Ion Pair Chromatoghrapy

8/12/2019 Ion Pair Chromatoghrapy

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How Theory and Practice Meet inIon Pair Chromatography

Jan StåhlbergAcademy of Chromatography

SWEE!



"#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 $implified model ande+periment$(

● Concl#$ion$(



What i$ Ion Pair Chromatography'

● Ion Pair Chromatography %IPC& i$ #$ed to$eparate ionic analyte$ on a re,er$ed pha$ecol#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

+ 0 _

/etention time




!o IP.reagent

+ 0 _

/etention time

0 chargedIP.reagent

+ 0 _




!o IP.reagent

+ 0 _

/etention time

0 chargedIP.reagent

+ 0 _

+0 _ . charged

IP.reagent



A0

A0

A0A0

A0A0

A0

A0

A+

A0A0

SOLIDPH

ASE

A+A+

A+

A+A+

A+

A+A+

A+

A-A-

A-

A-

A-A-

A-

A-

A-

A-

Scheatic e!"lanation of the effect ofthe IP-reagent on retention

When noIP regent i$added thedi$trib#tion i$ 1-1 for

all analyte$



A0

A0

A0A0

A0A0

A0

A0

A+

A0A0

SOLIDPH

ASE

A+A+

A+

A+A+

A+

A+A+

A+

A-A-

A-

A-

A-A-

A-

A-

A-

A-

Scheatic e!"lanation of the effect ofthe IP-reagent on retention

The IP.reagentwill ad$orb tothe $tationarypha$e(

IP+ IP+



Scheatic e!"lanation of the effect ofthe IP-reagent on retention#

A0

A0

A0A0

A0A0

A0

A0

A+A0

A0

SOLIDPHAS

E

A+

A+

A+A+

A+

A+

A+ A+

A+

A-A-

A-

A-A-A-

A-

A-

A-

A-

IP+

IP+

IP+

IP+

IP+

IP+

IP+

IP+The chargeof the ad$orbed

IP.reagentinteract$electro$taticallywith the chargeof the analyte$



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 $hall2#antitati,ely de$cribe how retention i$ affectedby-

●

The 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#tedclo$e to acharged$#rface'



The Electrical o#ble 3ayer

+

+

++

+

++

+

+

+

+

+

+

+

+

+ _

_

_

_

_

_

_

_ _

_

_

_

_

_

_

∆Ψ 4

At e2#ilibri#m a

minim#m in freeenergy i$ achie,ed(

A difference)∆Ψ) in

electro$tatic potentialbetween the $#rfaceand the electrolytei$ created(



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

+

+

++

+

++

+

+

+

+

+

+

+

+

+ _

_

_

_

_

_

_

_ _

_

_

_

_

_

_

∆5tot6∆5chem+47∆ΨA4 A4

88

IP IP4 4∆5tot6∆5chem+47∆Ψ8 8



Implication$ of the electro$taticretention model(

● ∆Ψ depend$ only on the concentration of IP.reagent on the $#rface and i$ independent ofthe type(

● ∆5chem i$ more or le$$ independent of the typeand concentration of the IP.reagent(

● The retention factor i$ de$cribed by the

e2#ation- *6*89e+p%.47∆Ψ:/T&

where *8 i$ the retention factor in ab$ence of

the IP.reagent(



Implication$ of the electro$taticretention model(

● 7or a gi,en analyte doe$ retention depend$ onthe $#rface concentration of the IP.reagentonly( It i$ independent of the type of IP.reagent)

●

E+ample- /etention factor of adrenaline a$ af#nction of the $#rface concentration of differentanionic IP.reagent$(% ;artha et(al( J( Chromatogr() <8<) %1=>?& @=(



E+perimental datafrom ;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 pointcharge$(

● 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 ofthe electrolyte(



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

● ∆Ψ depend$ linearly on the $#rface concentration of IP.reagent and on the in,er$e of the ionic $trength of themobile pha$e(

● ;oth the IP.reagent and the analyte are f#lly ioni4ed(

● 7or a gi,en col#mn doe$ ∆5chem for both the IP.reagentand the analyte depend only on the mobile pha$ecompo$ition) i(e( not on ∆Ψ(

