How Theory and Practice Meet in Ion Pair Chromatography

Jan Sthlberg Academy of Chromatography SWE E!

"#tline of the pre$entation

What i$ Ion Pair Chromatography %IPC&' Introd#ction to the theoretical concept$( A $implified) b#t practical) model for IPC( How to #$e the $imple model in practical wor*( Compari$on between the $implified model and e+periment$( Concl#$ion$(

What i$ Ion Pair Chromatography'

Ion Pair Chromatography %IPC& i$ #$ed to $eparate ionic analyte$ on a re,er$ed pha$e col#mn( An Ion Pair reagent i$ added to mod#late retention of the ionic analyte$( Typical IP reagent$ are-al*yl$#lphonate$) al*yl$#lphate$) tetra.al*ylammoni#m ion$ etc(

What is the effect of the IP-reagent on retention of analytes of different charge?

!o IP.reagent

_
/etention time

What is the effect of the IP-reagent on retention of analytes of different charge?

!o IP.reagent 0 charged IP.reagent

+

_
/etention time

What is the effect of the IP-reagent on retention of analytes of different charge?

!o IP.reagent 0 charged IP.reagent . charged IP.reagent

+

_
/etention time

Sche atic e!"lanation of the effect of the IP-reagent on retention

A0 A0 When no IP regent i$ added the di$trib#tion i$ 1-1 for all analyte$ A0 A0 A0 A+ A+ A+ A+ AAAA0 A0 A0 A0 A0 A+ A+ A+ A+ A+ AAAAA-

A+

AA-

S O L I D P H A S E

Sche atic e!"lanation of the effect of the IP-reagent on retention

A0 A0 The IP.reagent will ad$orb to the $tationary pha$e( IP+ AAAAAA0 A0 A0 A+ A+ A+ A+ A0 A0 A0 A0 A0 A+ A+ A+ A+ A+ IP+ AAAAA-

A+

S O L I D P H A S E

Sche atic e!"lanation of the effect of the IP-reagent on retention#

A0 A0 IP+ A+ A+ A+ A0 A0 A0 A+ A+ A+ A+ A+ A+ A0 IP+ A0 A0 IP+ A0 IP+ A0 A+ IP+ IP+ AAAAAAAAAIP+

The charge of the ad$orbed IP.reagent interact$ electro$tatically with the charge of the analyte$

IP+ A-

S O L I D P H A S E

E+perimental parameter$ in IP. chromatography

Type of analyte %charge) hydrophobicity) etc&( Type of IP.reagent %charge) hydrophobicity etc(& Concentration of IP.reagent( Type of organic modifier in the mobile pha$e( Concentration of organic modifier in the mobile pha$e( Ionic $trength and type of ion$ in the mobile pha$e(

How theory meet practice in IP. chromatography(

A theory for IP.chromatography $hall 2#antitati,ely de$cribe how retention i$ affected byThe charge of the analyte( The type and concentration of IP.reagent The type and concentration of organic modifier The ionic $trength( The theory $hall be phy$ically acceptable(

Charged $#rface in contact with an electrolyte(

(

_ +

How are ion$ di$trib#ted clo$e to a charged $#rface'

+ + _

+ _ + _

_ _ _ _ _ _ _

+ + + + + + +

_ + _ + _ + _ +

Charged $#rface in contact with an electrolyte(

(

_ +

How are ion$ di$trib#ted clo$e to a charged $#rface'

+ + _

+ _ + _

_ _ _ _ _ _ _

+ + + + + + +

_ + _ + _ + _ +

The Electrical
At e2#ilibri#m a minim#m in free + energy i$ achie,ed( A difference)) in electro$tatic potential between the $#rface and the electrolyte i$ created(

o#ble 3ayer

_ + _ + + + _ + _ + _ + _ + 4 _ _ + + _ + _ _ + _ _ + _ + _ +

Thermodynamic$ of ad$orption of the IP.reagent and retention(

_ + _ + + + _ + _ + + _ 8 8 + _ + 4 5tot65chem+47 4 _ _ + + 8 8 4 _ + 4 5tot65chem+47 _ _ + _ _ + _ + _ +

IP

IP

Implication$ of the electro$tatic retention model(

depend$ only on the concentration of IP. reagent on the $#rface and i$ independent of the type( 5chem i$ more or le$$ independent of the type and concentration of the IP.reagent( The retention factor i$ de$cribed by the e2#ation*6*89e+p%.47:/T& where *8 i$ the retention factor in ab$ence of the IP.reagent(

Implication$ of the electro$tatic retention model(

7or a gi,en analyte doe$ retention depend$ on the $#rface concentration of the IP.reagent only( It i$ independent of the type of IP.reagent) E+ample- /etention factor of adrenaline a$ a f#nction of the $#rface concentration of different anionic IP.reagent$( % ;artha et(al( J( Chromatogr() <8<) %1=>?& @=(

E+perimental data from ;artha et(al) J( Chromatogr( <8< %1=>?& @=

The 5o#y.Chapman Theory

A phy$ically ba$ed theory for a planar charged $#rface in contact with an electrolyte $ol#tion% a $ol#tion of the Poi$$on.;olt4mann e2(&( The electrolyte i$ a$$#med to con$i$t$ of point charge$( The $#rface charge$ are a$$#med to be e,enly $meared o#t( e$cribe$ the $#rface potential a$ a f#nction of $#rface charge den$ity and the ionic $trength of the electrolyte(

