Ch12 - Ion Chromatography

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199

Chapter 12

Ion Chromatography

John Statler

Dionex Corporation

Summary

General Uses

Separation and detection of ions and ionizable species

Profiling, that is, determining the qualitative distribution of mixtures of oligomeric ions Simultaneous determination of several ions in mixtures

Concentration of ionic species in samples of low concentration, often with simultaneous elimi-

nation of interfering matrix

Common Applications

Determination of inorganic anions or cations, organic acids, amines, amino acids, carbohy-

drates, or nucleic acids in a variety of samples

Monitoring water quality

Determination of the composition of industrial wastes

Monitoring the quality of intermediates in industrial processes

Determination of ionic composition of biological solutions

Separation of components of mixtures before mass spectrometry or other spectroscopic tech-

niques

Identification of ionic impurities

Purification of components from mixtures


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200 Handbook of Instrumental Techniques for Analytical Chemistry

Samples

StateLiquids that are aqueous or water-miscible solvents can be analyzed directly. Water-immiscible liquids,

solids, and gases must be extracted into or dissolved in aqueous solution before analysis.

Amount

Samples are usually introduced as 5- to 200-L volumes, although volumes as large as 100 mL can be

introduced using chromatographic preconcentration techniques when additional sensitivity is needed.

Preparation

Dilution or dissolution and filtration are the most common sample preparation procedures. Extraction

may be required for nonaqueous samples or preconcentration for dilute samples. The need for precol-

umn derivatization is rare.

Analysis Time

Excluding sample preparation, analysis times range from less than 3 min to more than 2 hr. Most com-

monly, however, analysis time is 10 to 15 min.

Limitations

General

Analyses are performed sequentially.

Analytes can be misidentified or their quantities incorrectly determined if other components are

not well separated.

The analysis consumes eluent, which must be replenished regularly.

Accuracy

For routine analysis, accuracy is about 3%, but under carefully controlled conditions and with the use

of an internal standard, relative standard deviation less than 1% is possible.

Sensitivity and Detection Limits

For a standard sample size of 25 L, detection limits are about 1 to 5 g/L (ppb) for most common in-

organic ions. However, this can vary a great deal depending on the detector response of the analytes,

the nature of the separation method, and interfering components in the sample.


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Ion Chromatography 201

Complementary or Related Techniques

Atomic absorption, atomic emission, and inductively coupled plasma spectroscopies, used for

determining the total amount of a metal rather than the amount of a certain ionic form of thatmetal

Mass spectrometry, used to obtain information on chemical structure and molecular mass

Nuclear magnetic resonance, used to obtain information on chemical structure

Infrared spectroscopy, used to obtain information on chemical structure, particularly before ion

chromatography to identify functional groups that can offer a key to separation or detection

Capillary electrophoresis, used as a confirmatory technique because it relies on an unrelated

separation mechanism

Introduction

Definition

In his comprehensive book on the subject, Small defines ion chromatography (IC) as the chromato-

graphic separation and measurement of ionic species (1). Common applications of ion chromatogra-

phy include the determination of simple anions, such as chloride and sulfate, simple cations, such as

sodium and calcium, transition metals, lanthanide and actinide metals, organic acids, amines, amino ac-

ids, and carbohydrates.

Early Developments

Modern ion chromatography began with a report by Small, Stevens, and Bauman (2) wherein they

described a way to combine an ion exchange chromatographic separation with simultaneous conduc-

tometric detection for the determination of anions including chloride, sulfate, nitrate, and phosphate,

or of cations including sodium, ammonium, potassium, and calcium. The key element was their de-

velopment of a device, later known as a suppressor, to lower the background conductometric signal

resulting from the liquid mobile phase, or eluent, while enhancing the conductometric signal from

the analyte ions. Originally, this pairing of ion exchange chromatography with suppressed conduc-

tometric detection was synonymous with IC. Eventually, however, the term expanded to include oth-

er detection methods and other chromatographic modes.

The first of the alternative detection techniques, nonsuppressed conductometric detection, emerged

in the late 1970s (3). Other routine techniques, including amperometry, optical absorbance, and fluo-rescence (4), as well as less common techniques such as inductively coupled plasma spectroscopy and

mass spectroscopy, have been used in the years since then. Likewise, ion exclusion, ion pairing, and

chelation chromatographies have been used in addition to ion exchange for the separation of ions (5).

Nevertheless, even with all these methods of separation and detection falling within the modern

definition of ion chromatography, IC as it is most often practiced today is still largely an ion exchange

separation with suppressed conductometric detection.


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Improvements in Separation

Synthetic ion exchange resins have been available for many decades, but the first resins used in modern

IC applications were surface-functionalized styrene divinylbenzene, in many ways very much likethose in common use today. Common anions were separated in about 30 min. The alkali metal cations

could be separated in about 25 min, but the alkaline earth cations required a separate analysis. These

analyses, especially for the cations, seem slow by todays standards, but were vastly superior to the la-

borious, single ion, wet chemical options. On todays high-efficiency ion exchange resins, common an-

ions can be resolved in less than 3 min, and the common group I and II cations can be determined

together in about 10 min.

