Measurement of 2 %26apos;,3 %26apos;-cyclic nucleotides by liquid chromatography-tandem mass spectrometry in cells

作者：Baehre Heike; Kdever Volkhard

来源：Journal of Chromatography B-Analytical Technologies in the Biomedical and Life Sciences, 2014, 964: 208-211.

DOI：10.1016/j.jchromb.2014.02.046

• Cells
• Liquid chromatography-tandem mass spectrometry
• 2 %26apos
• ,3 %26apos
• -Cyclic nucleotide monophosphates

Recently, the occurrence of 2%26apos;,3%26apos;-cyclic nucleoside monophosphates (2%26apos;,3%26apos;-cNMPs) in addition to 3%26apos;,5%26apos;-cNMPs in mammalian tissues was reported. We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the measurement of four 2%26apos;,3%26apos;-cyclic nucleotides, i.e., 2%26apos;,3%26apos;-cAMP, 2%26apos;,3%26apos;-cCMP, 2%26apos;,3%26apos;-cGMP, 2%26apos;,3%26apos;-cUMP, in cell samples. Chromatographic separation was achieved using a Zorbax eclipse XCB-C18 (50 mm x 4.6 mm; 1.8 mu m column; Agilent) connected to a QTRAP5500 system (AB Sciex) operating in positive ionization mode. Calibration curves were constructed in the range 0.41 fmol/mu L to 1666.6 fmol/mu L for 2%26apos;,3%26apos;-cAMP, 2%26apos;,3%26apos;-cCMP, and 2%26apos;,3%26apos;-cGMP, and 3.3-1666.6 fmol/mu L for 2%26apos;,3%26apos;-cUMP, respectively, showing squared correlation coefficients %26gt;0.9992. Accuracy and inter- and intra-day precision lay within the required ranges of %26lt;20% for LLOQ and %26lt;15% for higher concentration levels. The method was applied to the analysis of nucleotides in two different cell lines (Hek293T and HuT-78).

• 出版日期2014-8-1