ANALYSIS OF TISSUES 583 



(l>) Muscle and Tissues. 



5 gm. are cut into strips immediately after removal and put into 50 

 c.c. of alcohol in a conical flask. They are ground up and treated with fresh 

 alcohol for 12-16 hours. The alcoholic solutions are combined and put into 

 a 100 c.c. measuring flask, a few drops of alcoholic zinc chloride solution are 

 added, the volume made up to the mark and the solution filtered. 



Total Nitrogen, Ammonia and Urea. 



10 c.c. of the filtrate are used for the analysis of total nitrogen, ammonia 

 and urea which are carried out in the same way as described under blood. 



Creatinine. 



10 gm. of fresh muscle are put into a mortar 10-15 cm. in diameter, 

 cut up into small pieces with scissors and ground up with 20 gm. of sand 

 into a fairly uniform thick paste. 43 c.c. of saturated picric acid solution are 

 gradually added, whilst the rubbing is continued and finally about i gm. of solid 

 picric acid. The rubbing and stirring are continued for 5-10 minutes after 

 the last addition of picric acid. The proteins are thus converted into insoluble 

 picrates. The object of adding 43 c.c. of picric acid is .to allow for 75 per 

 cent, of water in the muscle and so as to have as nearly as possible 50 c.c. of 

 solution. 



In the case of other tissues, liver and brain especially, it is advisable to add 2 

 c.c. of formalin to the 10 gm. of tissue and to allow to stand for 10 minutes be- 

 fore treating as above with picric acid solution. The mixture is poured upon 

 a filter ; 20 c.c. of the filtrate are put into a dry measuring cylinder, i c.c. of 

 10 per cent, sodium hydrate is added and the mixture allowed to stand to 

 develop the colour. 



20 c.c. of a standard solution of *5 mgm. creatinine in 100 c.c. picric 

 acid are treated with i c.c. of 10 per cent sodium hydrate. 



The colours are compared setting the standard at 20 mm. 



20 x 2-5 



-p = mgm. creatinine m 100 gm. of muscle. 



. Creatine. 



5 gm. of muscle or tissue are cut up finely with scissors or a meat grinder 

 and placed in a 200 c.c. conical flask ; 100 c.c. of ^N sulphuric acid are added. 

 The flask is covered with tin foil and put in a autoclave at 130-135 for 30- 

 40 minutes. The autoclave is opened when it has cooled below 100 and 

 the contents of the flask transferred to a 200 c.c. measuring flask. The floccu- 

 lent masses of tissue are broken by shaking and the volume made up to the 

 mark. The solution is filtered. 



10 c.c. of the solution are titrated with 10 per cent, sodium hydrate with 

 phenolphthalein as indicator. 



Another 10 c.c. are placed in a 100 c.c. measuring flask and 1*5 c.c. + the 

 amount of i o per cent, sodium hydrate used to neutralise the solution are added, 

 the colour developed and the solution is diluted to the mark. 



As standard for muscle i mgm. of creatine per c c. ( = 1*389 gm. of creat- 

 inine zinc chloride per 1000 c.c.) and as standard for other tissues "5 mgm. of 

 creatine per c.c. is used. 



In the former case the standard is put at 10, in the latter at 20 mm. 



Under these conditions 



4000 

 reading = m S m * creatme per IO g m - of muscle. 



(c) Milk. 



The creatinine and creatine estimations are carried out in the same way 

 as is described for blood. 