Much of this improvement in speed has been a result of improvements in resin selectivity and unifor-

mity. Most resins used for IC have a thin surface region of ion exchange sites, minimizing band broaden-

ing caused by diffusion of the ions into the resin interior. In some cases, analysis speed can also be

increased with the use of gradient ion chromatography (6). This technique allows increasing eluent con-

centration to separate very weakly and very strongly retained ions in a short period of time.

Selectivity

The ideal ion exchange site is a permanent charge (that is, one whose charge does not change with pH)

that does not have secondary (nonion-exchange) interactions.

Since the beginning of IC, the most common functional group for anion exchange has been some

sort of quaternary alkyl ammonium group. This group has a permanent positive charge, regardless of

the pH of the surrounding eluent. The alkyl groups on the ammonium and the composition of the sur-

rounding resin can be varied to create different anion exchange environments. This allows the manu-

facturing of columns to produce resins with different selectivities for specialized applications.

Early cation exchangers usually had sulfonic acid groups as the cation exchange sites. The princi-

pal advantage of the sulfonic acid is that even at low pH it remains charged. However, these columns

are most often used with column switching techniques or with eluents containing both monovalent anddivalent ions to elute the monovalent and divalent analyte ions at similar times. In recent years, carbox-

ylic acids have become popular because the affinities of mono- and divalent analyte cations for the

functional group are more similar, so simple, unchanging isocratic eluents are capable of separating all

cations in similar times.

Improvements in Detection

Conductivity, the most common detection method in IC, has advanced by reducing noise through im-

provements in electronics and better isolation and control of cell temperature, by improved response

through detector cell design, and, in the case of suppressed conductometric detection, by improved sup-

pressor design to decrease internal volume and dispersion. All of these have helped lower detection lim-

its for many common ions to about 0.1 ng, or less than 10 g/L (ppb) for a 25-L injection.Electrochemical or amperometric detection as it was first used in IC was single-potential or DC

amperometry, useful for certain electrochemically active ions such as cyanide, sulfite, and iodide. But

the development of pulsed amperometric detection (PAD) for analytes that fouled electrode surfaces

when detected eventually helped create a new category of IC for the determination of carbohydrates.

Another advancement, known as integrated amperometry, has increased the sensitivity for other elec-

trochemically active species, such as amines and many compounds containing reduced sulfur groups,

that are sometimes weakly detected by PAD (7).


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Current Use

Types of SamplesIC is most often applied to aqueous samples or to solid samples that will dissolve in or can be extracted

into aqueous solutions. For aqueous samples, sample preparation may be unnecessary or may consist

of dilution or filtration before introducing the sample into the IC system. Insoluble solid samples, such

as soil or air filters, are usually extracted into an aqueous solution for analysis. Analysis of gaseous sam-

ples presents some interesting challenges in sample preparation (8) and is currently an active area of

research. Nonaqueous liquid samples, such as organic solvents, require either little sample preparation

if the solvent is miscible with water, or a liquidliquid extraction to an aqueous solution if the solvent

is immiscible.

Types of Analytes

What sets IC apart from most other techniques for the determination of ions is the ease with which it

can determine several ionic analytes simultaneously. In principle, any species that can exist as an ioncan be determined by IC. In addition to the common anions, such as chloride, bromide, sulfate, nitrate,

and phosphate, and common cations, such as lithium, ammonium, magnesium, and calcium, a multi-

tude of weak acids and bases can be ionized by adjusting pH. Amines are cationic below a pH of about

9. Carboxylic acids are anionic above a pH of about 3. Amino acids may be anionic at high pH or cat-

ionic at low pH. Sugars and similar carbohydrates, though not commonly considered ions, are actually

very weak acids and can be chromatographed as anions above a pH of 12 or 13. Transition and lan-

thanide metals, though often considered cations, readily form complexes with chelating anions and are

often chromatographed as anionic complexes.

How It Works

IC is the merging of a chromatographic technique for separating ions with a technique for detecting ions

and determining concentrations. Although the separation and detection are closely linked in practice,

the two processes can be conceptualized independently. Anion exchange and cation exchange are by

far the two most common separation techniques, but the alternative chromatographic methods of ion

exclusion, ion pairing, and chelation have some advantages in certain cases. Ions can be detected and

measured using several methods depending on the sensitivity and specificity needed. Species that are

ionic at or near neutrality can usually be detected using conductivity, the most common form of detec-

tion in IC. In many cases, however, amperometric, optical, ICP, or mass spectrometric methods may be

preferred.

Separation: Ion Exchange

Ion exchange is a process in which a charged analyte (also called a solute or eluite) in a flowing solution

competes with an eluent (mobile phase) ion of like charge for sites having the opposite charge on a sta-

tionary phase (Fig. 12.1). The sites of opposite charge are often called functional groups. Before intro-

ducing analyte ions into the systemthat is, before injectionthe functional group ions are paired with


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the eluent ions, maintaining electrical neutrality in the stationary phase. When a sample is injected, new

ions compete with the eluent ions at the functional group site. Analyte ions that compete successfully

(that is, those that have a high affinity for the ion-exchange site) are retained longer than ions that do not

compete well with the eluent ions. As with all chromatographic processes, ion exchange can be thoughtof as a process involving the rapid movement of the analyte between two phases, in this case a liquid

mobile phase and a solid stationary phase. In the stationary phase, the ions are immobilized. Ions travel

through the column only in the mobile phase. The more time an ion spends in the stationary phase, the

more slowly it moves through the column.

