Development and evaluation of a hydrophilic interaction liquid chromatography‐MS/MS method to quantify 19 nucleobases and nucleosides in rat plasma

Abstract

As essential endogenous compounds, nucleobases and nucleosides fulfill various functions in living organisms. This study presents the development and validation of a new hydrophilic interaction liquid chromatography tandem mass spectrometry method for simultaneous quantification of 19 nucleobases and nucleosides in rat plasma. For the sample preparation, 15 kinds of protein precipitants were evaluated according to the chromatographic profile and ion response of analytes. The optimization of chromatographic separation was respectively performed using reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography mode; each separation mode included two test columns with different stationary phases. The chromatographic profile and parameters such as half-width (W 1/2 ), capacity factor (K′) and tailing factor (f t ) were used to evaluate the separation efficiencies. Furthermore, the adopted composition of two mobile phase systems and the concentrations of the additives in the optimum buffer system were also investigated. The developed method was fully validated and successfully applied quantitatively to determine 19 nucleobases and nucleosides in plasma from normal and diabetic nephropathy (DN) rats. Significant differences between normal and DN rats were found in plasma levels of cytosine, xanthine, thymidine, adenosine, guanosine, inosine and 8-hydroxy-2′-deoxyguanosine. This information may provide a useful reference for the discovery of potential biomarkers of DN.

Orally delivered MK-4482 inhibits SARS-CoV-2 replication in the Syrian hamster model

“…To ensure signal fidelity triggered spectra were compared back to synthetic standards. Previously published MRM signals for biological nucleosides were utilized to confirm minimal interference at the retention time of interest 42 . All analytes were quantified against an 8-point calibration curve of the respective synthetic standard prepared in the target matrix and processed in the same manner as experimental samples.…”

Orally delivered MK-4482 inhibits SARS-CoV-2 replication in the Syrian hamster model

“…To ensure signal fidelity triggered spectra were compared back to synthetic standards. Previously published MRM signals for biological nucleosides were utilized to confirm minimal interference at the retention time of interest 35 . All analytes were quantified against an 8-point calibration curve of the respective synthetic standard prepared in the target matrix and processed in the same manner as experimental samples.…”

A comprehensive approach to detecting multitudinous bioactive peptides in equine plasma and urine using hydrophilic interaction liquid chromatography coupled to high resolution mass spectrometry

“…Hydrophilic substances that cannot be appropriately retained using RPLC, such as those hydrophilic peptides mentioned above, can be well retained by HILIC. HILIC has been used for the retention of a wide range of polar substances such as amino acids and peptides, carbohydrates, endogenous compounds, and nucleosides . A few recent review articles provide insight into the retention mechanism, column classification, and factors that affect retention and selectivity in HILIC .…”