Ion Chromatography: Working Principle and Applications

Dr. C. K. Jain
Scientist ‘F’ & Head
Environmental Hydrology Division
National Institute of Hydrology
Roorkee – 247 667
Email: [email protected]

1. Introduction
Ion Chromatography is a relatively new method for analyzing anions, cations and
polar substances and is based on the equilibration of solute ions between the solvent,
acting as the Mobile Phase and charged ion-exchange sites on a solid surface acting as
the Stationary Phase. The technique is being used extensively in the modern chemical
analysis laboratories. The development of new detection methods and advances in
separation materials continues to expand the application of ion chromatography. It can be
used to reliably quantify substances throughout a wide range of concentrations. A whole
variety of ions can be analyzed in a single determination. Complete automation of ion
chromatography is easy and help save time and reduce costs. Due to its reliability and
robustness, ion chromatography is used in many different fields of application.

2. Structural Definitions
The Mobile Phase is usually an aqueous solution of salts of weak or strong
Bronstead acids and bases.

The Stationary Phase is usually a “bed” of either organic plastic beads or

inorganic particles. In both cases, the surfaces of the Solid Phase carry fixed charged
functional groups of weak or strong Bronstead acids or bases.

The organic plastic Stationary Phase, often called a “resin bed”, consists of
small, amorphous non-crystalline particles of styrene and divinylbenzene (1%-16%)
copolymers. This material can be used over a pH range 2-12, for small atomic or
molecular ions (M.W.<500) which can penetrate the small pores of the resin. The resin
can be made with “dissociable functional groups” fixed to the surface. These groups can
be designed to act as either Bronstead acids or bases and react with water to give either
negatively or positively charged groups respectively.

The inorganic particulate Stationary Phase usually consists of small,

amorphous non-crystalline particles of “silica gel” or “alumina”. Silica gel is chemically
acidic and acts as a cation exchange medium over a pH range pH 2-8. Alumina is
chemically basic and acts primarily as an anion exchange medium over the pH range 6 to
11. Alumino-silicate materials called “zeolites” can be used over almost the full pH range
and exchange cations or anions as needed.

101
3. Functional Definiitions

Retention
n is based onn the attracttion betweenn solute ionss and charged sites on thhe
statioonary phase. The Reten ntion Time is a function of Stationnary Phase retention.
r Thhe
stronger the reteention, of ann anion/catiion exchangger Stationarry Phase, thhe longer thhe
Retenntion Time on
o the IC collumn.

For ions of
o the same radius.
r Retention depends directly on o ion chargge. In generaal,
the greater
g the ion charge, the
t greater the attractioon for ion exchange
e sittes. Typicallly
retenttion increasees from monnovalent, to divalent
d to trrivalent ion charge.
c

For ions of equal chaarge, retention also depends directlyly on ion sizze. The largeer
the ioon radius, th
he more polaarizable the ion, and thee more stronngly it is atttracted to ioon
exchaange sites.

The Eluttion Order depends onn the Selecttivity of a StationaryS P

Phase. Stronng
versuus weak excchanger resins can prodduce quite different
d eluution orders.. The elutioon
orderr of halides is F-, Cl-, Br- and I-, whicch corresponnds to increasing size of these ions.

The Eluen nt Ionic Strrength is deffined by the eluent buffeer concentrattion at a fixeed
pH. Since
S bufferr ions replacce analyte ioons on the Stationary Phhase, the higgher the Ioniic
Strenngth, the shoorter their retention timmes. Since buffers
b are prepared
p witth polyvalennt
weakk acids, the ioonic strengthh varies withh pH accordiing to the buuffer pKA;
HPO42-(aq)) + H+(aq) ⇆ H2PO4-(aq))
divalent monovalentt

Thus the retention

r tim
me of analytee ions can bee changed wiith pH.

4. Applica
ations
The vario
ous importannt fields of appplication foor ion chrom
matography innclude:

1) The ro outine invesstigation of aqueous sysstems such as a drinking water, rivers,

efflueents and rain water
2) For thet analysiss of ions in chemicaal products,, foods, coosmetics annd
pharmmaceuticals
3) Ultra--trace analyssis such as inn the semi-coonductor andd power induustry.

