在这方面，轻和重标记的化合物在相同条件下电离，但在MS Spectra上分别记录，这可以显著提高检测特异性并有助于鉴定核糖缀合物。采用DSPE-SIL-LC-DNLS-MS方法，对人尿液中核糖结合物进行了分析，共鉴定出49种核糖结合物，其中7种核糖结合物在正常人和淋巴瘤患者尿液中的含量有显著性差异。DSPE-SIL-LC-DNLS-MS策略结合了选择性富集、稳定同位素标记和双中性丢失扫描- MS分析，因此可以有效地减少假阳性结果，促进相对定量，并显著增加生物流体样品中鉴定的核糖缀合物的数量。总之，本研究建立了一个有前途的策略，有效的分析尿修饰核糖核苷，和多个修饰核糖核苷的含量变化的同时评估将提供更准确和结论性的结果，尿修饰核糖核苷作为癌症的指标。
Some modified ribonucleosides in biological fluids have been evaluated as cancer-related metabolites. Detection of endogenous modified ribonucleosides in biological fluids may serve as a noninvasive cancers diagnostic method: However, determination of modified ribonucleosides is still challenging because of their low abundance and serious Matrix interferences in biological fluids. Here) we developed a novel strategy for comprehensive, profiling of ribose conjugates from biological fluids using metal oxide-based dispersive solid-phase extraction (DSPE) followed With in vitro stable isotope labeling and double neutral loss scan-mass spectrometry analysis (DSPE-SIL-LC-DNLS-MS). Cerium dioxide (CeO2) was used to selectively recognize and capture ribose conjugates from complex biological samples under basic environment. The enriched ribose conjugates were subsequently labeled with a pair of isotope labeling reagents (acetone and acetone-d(6)); The glucosidic bond of acetone labeled ribose conjugates is readily ruptured, and the generated ribose that carries an isotope tag can be lost as a neutral fragment under collision induced dissociation (CID). Since the light (acetone) and heavy (acetone-d6) labeled compounds have the same chemical structures and can generate different neutral loss fragments (NL 172 and 178 Da), it is therefore highly convenient to profile ribose conjugates by double neutral loss scan mode in mass spectrometry analysis. In this respect, the light and heavy labeled compounds were ionized at the same condition but recorded separately on MS Spectra, which can significantly improve the detection specificity and facilitate the identification of ribose conjugates. Using the developed DSPE-SIL-LC-DNLS-MS strategy, we profiled the ribose conjugates in human urine, and 49 ribose conjugates were readily identified, among which 7 ribose conjugates exhibited Significant contents,change between healthy controls and lymphoma patients. The DSPE-SIL-LC-DNLS-MS strategy combines the selective enrichment, stable isotope labeling, and double neutral loss scan - MS analysis, which therefore can efficiently minimize false positive results, facilitate the relative quantification, and notably increase the numbers of identified ribose conjugates in biological fluids samples. Taken together, this study established a promising strategy for the effective profiling of urinary Modified ribonucleosides, and simultaneous evaluation of the contents change of multiple modified ribonucleosides should provide more accurate and conclusive results for the use of urinary modified ribonucleosides as indicators of cancers.