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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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    202 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 203

    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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    204 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 205

    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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    206 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 207

    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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    Ion Chromatography 209

    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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    210 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 211

    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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    212 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 213

    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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    214 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 215

    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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    216 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 217

    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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    218 Handbook of Instrumental Techniques for Analytical Chemistry

    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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    Ion Chromatography 219

    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

    email: [email protected]

    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

    email: [email protected]

    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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    220 Handbook of Instrumental Techniques for Analytical Chemistry

    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.