n6  Assay  of  Belladonna  Leaves.         {A'Va?cii, ffm' 
Into  this  percolator  is  first  introduced  a  plug  of  cotton  of  such  size 
as  to  practically  fill  up  the  neck  of  the  percolator ;  place  20  grammes 
of  the  powdered  leaves  in  the  percolator,  add  50  c.c.  menstruum 
(alcohol  2  volumes,  water  1  volume)  and  stir  with  a  heavy  iron  wire 
until  a  homogeneous  mixture  results  and  in  such  a  manner  that  the 
air-bubbles  are  brought  as  completely  as  possible  to  the  surface ; 
rinse  the  wire  and  sides  of  the  percolator  with  25  c.c.  of  the  men- 
struum and  allow  to  macerate  for  5-6  hours  or  over  night.  Perco- 
lation is  then  allowed  to  slowly  proceed  until  about  75  0  C.  percolate 
are  obtained  when  the  percolation  is  stopped  for  a  5-6  hours'  macera- 
tion; percolation  and  maceration  alternate  each  other  until  300  c.c. 
menstruum  have  been  used;  by  evaporating  the  percolate  in  a 
weighed  porcelain  dish  (about  150  c.c.  capacity)  until  the  extract 
could  be  no  longer  stirred  with  a  glass  rod,  the  yield  of  extract  was 
obtained. 
From  this  point  on  the  assay  of  the  leaves  and  of  belladonna  ex- 
tract are  identical;  the  extract  in  the  dish  obtained  from  the  20 
grammes  leaves  (or  about  5  grammes  belladonna  extract  weighed 
into  a  beaker)  are  disintegrated  by  using  10  c.c.  acidulated  water 
(2  c.c.  H2S04,  U.S.P.,  to  make  500  c.c.)  and  stirring  with  a  glass 
rod;  when  accomplished  the  mixture  is  transferred  to  a  separatory 
funnel  and  the  vessels  rinsed  with  several  small  portions  of  acidu- 
lated water  of  2  c.c.  each;  then  dissolve  and  remove  to  the  sepa- 
rator chlorophyll  and  other  substances  insoluble  in  the  acid  water 
by  using  three  portions  of  10  c.c.  each  of  a  mixture  of  ether-chloro- 
form (ether  4  parts  and  chloroform  i  part  by  weight)  ;  now  rinse  the 
vessels  (dish  or  beaker)  with  sufficient  acidulated  water  to  make  the 
total  quantity  of  this  25  c.c,  adding  this  to  the  liquids  in  the  separator, 
and  finally  add  20  c.c.  more  of  the  ether-chloroform.  Agitate  thor- 
oughly and,  after  separation  of  the  liquids,  allow  the  acid  solution 
to  run  into  a  beaker;  repeat  with  15,  10  and  5  c.c.  acidulated  water* 
Collect  the  first  three  portions  together,  reserving  the  last  portion 
to  rinse  the  beaker  containing  the  first  portions;  clean  the  sepa- 
rator, introduce  the  acid  solutions,  finally  rinsing  the  beaker  with  5 
c.c.  water,  add  25  c.c.  ether-chloroform  (ether  2  parts,  chloroform  4 
parts  by  weight ;  this  mixture  was  found  preferable  in  those  extrac- 
tions in  which  stearic  acid  was  necessary  to  break  up  emulsions)  and 
agitate  thoroughly;  after  separation  transfer  the  ether-chloroform 
solution  to  a  smaller  separator  (about  100  c.c.  capacity)  and  repeat 
