表观遗传修饰与机体的某些疾病密切相关，包括肿瘤的发展。其中一种主要的表观遗传修饰是DNA胞嘧啶的甲基化，5-甲基-2'-脱氧胞嘧啶。此外，5-mdC还可以氧化形成三种新的修饰物，即5-(羟甲基)-2'-脱氧胞嘧啶、5-甲酰基-2'-脱氧胞嘧啶和5-羧基-2'-脱氧胞嘧啶。液相色谱串联质谱法的耦合已广泛应用于生物和临床样品中甲基化DNA胞嘧啶的全面测定。这些方法基于测定游离化合物（例如尿液）或DNA的完全水解产物（例如组织）后的预选浓缩、衍生化和/或净化步骤。本综述重点介绍了过去20年来使用同位素稀释法结合液相色谱或气相色谱-质谱联用进行修饰核苷酸和核苷的定量测定的主要进展。给出了不同可能的标记化合物来源。特别强调了常用的不同类型色谱法（反相色谱和亲水相互作用色谱）和衍生化方法的开发，以提高色谱分离和电离效率。我们还回顾了双向色谱的应用，并指出了这些测定的重要生物学和临床应用。© 2023 年作者。由 Wiley & Sons Ltd.出版的《质谱测定评论》发表。

Epigenetic modifications are closely related to certain disorders of the organism, including the development of tumors. One of the main epigenetic modifications is the methylation of DNA cytosines, 5-methyl-2'-deoxycycytidine. Furthermore, 5-mdC can be oxidized to form three new modifications, 5-(hydroxymethyl)-2'-deoxycytidine, 5-formyl-2'-deoxycytidine, and 5-carboxy-2'-deoxycytidine. The coupling of liquid chromatography with tandem mass spectrometry has been widely used for the total determination of methylated DNA cytosines in samples of biological and clinical interest. These methods are based on the measurement of the free compounds (e.g., urine) or after complete hydrolysis of the DNA (e.g., tissues) followed by a preconcentration, derivatization, and/or clean-up step. This review highlights the main advances in the quantification of modified nucleotides and nucleosides by isotope dilution using isotopically labeled analogs combined with liquid or gas chromatography coupled to mass spectrometry reported in the last 20 years. The different possible sources of labeled compounds are indicated. Special emphasis has been placed on the different types of chromatography commonly used (reverse phase and hydrophilic interaction liquid chromatography) and the derivatization methods developed to enhance chromatographic resolution and ionization efficiency. We have also revised the application of bidimensional chromatography and indicated significant biological and clinical applications of these determinations.© 2023 The Authors. Mass Spectrometry Reviews published by John Wiley & Sons Ltd.