The   molecular   rota-onal   spectrum   is   highly   sensi-ve   to   the  three   dimensional   geometry   of   the   molecule.     The   simulated  molecular   rota-onal   spectra   of   two   artemisinin   diastereomers  that   have   been   iden-fied   in   the   natural   product   are   shown.    These   two   structures   differ   in   the   stereochemistry   at   a   single  asymmetric   carbon   (the   structures   are   epimers).     As   the  expanded   scale   view   on   the   right   makes   clear,   there   is  essen-ally  no  spectral  overlap   in   the  rota-onal  spectra  so   that  the   diastereomers   can   be   readily   iden-fied   in   the   FT-­‐MRR  spectrum.  

Differen'a'on  of  Diastereomers  without  Chromatography  

BrightSpec   is   developing   instruments   based   on   Fourier  transform   molecular   rota8onal   resonance   (FT-­‐MRR)  spectroscopy  with  full  capabili8es  for  chiral  analysis:        •  Quan'ta've   determina'on   of   diastereomer  

composi'on   for   a   molecule   with   mul'ple   chiral  centers.  

•  Determina'on  of  the  absolute  configura'on  of  chiral  molecules.  

•  Quan'ta've   measurement   of   the   enan'omeric  excess   for  a  mixture  of   le?  and   right  handed   forms  of  a  single,  chiral  diastereomer.      

The   technique   does   not   require   chromatography   to  separate  diastereomers  and  enan8omers  and  it  does  not  require   chiral   deriva8zing   agents   to   dis8nguish   the  enan8omers.    Chiral  analysis  of  molecules  of  interest  can  be  performed  directly  on  complex  sample  mixtures.    The  technique   can   provide   real-­‐'me   monitoring   of  diastereomer  composi8on  and  enan8omeric  excess.      BrightSpec  FT-­‐MRR  spectrometers  for  chiral  analysis  are  enabled  by  two  recent  technology  advances   in  the  field  of   molecular   rota8onal   spectroscopy.[1]     Broadband  Fourier   transform   rota8onal   spectroscopy   has   been  extended   to   low   frequency   opera8on   so   that   large  molecules   can   be   studied   using   pulsed   jet   sample  methods.[2,3]     This   advance   has   made   it   possible   to  determine   the   structure   of   difficult   chemical   systems,  like  water  clusters  [4]  and  molecular  complexes  [5].    The  high  spectral   resolu8on  of   the   instrument  coupled  with  the   extreme   sensi8vity   of   the   spectral   paSern   to   small  changes   in   the   three   dimensional   geometry   make   it  possible   to   resolve   diastereomers   of   molecules   with  mul8ple  chiral  centers  without  prior  separa8on.  

FT-­‐MRR  Spectrometers  for    Chiral  Analysis  

Early  Access  Program  Analy-cal  services    and  on-­‐site  spectrometers  

Contact  BrightSpec  

BrightSpec  is  ac'vely  seeking  customers  with  challenging  problems  in  chiral  analysis  who  are  interested  in  working  with  us  to  develop  this  new  measurement  technology.    Please  contact    

jus'n.neill@brightspec.com  

©2014,  BrightSpec,  Inc.  

Literature  References  (1)  B.H.  Pate,  “Taking  the  Pulse  of  Molecular  Rota8onal  Spectroscopy”,  Perspec8ves,  Science  333,  947-­‐948  (2011).      (2)  Cristóbal  Pérez,  Simon  Lobsiger,  Nathan  A.  Seifert,  Daniel  P.  Zaleski,  Berhane  Temelso,  George  C.  Shields,  Zbigniew  Kisiel,    and  Brooks  H.  Pate,  “Broadband  Fourier  Transform  Rota8onal  Spectroscopy  for  Structure  Determina8on:  The  Water  Heptamer  (Fron8ers  Ar8cle)”,  Chem.  Phys.  LeJ.  571,  1-­‐15  (2013).      (3)  Steven  T.  Shipman,  Jus8n  L.  Neill,  Richard  D.  Suenram,  MaS  T.  Muckle,  Brooks  H.  Pate,  “Structure  Determina8on  of  Strawberry  Aldehyde  by  Broadband  Microwave  Spectroscopy:  Conforma8onal  Stabiliza8on  by  Dispersive  Interac8ons”,  J.  Phys.  Chem.  LeJ.  2,  443-­‐448  (2011).    (4)  Cristóbal  Pérez,  MaS  T.  Muckle,  Daniel  P.  Zaleski,  Nathan  A.  Seifert,  Berhane  Temelso,  George  C.  Shields,  Zbigniew  Kisiel,  and  Brooks  H.  Pate,  “Structures  of  Cage,  Prism,  and  Book  Isomers  of  Water  Hexamer  from  Broadband  Rota8onal  Spectroscopy”,  Science  336,  897-­‐901  (2012).  

