464 
Assay  of  Heart  Tonics. 
f  Am.  Jour.  Pharm. 
I     October,  1909. 
animals.  The  water  should  cover  the  bottom  of  the  baskets  or  plates 
about  one-fourth  inch.  Small  balance  for  Weighing  the  frogs. 
Volumetric  flasks  for  diluting  the  preparations  to  be  assayed,  and 
glass  pipettes,  graduated  into  hundredths  of  a  cubic  centimetre,  with 
long  narrow  points,  or,  better,  supplied  with  hypodermic  needle- 
points for  injecting  the  frogs. 
12.  The  frogs  should  be  handled  with  much  care,  avoiding 
squeezing,  pinching,  and  such  while  making  the  injection  in  the  ab- 
dominal lymph-sac.  The  amount  of  the  heart  tonic  preparation 
should  be  very  carefully  diluted  with  physiological  salt  solution,  so 
that  the  total  quantity  injected  into  each  frog  will  be  about  0.5  c.c. 
13.  It  is  necessary  in  making  an  assay  of  the  heart  tonics  to  inject 
several  series  of  frogs  (see  Table  I). 
14.  Death  of  the  frog  should  be  the  criterion  for  reading  end 
results,  a  final  examination  of  the  frog  being  made  at  the  end  of 
twelve  hours ;  those  dying  prior  to  the  end  of  the  twelve-hour  period 
art  considered  as  killed  by  the  drug,  unless  some  accident  has 
occurred. 
15.  Ten  times  the  normal  minimum  fatal  dose  (M.F.D.)  per 
gramme  body  weight  of  frog  of  the  standard  preparation  properly 
diluted  and  injected  is  considered  as  a  heart  tonic  unit  (H.T.U.),. 
consequently  the  number  of  H.T.U.s  in  a  given  cubic  centi- 
metre of  a  preparation  is  one-tenth  of  the  quotient  obtained  by 
dividing  one  by  such  M.F.D.,  i.e.,  the  number  of  H.T.U.s  in  each 
cubic  centimetre  of  fluid  is  one-tenth  of  the  reciprocal  of  the  M.F.D. 
To  illustrate :  if  the  minimum  fatal  dose  of  a  given  drug  per  gramme 
body  weight  of  frog  is  found  to  be  equivalent  to  0.01,  then  the 
given  substance,  assuming  that  it  belongs  to  the  group  of  heart 
tonics,  would  contain  10  heart  tonic  units:  (M.F.D.  — —   -f-  10  = 
0.0 1 
10  H.T.U.s).  This  rule  can  be  applied  to  any  of  the  heart  tonics 
as  a  means  of  expressing  such  values  in  whole  numbers.  Definition : 
A  heart  tonic  unit  (H.T.U.)  is  ten  times  the  normal  minimum 
fatal  dose  per  gramme  body  weight  of  standard  test  frogs  kept  under 
proper  test  conditions. 
It  is  to  be  understood  that  the  H.T.U.  is  a  measure  of  toxic 
value,  although  such  toxic  values  may  be  translated  directly  into 
therapeutic  values,  based  on  the  U.S. P.  average  dose  or  any  other 
dose. 
