ANALYSIS OF TISSUES 589 



The procedure, which takes 15 minutes, is the following: 



i c.c. of blood is collected in an accurately graduated i c.c. pipette, or in 

 a special Hawksley pipette graduated to contain i c.c. of blood under standard 

 conditons, and run into 20 c.c. of distilled water contained in a mortar. The 

 blood sinks and the pipette is rinsed out several times with the supernatant 

 water so as to remove the last traces of blood from the pipette. The mixture 

 is stirred with a pestle and 2 c.c. of the phosphoric acid solution are added. 

 After stirring, 15 c.c. of dialysed iron are run in from a burette and well mixed 

 and then 2 c.c. of sodium sulphate solution. The total volume is now 40 c.c. 

 On stirring this mixture it forms a thick paste, which gradually becomes more 

 fluid. The liquid and solid are separated by pouring upon a filter paper con- 

 tained in a square of calico in a funnel, collecting the ends of the calico 

 together and gradually squeezing. Most of the liquid can thus be separated 

 in a clear condition, if the squeezing be carefully done, but it is better to press 

 the liquid through quickly and to filter the turbid solution through a small 

 filter. 30 c.c. can be obtained. 20 c.c., or 15 c.c. if the blood contain large 

 amounts of sugar + 5 c.c. of water, are used for the estimation. 3 c.c. of the 

 alkaline copper iodine solution are added to 20 c.c. of the clear solution in a 

 100 c.c. conical flask and the mixture boiled under a reflux condenser. The 

 reflux should not be attached by a cork, but its lower end is rested on the 

 neck of the flask, or the test tube with circulating water is put in the open end. 



The boiling is continued for exactly 10 minutes from the time at which 

 the flask is placed over the flame. The time taken to reach the boiling-point 

 should be from i minute 30 seconds to i minute 50 seconds. A small mano- 

 meter to regulate the gas pressure, such as described by Cole, 1 is useful if many 

 determinations have to be made. The flask is removed and thoroughly cooled 

 in running water for i -5 minutes. 5 c.c. of hydrochloric acid are added to 

 the cold solution and after effervescence has subsided it is shaken for -5 minute. 

 The whole of this operation should not exceed i minutfe in duration. A few 

 drops of starch solution are added and the solution titrated with -oo^N thio- 

 sulphate solution. The difference between this figure and that obtained in a 

 determination with 3 c.c. of copper iodine solution in 20 c.c. of water gives 

 the amount of thiosulphate required for the iodine in the cuprous iodide. The 

 corresponding amount of glucose is given in the table on p. 606. The result 

 must be multiplied by 2, since 20 c.c. of the total 40 c.c. were used. It is 

 the amount in i c.c. of blood. 



The results are about -01 per cent, too high. This is due to the presence in 

 blood of an iodine absorbing substance. Its amount can be determined by 

 carrying out an experiment with 20 c.c. of blood filtrate and 3 c.c. of an 

 alkaline iodine solution containing no copper. This amount is deducted. 

 In still more accurate determinations, allowance must be made for the small 

 amount of substance produced by boiling glucose with alkali, which also 

 absorbs iodine and accounts for -^ of the total absorption. This small amount 

 is determined by an experiment in which 20 c.c. of the filtrate are boiled with 

 the alkaline iodine solution containing no copper. 



By this method Maclean was able to show that diabetic blood showed no 

 decrease in glycolytic power and inferred that diabetic tissues, like normal 

 tissues, possessed the power of destroying glucose. 



Bang's Micro-Method. 



Bang 2 has adapted his second method (p. 230) for estimating glucose in 

 2-3 drops of blood. This method has been found to be satisfactory by Gardner 

 and Maclean who have made one or two important modifications. It is 

 carried out as follows : 



*Biochem, J,, 1914, 8, 137, 3 Biochem, 2s,, 1913, 49, i ; 1914, 57, 300. 