● The $implification$ are rea$onable when the ratio *cIP:*8 i$between @ . 18



Simplified theory) cont(

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



. 4A94IP

4IP01@

9ln%%n8

BIP

cIP

:κ &0con$tln *cA 6 ln *8A 0

. 4A94IP

4IP01@

9ln%%n8

BIP

cIP

:κ &0con$t&

Simplified theory) cont● $ing appro+imation$ of the 5.C theory and $ome

algebra the following e2#ation can be obtained-

Capacity factor at concentrationcIP

Capacity factor at concentration

cIP68

Electro$tatic contrib#tiona$ a f#nction analyte charge) 4A)IP reagent charge 4IP) it$ bindingcon$tant BIP) the conc( ofthe IP.reagent CIP and ionic$trength( κ i$ in,er$ely prop.otional to I

1:@



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

● The $implified theory predict$ that the $lope of aln *A ,$ ln cIP plot i$-

● for monocharged analyte$ and IP.reagent$ of

oppo$ite charge(● . for monocharged analyte$ and IP.reagent$

of the $ame charge(

●

1 for dicharged analyte$ and monocharged IPreagent$ of oppo$ite charge(



Slope 01:@D Antra2#inone$#lphonate and!aphtalene$#lphonate(

Slope 1:@D ;en4yltrib#tylammoni#mion and;#pi,acaine(

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

E+perimental data from- J( Ståhlberg andA( 7#rGngen) Chromatographia) @? %1=>&><(

Compari$on of theoretical and e+perimental re$#lt$ forthe $lope of the ln *A ,$ ln cIP) monocharged IP reagent(



When 4IP6 0@and 4A6 .1the theoretical

$lope i$ 8(?and when4A6 .@ it i$ 8(>(

E+perimental data from

C( Petter$$on and 5(Schill) Chromatographia)@> %1=>=& ?<(

$lope68(?

$lope 6 8(>



. 4A94IP

4IP01@ 9ln%%n8BIPcIP:

κ &0con$tln *cA 6 ln *8A 0

. 4A94IP

4IP01@ 9ln%%n8BIPcIP:

κ &0con$t&

Simplified theory) cont● $ing appro+imation$ of the 5.C theory and $ome

algebra the following e2#ation can be obtained-

/etention factor of the analyte( epend$ on the concentration of the organic

modifier in the mobile pha$e

/ t ti f ti f i



/etention a$ a f#nction of organicmodifier concentration(

7rom re,er$ed pha$e chromatography of non.charged analyte$ it i$ *nown that the retention

factor a$ f#nction of organic modifierconcentration follow$ the following e2#ation-

ln * 6 ln *0 S9φ

where *0 i$ the retention factor at modifierconcentration 4ero( S i$ a con$tant which i$

characteri$tic for the analyte and φ i$ the organicmodifier concentration(



Simplified theory) cont(

The appro+imate e2#ation deri,ed from the $implifiedtheory incl#de$ both electro$tatic effect$ and the effect of,arying the organic modifier content of the mobile pha$e(

The e2#ation can therefore be #$ed for predicti,ep#rpo$e$(

E+ample- The combined effect of ,arying the IP reagentand the organic modifier concentration(

Analyte- "ctopamine(IP reagent- "ctyl$#lfonate"rganic modifier- Methanol

P di ti f t ti b th



Prediction of retention by the$implified theory(

Two e+perimentalpoint$ at two

different organic

modifierconcentration$ canbe #$ed to predicta large n#mber of

e+perimentalre$#lt$(

@F Me"H

18 Me"H



Prediction of retention by the



Prediction of retention by the$implified theory) cont(

7rom the

difference in theordinate ,al#e$can the ,al#e for S

be calc#lated(





It i$ now ea$y topredict the log * ,$

log cIP at a thirdorganic modifierconcentration) inthi$ ca$e ?8

Me"H(

E+perimental data from-;artha et(al( J( Chromatogr(

<8< %1=>?& @=(





● Analogo#$ly) fromtwo e+perimentalpoint$) the retentionof !p.$#lphonate a$ af#nction of theoctyl$#lphonateconcentration can bepredicted for ,ario#$organic modifierconcentration$

● E+perimental data from- ;arthaet(al( J( Chromatogr( <8< %1=>?&

@=(

C l i 7 t f th El t t ti th



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

● The theory and it$ parameter$ are phy$icallywell 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 differente+perimental condition$(



Than*$ to

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



Than*$ to

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

● To K" for li$tening(

Ion Pair Chromatoghrapy

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