A low-cost mass spectrometry-based approach for quantifying purines in placental explants

Purine nucleotides serve as immediate building blocks for the synthesis of RNAs and DNA, energy carriers co-substrates in activation many metabolites, regulators cellular metabolism, signaling molecules. Knowledge actual tissue-specific concentration these metabolites their rate is essential to studies focused on a metabolic rewiring pathogenesis different diseases, including carcinogenesis. The existing data mainly characterize purine body fluids malignant cells. protocols estimating de novo biosynthesis usually required usage radioactive precursors costly facilities like HPLC with tandem mass spectrometry or nuclear magnetic resonance. We developed new simplified method measure level purines by HPLC/ESI-MS, using stable isotope glycine label newly synthesized purines. proved be highly sensitive, interday repeatable, intraday reproducible. applied placental explants from first third trimesters gestation MCF7 cells cultivated 20 μM 40 homocysteine. Corresponding homocysteine typical breast cancer pregnancy-associated complications affecting human placenta. content total trimester was around 9.0 μmol/g wet tissues at specified cultivation conditions. formed comprised approximately 1% decreased steadily mM They reached values characteristic grown without homocysteine, 4.0 tissues. In cells, 50% under treatment. A low-cost spectrometry-based approach may used estimate eukaryotes’ wide range concentrations. • HPLC/ESI-MS glycine. measured placenta pregnancy. Homocysteine induces decrease purine’s due shift methionine cycle.