Analysis of adenosine phosphates in HepG-2 cell by a HPLC-ESI-MS system with porous graphitic carbon as stationary phase

A high performance liquid chromatography coupled with electrospray ionization/mass spectrometry method was developed for the determination of adenosine 5'-monophosphate (AMP), adenosine 5'-diphosphate (ADP), and adenosine 5'-triphosphate (ATP) in the extract of HepG-2 cells. The chromatographic conditions were optimized by using porous graphitic carbon as the stationary phase for the retention and separation of the AMP, ADP and ATR Negative-ion mode ESI-MS in basic mobile phase was applied to improve the method sensitivity. An external calibration method with linear ranges from 0.22 to 57.80 mu M for AMP, from 0.59 to 117.37 mu M for ADP, and from 0.49 to 98.81 mu M for ATP was used for quantitative analysis. The levels of ATP, ADP, and AMP in HepG-2 cells treated with benzo[a]pyrene with different time periods were determined. Total adenine nucleotides and the energy charge potential were calculated for the investigation of the effect of benzo[a]pyrene on cell energy metabolism. (C) 2009 Elsevier B.V. All rights reserved.