684 STUDIES ON EOSIN HEMOLYSIS 



which the dye itself appears to catalyze the oxidation which is accel- 

 erated by the sun's rays. We have noted that the bleaching of eosin 

 solutions when exposed to sunlight may be markedly inhibited by the 

 addition of tryptophane, while alanine, glycocoll, and phenylalanine 

 afford little or no protection. 



Three possible reactions may conceivably take place in the oxidation 

 of the protective protein or amino-acid. The first is oxidative deami- 

 nation which yields an a-ketonic acid. For tyrosine the reaction is 

 represented by the equation: 



CeH^OH-CHz-CHNHzCOOH + = CeHiOH-CHa-CO-COOH + NH, 



This is a universal reaction and represents a step in the normal catab- 

 olism of the amino-acids. The protective action against eosin hemo- 

 lysis afforded by tyrosine and tryptophane cannot be due to this 

 reaction, since it is not at all specific for these two amino-acids. More- 

 over, ammonia, which itself possesses hemolytic properties, is set free 

 in the reaction. It is possible that the latter substance may be a 

 factor which is concerned in eosin hemolysis. Experimental work, 

 however, does not appear to support this hypothesis. A series of test- 

 tubes containing 100 mg. each of glycocoll, alanine, and tryptophane, 

 dissolved in 5 cc. of normal salt solution and 1 cc. of 1 : 10,000 eosin 

 was exposed to sunlight for 1 hour. At the end of this time the eosin 

 was decolorized in the tubes containing glycocoll and alanine. The 

 ammonia was taken up with permutit and subsequently set free by the 

 addition of NaOH and the solutions were Nesslerized. A trace of 

 ammonia was found in each instance while none was evident in the 

 control tubes which had been kept in the dark. After 4 hours exposure, 

 the tr3^tophane solution gave an ammonia content which corresponds 

 to approximately 0.1 cc. of a 0.03 normal solution, about half of the 

 amount necessary to hemolyze completely the dosage of red cells. A 

 series of test-tubes, each containing 1 cc. of a 0.1 normal solution 

 respectively of glycocoll, alanine, phenylalanine and tryptophane, 

 and 1 cc. of 1 : 10,000 eosin solution was exposed to sunlight for 1 hour, 

 and after exposure 0.5 cc. of a 5 per cent saline suspension of washed 

 sheep cells was added to each tube, the mixtures were shaken and 

 placed in the ice chest. Only a trace of hemolysis was shown by 

 the tubes after standing over night. In this connection it might be 