Schemaatic of an Ion
n Chromatoography Sysstem

102
 The abov ve schematicc represents a non-supppressed ion chromatography system m.
The sample
s is inttroduced ontto the system
m via a sampple loop on the injector.. When in thhe
injectt position the sample is pumped
p withh the eluent onto the collumn and thee sample ionns
are thhen attracted to the charged statioonary phase of the coluumn. The chharged eluennt
elutess the retaineed ions which then go through thee detector (w which is most commonlly
conduuctivity) and d are depicteed as peaks on
o a chromattogram.

2. Three Main
M Modes of Ion Chromato
C ography Columns
The diffeerent modes of chromattography (annion exchannge, cation exchange
e annd
ion exclusion)
e simply
s relate to the diifferent typees of colum
mns used to achieve thhe
separration of thhe ions. Thee eluent moode of deteection - how wever unlesss stated thhe
follow
wing is all based on condductivity dettection.

2.1 Ion exch

hange
Ion exchaange chromaatography (IC C) is based on
o a stoichioometric chem
mical reactioon
betweeen ions in a solution and
a the oppoositely chargges groups functional groups
g on thhe
colum
mn resin. In n the simpleest case in cation chroomatographyy these are sulfonic aciid
groupps or carbox xylic acid grroups and inn anion chroomatographyy quarternaryy ammonium m
groupps.

2.2 Anion exchange

e
Anion exchange chroomatographyy forms the largest grouup of IC meethods mainlly
becauuse there arre few alteernatives with such sim mplicity, sennsitivity or selectivity -
particcularly for su
ulfate. Anion exchange can exist wiith or withouut suppression and of thhe
two, suppressed methods aree the most widely
w usedd. Eluents foor suppressedd chemistriees
tend to
t be either carbonate
c baased which give
g greater flexibility orr hydroxide based.

C
Chromatogr
ram Showin
ng an Anionn Exchange Separation Followed by
b Direct
Condu
uctivity Dettection (Non
n-Suppresseed)

It is sometimes a beenefit to worrk non-supppressed for example

e whhen analyzinng
weakk acids like cyanide
c or boorate that woould not be seen
s if a supppressor is ussed.

103
 Sup
ppression in
n Ion Chrom
matography

Often, a device
d calledd a suppressor is used annd is placed between thee column annd
detecctor as show wn above. When
W supprression is ussed the deteector is alm
most certainlly
conduuctivity. The chromatoggram below shows a saample with a suppressoor unit placeed
betweeen the colu umn and deteector. The grreatest achieevement of suppression
s is to increasse
the seensitivity off the anion, however
h at thhe same tim
me the backgrround conduuctivity of thhe
eluennt is greatly reduced. The same supppressor unnits can also be used too increase thhe
sensittivity of organic acids ussing a techniique known as inverse suuppression.

Chrromatogram
m Showing an
a Anion Exxchange Sepparation Foollowed by Suppression
S n
and then Con
nductivity Detection
D

The supprressor used in

i anion chroomatographyy is simply a cation exchhanger and itts
job is to removee cations annd replace thhem with ann H+. So a sodium carbbonate eluennt
(~8000uS) would be
b convertedd to carbonicc acid (~18uuS) by the suuppressor and the analytee,
for example
e NaCl (~126uS S without suuppression) would becoome HCl (~ ~426uS witth
supprression). Thhe suppressor convertss the analytte to the free f acid foorm and thhe
backgground is red
duced whilstt the sample signal is enhhanced.

The Mettrohm MSM M (Metrohm m Suppressor Modulee) contains 3 separatte

supprressor units. At any onee time, one will be in-liine with thee eluent andd conductivitty
detecctor, one will be in-line with
w dilute sulfuric
s acidd (replacing the
t removedd cations witth
H+) and
a the third d is washed with
w water. The
T benefits of this technnique are lacck of baselinne
noisee and a rugg
gedness that is reflected in the fact that
t the MSM M comes wiith a ten yeaar
warraanty and is not
n adverselyy affected byy the transittion metals or
o amines whhich can be a
probllem for otherr manufacturrers.

