246 PHYSIOLOGICAL CHEMISTRY 



paratus with good stream of water in the condenser. Heat carefully for a few 

 minutes until danger of foaming is past and then allow to boil vigorously. Distill 

 over 500 c.c. of liquid, bringing the volume of the fecal suspension down to about 

 100 c.c. toward the end of the distillation. 



Transfer the distillate to a clean Kjeldahl flask, add 2 drops of phenolphthalein 

 as an indicator. Make neutral with N sulphuric acid and add i c.c. excess. Dis- 

 till with steam as before, collecting the first 500 c.c. of distillate and bringing the 

 residue finally to about 100 c.c. Mix distillate well by shaking. 



Take an aliquot portion of the distillate (100 c.c ); add i c.c. of a 2 per cent 

 solution of /3-naphthaquinone sodium monosulphonate solution. Then add 2 c.c. 

 of 10 per cent KOH. Shake and let stand for 1 5 minutes. This is best carried out 

 in a 150 c.c. Squibb shape separatory funnel. Extract with chloroform, shaking 

 vigorously, using a 10 c.c. and a 7 c.c. portion which will bring the total volume of 

 the extract to the mark of a 15 c.c. graduated tube. Mix thoroughly. Run at 

 the same time and in the same way a standard using i c.c. of a solution of indol o.i 

 mg. of indol per c.c. Compare the extract with this standard in a colorimeter, 

 using the standard ordinarily at the 30-mm. mark. Calculate the indol to the 

 basis of milligrams of indol per gram of moist feces. 



Indol and naphthaquinone solutions should be freshly prepared or may be 

 kept in the ice-box for some days. Indol distillates should be kept in the ice-box 

 if not used at once, especially in hot weather. The feces must be fresh. 



24. Quantitative Determination of Fat in Feces. Principle. The 

 determination of fat in dried feces is a more or less tedious process, and 

 one which is somewhat dangerous if applied to pathological feces. Most 

 of the methods for the determination of fat in the moist feces are accurate, 

 but require a long time. Saxon 1 has proposed a method for the deter- 

 mination of fat in moist feces, which is speedy, convenient, and accur- 

 ate. The soaps of the feces are converted into free fatty acids by 

 means of hydrochloric acid, and the material is then extracted by shak- 

 ing with ether. The ether removes the neutral fat, the fatty acids 

 which were present as such, the fatty acids derived from the soaps, and 

 the cholesterol. The ether is removed by distillation, the crude fat 

 purified by means of petroleum ether, and the weight of the total fat 

 obtained. The fat is the:nf dissolved in benzene and titrated with tenth- 

 normal sodium alcoholate solution, using phenolphthalein as an indi- 

 cator. The fatty acid is calculated, from the titration, to stearic acid. 



Procedure. Place about 5 grams (accurately weighed) of the thoroughly 

 mixed feces in a 100 c.c. glass-stoppered graduated cylinder. 2 



Add 20 c.c. of distilled water, i to 2.5 c.c. of concentrated hydrochloric acid 

 (depending upon the amount of the sample) and again, sufficient distilled water 

 to make a total bulk of 30 c.c. Add exactly 20 c.c. of ether, stopper, and shake 



1 Saxon: /. Biol. Chem., 17, 99, 1914. 



1 Care must be taken not to smear the neck of the cylinder. This may be avoided by 

 removing the feces from the weighing bottle by means of a glass rod, the end of which is 

 flattened, and bent in the shape of a hoe, and transferring small bits of the feces from the hoe 

 to the cylinder, using short pieces of glass rod, which are dropped into the cylinder together 

 with the feces. 