The key to the separation of analyte ions is the differential affinities a functional group has for dif-

ferent analyte ions. For example, if analyte A has a higher affinity for the stationary phase site than does

analyte B, A will compete with the eluent ions more successfully for those sites and be retained longer.

A will therefore elute from the column after B. The relative affinities of analytes for the stationary phase

are known as the selectivity. Selectivity is determined by many parameters of the separation, including

type of functional group, stationary phase environment near the functional group, characteristics of eluent

ions, eluent ion concentration, nonionic or oppositely charged eluent additives such as solvents or ion-

pairing agents, and temperature. The first two parameters are determined by the design of the ion ex-

change column and are usually optimized for a given class of analytes, such as common inorganic anions,organic acids, or inorganic cations. The other parameters can be adjusted by the analyst to tailor the sep-

Figure 12.1 Representation of the ion exchange process for an anion with hydroxide eluent.


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aration to specific requirements, but usually ion-exchange columns are designed with a specific set of

chromatographic conditions in mind.

The quantity of functional group sites in the stationary phase is known as the capacity. Capacity is

usually expressed as the number of equivalents per column or equivalents per gram of resin. A highercapacity results in longer retention of the analyte ions. Capacity is independent of selectivity (that is,

capacity can be increased or decreased without altering selectivity) and is determined by the resin man-

ufacturer.

Separation: Ion Exclusion

Ion exclusion, in many respects, is complementary to ion exchange. Like ion exchange, the stationary

phase is an ion exchange resin, although it has a very high ion exchange capacity and is the same charge

as the analyte ions. Its greatest utility is the separation of weakly ionized species while eluting strongly

ionized species in the void volume.

The ion exclusion process (Fig. 12.2) relies on the establishment of an electrical potential between

a dilute mobile phase and a stationary phase of high ion-exchange-site concentration. The relatively

high concentration of ion exchange sites in the resin dictates a high concentration of counterions in the

stationary phase to maintain electrical neutrality. However, diffusion forces tend toward equalizing

counterion concentrations in the mobile and stationary phases, a situation that leaves the stationary

phase charged the same as the analyte ions. This situation of high potential energy is often called the

Donnan potential. The Donnan potential permits neutral molecules to enter the stationary phase, where-

as analyte ions are repelled or excluded, hence the term ion exclusion. Weakly ionized species exhibit

Figure 12.2 Representation of the ion exclusion process for a carboxylic acid.


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intermediate behavior and are separated from one another based largely on their extent of ionization.

The most common application of ion exclusion is for the separation of organic acids using a sulfonated

macroporous cation exchange resin in the hydronium ion form. The separation of weak bases using a

macroporous anion exchange resin is possible, but is much less common.

Separation: Ion Pairing

Ion-pair chromatography, also known as ion-interaction or dynamic ion-exchange chromatography, is

a technique using a neutral hydrophobic stationary phase and a mobile phase containing a hydrophobic

ion, sometimes called the ion-pairing agent, having a charge opposite to that of the analyte. A common

explanation of the ion-pair mechanism is that the ion-pairing agent is associated with the stationary

phase and functions as an ion-exchange functional group. For anionic analytes, quaternary ammonium

ions, such as tetrabutylammonium, are most commonly used as ion-pairing agents. Alkylsulfonates,

such as octanesulfonate, are most commonly used for cationic analytes. The most common stationary

phases are alkyl-bonded porous silica resins, commonly used in reversed phase HPLC, and

macroporous styrenedivinylbenzene polymers.

Ion-pair chromatography, unlike ion-exchange chromatography, has some flexibility with respect to

selectivity and capacity. Selectivity and capacity can be altered by simply changing the ion-pairing agent,

and capacity can be increased by increasing the ion-pairing agents concentration. Historically, ion-pair

chromatography has also had the advantage of compatibility with certain organic solvent modifiers, such

as methanol or acetonitrile, added to the mobile phase to alter selectivity, but with the availability of sol-

vent-compatible ion exchange resins today, organic modifiers can be used to alter selectivity in ion ex-

change as well. The presence of ion-pairing agents can complicate the use of conductometric and

amperometric detections; nevertheless, both detection techniques are commonly used with ion-pair chro-

matography. The combination of ion-pair chromatography and suppressed conductivity detection is often

called mobile phase ion chromatography (MPIC).

Analytes most suited to ion-pair chromatography are large, hydrophobic ions because the slow

mass transfer or secondary hydrophobic interactions that these types of ions exhibit with typical ion-exchange stationary phases leads to poor efficiency, and often poor resolution.

