240 
Studies  on  Pepsin. 
[Am.  Jour.  Pharm. 
*•       April,  1919. 
heat.  From  the  fact  that  the  highest  strength  samples  still  give 
strong  tests  with  Molisch  reagent,  it  may  be  possible  that  the  pure 
enzyme  is  a  conjugated  protein,  probably  a  glycoprotein. 
Confirming  this  view,  the  mineral  matter  is  decidedly  less  in  the 
purified  samples  than  in  the  original  basic  material,  approaching 
almost  to  the  value  for  pure  proteins  in  the  case  of  the  strongest 
samples.  Both  sulphur  and  calcium  are  probably  unaffected  by  the 
purification,  but  there  is  a  decided  decrease  in  the  phosphorus  content 
and  seemingly  a  total  elimination  of  chlorides.  Other  than  the  in- 
crease which  would  obtain  by  removal  of  non-nitrogenous  impuri- 
ties, there  is  probably  not  much  change  in  the  content  of  total  nitro- 
gen as  a  result  of  pepsin  purification. 
The  manner  in  which  the  a-amino  acids  decrease  as  the  proteo- 
lytic activity  increases  is  striking,  and  seems  to  be  almost  proportional 
in  amount.  It  is  noteworthy  that  the  small  amount  of  a-amino  acid 
present  in  the  sample  tesing  1  : 40,000  (0.61  per  cent.)  very  nearly 
approaches  the  value  for  this  factor  due  to  lysin  as  found  present  by 
Van  Slyke  and  Birchard20  in  most  proteins  analyzed  by  the  nitrous 
acid  method. 
Results  of  optical  activity  determinations  are  apparently  of  no 
significance,  since  the  same  values  are  obtained  with  several  different 
strengths  of  pepsin.  As  already  mentioned  above,  the  reaction  in 
aqueous  solution  of  the  strongest  (1:40,000)  pepsin  is  significant 
because  of  its  very  slight  acidity.  It  would  seem  very  likely,  that 
the  concentration  of  hydogen  ions  in  solutions  of  the  pure  enzyme, 
when  isolated,  will  probably  show  only  the  slight  acidity  comparable 
to  that  given  by  other  proteins. 
In  connection  with  the  assays  of  proteolytic  strength  by  the  U.  S. 
P.  method,  it  was  deemed  of  interest  to  make  a  comparison  of  the 
rennetic  power  of  the  different  samples.  It  is  a  significant  fact  that 
throughout  the  entire  series,  from  1  : 2,000  to  1  : 40,000,  the  rennetic 
activity  and  proteolytic  strengths  are  found  to  go  hand  in  hand. 
This  is  being  investigated,  and  will  be  reported  upon  in  a  later  paper. 
Conclusions. 
1.  The  purification  of  pepsin  seems  to  consist  in  the  elimination 
of  secondary  protein  derivatives  including  a-amino  acids. 
2.  Calcium  and  sulphur  appear  to  be  unaltered  as  a  result  of 
20  Van  Slyke  and  Birchard,  /.  Biol.  Chem.,  vol.  16,  539,  1914. 
