Early changes of oxypurines in rat brain following focal cerebral ischemia

Cerebral hypoxanthine, xanthine, and uric acid levels were measured by high-performance liquid chromatography (HPLC) for up to 4 hours following focal cerebral ischemia in the rat. Fifty male Sprague-Dawley rats were subjected to occlusion of the left middle cerebral artery under halothane inhalation anesthesia. The animals were sacrificed with microwave at 30, 60, 120, and 240 minutes after surgery. The brains were removed and divided into right and left hemisphere. Each hemisphere was homogenized with perchloric acid and centrifuged. The supernates were filtrated with membrane filter. An aliquot of the filtrate was used for measurement of uric acid, xanthine, and hypoxanthine in both of the ischemic and contralateral hemisphere by a HPLC system. A HPLC with multiple ultraviolet spectroscopy was used for measuring hypoxanthine and xanthine. Identification of hypoxanthine and xanthine was made by parallel chromatography of standards, disappearance with xanthine oxidase, and the spectrum of UV absorption. Uric acid was measured by reversed-phase HPLC with electrochemical detection as reported previously. Hypoxanthine increased rapidly and arrived at a peak value at 60 minutes. Xanthine increased not so rapidly as hypoxanthine and showed the highest value at 120 minutes. Uric acid also increased significantly but very slowly and did not seem to reach the peak value during the observation period. Hypoxanthine is oxidized to xanthine and then xanthine is oxidized to uric acid at the terminal stage of purine degradation. The order of peak times of cerebral hypoxanthine, xanthine, and uric acid levels following cerebral ischemia corresponds to the order in purine metabolism. This result strongly suggests that hypoxanthine is degraded into uric acid in ischemic rat brain.(ABSTRACT TRUNCATED AT 250 WORDS)