522 PHYSIOLOGICAL CHEMISTRY 



Glucose 



i. Benedict's Method. 1 Principle.- Benedict's reagent for the esti- 

 mation of reducing sugars contains potassium thiocyanate as well as 

 copper sulphate, and in the presence of the former a white precipitate 

 of cuprous thiocyanate is formed on reduction instead of the usual red 

 precipitate of cuprous oxide. The small amount of potassium ferro- 

 cyanide also aids in keeping cuprous oxide in solution. As the pre- 

 cipitate formed is white the loss of all blue tint in the solution, indicating 

 complete reduction of the copper, is readily observed. The alkali 

 used is sodium carbonate, which has the advantage over the hydroxides 

 in that there is less danger of destruction of small amounts of sugar. 

 The solution also has the great advantage of being stable for an in- 

 definite length of time. The method is recommended for simplicity 

 and accuracy. 



Procedure. The urine, 10 c.c. of which should be diluted with water to 100 

 c.c. (unless the sugar content is believed to be low, when it may be used undiluted) , 

 is poured into a 50 c.c. burette up to the zero mark. Twenty-five c.c. of the 

 reagent 2 are measured with a pipette into a porcelain evaporation dish (25-30 cm. 

 in diameter), 10 to 20 grams of crystallized sodium carbonate (or one -half the 

 weight of the anhydrous salt) are added, together with a small quantity of pow- 

 dered pumice stone or talcum, and the mixture heated to boiling over a free 

 flame until the carbonate has entirely dissolved. The diluted urine is now run in 

 from the burette, rather rapidly, until a chalk-white precipitate forms and the 

 blue color of the mixture begins to lessen perceptibly, after which the solution 

 from the burette must be run in a few drops at a time, until the disappearance of 

 the last trace of blue color, which marks the end point. The solution must be 

 kept vigorously boiling throughout the entire titration. If the mixture becomes 

 too concentrated during the process, water may be added from time to time to 

 replace the volume lost by evaporation. 



Calculation. The calculation of the percentage of sugar in the original sample 

 of urine is very simple. The 25 c.c. of copper solution are reduced by exactly 50 

 mg. of glucose. Therefore the volume run out of the burette to effect the reduc- 

 tion contained 50 mg. of the sugar. When the urine is diluted i : 10, as in the 



Benedict: Jour. Am. Med. Ass'n, 57, 1193, 1911. 



2 Copper sulphate (crystallized*) 18 . o grams. 



Sodium carbonate (crystallized, one-half the weight of the 



the anhydrous salt may be used) 200 . o grams. 



Sodium or potassium citrate 200. o grams. 



Potassium thiocyanate 125 . o grams. 



Potassium ferrocyanide (5 per cent solution) 5.0 c.c. 



Distilled water to make a total volume of 1000. o c.c. 



With the aid of heat dissolve the carbonate, citrate and thiocyanate in enough water to 

 make about 800 c.c. of the mixture and filter if necessary. Dissolve the copper sulphate 

 separately in about 100 c.c. of water and pour the solution slowly into the other liquid, with 

 constant stirring. Add the ferrocyanide solution, cool and dilute to exactly i liter. Of the 

 various constituents, the copper salt only need be weighed with exactness. Twenty-five 

 c.c. of the reagent are reduced by 50 mg. of glucose. 



