Am.  Jour.  Pharm.  \ 
March,  19 19.  > 
Note  on  Trypsin. 
175 
from  albuminous  matter,  I  was  surprised  to  find  that  no  pre- 
cipitate was  formed,  i.  e.,  the  ordinary  preparations  of  trypsin,  and 
aqueous  extracts  of  pancreas,  gave  precipitates,  but  the  purified  prep- 
aration (the  strength  of  which  was  equal  to  that  of  the  crude  prep- 
aration) gave  no  precipitate.  It  appears,  therefore,  that  the  matter 
precipitated  by  safranine  is  an  albuminous  compound,  which  carries 
the  enzyme  down  with  it,  and  that  the  precipitation  by  this  method 
does  not  produce  an  enzyme-safranine  compound.  The  method  ap- 
pears to  be  analogous  to  that  used  by  Cohnhein5  for  the  precipita- 
tion of  diastase  from  saliva  by  means  of  calcium  phosphate,  in  which 
a  certain  quantity  of  phosphoric  acid  was  added  to  saliva  and  the 
liquid  neutralized  with  lime  water,  which  caused  a  copious  precipi- 
tate of  calcium  phosphate.  This  precipitate  carried  down  with  it  a 
large  proportion  of  the  proteins  of  the  saliva,"  together  with  the 
diastase  or  ptyalin.  The  precipitate  was  then  collected  by  filtra- 
tion, and  extracted  with  a  volume  of  water  equal  to  that  of  the 
saliva  originally  used.  The  diastase  is  slightly  more  soluble  than 
the  proteins,  and  accordingly  passes  into  solution  first.  Conheim 
repeated  this  process  several  times,  and  finally  precipitated  the 
last  extract  by  the  addition  of  alcohol.  This  precipitate  was  col- 
lected, washed  with  alcohol,  and  dried  over  sulphuric  acid,  appear- 
ing then  as  an  amorphous  white  powder,  freely  soluble  in  water. 
The  author  considered  it  to  be  free  from  proteins  as  its  solution  did 
not  yield  the  usual  reactions  characteristic  of  these  bodies.6 
I  have  not'  yet  had  the  opportunity  to  compare  the  nitrogen  con- 
tent of  the  purified  trypsin  with  that  of  the  trypsin  before  purifica- 
tion by  the  method  I  am  about  to  describe.  This  would  have  an 
important  bearing  on  the  constitution  of  the  enzyme. 
On  referring  to  Robertson's  book  on  the  proteins,7  I  find  that, 
speaking  of  his  own  work  two  years  earlier,  he  says,  in  the  course 
of  a  discussion  on  the  compounds  of  proteins  with  dyes,  "I  have 
found  that  in  faintly  alkaline  solution  trypsin  or  some  constituent  of 
Grubler's  trypsin  (nach  Spalteholz)  forms  an  insoluble  compound 
with  safranine  " ;  from  this  it  appears  that  Robertson  himself  was 
5  Virchon's  Archiv,  1863,  28,  241.  J.  Reynolds  Green^  The  Soluble  Fer- 
ments (Camb.  Univ.  Press,  1899)^  p.  45. 
6  Since  writing  this  paper  the  author  has  attempted  to  apply  Cohnheim's 
method  to  trypsin,  but  has  been  unsuccessful. 
7  The  Proteins  (Univ.  of  California  Publications  in  Physiology),  Octo- 
ber, 1909. 
