712 EXPERIMENT STATION RECORD. 



Instead of determining the end point of the reaction by noting the yellow color 

 or using starch and iodid of potash, it is advisable to 'employ methyl violet or 

 gentian violet in a 1 per cent solution in 70 per cent alcohol. 



"About 10 drops of this solution is added to 10 cc. of 5 per cent hydrochloric 

 acid and dotted over a white tile. The addition of sodium bromate or traces of 

 free bromin causes a change from olive green (the color of the dyestufCs in acid 

 solution) to a deep bluish- violet. The change is sharp, but the color fades after 

 a few minutes. Dibromotyrosin only is apparently formed and the absorption 

 at first is very rapid, but falls off considerably toward the end of the reaction, 

 as experiments cited show. The solution to be titrated is made of not more than 

 5 per cent acidity with hydrochloric acid ; a lower concentration than 2 per cent 

 turns the indicator blue, while more than 5 per cent turns it yellow. To the acid 

 solution is added 15 to 20 cc. of 20 per cent sodium bromid, and the liquid is then 

 titrated with twentieth-normal sodium bromate. Toward the end of the reac- 

 tion at least 30 seconds should elapse between successive additions of the bro- 

 mate, and the solution should be well shaken in a stoppered bottle. 



" Results quoted show a plus errox- varying from 0.4 to 2.3 per cent. In two of 

 the test liquids there were present, in addition to tyrosin, leucln, asparagin, and 

 ammonium chlorid. Experiments carried out with edestin digested in dilute so- 

 dium carbonate solution with trypsin, and corrected for the bromin absorbed by 

 the protein, confirmed Brown and Millar's contention that the method can be 

 used for determining tyrosin in presence of proteins and other early cleavage 

 products, and also that practically all the tyrosin is liberated in the first stages 

 of tryptic digestion." 



Accuracy of the estimation of 1-tyrosin in proteins, E. Abdebhalden and 

 D. FucHS (Hoppe-Seyler's Ztschr. Physiol. C'hem., 83 {1913), No. 6, pp. 468- 

 473; abs. in Analyst, 38 (1913), No. 446, pp. 219, 220).— The Folin and Denis 

 colorimetric method for determining tyrosin (E. S. R., 28, p. 805) in proteins is 

 said to yield high results because similar colors are produced by oxytryptophan 

 and tryptophan. 



The authors observed, as did Folin and Denis, that the separation of tyrosin 

 by simple crystallization is xevy difficult. " The cause of this has been traced 

 in certain cases to absorption of acid or ammonia fumes from the atmosphere 

 during evaporation in open basins. Much better results are obtained if the 

 evaporation be conducted under reduced pressure. A more interesting reason 

 for quite considerable quantities of tyrosin failing to crystallize is the forma- 

 tion of salt-like compounds with the basic constituents of proteins, such as the 

 compound of tyrosin with lysin observed by Fischer and Abderhalden. In 

 view of this experience, tests were made to determine whether it is possible to 

 recover quantitatively tyrosin added to gelatin. Tyrosin-free gelatin was dis- 

 solved in five times its weight of 25 per cent sulphuric acid, a known quantity of 

 tyrosin added, and boiled for 20 hours. Baryta was then added to remove the 

 sulphuric acid, and the barium sulphate extracted with boiling water until the 

 extract no longer gave a violet color with a 1 per cent aqueous solution of 

 triketohydrindene hydrate (ninhydrin). This reagent proved much more 

 delicate than Millon's reagent. By concentrating the combined filtrates at 45* 

 C, it was possible to recover as much as 90 per cent of the added tyrosin. In 

 certain cases, however, the yield amounted to only 60 to 70 per cent 



" It was found possible to recover the tyrosin almost quantitatively by the 

 following method : The liquid after hydrolysis was diluted until it contained 

 2.5 per cent sulphuric acid. A 10 per cent solution of phosphotungstic acid was 

 then added with constant stirring, great care being taken to avoid the addition 

 of any excess. The precipitate was filtered off and washed, and the filtrate 

 treated with baryta to remove phosphotungstic acid and filtered. To this filtrate 