Separation: Chelation

Certain organic groups, such as dicarboxylates, tend to form complexes with metal ions, but resins with

such chelating functional groups have not found wide use as stationary phases. Although these resins

have a high selectivity for certain metal ions, notably transition and lanthanide metals, the exchange

process is too slow for efficient chromatographic separations. Instead, metals are usually chromato-

graphed by anion exchange as anionic complexes of pyridine dicarboxylic acid or similar anionic

chelating agents.

Chelating resins are often used for metal concentration or matrix elimination. Many spectroscopic

techniques experience interferences from main group metals, such as sodium and calcium, which canbe present at concentrations that are several orders of magnitude higher than those of the metal analytes.

IC, on the other hand, has the selectivity for interference-free determination of many transition or lan-

thanide metals.


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Detection: Suppressed Conductivity

In the mid-1970s, Small and coworkers (2) recognized that the very nature of ion exchange chromatog-

raphy required an eluent ion to exchange with the analyte ion. Conductivity, a property shared by allions, would be the ideal choice as a universal detection method, but the background conductance of the

eluent would reduce the sensitivity of the technique. They solved this problem by using a second col-

umn, a suppressor column, having the opposite functionality as the analytical column and placing it be-

tween the analytical column and the detector. For determining anions, the suppressor column would be

a cation exchange column in the hydronium ion form and for determining cations the suppressor would

be an anion exchange column in the hydroxide ion form. When the salt of a weak acid or base is used

as the eluent, the suppressor reduces the conductometric signal from the eluent.

As an example, consider what would happen during suppression of an eluent containing sodium

bicarbonate and sodium carbonate, a common eluent in IC (Fig. 12.3). The carbonate and bicarbonate

anions exchange with the analyte anions (such as chloride) during the separation. The effluent then

passes through a cation exchange column in the hydronium ion form. Sodium ion is exchanged for hy-

dronium, forming carbonic acid, a weak acid having a very low conductometric signal. An analogous

reaction occurs in the case of cations. Another benefit of the suppression reaction is that the response

for the analyte ions is often increased. According to Kohlrauschs law, the measured conductivity of an

ionic compound is the sum of the equivalent conductances of the anion and cation. The equivalent con-

ductance of hydronium is the highest of all cations, so the measured conductance of each anion increas-

es after suppression because the hydronium counterion contributes more to the total conductance.

Similarly, hydroxide ion has the highest equivalent conductance of all anions, so the total conductance

for cations increases after suppression. The principal disadvantages to this approach are that the sup-

pressor column causes some dispersion of the analytes before detection and the suppressor must be dis-

carded or regenerated as the hydronium ions are depleted and the column can no longer suppress the

eluent.

The suppressor device has changed through its evolution (9). Today it is commercially available in

three versions: a packed column similar to the original one used about 20 years ago, a slurry of suspend-

ed ion exchange resin that is added to the effluent after the analytical column to accomplish the sup-pression reaction, and an ion-exchange membrane device that continuously supplies the suppressing

ion from a chemical source or from the electrolysis of the water in recycled, suppressed eluent. The

packed column suppressor is based on long-tested technology, but it must be regenerated or replaced

periodically as it becomes exhausted. The postcolumn reagent approach has the lowest startup costs,

but the suppressing resin is a consumable, discarded with the column effluent. The membrane devices

have low dead volume, so they lose little efficiency during suppression, and the electrolytic version of

the device has no consumable costs and almost no maintenance, but these types of suppressors have

higher startup costs.

Analytes detectable by suppressed conductance are those that, when suppressed, are ionic. Gen-

erally, this means that anions with pKas below about 5 and cations with pKbs below about 5 can usu-

ally be detected by this method. In other words, if the analyte is ionic at pH 7, the approximate pH

of most eluents after suppression, suppressed conductance is a viable detection method.

Detection: Nonsuppressed Conductivity

Nonsuppressed IC, sometimes called single-column IC, operates with a higher background conductance.

The higher background necessitates careful control of temperature, so conductivity detectors (Chap. 39)

designed for nonsuppressed systems generally have thermostated cells and sometimes insulated chambers

for the analytical column and the detector cell. Analytical columns designed for nonsuppressed IC are


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methods have the advantage that weak acid anions and weak base cations can be detected more readily.

For example, a cation with a pKb of 9 would be only about 1% ionic after suppression (pH of 7), but in

1 mM HCl (pH of 3) it would be about 99% ionic.

Detection: Single-Potential Amperometry

Any analyte that can be oxidized or reduced is a candidate for amperometric detection (Chap. 36). The

simplest form of amperometric detection is single-potential, or direct current (DC), amperometry. A

voltage (potential) is applied between two electrodes positioned in the column effluent. The measured

current changes as an electroactive analyte is oxidized at the anode or reduced at the cathode. Single-

potential amperometry has been used to detect weak acid anions, such as cyanide and sulfide, which

are problematic by conductometric methods. Another, possibly more important advantage of amperom-

etry over other detection methods for these and other ions, such as iodide, sulfite, and hydrazine, is

specificity. The applied potential can be adjusted to maximize the response for the analyte of interest

while minimizing the response for interfering analytes.

