1260 PHYSIOLOGY 



The disadvantage of this method is that other substances, such as ammonia, 

 creatinine, and uric acid, give off a certain amount of their nitrogen with sodium 

 hypo bro mite, so that the method is not strictly accurate, though enough so for 

 most clinical purposes. In actually carrying out the method 5 c.c. of urine are 

 treated with 25 c.c. of freshly prepared solution of sodium hypobromite, and 

 the nitrogen evolved is collected in a graduated tube over water. 



Folin's Method. In Kjeldahl's method all the nitrogenous constituents of 

 the urine are converted into ammonia by boiling with strong sulphuric acid. 

 This conversion occurs with extreme readiness in the case of urea, so that by 

 using a weaker acid and carefully regulating the temperature the hydrolysis may 

 be confined practically to the urea itself. This is the principle of Folin's method 

 of estimating urea. 



Five cubic centimetres of urine are measured into a 200 c.c. Erlenmeyer 

 flask. Five cubic centimetres of concentrated hydrochloric acid, 20 grm. 

 crystallised magnesium chloride, a piece of paraffin the size of a small hazel 

 nut, and finally 2 or 3 drops of a 1 per cent, solution of alizarin red in water 

 are added. A special safety tube is then inserted into the neck of the 

 flask and the mixture boiled until each returning drop from the safety tube 

 produces a very perceptible bump. The heat is then reduced somewhat, and the 

 heating is continued for a full hour. The alizarin red is used in order to ensure 

 that the contents of the flask do not become alkaline. At the end of an hour 

 the contents of the flask are put into a litre flask with about 700 c.c. water and 

 20 c.c. of a 10 per cent, sodium hydrate, and the ammonia is then distilled off 

 into a measured quantity of acid. The results obtained in this way will give us 

 the total amount of urea together with any ammonia which was preformed in 

 the urine. It is therefore necessary also to determine the amount of this pre- 

 formed ammonia. 



ESTIMATION OF AMMONIA. In Folin's method for the estimation of 

 ammonia, this is set free by the addition of weak alkali (sodium carbonate) and is 

 then removed from the urine at ordinary room temperature by passing a strong 

 current of air through the liquid. The issuing current of air carrying the ammonia 

 passes through a measured quantity of decinormal acid. If the air current be 

 sufficiently strong, one and a half hours is sufficient to remove the whole 

 of the ammonia from 25 c.c. of urine. The decinormal acid is then titrated 

 and the amount of ammonia reckoned. In carrying out the method 25 c.c. 

 of urine is measured into a cylinder 30 to 45 cm. high and about a 

 gramme of sodium carbonate and some petroleum (to prevent foaming) are 

 added. The upper end of the cylinder is then closed by a doubly per- 

 forated rubber stopper through which pass two glass tubes, only one of 

 which is long enough to reach below the surface of the liquid. The shorter 

 tube, about 10 cm. in length, is connected with a calcium chloride tube filled with 

 cotton, and this in turn is connected with a glass tube extending to the bottom 

 of a wide-mouthed bottle, capacity about 500 c.c., which contains 20 c.c. decinormal 

 acid in 200 c.c. of water. 



A more convenient method for the estimation of ammonia is that originally 

 proposed by Schiff and recently worked out by Malfatti. It depends on the fact 

 that when a neutral solution of an amironium salt is treated with formaldehyde, 

 combination occurs with the formation of hexamethylene tetramine (urotropine) 

 and the liberation of a corresponding amount of acid, which can be estimated 

 by titrating with decinormal alkali. The reaction which occurs is as follows : 



6CH 2 + 2(NH 4 ) 2 S0 4 - 6H 2 O + N 4 (CH 2 ) 6 + 2H 2 S0 4 . 

 Formaldehyde Hexamethylene tetramine 



In carrying out this method 25 c.c. of urine are measured by means of a pipette 