 (5)  Seifert,  N.  A.;  Zaleski,  D.  P.;  Pérez,  C.;  Neill,  J.  L.;  Pate,  B.  H.;  Vallejo-­‐López,  M.;  Lesarri,  A.;  Cocinero,  E.  J.;  Castaño,  F.;  Kleiner,  I.,  “Probing  the  C-­‐H·∙·∙·∙pi  Weak  Hydrogen  Bond  in  Anesthe8c  Binding:  The  Sevoflurane–Benzene  Cluster”,  Angew.  Chem.  2014,  126,  3274.      (6)  PaSerson,  D.;  Schnell,  M.;  Doyle,  J.  M.  Enan8omer-­‐Specific  Detec8on  of  Chiral  Molecules  via  Microwave  Spectroscopy.  Nature  2013,  497,  475–477.      (7)  PaSerson,  D.;  Doyle,  J.  M.  Sensi8ve  Chiral  Analysis  via  Microwave  Three-­‐Wave  Mixing.  Phys.  Rev.  LeJ.  2013,  111,  023008.      (8)  Simon  Lobsiger,  Cristobal  Perez,  Luca  Evangelis8,  Kevin  K.  Lehmann,  Brooks  H.  Pate,  “Molecular  Structure  and  Chirality  Detec8on  by  Microwave  Spectroscopy”,  J.  Phys.  Chem.  LeJ.  (submiSed).  

The  New  Physics  of  Absolute  Configura'on  Determina'on  by  FT-­‐MRR  Spectroscopy  

Analysis  of  the  chirality,  or  handedness,  of  the  dis8nct  diastereomers   is   possible   through   a   revolu8onary  measurement   technique   first   reported   in   Nature   in  2013.[6]     This   work   demonstrated   that   a   chiral  spectral   signature   can   be   measured   using   FT-­‐MRR  spectroscopy   when   a   three-­‐wave   mixing   scheme   is  employed.    The  key   insight   is   that  enan8omers  differ  in  the  way  that  the  molecular  dipole  moment  projects  onto   the   three   principal   axes   of   molecular   rota8on  and   that   the   three   wave   mixing   method   creates   a  signal   that   reflects   this   physical   difference.     The  handedness   of   the   molecule   is   obtained   from   the  phase   of   the   molecular   free   induc8on   decay   and  enan8omeric  excess   is  measured  by  the  amplitude  of  the   chiral   signal.[7]     Because   the  measurement   uses  three  precisely  defined  resonance  frequencies  for  the  molecule,   there   is   prac8cally   no   chance   to   record   an  interfering  signal  even  in  a  complex  sample  matrix.  

The   measurement   uses   the   fact   that   enan-omers   can   be  dis-nguished   by   the   proper-es   of   the  molecular   dipole  moment.    The  product  of  the  projec-ons  of  the  dipole  moment  on  the  three  principal  axes  for  molecular  rota-on  (denoted  a,  b,  and  c)  changes  signs   for   the   enan-omer   pair.     This   behavior   is   illustrated   for  solketal  (C6H12O3).[8]  

Three  Wave  Mixing  Measurements  Determine  the  Absolute  Configura'on  

Three   wave   mixing   experiments   use   two,   resonant   excita-on   pulses   to   generate   a   coherent   chiral   signal   that   is   detected   by   FT-­‐MRR  spectroscopy.     The   chiral   signal   is   propor-onal   to   the  product   of   the  dipole  moment   components   and,   therefore,   has   opposite   sign   (or  phase)  for  the  two  enan-omers.    The  chiral  signals  for  pure  R-­‐  and  S-­‐solketal  are  shown  in  the  middle  panel  to  illustrate  this  effect.[8]    More  generally,  the  amplitude  of  the  chiral  FT-­‐MRR  signal  measures  the  enan-omeric  excess  (ee)  providing  way  to  monitor  ee  in  real  -me.