3   Chapter 3: Karl Fischer Titration Method Utilizing THF Extraction 36

3.4   Test Method Assessment 39

3.4.1   Analytical Method 39

In  order  to  isolate  the  possible  sources  of  test  method  variability  in  the  THF  

extraction  method,  all  method  assessment  experiments  were  performed  without  the   presence  of  a  stopper  unless  stated  otherwise.    This  eliminated  the  possible  

variability  that  could  be  attributed  to  the  stopper  samples  themselves.    Tested   samples  consisted  solely  of  THF  to  mimic  the  blank  in  the  original  analytical   method. Sample  Container  

Cap  Type  

A  graduated  cylinder  was  used  to  aliquot  40  mL  of  tetrahydrofuran  (THF)  into  nine   125-­‐mL  sample  bottles.    Onto  three  bottles,  thick  septum  caps  were  added,  three   more  bottles  had  thin  septum  caps,  and  the  final  three  bottles  received  non-­‐septum   caps.    The  extraction  and  test  process  remained  the  same  as  the  original  test  method   except  samples  were  tested  at  4,  10,  24  and  32  hours  and  the  non-­‐septum  cap  


Image  3.1:  Septum  Caps  and  Non-­‐Septum  Caps  


In  a  second  experiment,  a  graduated  cylinder  was  used  to  aliquot  40  mL  of   tetrahydrofuran  (THF)  into  seven  125-­‐mL  sample  bottles  and  non-­‐septum  caps   were  added.    The  extraction  and  test  process  remained  the  same  as  the  original  test   method  except  samples  were  tested  at  0,  24  and  36  hours  and  at  each  of  the  test   intervals,  for  three  of  the  samples,  a  thick  septum  cap  was  added  through  which   sample  injection  aliquots  were  obtained.    For  the  other  samples,  the  non-­‐septum   caps  were  removed  in  a  dry  box  to  obtain  the  sample  injection  aliquots.    The  

samples  with  septum  caps  only  had  exposure  to  the  septum  caps  during  the  sample   injection  testing  periods  and  during  the  rest  of  the  extraction  time,  non-­‐septum  caps   were  present  on  these  samples.  

Sample  Bottle  

Six  125-­‐mL  bottles  and  PTFE-­‐lined  non-­‐septum  caps  were  removed  from  the   laboratory  cabinets  where  they  were  stored  at  ambient  conditions  after  washing   and  drying.    Three  of  the  bottles  and  caps  were  placed  in  a  100oC  dry  heat  oven  to  

further  dry  for  one  hour  and  then  were  cooled  in  a  desiccator  for  another  hour.     Next,  a  pipette  was  used  to  aliquot  40  mL  of  tetrahydrofuran  (THF)  into  all  of  the   sample  bottles  and  the  caps  were  added.    The  extraction  and  test  process  remained   the  same  as  the  original  test  method  except  samples  were  tested  at  0,  24,  and  48   hours  and  for  all  samples,  the  non-­‐septum  caps  were  removed  in  a  dry  box  to  obtain   the  sample  injection  aliquots  and  the  coulometric  cell  was  not  swirled  prior  to   titration  initiation. Equipment  for  Measurement  

Three  40-­‐mL  samples  of  THF  were  aliquoted  into  125-­‐mL  bottles  with  non-­‐septum   caps  using  a  funnel  and  graduated  cylinder.    An  additional  three  40-­‐mL  samples  of   THF  were  aliquoted  into  125-­‐mL  bottles  with  non-­‐septum  caps  using  a  glass  pipette.     The  extraction  and  test  process  remained  the  same  as  the  original  test  method   except  samples  were  tested  at  0,  24  and  33  hours  and  for  all  samples,  the  non-­‐ septum  caps  were  removed  in  a  dry  box  to  obtain  the  sample  injection  aliquots. Location  of  Sample  Injection  Aliquoting    

Six  40-­‐mL  samples  of  THF  were  aliquoted  into  125-­‐mL  bottles  with  non-­‐septum   caps  using  a  glass  pipette.    The  extraction  and  test  process  remained  the  same  as  the   original  analytical  method  except  samples  were  tested  at  0,  6,  10,  24,  and  31  hours   and  no  swirling  of  the  titration  cell  occurred  after  sample  injection.    Three  of  the   samples’  injections  were  aliquoted  by  removing  the  non-­‐septum  caps  in  a  dry  box.     The  other  three  samples’  injections  were  aliquoted  by  removing  the  non-­‐septum   caps  on  a  lab  bench. Titration  Methodology  

Three  40-­‐mL  samples  of  THF  were  aliquoted  into  125-­‐mL  bottles  with  non-­‐septum   PTFE-­‐lined  caps  using  a  glass  pipette.    The  samples  were  then  analyzed  twice  at   time  zero  using  the  same  test  process  as  the  original  test  method  except  the  caps   were  removed  in  the  dry  box  to  obtain  the  sample  injection  aliquots.    In  addition  for   the  first  test,  the  titration  cell  was  swirled  between  each  sample,  but  no  shaking   occurred  after  sample  injection  and  prior  to  titration  initiation.    For  the  second  test,   the  titration  cell  was  swirled  between  each  sample  and  after  each  sample  injection   prior  to  titration  initiation. Extraction  Process  

Duration  of  Shaking  

Six  stoppers  were  each  placed  in  a  125-­‐mL  bottle  with  PTFE-­‐lined  cap  and  covered   with  40  mL  of  THF  aliquoted  using  a  glass  pipette.    Three  of  the  stopper  samples   were  placed  on  a  shaker  and  shook  for  a  total  of  8  hours  while  the  other  three   stopper  samples  remained  on  the  lab  bench  for  the  8  hour  period.    Both  sets  of   stoppers  were  tested  at  zero,  2,  4,  6,  and  8  hours.    Shaker  stopper  samples  were   removed  from  the  shaker  one  at  a  time  for  testing  to  ensure  the  maximum  amount   of  time  for  each  stopper  on  the  shaker  over  the  eight  hours.    The  test  process   remained  the  same  as  the  original  analytical  method  except  the  non-­‐septum  caps   were  removed  in  the  dry  box  to  obtain  the  sample  injection  aliquots  and  no  swirling   of  the  titration  cell  occurred  after  sample  injection  and  before  titration  initiation.        

Length  of  Extraction  

Three  stoppers  were  each  placed  in  a  125-­‐mL  bottle  with  PTFE-­‐lined  cap  and   covered  with  40  mL  of  THF  aliquoted  using  a  glass  pipette.    A  second  set  of  three   bottles  was  prepared  in  the  same  fashion  without  the  presence  of  stoppers  to  serve   as  the  blank  controls.    The  extraction  and  test  process  remained  the  same  as  the   original  test  method  except  samples  were  tested  at  0,  6,  10,  24,  and  31  hours.    In   addition,  the  non-­‐septum  caps  were  used  and  removed  in  the  dry  box  to  obtain  the   sample  injection  aliquots  and  no  swirling  of  the  titration  cell  occurred  after  sample   injection  and  before  titration  initiation.      

3.4.2 Test  Method  Assessment  Results/Discussion  

In document Water Content Determination of Rubber Stoppers Utilized for Sealing Lyophilized Pharmaceutical Products: Assessment of Two Karl Fischer Titration Methods (Page 49-53)