^i^SST}  Enzymes  in  Milk.  57 
THE  DIFFERENTIATION  OF  THE  ENZYMES  IN  MILK 
BY  HYDROGEN  DIOXIDE  AND  ITS  TESTS. 
By  Charles  H.  La  Wall. 
In  a  sample  of  milk  recently  examined,  which  was  suspected  of 
having  been  preserved  with  hydrogen  dioxide,  tests  were  made  for 
that  substance  by  three  different  methods.  Two  of  these  methods 
were  reversals  or  adaptations  of  the  methods  commonly  employed 
for  the  detection  of  boiled  or  sterilized  milk,  or  milk  in  which  the 
enzymes  had  been  destroyed  by  heating. 
In  Dupouy's  method,  paradiamidobenzene  is  used  in  aqueous 
solution,  a  few  drops  being  added  to  5  c.c.  of  the  sample,  after 
which  a  few  drops  of  hydrogen  dioxide  solution  are  added.  In  the 
presence  of  raw  milk  a  strong  blue  coloration  is  obtained;  milk 
that  has  been  heated  to  over  79 0  C.  gives  no  coloration.  Check  ex- 
periments with  this  method  showed  that  in  the  presence  of  varying 
amounts  of  hydrogen  dioxide  solution,  down  to  0.5  per  cent,  (corre- 
sponding to  15  parts  in  100.000.  of  absolute  H202),  a  strong  blue 
coloration  appeared  as  soon  as  the  paradiamidobenzene  solution  was 
added. 
In  the  method  recently  described  by  Wilkinson  and  Peters  (Chem. 
Abs.j  1908,  p.  3375)  the  test  for  sterilized  or  heated  milk  is  carried 
out  with  benzidine  fparadiaminodiphenyl)  as  follows  :  To  10  c.c.  of 
the  milk  is  added  2  c.c.  of  a  4  per  cent,  alcoholic  solution  of  benzi- 
dine, then  two  or  three  drops  of  acetic  acid  are  added  to  produce 
coagulation,  and  finally  2  c.c.  of  a  3  per  cent,  solution  of  hydrogen 
dioxide. 
The  test  as  thus  applied  detects  raw  milk  by  the  production  of  a 
blue  zone.  When  employed  to  detect  hydrogen  dioxide,  raw  milk 
is  used  and  the  test  is  applied  as  described,  except  that  instead  of 
adding  the  solution  of  hydrogen  dioxide,  the  milk  suspected  of 
containing  the  hydrogen  dioxide  is  added.  It  was  found  that  with 
quantities  varying  from  3  parts  absolute  H202  in  1000,  down  to 
15  parts  in  100,000,  the  reaction  was  prompt  and  unmistakable.  So 
-far  as  the  detection  of  the  hvdrogen  dioxide  was  concerned,  there- 
fore, either  of  the  methods  was  equally  applicable  and  both  were 
found  to  be  superior  to  the  method  of  adding  potassium  chromate 
and  diluted  sulphuric  acid,  which  causes  the  development  of  a  blue 
color  in  presence  of  H202. 
