236 PHYSIOLOGICAL CHEMISTRY 



Allow to separate, pour or pipette off the ethereal solution. Filter and remove the 

 ether by evaporation. The residue contains cholesterol and the mixed fats of 

 the feces. For every gram of fat add about 1.5 grams of solid potassium hy- 

 droxide and 25 c.c. of 95 per cent alcohol and boil in a flask on a water-bath for 

 one-half hour, maintaining the volume of alcohol constant. This alcoholic- 

 potash has saponified the mixed fats and we now have a mixture of soaps, 

 cholesterol and koprosterol. Add sodium chloride, in substance, to the mixture 

 and extract with ether to dissolve out the cholesterol and koprosterol. Remove 

 the ether by evaporation and examine the residue microscopically for cholesterol 

 and koprosterol crystals. Try any of the other tests for cholesterol as given on 

 page 212. 



6. Blood. Undecomposed blood may be detected macroscopically. 

 If uncertain, look for erythrocytes under the microscope, and spectro- 

 scopically for the spectrum of oxyhemoglobin (see Absorption Spectra, 

 Plate I). 



In case the blood has been altered or is present in minute amount 

 ("occult blood")? and cannot be detected by the means just mentioned, 

 the following tests may be tried: 



(a) Benzidine Reaction. Make a thin fecal suspension using about 5 c.c. of 

 distilled water, and heat it to boiling to render oxidizing enzymes inactive. 

 To 2 c.c. of a saturated solution of benzidine in glacial acetic acid add 3 c.c. of 

 3 per cent hydrogen peroxide and 2-3 drops of the cooled fecal suspension. 

 A clear blue color appears within one to two minutes in the presence of blood. 

 If the mixture is not shaken a ring of color will form at the top. Minute traces of 

 blood are more easily detected by the latter procedure. 



Wagner 1 has simplified the benzidin test so that it can be applied 

 much more conveniently. 



Slide Modification. Take up a little of the solid stool on a match, smear it 

 on an object glass and pour the reagent over it. It turns blue if there is blood 

 present and there is no misleading green tint from fluid. Make the solution 

 as follows: Add a knife-tip of benzidine to 2 c.c. of glacial acetic acid, and 

 add 20 drops of a 3 per cent solution of hydrogen peroxide. 



By this dry technic there is no danger of soiling the fingers, and the 

 test is more sensitive than the usual "wet" benzidine test. The smear 

 of stool is either blue or it is not blue. The rapidity of the color change 

 gives some idea as to the proportion of blood in the stool; with much 

 blood present the change to blue is instantaneous. It is claimed by 

 Vaughn 2 that pus and the usual drugs and foods ingested do not 

 interfere with the reaction. 



(b) Lyle-Curtman Guaiac Procedure. 3 Approximately 10 gm. of the stool 

 are transferred to a beaker, 25 c-.c. of distilled water are added, and the mixture 

 is stirred until of uniform consistency. Over a low flame, the mixture is heated 



1 Wagner: Zentbl.fur Chirurgie, 41, No. 28, 1914. 



2 Vaughn: Jour, of Lab., Clin. Med., 2, 437, 1917. 

 3 Lyle and Curtman: Jour. Biol. Chem., 33, i, 1918. 