Detection: Pulsed Amperometry

An extension of single-potential amperometry, (Chap. 36) is pulsed amperometry, most commonly

used for analytes that tend to foul electrodes. Analytes that foul electrodes reduce the signal with each

analysis and necessitate frequent cleaning of the electrode. In pulsed amperometric detection (PAD), a

working potential is applied for a short time (usually a few hundred milliseconds), followed by higher

or lower potentials that are used for cleaning the electrode. The current is measured only while the

working potential is applied, then sequential current measurements are processed by the detector to pro-

duce a smooth output. PAD is most often used for detection of carbohydrates after an anion exchange

separation, but further developments of related techniques (7) show promise for amines, reduced sulfur

species, and other electroactive compounds.

Detection: Optical

The most commonly used optical detectors are absorbance detectors and fluorescence detectors (Chap.

25 and 26). Fluorescence detectors are rarely used in IC, but absorbance detectors are quite common.

Absorbance is used for detection under three circumstances, direct photometric detection, indirect pho-

tometric detection, and photometric detection after a postcolumn derivatization.

Some ions, most importantly certain anions, are chromophoric; that is they absorb light. Such is

the case with the common anions nitrite and nitrate. These can be detected in the ultraviolet region, by

monitoring absorbance at 215 nm, without detecting nonchromophoric anions such as chloride. Absor-

bance can also be used for detection of many organic ions, such as aryl amines and organic acids, fol-

lowing ion exchange chromatography.More commonly, however, ions are not chromophoric. Nevertheless, they can be detected indirect-

ly using a chromophoric eluent ion of the same charge, a technique known as indirect photometric de-

tection. During the ion-exchange process, ionic concentration remains constant at the stationary phase

functional groups, so at the detector the presence of each equivalent of analyte requires the absence of

an equivalent of chromophoric eluent ion. The use of indirect photometric detection is not very com-

mon today.

By far, the most common use of absorbance detection in IC is with an ion-exchange separation fol-


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lowed by a derivatization reaction that renders the analytes chromophoric or, in some cases, more chro-

mophoric. Ideally, the postcolumn reaction is fast (complete within seconds) and does not produce

interferences (for example, by reaction with eluent components) in the absence of analytes. This tech-

nique is commonly used for the determination of transition metals and amino acids. As was mentionedearlier, transition metals are often chromatographed as anion complexes. By adding 4-(2-pyridylazo)re-

sorcinol (PAR) to the column effluent, PAR complexes of the metals rapidly form and are detected by

absorbance. Similarly, amino acids can be chromatographed by either anion or cation exchange, deriva-

tized after separation with ninhydrin and detected by absorbance or, if lower detection limits are de-

sired, derivatization with ortho-phthalaldehyde allows fluorescence detection.

Detection: Specialized Detectors

Recently, the scientific community has begun coupling ion-exchange separations with specialized tech-

niques for detecting the analytes. The two most common are inductively coupled plasma (ICP) spec-

troscopy, useful for the determination of metals, and mass spectroscopy (MS).

Coupled to ICP, IC (Chap. 21 and 22) is more a sample preparation step than a chromatographic

separation. A column containing a chelating stationary phase is used to selectively concentrate metals

from a sample matrix. An intermediate column wash step can selectively remove interfering metals if

necessary. Then the column is washed with a strong eluent, delivering the metals of interest to the ICP.

MS (Chap. 33) is not generally compatible with the highly ionic eluents commonly used in IC.

However, just as with conductivity detection, eluent ions can be eliminated from the effluent by the use

of a suppressor before it enters the MS (10). In these cases, the goal of IC-MS is not the quantification

of the analytes, but rather their mass spectral identification and characterization.

What It Does

Instrumentation

Minimally, an IC system consists of an eluent reservoir, an analytical pump to deliver the eluent to the

analytical column, an injection valve or other means of introducing the sample, an analytical chromato-

graphic column, a detector, and a data processing device (Fig. 12.4). These components are the same

components in an HPLC system. Here, however, we discuss each as it relates to IC.

The eluent reservoir can be as simple as a bottle with a fluid line that leads to the analytical pump.

Usually, however, the eluent is kept under a pressure of about 1/3 atm so that eluent is delivered to the

analytical pump without fluid interruption. Eluents at high pH, such as sodium hydroxide, must be kept

under an inert atmosphere to prevent carbon dioxide in the air from forming carbonate in the eluent and

thereby altering the eluting ion.The pump used in IC is usually nonmetallic. This is because many IC methods use strong acids,

strong bases, or high concentrations of salts that may corrode metallic systems. Materials such as poly-

etheretherketone (PEEK) are compatible with the vast majority of IC applications. Metallic pumps may

be used if care is taken to maintain them properly. Many manufacturers of metallic pumps for IC rec-

ommend regular passivation of the system.

High-sensitivity applications usually require a dual-piston pump for relatively pulse-free opera-

tion, although single-piston pumps with pulse dampers are usually sufficient for routine, mg/L-level


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analysis. The most flexible systems have dual-piston gradient pumps, which allow the analyst to per-

form gradient methods or to mix eluents isocratically, aiding method development.

