Ion Exchange Chromatography
The ion chromatography involves the separation of polar molecules and ions based on their charge. This separation technique can be used to purify any kind of charged molecules
The ionic functional groups in the stationary phase interact with the analytes of opposite charge
Ion exchange Chromatography is a Adsorption Chromatography
The earliest report of ion-exchange chromatography date back to 1850, Thompson studied the adsorption of ammonium ions to soils. Spedding and Powell published a series of papers describing practical methods for preparative separation of the rare earths by displacement ionexchange chromatography in 1947
Beginning in the 1950s, Kraus and Nelson reported numerous analytical methods which are used for metal ions based on separation of their chloride, fluoride, nitrate or sulfate complexes by anion chromatography
Type of Ion Chromatography
Based on the charge of analyte to be exchanged, ion - exchange chromatography can be sub-divided as:
- Cationic Exchanger Chromatography
- Anion Exchanger Chromatography
The stationary phase of cation exchange chromatography contains negatively charged functional groups which retain positively charged ions, while stationary phase of anion exchange contains positively charged functional groups and they retain negatively charged analytes
Ion exchange chromatography is a valuable method due to its mild separation conditions, low cost, high capacity, versatility, high resolving power, and wide range applicability
Basics of Ion Chromatography
Phases of Ion Chromatography
The first stage is equilibration in which the ion exchanger is brought to a starting state, in terms of pH and ionic strength, which allows the binding of the desired solute molecules. The exchanger groups are associated at this time with exchangeable counter-ions (usually simple anions or cations, such as chloride or sodium).
The second stage is sample application and adsorption, in which solute molecules carrying the appropriate charge displace counter-ions and bind reversibly to the gel. Unbound substances can be washed out from the exchanger bed using starting buffer
In the third stage, substances are removed from the column by changing to elution conditions unfavourable for ionic bonding of the solute molecules. This normally involves increasing the ionic strength of the eluting buffer or changing its pH
Desorption is achieved by the introduction of an increasing salt concentration gradient and solute molecules are released from the column in the order of their strengths of binding, the most weakly bound substances being eluted first
The fourth and fifth stages are the removal from the column of substances not eluted under the previous experimental conditions and re-equilibration at the starting conditions for the next purification.
Separation is obtained since different substances have different degrees of interaction with the ion exchanger due to differences in their charges, charge densities and distribution of charge on their surfaces
These interactions can be controlled by varying conditions such as ionic strength and pH
Factors Affecting Chromatographic Separation
Ion Exchange Resin: The swelling factor and cross linking is important for the effective separation. The cross linking should be controlled as its affects the exchanger’s capacity
Swelling helps in proper exposure of charged functional groups for exchange of ions
Sample: The concentration and charge of ions
Buffer: The pH of the buffer should impart the same charge to the sample ions as present in the Column. Anionic Exchange Chromatography should be carried out with cationic buffers and vice versa because buffer ion will indulge in ion exchange, which will be of no use
The best known stationary phases are issued from copolymers of styrene and divinylbenzene, in order to obtain packings hard enough to resist pressure in the column. They are made of spherical particles with diameters of 5 to 15µm that are modified on the surface in order to introduce functional groups with acidic or basic properties
Porous silica particles can serve to support, through covalent bonding, alkylphenyl chains carrying sulfonated groups or quaternary ammonium groups
A polymer called ‘latex’, prepared from a monomer that contains organic groups, is deposited as an array of tiny beads 0.1–0.2µm in diameter) on an waterproof support to form a continuous film-like layer about 1–2m thickness. The support is made of micro-spheres of silica or glass or polystyrene of about 25µm diameter
Preparation of Ion exchangers
Preparation of the exchange medium is essential for satisfactory performance of ion exchange chromatography. Apart from removing impurities, there are three major steps that are absolutely important in ion exchanger preparation.
Swelling: For anion Exchangers it is done by treating it first with an acid (0.5 N HCl) and then with base (0.5 N NaOH). Reverse is true for preparation of cation exchangers.
Removal of fine particles: Large number of such particle can result in decreased flow rate and improper resolution. For this the exchangers are repeatedly suspended in large volume of water
Addition of counter ions: Accomplished by washing the exchanger with suitable reagent depending upon which counter ion to be introduced. For e.g., NaOH (Na+ ), HCl (H+ ), etc,.
Advances in IC
Schematic of an ion chromatograph instrument
The mobile phase contains ions that create a background conductivity, making it difficult to measure the conductivity due only to the analyte ions as they exit the column
To improve the signal to noise ratio, when using a conductivity detector, a device called a suppressor, designed to selectively remove the mobile phase ions is placed after the analytical column and before the detector
The principle consists to convert the mobile phase ions to a neutral form or replacing them by others of higher conductivity. Suppressor-based detection is more useful for anion analysis than for cation analysis
Ion chromatography can be also used for the analysis of ions in natural brine waters, which include seawaters, subsurface brines, geothermal brines, and high salinity ground waters
Toxins – Cyanide and Chromium
Air Pollutants - HF, HCl, NOx and SOx on stationary emission
The most popular applications of ion chromatography are determination of common anions (F-, Cl-, Br-, NO2-, NO3-, PO43-, SO42-) and cations (Na+, K+, NH4+, Mg2+, Ca2+) in water and wastewater
In this application, ion-exchange resins are used to remove poisonous (e.g. copper) and heavy-metal (e.g. lead or cadmium) ions from solution, replacing them with more innocuous ions, such as sodium and potassium
There are some ion-exchange resins that do remove organic ions, such as MIEX (magnetic ion-exchange) resins. Domestic water purification resin is not usually recharged – the resin is discarded when it can no longer be used.