A simple method for the simultaneous determination of guanine, xanthine and hypoxanthine by RP-HPLC with UV detection

Department of Marine Science, Calcutta University, 35, Ballygunge Circular Road, Kolkata-700 019, India

E-mail : natasha.majumder@gmail.com

Manuscript received online 14 August 2014, accepted 06 February 2015

A simple, rapid and accurate method for the simultaneous determination of guanine, xanthine and hypoxanthine has been developed. The quantitative determination of these bases was accomplished by reverse phase high-performance liquid chromatography (RP-HPLC) with UV detection based on their different retention time without chemical suppression. Separation in the order of guanine, hypoxanthine and xanthine were obtained on ODS hypersil column using mobile phase containing 1% ACN in acetic acid (50 mM), ammonium acetate (85 mM) and 0.5% methanol at 0.5 ml min^–1 flow rate. A portion of the natural DNA isolated from Avicennia marina was spiked with known amount of hypoxanthine and its concentration was found to be 1.4 µM.