THE NATURE OF THE PEROXIDE-LIKE 

 SUBSTANCES FORMED IN MICE BY X-RAYS 



V. J. HoRGAN and J. St. L. Philpot 

 Medical Research Council Radiobiological Research Unit, Harwell 



In our last publication ^ we gave evidence that a lethal dose of X-rays in 

 a mouse caused an immediate increase in peroxide-like material (called 

 'peroxide' for short) with mean ionic yield about 80, suggesting that the 

 well-known chain reaction of autoxidation could be initiated by X-rays 

 in vivo. We noted, however, that the peroxide was not as reactive as 

 expected in that the cuprous-catalysed reaction with aerobic leucoindo- 

 phenol was far from complete in 10 minutes at room temperature. The 

 necessity for air as well as peroxide in the reaction with leucoindophenol 

 complicated the interpretation and we looked for an anaerobic reagent. 

 Stannous chloride reacted anaerobically but required heating even with 

 ordinary 'reactive' peroxides (hydroperoxides) of the expected sort, so was 

 useless for distinguishing 'reactive' peroxides from the 'unreactive' ones 

 apparently present in the mouse. Leuco-brilliant cresyl blue, catalysed by 

 alkaline cuproiodide, was mentioned as a promising new reagent but was 

 awaiting a proper test. 



EXPERIMENTAL 



Mice, irradiated or not with lOOOr of 250 kV X-rays, were killed and 

 extracted with 100 ml of peroxide-free n-butanol in a Waring blender or 

 top-drive macerator, sometimes in an atmosphere of nitrogen. After filter- 

 ing 2ml were diluted to 9 ml and deoxygenated by disdllation of the solvent ^ 

 or better by 3 minutes' bubbling with nitrogen in a vessel that had been 

 heated at 100° in vacuo to remove absorbed oxygen. The peroxide reagent 

 was then added, consisting of 1 ml of a 0-002M solution of buffered leuco- 

 brilliant cresyl blue in «-butanol. The leuco-dye was made by running 

 the dye through a column of copper or cadmium powder, with lithium 

 iodide or other complexing agent in the soludon to combine with the metal 

 ions. With the copper column an amount of cuproiodide catalyst equivalent 

 to the dye was automatically introduced in this way. After 10 minutes at 

 room temperature or 100° C the reaction mixture was introduced anaero- 

 bically into the cell of a Hilger Uvispek Spectrophotometer and measured 

 at 600 m[j. where the molar extinction coefficient of brilliant cresyl blue 

 was found to be 18,800. 



RESULTS AND DISCUSSION 



The present communication is even more of an interim report than the 

 last, and condnual changes in technique make it even harder to quote 

 satisfactory experiments, but the following generalizations will give some 

 idea of the trend of the research. 



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