104
 Schem
matic of Metrrohm Suppressor Mod
dule

The anion n chemical suppressionn can be takken a stage further

f withh a secondarry
CO2 suppressor instrument
i (
(fitted after the MSM) which
w removes carbon dioxide from m
the suppressor reeaction and carbonate fromf the sample, whichh means thaat the mobille
phasee is convertted to wateer instead of carbonic acid so a background
b conductivitty
approoaching 1uS is then achhieved. This is known ass Sequentiall Suppressioon. Sequentiaal
supprression givees a greatly minimized injection peak, no sysstem peak (ffrom eluentt),
superrb linearity and
a an enhanncement in thet peak areeas allowing lower limitss of detectioon
to be achieved.

Gradient chemistry can be realized using thee MSM-HC (high capaccity) and usees
the saame cation exchanger
e as the MSM but in a largger form. Thhe MSM-HCC can be useed
in coombination with
w a low pressure
p or high
h pressurre pump system. Both thhe MSM annd
MSM M-HC have a compact deesign.

105
 Chro
omatogram of a Carbonate Gradieent Using th
he MSM-HC
C

2.3 Cation Exchange

E
There is a variety off cation coluums availablee; however the modern ones contaiin
carbooxylic acid functional
fu grroups. A largge number of applicationns for silica--gel based ioon
exchaangers existt. These collumns allow w simultaneoous separatiion of alkalli metals annd
alkaliine earths plus the sepaaration of soome transitioon metals. Small
S aminess can also be b
analyyzed using caation exchannge columns.

Chromatogra
am of Group I/II Metalls with Nitrric Acid/Dip
picolinic Aciid Eluents

The eluennts used forr non-suppreesed cationss exchange are week accids with thhe
compplexing agen nt such as dipicolinic
d a
acid, the conncentration of which can
c effect thhe
elutioon of calcium
m and heavyy metals succh as iron, zinc z and cobbalt. This caan be used to
t
great effect to chaange the seleectivity of thhe separationn.

Cations can
c become less sensitiive when suuppressed annd so are analyseda witth
directt conductiviity detectionn. It makes sense also because caation suppreessors can be b
notorriously unrelliable and addd to the coosts of runninng the instruument. The stability annd
efficiiency of thee Metrohm conductivity
c y detector cooupled with a very low w pump noisse
meann that Metro ohm can work
w non-supppressed eaasily with fast
fa instrumeent hardwarre
equilibration timee.

106
2.4 Ion excllusion
Ion excluusion chromaatography (IIEC) is mainly used forr the separaation of weaak
acids or bases. The
T greatest importance of IEC is foor the analyssis of weak acids such asa
carbooxylic acids, carbohydrattes, phenols or amino accids.

atogram Shoowing Ion Exclusion

Chroma E with an Mineeral Acid Elluent

3. Other Modes
M of Detection
D

3.1 Ampero
ometric Deetection
In princip
ple voltammmetric detectors can be used
u for all compoundss which havve
functtional groupss which are easily reducced or oxidizzed and is a very sensitive techniquee.
Apartt from a few w cations (FFe3+, CO2+) it
i is chiefly anions suchh as cyanidee, sulfide annd
nitritee which caan be deterrmined in the ion analysis sectoor. The moost importannt
applications lie however inn the analyssis of sugars by anion chromatogrraphy and in i
cliniccal analysis using a form
m of amperoometric detection know as Pulsed Amperometri
A ic
Detecction (PAD).

3.2 Photom
metric Deteection

Chromatogr
C ram Showin
ng Analysis of 1 PPM Chromate
C

Because of its extreemely wide range of application

a photometricc or UV/VIIS
detecction is the most impoortant detecttion methodd used in HPLC,
H as many
m organiic

107
molecules contain chromophhore groups, or can havee one introduuced or addeed, which arre
able to
t absorb in the UV or VIS V spectrum m. In the fieeld of inorgaanic ion anallysis UV/VIIS
detecction plays a smaller rolle. While off the simple anions onlyy analytes suuch as nitratee,
brommide or iodid de absorb, immportant anaalytes such asa fluoride, sulfate or phosphate
p caan
only be measureed indirectlyy. Many cattions do nott absorb at all, but muultivalent annd
transiitional metaals in particuular can be converted ini a post-column derivaatization witth
chelaating agents formers suuch as 4-(2-pyridylazo))-resorcinol (PAR) to form f coloreed
compplexes. Redo ox-active annalytes suchh as bromatee and other oxy-halide ions can be b
analyyzed by UV V/VIS detection after undergoingg a post-coolumn reacttion with an a
electrrochemicallyy active indicator.