The injection valve must introduce a reproducible sample volume into the IC system. The mostcommon means of accomplishing this is by the use of a fixed-loop injection. A length of tubing of

known volume is attached to the valve and switched into the eluent stream during injection.

Many injection systems include an autosampler for unattended, high-volume work. The autosam-

pler may have an injection valve built in, or it may deliver the sample to a remote injection valve. The

former type is usually chosen if sample sizes are small because having an injection valve in the au-

tosampler shortens the distance that the sample travels between sample vial and injection valve.

Certain low-level IC methods use a concentration column in place of the injection loop on the in-

jection valve. This concentration column may be a guard column of the type used for the analytical sep-

aration or it may be a column specifically designed for concentration in a particular application. If the

concentration column has a high backpressure, an auxiliary pump is usually required to deliver the sam-

ple to that column.

The chromatographic separation takes place on the analytical column. Usually, a guard column is

placed before the analytical column to extend its lifetime. Most commonly, the guard column is simplya short, inexpensive version of the analytical column that is replaced as needed. Typically, the guard

column also adds about 20% to the capacity of the analytical system.

The detector ordinarily is one of the options discussed previously, namely conductometric, amper-

ometric, optical, or some other method of measuring ions, along with any system used to facilitate de-

tection. These systems include suppressors and postcolumn reaction devices.

The simplest data processing device is a chart recorder that converts a detector output, usually a

voltage within a predetermined range, into a timevoltage profile, called a chromatogram. However,

Figure 12.4 The ion chromatographic system.


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this device relies on the analyst to convert the area or height of a peak drawn on the paper to a quantity

of analyte, a labor-intensive step.

A recording integrator does much the same thing as a chart recorder, but calculates the peak area

automatically and may even calculate analyte quantities based on standards that were injected earlier.Some integrators can also send signals to the instruments, thereby controlling simple functions such as

injections and detector range changes.

Data processing through a computer offers the greatest flexibility and automation. As with the in-

tegrator, analyte quantities can be determined automatically, but the raw data can also be stored and

reprocessed using different parameters at a later time. Computer-based systems usually have complete

instrument control as well, so that complete operating conditions can be stored on computer and used

later to reproduce the analytical method.

Analytical Information

Qualitative

Like other forms of liquid chromatography, IC can indicate sample composition. The components

present elute at nearly unique retention times, determined either by separate injections of known stan-

dards of each ion, or more accurately by adding a small amount of a known standard to the sample and

identifying the component that experiences an increase in peak size. An estimate of relative concentra-

tions can also be made, but accurate concentrations can be determined only after calibrating the detector

response.

One can also obtain information about the distribution of components that differ in the number of

repeating units. Examples are samples containing polyphosphates, such as polyphosphoric acid (Figure

12.5), linear oligosaccharides, such as hydrolyzed starch, oligonucleotides, and ionic surfactants, suchas linear alkyl benzenesulfonic acid. Although each component may be completely resolved from other

components, individual pure standards of each component are difficult if not impossible to obtain, so

estimates of concentrations are only qualitative.

Quantitative

IC is first and foremost a quantitative technique. The purity of standards determines the accuracy of the

technique, and because pure standards of most ions, usually in the form of salts, typically are easy to

obtain, accuracy can be 1 to 2%. Analyte concentrations are determined by establishing a relationship

between known concentrations of standards and their responses, in terms of either peak height or peak

area. This is commonly known as the calibration curve. Such a calibration using independently chro-

matographed standards and samples, an external calibration, usually exhibits precision of less than 3%relative standard deviation or coefficient of variation (CV). The CV often can be reduced to below 1%

by the use of an internal standard, a fully resolved component added to the injection of sample or stan-

dard. Peak responses are then normalized to the internal standard.

Depending on the exact conditions used, and especially on how well the analyte and detection

method are matched, detection limits can vary from below 1 g/L for a 25-L sample to above 1 mg/

L. Detection limits are sometimes lowered by the use of a concentrator column so that large volumes

(as much as 100 mL) can be injected. For the most common applications, inorganic anions and cations

using suppressed conductivity detection, detection limits are about 1 to 5 g/L for a 25-L sample.


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It is usually desirable, though not necessary, to have a linear relationship between analyte concen-

tration and response. The linearity is usually expressed by the coefficient of determination (r2) over the

calibration range. Beers law for absorbance detection, and the high degree of dissociation of many spe-cies in dilute solutions for conductometric detection, are reasons why calibration curves are usually lin-

ear (r2 > 0.999) over three, and sometimes four, orders of magnitude. Similar linearities are common

for amperometric and fluorimetric detection, but because exact conditions for detection can vary, so can

linearity.

For IC the dynamic rangethat is, the concentration range over which analytes can be deter-

minedis a function not only of detection but also of separation. Although it is true that better detection

limits can extend the dynamic range at low concentrations, it is column capacity that extends the range

at high concentrations. The more analyte that can be injected into a system without overloading the col-

umn, the greater is the dynamic range. With most detection methods, there is little reason to use low-

capacity columns; however, nonsuppressed conductometric detection often uses a low-capacity column

to allow the use of low eluent concentrations.

