292 PRACTICAL PHYSIOLOGY 



let stand at room temperature for thirty to forty minutes. Add 5 c.c. 



N 

 cone. HC1 and titrate excess of iodine with y^ sodium thiosulphate 



N 

 solution. 1 c.c. of y^ iodine solution = 1-567 mg. phenol. On the 



basis of the results dilute phenol solution so that 10 c.c. = 1 mg. of 

 phenol. (Benedict and Theis (J. Biol. Chem. 36, 1918) recommend 

 resorcinol as standard as the solution is easily prepared and keeps well. ) 

 Add to 10 c.c. of this standard in a 100 c.c. flask 0-5 c.c. HC1 and 10 c.c. 

 of the silver lactate-lactic acid solution (as lactic acid also gives a blue 

 colour with the reagent), shake well and filter. To the filtrate add 

 10 c.c. phenol reagent 1 and 2 5 c.c. saturated sodium carbonate solution. 

 (For free phenols best 5 c.c. reagent and 15 c.c. carbonate solution.) 

 Fill to the 100 c.c. mark with water about 30 C. and allow to stand 

 about twenty minutes. Set standard in Duboscq at 20 mm. 



Estimation of Sugar. Pavy's Method. The standard solution con- 

 tains 120 c.c. Fehling's solution and 300 c.c. strong ammonia per litre. 



The nozzle of a burette is fitted to a small round-bottomed flask 

 by means of a cork through which is also passed a short bent tube 

 to allow of the escape of steam and ammonia, when the flask is boiled. 

 The urine is diluted exactly 10 to 50 times according to the amount 

 of sugar present. The burette is filled with diluted urine, care being 

 taken to see that there are no bubbles in the nozzle. 10 c.c. of Pavy's 

 solution and about an equal volume of water are placed in the flask. 

 The flask is now heated till it boils. The heating is continued and the 

 urine allowed to drop in from the burette at such a rate that boiling 

 does not cease. When the colour of the solution in the flask is per- 

 ceptibly less the rate of addition of drops is reduced, but is continued 

 until all the blue colour has disappeared. The first reading will be 

 almost certainly too high so that repeat determinations must be 

 made. In the later determinations it is recommended to run in fairly 

 rapidly the quantity of urine which will almost suffice, then wait till 

 the colour is constant before adding the rest of the urine required 

 drop by drop till the blue colour has completely disappeared. The 

 amount of diluted urine employed should not be less than 2 c.c. or 

 more than 5 c.c. Calculation : 10 c.c. Pavy solution are equivalent to 

 0-005 gm. dextrose. 



Quantitative Estimation of Beta-Oxybutyric Acid, Acetoacetic Acid 

 and Acetone (van Slyke). In this method all three substances 

 total acetone bodies can be determined at once, a separate determina- 

 tion of the beta oxybutyric acid can be made by boiling off the aceto- 

 acetic acid and acetone from the treated urine which has been acidified 

 with sulphuric acid. 



Place 25 c.c. of the urine to be tested in a 250 c.c. measuring flask, 

 add 100 c.c. distilled water, 50 c.c. copper sulphate solution (200 gms. 

 copper sulphate, CuSO 4 ,5H 2 O, dissolved in water and made up to 

 1,000 c.c.) and mix well. Next add 50 c.c. of a well-shaken suspension 

 of calcium hydroxide (100 gms. light calcium hydroxide in 1,000 c.c. 

 water) and shake the whole mixture well. The copper and lime are 

 added to remove sugar and other interfering substances. If the urine 

 contains more than 8 per cent, sugar it must be diluted so that the 

 sugar present does not exceed this amount. The reaction of the 

 1 See footnote on page 291. 