4. Sample Preparatiion Techn

niques for Ion Chrom
matograph
hy
Quite ofteen with probblematic ionn chromatogrraphy appliccations, the matrix of thhe
sampple makes it difficult to accurately quantify
q the species of interest
i withh the standarrd
ion chromatogra
c aphy set-upp and somee form of sample preeparation thhen becomees
necesssary.

Scheematic Diagram of Stan

ndard Ion Chromatogr
C raphy Set-up
p

The samp ple preparatiion may be as

a straightfoorward as simmply dilutinng the samplle
with deionised water
w or caan involve injection
i off the samplee through a solid phasse
extraction cartrid
dge to remoove the interrference. In the case off more difficcult forms of
o
sampple matrices it may be necessary to t add addittional dediccated samplee preparatioon
moduules to the sttandard ion chromatogra
c aphy configuuration.

108
4.1 Dilution of the Sample
Dilution of the sample is performed when the concentration of the analytes of
interest either exceed the working capacity of the separation column chosen, or there are
sample matrix effects that can often be minimized by a dilution usually with water but
eluent can also be used.

4.2 Filtration of the Sample

It is recommended to filter all samples prior to injection with 0.45mm filters to
ensure that any particulate material from the samples don’t make their way onto the
injection valve or the analytical column where they can cause blockages and considerably
reduce the lifetime of the column(s).

4.3 Solid Phase Extraction Cartridges

Passage of the sample through one or more solid phase extraction cartridges prior
to injection will often retain selectively certain species within the homogeneous sample.
Quite often the retained species are substances that would interfere with the
chromatography had they not been previously removed. There are a number of different
cartridges whose suitability depends upon the type of chemistry undertaken.

For anion analysis, the sample can be treated with a cation exchanger in the H+
form that removes divalent cations that can mask any fast eluting anions. This type of
exchange cartridge removes carbonate/bicarbonate and is also useful for the removal of
cations from samples being determined by ion exclusion chromatography. Another option
is the use of a cation exchanger in the Ag+ form for the removal of any halides present in
the sample.

Similarly for cation analysis, one can employ an anion exchanger in the OH- form
to remove any interfering anions present in the sample. Another common type is the non-
polar exchange cartridge (reversed phase) that often utilizes C18 groups to remove
organic substances that would otherwise interfere with the chromatography.

4.4 Digestion of the Sample

If digestion techniques are to be employed then analyte content should be changed
as little as possible and any organic matter present should ideally be destroyed
completely. One can obtain analytical inaccuracies due to an exaggerated blank value as a
result of the chemicals used during the digestion. Different types of digestion include
wet, microwave and UV, the suitability of each depends on both the sample matrix and
the analytes of interest being determined.

4.5 Instrumental Sample Preparation Modules

109
 Often witth more com mplex samplle matrices, one has to add additionnal dedicateed
sampple preparatioon modules to the standard ion chroomatographyy configuration. There arre
a num mber of diffe
ferent instrum
ment optionss available within
w the Metrohm
M rangge dependinng
on thhe type of saample treatm
ment requiredd prior to annalysis. Metrrohm has acctually been a
pioneeer of inline sample prepparation moddules with thhe release off the 754 IC Dialysis Unnit
as long ago as 1997,
1 since then the technology haas been opttimized and considerablly
improoved so thaat today thee Metrohm IC portfollio contains many diffferent samplle
prepaaration instrruments to automate annd improvee analysis tiimes and thhroughput of o
difficcult sample matrices
m suchh as emulsioons or diary products.
p

4.6 Dialysiss
Dialysis is
i based on the selectivee diffusion ofo moleculess or ions froom one liquiid
(donoor or samplee solution) to
t another (aacceptor sollution) via a membrane. The drivinng
force for the traansfer is thee concentratiion gradientt across the membrane.. Contrary to t
dynam mic dialysis, where twoo solutions continuously pass througgh the dialyssis module, at a
least one solution
n is temporarrily stopped until the conncentration in
i the accepttor solution is
i
the same
s as thaat in the doonor solutionn. This pateented stopped-flow proocedure takees
betweeen 10 and 14
1 minutes anda can be diirectly couplled to an IC setup.

