Theory of Ion Chromatography from MetrohmThere are many different ways of determining ions qualitatively and quantitatively. One such technique that is widely used is ion chromatography. Ion chromatography is one member of the large family of chromatographic methods. It is used to determine all ions which carry one or two charges. In the past ion chromatography (IC) was a very expensive method but today it is much more favorably priced, thanks to the quality Compact IC's available from Metrohm. Which Ion Chromatography Column? There are many important fields of application today for ion chromatography such as: 1. the routine investigation of aqueous systems such as drinking water, rivers, effluents and rain water. 2. for the analysis of ions in chemical products, foods, cosmetics, pharmaceuticals etc 3. ultratrace analysis such as in the semi-conductor and power industry. Ion Chromatography can be used for the analysis of anions, cations, organic acids and amines plus analytes such as carbohyrates Schematic of an Ion Chromatography System The above schematic represents a non -suppressed ion chromatography system. The sample is introduced onto the system via a sample loop on the injector. When in the inject position the sample is pumped onto the column by the eluent and the sample ions are then attracted to the charged stationary phase of the column. The charged eluent elutes the retained ions which then go through the detector (which is most commonly conduct ivity) and are depicted as peaks on a chromatogram. Three main modes of Ion Chromatography Columns. The different modes of chromatography (anion exchange, cation exchange and ion exclusion) simply relate to the different types of columns used to achieve the separation of the ions. The eluent used depends on the column type and also the mode of detection - however unless stated the following is all based on conductivity detection. The chromatogram below shows a sample with a suppressor unit placed between the column and detector. The greatest achievment of suppression is to increase the sensitivity of the anion. . The two forms are anion exchange with or without suppression and of these two suppressed methods are the most widely used. Eluents for suppressed chemistries tend to be either carbonate based or hydroxide.particularly for sulphate. When suppression is used the detector is almost certainly conductivity.Ion exchange Ion exchange chromatography (IC) is based on a stoichiometric che mical reaction between ions in a solution and the oppositely charges groups functional groups on the column resin. In the simplest case in cation chromatography these are sulfonic acid groups or carboxylic acid groups (such as maleic acid) and in anion chr omatography quaternary ammonium groups. however at the same time the background conductivity of th e eluent is greatly reduced. chromatogram showing an anion exchange separation followed by direct conductivity detection (non -suppressed) Suppression in Ion Chromatograph Often. sensitivity or selectivity . Anion exchange Anion exchange chromatography forms the largest group of IC methods mainly because there are few alternatives with such simplicity. a device called a suppressor is used and is placed between the column and detector as shown above. The same suppressor units can also be used to increase the sensitivity of organic acids. . one will be in-line with the eluent and conductivity detector. no system peak. which means that the sodium carbonate/sodium bicarbonate mobile phase is converted to water instead of carbonic acid so a background conductivity approaching 1uS is achieved. The 853 MCS can be fitted to all Modular and 761/861 Compact IC·s and the benefits include no injection peak. for example NaCl (~126uS without suppression) would become HCl (~426uS with suppression). for example. The anion chemical suppression can be taken a stage further with the new 853 MCS instrument (fitted inline after the MSM) which removes carbon dioxide from the suppressor reaction and carbonate from the sample. adversely affected by the transition metals which can cause precipitation problems for other types of suppressor technology. and 861 Compact) and is available in the modular system as the 833 Advanced IC Liquid Handling Unit. This suppressor module is present in all the Metrohm Compact ICs (792 Basic. superb linearity and an enhancement in the peak areas allowing lower limits of detection to be achiev ed. one will be in -line with dilute sulphuric acid (replacing the removed cations with H+) and the third is washed with water. 790 Personal. The ways in which this can be done are varied but the two common ways are as follows: 1: The Metrohm MSM (Metrohm Suppressor Module) contains 3 separate suppressor units. So a sodium carbonate eluent (~800uS) would be converted to carbonic acid (~18uS) by the suppressor and the analyte. The benefits of this technique are lack of baseline noise and a ruggedness that is reflected in the fact that the MSM comes with a ten year warranty and is not. At any one time.Chromatogram showing an anion exchange separation followed by suppression and then conductivity detection The suppressor used in anion chromatography is simply a cation exchanger and its job is to remove cations and replace them with an H+. The import ance of this is that carbonate eluents can now be used for gradient elution of anions which means more versatility. A large number of applications for silica-gel-based ion exchangers exist. These columns allow simultaneous separation of alkali metals and alkaline earths plus the separation of transitional metal and heavy metal ions is also possible. As the concentration of the eluent increases throughout the gradient. Chromatogram showing gradient elution with carbonate eluents. S mall amines can also be analysed using cation exchange columns. . Schematic of the Metrohm Gradient System with two 818 Advanced IC Pumps forming the high pressure gradient pump (P1 and P2) and the 828 IC Dual Suppressor. The Dual Suppressor then reduces the conductivity of the carbonic acid (~18uS) by removing it to leave water (~1uS). Cation Exchange There is a variety of cation columns available. the baseline rise is a result of the increasing conductivity of the eluents suppression product. The Metrohm Dual Suppressor is a continuous suppression device which removes the cations and replaces them with H+ (which is provided by electrolysis of water) so with a carbonate eluent it forms carbonic acid. however the modern ones contain carboxylic acid functional groups.2. no system peak. less corrosive eluents and no gases required. Chromatogram showing ion exclusion with an acid eluent For a more detailed explanation of the theory of ion chromatography and detection see the Metrohm Monograph 'Practical Ion Chromatography' available free of charge from Metrohm UK. Ion exclusion Ion exclusion chromatography (IEC) is mainly used for the separation of weak acids or bases. Cations become less sensitive when suppressed and so are analysed with direct conductivity detection which also allows heav y metals to be analysed as shown above. 