"#tline of a $implified electro$tatic theory for IP.chromatography(

depend$ linearly on the $#rface concentration of IP. reagent and on the in,er$e of the ionic $trength of the mobile pha$e( ;oth the IP.reagent and the analyte are f#lly ioni4ed( 7or a gi,en col#mn doe$ 5chem for both the IP.reagent and the analyte depend only on the mobile pha$e compo$ition) i(e( not on ( The $implification$ are rea$onable when the ratio *cIP:*8 i$ between @ . 18

Simplified theory) cont(

A$ing appro+imation$ of the 5.C theory and $ome algebra the following e2#ation can be obtained-

Simplified theory) cont

A$ing appro+imation$ of the 5.C theory and $ome algebra the following e2#ation can be obtained-

. 4A94IP ln *cA 6 ln *8A 0 @ 4IP01

9ln%%n8BIPcIP:&0con$t& &0con$t

Capacity factor Capacity factor at concentration at concentration Electro$tatic contrib#tion cIP a$ a f#nction analyte charge) 4A) cIP68 IP reagent charge 4IP) it$ binding con$tant BIP) the conc( of the IP.reagent CIP and ionic $trength( i$ in,er$ely prop. otional to I 1:@

/etention a$ a f#nction of the charge of the IP.reagent and the analyte(

The $implified theory predict$ that the $lope of a ln *A ,$ ln cIP plot i$C for monocharged analyte$ and IP.reagent$ of oppo$ite charge( . C for monocharged analyte$ and IP.reagent$ of the $ame charge( 1 for dicharged analyte$ and monocharged IP reagent$ of oppo$ite charge(

Compari$on of theoretical and e+perimental re$#lt$ for the $lope of the ln *A ,$ ln cIP) monocharged IP reagent(
Slope 01:@D Antra2#inone$#lphonate and !aphtalene$#lphonate( Slope E 1:@D ;en4yltrib#tylammoni#mion and ;#pi,acaine(

Slope 01D 1)F.!aphtalene.di.$#lphonate(

E+perimental data from- J( Sthlberg and A( 7#rGngen) Chromatographia) @? %1=>H& H><(

When 4IP6 0@ and 4A6 .1 the theoretical $lope i$ 8(? and when 4A6 .@ it i$ 8(>(
E+perimental data from C( Petter$$on and 5( Schill) Chromatographia) @> %1=>=& ?<H(

$lope 6 8(> $lope68(?

Simplified theory) cont

A$ing appro+imation$ of the 5.C theory and $ome algebra the following e2#ation can be obtained-

. 4A94IP ln *cA 6 ln *8A 0 @ 4IP01

/etention factor of the analyte(

9ln%%n8BIPcIP:&0con$t& &0con$t

epend$ on the concentration of the organic modifier in the mobile pha$e

/etention a$ a f#nction of organic modifier concentration(

7rom re,er$ed pha$e chromatography of non. charged analyte$ it i$ *nown that the retention factor a$ f#nction of organic modifier concentration follow$ the following e2#ationln * 6 ln *0 E S9 where *0 i$ the retention factor at modifier concentration 4ero( S i$ a con$tant which i$ characteri$tic for the analyte and i$ the organic modifier concentration(

Simplified theory) cont(

The appro+imate e2#ation deri,ed from the $implified theory incl#de$ both electro$tatic effect$ and the effect of ,arying the organic modifier content of the mobile pha$e( The e2#ation can therefore be #$ed for predicti,e p#rpo$e$( E+ample- The combined effect of ,arying the IP reagent and the organic modifier concentration( Analyte- "ctopamine( IP reagent- "ctyl$#lfonate "rganic modifier- Methanol

Prediction of retention by the $implified theory(

Two e+perimental point$ at two different organic modifier concentration$ can be #$ed to predict a large n#mber of e+perimental re$#lt$(

18I Me"H

@FI Me"H

Prediction of retention by the $implified theory) cont(

At both modifier concentration$ the $lope of log * ,$ log cIP i$ 1:@(

Prediction of retention by the $implified theory) cont(

7rom the difference in the ordinate ,al#e$ can the ,al#e for S be calc#lated(

Prediction of retention by the $implified theory) cont(

It i$ now ea$y to predict the log * ,$ log cIP at a third organic modifier concentration) in thi$ ca$e ?8I Me"H(
E+perimental data from;artha et(al( J( Chromatogr( <8< %1=>?& @=(

Prediction of retention by the $implified theory) cont(

Analogo#$ly) from two e+perimental point$) the retention of !p.$#lphonate a$ a f#nction of the octyl$#lphonate concentration can be predicted for ,ario#$ organic modifier concentration$
E+perimental data from- ;artha et(al( J( Chromatogr( <8< %1=>?& @=(

Concl#$ion$- 7eat#re$ of the Electro$tatic theory for Ion Pair Chromatography

The theory and it$ parameter$ are phy$ically well defined It encompa$$e$ all po$$ible combination$ of charge of the IP.reagent and the analyte( There i$ good agreement between theory and e+periment$) A $implified form of the theory i$ $imple to #$e( The $implified form can be #$ed for prediction of capacity factor$ #nder many different e+perimental condition$(

Than*$ to

r( J*o$ ;artha for hi$ important contrib#tion$ d#ring the co#r$e of thi$ wor*(

Than*$ to

r( J*o$ ;artha for hi$ important contrib#tion$ d#ring the co#r$e of thi$ wor*(

To K"A for li$tening(