As the dialysis
d is performed
p d
during the recording
r off the previoous sampless’
chrommatogram, th he overall analysis
a timee is not prollonged. Wheereas in the conventionaal
setupp 2 two channnel peristaltic pumps trransport the sample andd the acceptoor solution to t
and from
f i compact dialysis Dosinos (accurrate liquid pipette)
the diaalysis cell, in p dosees
ultra pure water through
t the acceptor commpartment of
o the cell. The
T stopped-flow status is i
achieeved by stoppping the Dossino and bloocking the ouutlet capillarry of the celll by feeding it

110
throuugh the valvee of the sampple processoor. The reporrted dialysis recovery rattes have beeen
foundd to be in excess of 98%% using the paatented stoppped flow meethod.

4.7 Ultra-Fiiltration
The aim of samplee filtration is to prottect the seeparation coolumns from m
contaamination an nd blockagee from particculates that may be preesent in the sample. Thhe
ultra--filtration kiit in combinnation with the 858 Advanced
A Saample Proceessor ensurees
autommatic inline filtration
f witth sample inj
njection. It iss eminently suitable
s for those
t samplees
with a light to meedium load such
s as surfaace waters annd digestion solutions.

The sam mples are pllaced onto the samplee carousel before beinng processeed
matically. Saample filtrattion and inttroduction too the injectiion valve is achieved by
autom b
meanns of an inteegrated doubble channel peristaltic pump
p meanning that it is
i possible to
t
aspiraate slightly viscous
v sampples.

The samp ple is conveyyed by one channel of the pump thhrough the ultra-filtratio
u on
cell passing
p the membrane.
m A the same time the filttrate is aspirrated off froom the rear of
At o
this membrane
m and
a transferrred to the saample loop by b the seconnd channel of the pumpp.
Only a small fraaction of thee sample is removed ass filtrate so the contamiinants remaiin
mainlly in the sample
s streaam preventiing the reggenerated ceellulose mem mbrane fromm
becomming blockeed too quicklly.

5. Metrohm Samplee Preparattion Modu

ule
The 850 Professionaal IC Anioon – MCS – Prep 3 is the proffessional ioon
chrommatography system for the determ mination of anions withh simultaneoous Metrohm m
inlinee neutralizattion or Mettrohm inline cation rem moval for analysis
a of for examplle
alkaliine solutionss. In the botttom segmennt of the 850 Professionnal IC system
m there is an
a
additional anion sample
s preparation moddule «MSM XL»X and a bidirectional
b dual channeel
peristtaltic pump. Both these components
c are used forr Metrohm Inline Samplle Preparatioon
(MISSP).

111
 The modu ules consistss of a reactoor block that houses the cation exchaangers with a
contrrol unit that contains a twwo channel peristaltic puump that coonveys the reegenerant annd
rinse solutions.

The matrix elimination occurs innline whilst the regenerration and rinsing
r of thhe
packeed bed supp pressor insidde the <<MS SM XL>> occuro simultaaneously offfline. A fressh
supprressor channnel is used foor each new
w analysis annd because thhe rinse andd regeneratioon
occurrs after each determination, the capaacity is practiically unlimited.

The samp ple solution is

i transferredd to the sam
mple preparattion module from an autto
samppler via a looop injectionn and rinsedd with deionnised water. The samplle cations arre
exchaanged again nst protons (H+). If soddium hydrooxide constittutes the saample matrixx,
waterr is formed d by neutraalization. Thhe sample solution
s theen passes onto
o the pree-
conceentration coolumn wheree the trace anions to be b determineed are retainned and theen
elutedd by the eluuent flowingg in a counnter flow dirrection. The analyte aniions are theen
separrated on the analytical coolumn beforre quantificattion using chhemical supppression witth
conduuctivity deteection.

Ch
hromatogrram Show
wing Matriix Eliminaation on a
Sample of
o Causticc Soda

-----

112