Figure 12.5 Anion profile of polyphosphoric acid.


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Applications

Conductivity

The determination of most anions and cations, both inorganic and organic, is the mainstay of IC. It is

used to analyze drinking water, rain water, soil, foods, chemicals, and countless other samples. Many

municipal water facilities monitor ionic content of drinking water to ensure quality and safety (Fig.

12.6). The anion composition of rain and other forms of precipitation tells us a lot about our air quality

(Fig. 12.7). Generally, high concentrations of anions in rain indicate high acid content. The ion compo-

sition of soil can help determine its suitability for agriculture. Proper reporting of the ionic content of

foods and beverages is crucial to helping us maintain healthy diets. Several industries rely on high-pu-

rity reagents for their processes and IC is often used to monitor that purity. If ionic impurities are too

high, poor-quality products may be produced. IC has also been used for more exotic applications such

as determining the composition of moon rocks or the ionic content of ice found in Antarctica that is

thousands of years old.Thousands of publications cover applications of IC with conductivity detection (Chap. 39). The an-

alyst may want to consult one or more of the books listed in the References for a more complete dis-

cussion of the topic.

Amperometry

Although it is a relatively new technique, the determination of carbohydrates is one of the most com-

mon applications of IC with amperometric detection (Chap. 36). It is the pairing of the most selective

separation technique with the most specific and sensitive detection technique for this class of com-

Figure 12.6 Cations in drinking water.


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pounds. It has been applied to the analysis of foods (Fig. 12.8), oligosaccharide and polysaccharide pro-

filing, and monosaccharide compositional analysis and oligosaccharide analysis of carbohydrates from

glycoproteins.

Amperometry is also used with more traditional ions, such as iodide and sulfite. Each of these ions

can be detected by conductivity, but detection limits are much lower using single-potential or pulsed

amperometry and, depending on the sample matrix, amperometry may be more specific for these ions

than for other ions that are present. Examples of sulfite in beer and wine and iodide in foods have been

reported.

An interesting application of amperometric detection in IC is the detection of cyanide or sulfideusing a silver working electrode. In these cases, it is not the analyte that undergoes oxidation but the

working electrode. A very low potential is applied (0.05 or 0.00 V) and the current measured from the

formation of silver complexes according to the reactions

Because these are weak anions, they can be chromatographed by either anion exchange or ion exclu-

2CN

Ag0

Ag(CN)2

e

S=

2Ag0

Ag2S+2e

+

+

+

Figure 12.7 Anions in rain water.


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sion. In cases where high concentrations of chloride or other ions that react with silver may be present,

ion exclusion is usually the better choice.

Optical

Certain inorganic ions, such as nitrate and nitrite, are chromophoric and can therefore be specifically

detected using UV absorbance (Chap. 25) in complex samples such as waste water. Usually, however,

the analyst is also interested in the nonchromophoric ions in the sample, so conductivity may be the

better choice. Of course, there are a wide variety of applications for chromophoric amines and carbox-

ylic acids. Aromatic amines and carboxylates can be detected by either UV absorbance or conductivity

but, in general, as their molecular weights increase their conductometric responses decrease and their

UV responses remain the same or increase.

Metals, especially transition metals and lanthanide metals, are most commonly determined with

IC by chromatographing them as anionic complexes, of oxalate or pyridinedicarboxylate for instance,

and detecting them by absorbance after a postcolumn reaction (Fig. 12.9). In theory, this approach

has the advantage over spectroscopic methods of being able to separately determine different oxida-

tion states of a given metal. In practice, this works very well for Cr(III)/Cr(VI) and with some care

can work for Fe(II)/Fe(III). However, because many eluents are prone to upset any redox equilibrium

that may exist in the sample and because of the care needed in preventing standards from oxidizing

or reducing, IC does not have wide application for oxidation state speciation.

The determination of amino acids was perhaps the first widely used IC application. HPLC methods

using precolumn derivatization and a reversed phase separation have replaced IC for amino acid anal-

Figure 12.8 Carbohydrate components of orange juice.


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ysis of purified proteins, but IC is still commonly used for physiological and other complex samples

that are prone to interferences by reversed phase methods.

Hyphenated Techniques

IC Inductively Coupled Plasma Optical Emission Spectrometry

The coupling of IC, or more accurately chelation concentration, to ICP spectroscopy is a young tech-

nique and not widely used, but it has advantages over simple ICP for difficult samples. One of the most

challenging types of samples for ICP is one with metals of interest at low concentration (5 to 10 g/L)

and interfering metals (iron, aluminum, calcium) at much higher concentrations of about 50 to 100 mg/

L. Chelation concentration can be an automated sample preparation technique that concentrates the

metals at low concentration, eliminates the interfering metal, and delivers the sample to the ICP in the

column effluent. Standards are concentrated by the same process and delivered to the ICP in the same

effluent. The technique has been used for mg/kg-level lanthanide metals in acid digested rock, g/L-

level transition metals in sea water, and a variety of other samples. See Chap. 21 and 22 for further dis-

cussion.

