Methods for Testing Stains 293 



in the ratio. More elaborate methods have been used to express the shape of the 

 curve independently of the peak position (Lillie and Roe, 1942) but we have chosen 

 the simple ratio of the color densities at minus and plus 15 xn/j, of the peak wave- 

 length found with the given sample. These densities can be estimated from the 

 plotted spectrmn or an absolute figure actually measured after location of the peak. 



TiTANOus Chloride Assay 



A study of the titanous chloride standardization, use of the solution, and the dye 

 titration itself led to various modifications. Titanous chloride solutions are standard- 

 ized against eerie sulfate, which in turn is standardized against arsenious oxide. Both 

 titrations employ ferrous-o-phenanthroline indicator which gives sharp endpoints. 



Sodium Arsenite — Dissolve 4.947 g. of U. S. Bureau of Standards arsenious 

 oxide (AS2O3) in a 600 ml. beaker containing 200 ml. of distilled water and 5 g. 

 of sodium hydroxide pellets. After complete solution add 13 ml. of concentrated 

 hydrochloric acid, mix, and then add 10 g. of sodium bicarbonate slowly to prevent 

 loss by spray. Transfer the solution quantitatively to a 1-liter volumetric flask, 

 dilute to the mark, and mix. The resulting solution is O.liV. 



Ceric Sulfate — Weigh 66.5 g. of anhydrous eerie sulfate [Ce(S04)2] into a 600 ml. 

 beaker and add 28 ml. of concentrated sulfuric acid. Add cautiously about 30 ml. 

 of distilled water with stirring. Heat the solution, stir, and add successive portions 

 of water until the salt is completely dissolved. Cool, transfer the solution to a 

 1-liter volumetric flask and dilute to the mark. This approximately 0.1 iV eerie 

 sulfate solution is completely stable. 



Iodine Monochloride Catalyst — Dissolve 10 g. of potassium iodide (KI) and 6.74 g. 

 of potassium iodate (KIO3) in 90 ml. of distilled water. Add 90 ml. of concentrated 

 hydrochloric acid and mix. This solution is 0.5 molar in iodine monochloride. It 

 should be stored in the dark. 



Standardization of Ceric Sulfate — Pipette exactly 25.0 ml. of 0.1 N sodium ar- 

 senite into a 200 ml. Erlenraeyer flask, add 20 ml. of concentrated hydrochloric acid, 

 and 2.5 ml. of 0.005 M iodine chloride (dilute above stock). Dilute the mixture to 

 about 100 ml., add 1 drop of ferrous-o-phenanthroline indicator^ and titrate with 

 the ceric sulfate until the orange color of the indicator returns only slowly after 

 dropwise addition of the ceric solution. At this stage warm the solution to 50° C, 

 add another drop of indicator, and continue the titration dropwise until a single 

 drop produces a light green solution with no return of the orange color for at least 

 one minute. The endpoint is sharp and reproducible. The normality of the ceric 

 sulfate solution is calculated as follows: 



25 X 0.1 



, N ceric = 



ml. eerie 



Titanous Chloride Solution— The stock solution is 20% titanous chloride sta- 

 bilized with hydrochloric acid'. An approximately 0.05 A^ solution is prepared by 

 diluting 8.0-8.5 ml. of the stock solution to 200 ml. with distilled water. Storage 



2"Ortho-Phenanthroline Ferrous Complex (Ferroin) 0.025 M". Purchased from 

 the G. Frederick Smith Chemical Co., Columbus, Ohio. 

 'LaMotte Chemical Products Co., Baltimore, Md. 



