An Isocratic, Reversed-Phase HPLC Method for the Determination of Postischemic Efflux of Purines and Pyrimidines during Reperfusion of Isolated Liver

V. Yu. Toshchakov^*, L. V. Bashkina, N. A. Onishchenko, and V. I. Shumakov

Institute of Transplantation and Artificial Organs, Ministry of Health Care of the Russian Federation, ul. Shchukinskaya 1, Moscow, 123182 Russia; fax: (095) 943-0008.

^* To whom correspondence should be addressed.

Received August 12, 1997; Revision received October 15, 1997

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An isocratic, reversed-phase HPLC method was developed for the determination of the rates of purine and pyrimidine efflux during early reperfusion of isolated organs after non-perfusion cold conservation. The method enables determination of uric acid, cytidine, xanthine, hypoxanthine, uridine, AMP, inosine, and adenosine in liver perfusate using a standard C-18 column (25 cm length). Peaks are resolved by elution with buffer containing 1% acetonitrile, 20 mM potassium citrate (pH 6.25), and 25-55 mM tetramethylammonium. The effects of pH and solvents on peak retention times are described. As an example of the application of the method, the effects of allopurinol on the rates of postischemic efflux of purines and pyrimidines during reperfusion of liver stored in the cold for 24 h in Euro-Collins solution was studied.

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KEY WORDS: HPLC, purines, pyrimidines, organ conservation, ischemia, reperfusion, liver, allopurinol

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