Determination of concentrations of adenosine and other purines in human term placenta by reversed-phase high-performance liquid chromatography with photodiode-array detection: evidence for pathways of purine metabolism in the placenta

A robust analytical method, using reversed-phase high-performance liquid chromatography with gradient elution and photodiode-array detection, was used to measure six purines and β-NAD⁺ in acid-soluble extracts of samples taken from six different regions of human term placenta. Resolution of the analyte peaks in chromatographic profiles of the extracts, and the use of optimized integration, allowed simultaneous quantitation of all seven analytes from a single chromatogram. Peak purity was confirmed via on-line analysis of peak spectra, utilizing the purity parameter treatment of spectral data. Major placental purines were adenosine, inosine, hypoxanthine and adenine. Except for adenine, concentrations of the purines varied by two-fold or more between different regions of each placenta, but concentration ratios, i.e., adenosine/inosine and inosine/hypoxanthine, were similar. The findings indicate that the pathway of ATP breakdown to hypoxanthine in ischemic human term placenta is via adenosine, and that regional differences in placental concentrations of adenosine and its metabolites may result from regional differences in degree of ischemia.