SIMULTANEOUS DETERMINATION OF ADENOSINE, INOSINE, HYPOXANTHINE, XANTHINE, AND URIC-ACID IN MICRODIALYSIS SAMPLES USING MICROBORE COLUMN HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH A DIODE-ARRAY DETECTOR

In the myocardial interstitial space, adenosine and its metabolites ar e important markers of ischemia, regulators of blood flow, and may pro duce cardioprotection against ischemia. A fast and sensitive method to assess the concentrations of adenosine and its metabolites is necessa ry to determine their involvement in mediating these effects. A method for the simultaneous determination of adenosine, inosine, hypoxanthin e, xanthine, and uric acid in the interstitial fluid of the canine myo cardium was developed using microdialysis, microbore column high-perfo rmance liquid chromatography, and a photo diode array detector (DAD), The microdialysis samples were injected directly onto a microbore C-18 reverse-phase column without any prior sample preparation. Use of a D AD in this method provided many advantages. First, a DAD allowed the s imultaneous detection of UV absorbance at multiple wavelengths, allowi ng the detection of each compound at their maximal UV absorbance. Furt her, the full UV absorption spectrum was recorded for each detected pe ak, confirming peak purity and identity. Using a microbore HPLC column and detection of UV absorbance at the maximal absorbance for each com pound improve the sensitivity for all compounds. The detection limit o f these compounds is 50 fmol (signal-to-noise ratio, S/N = 3). This me thod is useful in analyzing the temporal effect of a prolonged period of myocardial ischemia and reperfusion upon interstitial adenosine, in osine, hypoxanthine, xanthine, and uric acid concentrations in an in v ivo canine model. (C) 1996 Academic Press, Inc.