METHODS FOR STUDY OF DIGESTIBILITY OF LIPIDS 211 



Van de Kamer et al. i4 have proposed two procedures by which the fat 

 can be measured within thirty-five to forty-five minutes, with an error 

 which does not exceed 2%. The procedures are modifications of the 

 methods of von Liebermann and Szekely 46 and of Saxon. 47 In the first 

 procedure (Method A), feces are saponified with concentrated potassium 

 hydroxide in ethanol; HC1 is added to acidify the solution, additional 

 ethanol is added, and the fatty acids are extracted with petroleum ether. 

 The concentration of ethanol is so chosen that the petroleum ether and 

 ethanol solutions readily separate. This can be expedited by the addi- 

 tion of sodium chloride and amyl alcohol. The separation usually re- 

 quires five to ten minutes. The total fatty acids are estimated by titra- 

 tion of an aliquot sample of the petroleum ether extract with alkali, 

 using thymol blue as an indicator. This entire determination requires 

 only approximately thirty-five minutes. 



By means of the second procedure of van de Kamer et al. u (Method B) 

 a determination of split and of unsplit fat can be made separately. In 

 this case, the sample of feces is not treated with alkali ; instead it is boiled 

 for one minute with hydrochloric acid to convert the soaps into free fatty 

 acids, as in the method of Saxon. 47 Ethanol, sodium chloride, and amyl 

 alcohol are then added, and the solution is extracted with petroleum ether 

 as in Method A. An aliquot sample of the petroleum ether extract is 

 evaporated to dryness, and the free fatty acids are estimated by titration 

 with 0.1 iV isobutyl alcoholic KOH. Thereupon, an excess of the same 

 solution is added, and the unsplit fat is hydrolyzed by boiling. The ex- 

 cess alkali remaining is determined by back titration with 0.1 iV hydro- 

 chloric acid, again using thymol blue as an indicator. It is, of course, 

 obvious that neither Method A nor Method B of van de Kamer and col- 

 laborators 44 is useful in the determination of fecal lipids other than fats. 



(7) Methods for the Determination of Inert Chemical Substances 



a. Lignin. Lignin can be accurately determined by the procedure of 

 Ellis, Matrone, and Maynard. 16 This involves the four-hour extraction 

 of a 1 g. sample of food or feces ground to pass through a forty-mesh 

 screen, with an ethanol-benzene mixture (32 parts of 95% ethanol to 68 

 parts of benzene by weight); a coarse alundum thimble is used. After 

 two washings, each with 95% ethanol and with diethyl ether (using suc- 

 tion), the material is dried, transferred to a 50 ml. stoppered Erlenmeyer 

 flask, and digested overnight at 40°C. with a 1% pepsin solution in 0.1 iV 

 HC1. After the residue has been thoroughly washed with hot water, 



