URINE 441 



and then always in conjunction with either acetone or acetoacetic acid. 

 Either of these bodies may be formed from /3-hydroxybutyric acid under 

 proper conditions. It is present in especially large amount in severe 

 cases of diabetes and has also been detected in digestive disturbances, 

 continued fevers, scurvy, measles, and in fasting. It is probable 

 that, in man, 0-hydroxybutyric acid, in common with acetone and 

 acetoacetic acid, arises principally from the breaking down of fatty 

 tissues within the organism. Any condition in which large amounts 

 of acetone and acetoacetic acid, and in severe cases 0-hydroxybutyric 

 acid also, are excreted in the urine is known as an "acidosis." In 

 diabetes the deranged metabolic conditions cause the production of 

 great quantities of these suDstances which lead to an acid intoxication 

 and ultimately to diabetic coma. In severe diabetes 50-100 grams or 

 over per day may be excreted. In. such conditions the /3-hydroxy- 

 butyric acid may constitute 60-80 per cent of the total acetone bodies. 

 In rare cases we may have an excretion of large amounts of /3-hydroxy- 

 butyric acid with a low acetone output. An acidosis may also occur 

 under certain physiological conditions (see Chapter XXVII on Metab- 

 olism) . 



Ordinarily /3-hydroxybutyric acid is an odorless, transparent syrup 

 which is levorotatory and easily soluble in water, alcohol, and ether; 

 it may be obtained in crystalline form. 



EXPERIMENTS 



i. Black's Reaction. 1 Inasmuch as the urinary pigments as well as any 

 contained sugar or acetoacetic acid will interfere with the delicacy of this test 

 when applied to the urine directly, the following preliminary procedure is neces- 

 sary : Concentrate 10 c.c. of the urine under examination to one-third or one- 

 fourth of its original volume in an evaporating dish at a gentle heat. Acidify 

 the residue with a few drops of concentrated hydrochloric acid, add sufficient 

 plaster of Paris to make a thick paste and allow the mixture to stand until it 

 begins to "set." It should now be stirred and broken up in the dish by means 

 of a stirring rod with a blunt end. Extract the porous meal thus produced twice 

 with ether by stirring and decantation. Any /3-hydroxybutyric acid present 

 will be extracted by the ether. Evaporate the ether extract spontaneously or 

 on a water-bath, dissolve the residue in water, and neutralize it with barium 

 carbonate. To 5 to 10 c.c. of this neutral fluid in a test-tube add 2 to 3 

 drops of ordinary commercial acid hydrogen peroxide. Mix by shaking and 

 add a few drops of Black's reagent. 2 Permit the tube to stand and note the 

 gradual development of a rose color which increases to its maximum intensity 

 and then gradually fades. 3 



1 Black: Jour. Biol. Chem., 5, 207, 1908. 



2 Made by dissolving 5 grams of ferric chloride and 0.4 gram of ferrous chloride in 100 

 c.c. of water. 



3 This disappearance of color is due to the further oxidation of the acetcacetic acid. 



