Detection of free radicals in ischemic and reperfused canine gracilis muscle flaps by use of spin-trapping electron paramagnetic resonance spectroscopy

Objective-To determine whether free radicals are produced in ischemic and r eperfused canine skeletal muscle. whether free radicals can be detected fro m effluent blood by use of spin-trapping electron paramagnetic resonance (E PR) spectroscopy, and whether free radical-induced skeletal muscle damage i s detectable by use of light microscopy. Animals-6 healthy mixed-breed dogs. Procedures-Dogs were anesthetized and both gracilis muscles were isolated, leaving only the major vascular pedicle intact. Ischemia was induced in 1 f lap for 4 hours; the contralateral flap served as the control. Ischemic fla ps were then reperfused for 15 minutes, alpha -Phenyl-N-tert-butyinitrone, a spin-trapping agent, was administered intravenously to each dog 1 hour pr ior to reperfusion. Following reperfusion, effluent blood samples from musc le flaps were obtained and processed for EPR spectroscopy. Muscle biopsy sp ecimens were obtained for histologic evaluation, and dogs were euthanatized . Results-Spin adducts were not detected in blood from control flaps. However . spin adducts were detected in ail ischemic-reperfused muscle flaps. Princ ipal signals identified were characteristic of oxygen- and carbon-centered radicals. Significantly more muscle damage was detected in ischemic-reperfu sed flaps, compared with control flaps. Conclusions and Clinical Relevance-Free radicals may be an important compon ent of injury induced by ischemia and reperfusion of canine skeletal muscle . Spin-trap adducts of free radicals can be detected in effluent blood of c anine muscle flaps by use of spin-trapping EPR spectroscopy. Spin-trapping EPR spectroscopy may be useful for the study of antioxidants and free radic al scavengers in attenuating ischemia and reperfusion-mediated skeletal mus cle damage.