IC Mass Spectrometry

IC has been interfaced with MS not for quantitative analysis, but for analyte identification. An ion chro-

matographic separation, with either a volatile salt eluent or one that can be suppressed before entering

the MS, is used to resolve the components. The technique has been applied to oligosaccharides and sul-

fonic acids (10). There are no commercial instruments dedicated to this technique, so the analyst want-

ing to work in this research area should be knowledgeable about both IC and MS. See also Chap. 33.

Figure 12.9 Transition metals concentrated from sea

water using chelation concentration followed by IC.


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Nuts and Bolts

Relative Costs

Complete system $$ to $$$

Components

Single-piston pump $

Dual-piston pump $$

Conductivity detector $ to $$

Amperometric detector $ to $$

Absorbance detector $ to $$

Fluorescence detector $ to $$

Autosamplers $ to $$Data systems

Recorders and integrators $ to $$

Computer-based $$

Columns


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Deerfield, IL 60015

phone: 708-948-8600, 800-255-8324

fax: 708-948-1078

email: [email protected]: http://www.alltechweb.com/

Bio-Rad Laboratories,

Life Science Group (C)

2000 Alfred Nobel Dr.

Hercules, CA 94547

phone: 510-741-1000, 800-424-6723

fax: 800-879-2289

Internet: http://www.biorad.com

Dionex Corp. (I & C)

P.O. Box 3603, 1228 Titan Way

Sunnyvale, CA 94088-3603

phone: 408-737-0700, 800-723-1161fax: 408-730-9403

email: [email protected]

Internet: http://www.dionex.com

EM Separation Technology (I & C)

480 S. Democrat Rd.

Gibbstown, NJ 08027-1297

phone: 609-224-0742, 800-922-1084

fax: 609-423-4389

Interaction Chromatography (I & C)

2032 Concourse Dr.

San Jose, CA 95131

phone: 408-894-9200

fax: 408-894-0405

SaraSep, Inc. (I & C)

2032 Concourse Dr.

San Jose, CA 95131

phone: 408-432-8536

fax: 408-432-8713


Internet: http://www.sarasep.com

Waters Corp. (I & C)

34 Maple Street

Milford, MA 01757

phone: 508-478-2000, 800-254-4752

fax: 508-872-1990


Internet: http://www.waters.com

Required Level of Training

Operation of the IC system can be performed by anyone with a basic, high-school level understanding


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of chemistry. Instrument troubleshooting is a skill that is usually gained through experience. Many IC

manufacturers offer basic courses on the operation and maintenance of their instruments for those who

want some additional training.

Integrators and computer-based data systems for IC are usually sufficient for routine quantitativeanalysis. Only minimal training is necessary to obtain accurate quantitative data, but a sound under-

standing of analytical chemistry and chromatography is needed to develop a method and to verify ini-

tially that the data system is programmed properly.

Service and Maintenance

Many modern IC components have internal diagnostics that can alert the analyst to the source of prob-

lems. Some systems even keep records of when maintenance tasks were last performed. The most com-

mon maintenance tasks are replacing the piston seals in the analytical pump, replacing lamps in the

optical detectors, and replacing the reference electrode in the amperometric detector cell. Regular

cleaning of the analytical column may also be necessary if sample matrices that may foul the column

are injected. The operator manual for the column usually has instructions from the manufacturer on how

best to clean the column.

Suggested Readings

SMALL, H.,Ion Chromatography, New York: Plenum Press, 1990.

WALTON, H. F., and R. D. ROCKLIN,Ion Exchange in Analytical Chemistry,Boca Raton, FL: CRC Press, 1990.

WEISS, J.,Ion Chromatography, 2nd ed.Weinheim, Germany: VCH. Verlagsgesellschaft mbH, 1995 (English

translation).

References

1. H. Small,Ion Chromatography(New York: Plenum Press, 1989).

2. H. Small, T. S. Stevens, and W. C. Bauman,Analytical Chemistry, 47 (1975), 18019.

3. J. S. Fritz,Analytical Chemistry, 59 (1987), 335A44A.

4. R. D. Rocklin,Journal of Chromatography,546 (1991), 17587.

5. H. F. Walton and R. D. Rocklin,Ion Exchange in Analytical Chemistry(Boca Raton, FL: CRC Press, 1990);

J. T. Gjerde and J. S. Fritz,Ion Chromatography, 2nd ed. (New York: Heuthig, 1987).

6. R. D. Rocklin, C. A. Pohl, and J. A. Schibler,Journal of Chromatography,411 (1987), 10719; W. R. Jones,

P. Jandik, and A. L. Heckenberg,Analytical Chemistry, 60 (1988), 19779.

7. D. C. Johnson and W. R. LaCourse,Analytical Chemistry, 62 (1990), 589A97A.

8. P. K. Dasgupta,Analytical Chemistry, 64 (1992), 775A83A.

9. S. Rabin and others,Journal of Chromatography,640 (1993), 97109.

10. R. A. M. van der Hoeven and others,Journal of Chromatography,627 (1992), 6373; J. Hsu,Analytical

Chemistry, 64 (1992), 43443.

Ch12 - Ion Chromatography

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