790 Personal IC and 861 Compact IC. 833 Advanced IC Liquid Handling Unit and 820 Advanced IC Separation Centre. . HYPERLINK enquiry@metrohm. For modular IC see 819 Advanced IC Detector. 818 Advanced IC Pump. For on -line IC see the 811 Online IC and the 821 Compact Online IC.co. The greatest importance of IEC is for the analysis of weak acids such as carboxylic acids. zinc and cobalt. carbohydrates. the concentration of which can effect the elution of calcium and heavy metals such as iron.Chromatograms showing cation exchange with two different tartaric acid/dipicolinic acid eluents The eluents used for non -suppressed cation exchange are weak acids with a complexing agent such as dipicolinic acid. phe nols or amino acids.uk For compact ion chromatography units containing all components see 792 Basic IC. Photometric detection (see UV-Vis Spectrophotometer. Amperometry is very sensitive. however. as many organic molecules contain chromophore groups. important analytes such as fluoride. . While of the simple anions only analytes such as nitrate. in the analysis of sugars by anion chromatography and in clinical analysis. but multivalent and transitional metals in particular can be converted in a post -column derivatization with chelate fo rmers such as 4-(2-pyridylazo)-resorcinol (PAR) or Tiron to form colored complexes. nitrate. or can have one introduced or added. which are able to absorb in the UV or VIS spectrum. The amperometric detector is the most important version. Co2+) it is chiefly anions such as nitrite. The most important applications lie.Other modes of Detection Amperometric detection (see 791 IC-VA Detector and 817 Bioscan) (Amperometric detection (see 791 IC -VA Detector and 817 Bioscan) In principle voltammetric detectors can be used for all compounds which have functional groups which are easily reduced or oxidized. sulfate or phosphate can only be measured indirectly. 844 Compact UV-Vis and Post Column Reactor) Chromatogram showing analysis of 1ppb chromate using the 844 Compact UV-Vis Because of its extremely wide range of application photometric or UV/VIS detection is the most important detection method used in HPLC. In the field of inorganic ion analysis UV/VIS detection plays a smaller role. bromide or iod ide absorb. Redox-active analytes such as bromate and other oxohalide ions can be analyzed by UV/VIS detection after undergoing a post-column reaction with an electrochemically active indicator. Many cations do not absorb at all. thiosulfate as well as halogens and pseudo -halogens which can be determined in the ion analysis sector. Apart from a few cations (Fe3+. C) is a subdivision of High Performance Liquid Chromatography (H. the matrix of the sample makes it difficult to accurately quantify the species of interest with the standard ion chromatography set -up and some form of sample preparation then becomes necessary. Chromatograph y techniques are divided up according to the physical states of the two participating phases. A general definition of ion chromatography can be applied as follows. In the case of more difficult forms of sample matrices it may be necessary to add additional dedicated sample preparation modules to the standard ion chromatography configuration. one of which is stationary and the other mobile moving in a particular direction. Schematic Diagram of Standard Ion Chromatography Set -up The sample preparation may be as straightforward as simply diluting the sample with deionised water or can involve injection of the sample through a solid phase extraction cartridge to remove the interference.Sample Preparation Techniques for Ion Chromatography Introduction to Sample Preparation Techniques Quite often with problematic ion chromatography applications.µ ion chromatography includes all rapid liquid chromatography separations of ions .L.C). What is Ion Chromatography? Chromatography is a method for separating mixtures of substances using two phases. The term Ion Exchange Chromatography or Ion Chromatography (I.P. The process of ion exchange leads to a condition of equilibrium. Another option is the use of a cation exchanger in the Ag+ form for the removal of any halides present in the sample. the side to which the equilibrium lies depends on the affinity of the participating ions to the funct ional groups of the stationary phases. . This type of exchange cartridge removes carbonate/bicarbonate and is also useful for the removal of cations from samples being determined by ion exclusion chromatography. A stoichiometric chemical reaction occurs between ions in a solution and a solid substance carrying functional groups that can fix ions as a result of electrostatic forces. Solid Phase Extraction Cartridges Passage of the sample through one or more solid phase extraction cartridges prior to injection will often retain selectively certain species within the homogeneous sample. There are a number of different cartridges whose suitability depends upon the type of chemistry undertaken. or there are sample matrix effects that can often be minimised by a dilution usually with water but eluent can also be used. In theory ions with the same charge can be exchanged completely reversibly between the two phases. Another common type is the non-polar exchange cartridge (reversed phase) that often utilises C18 groups to remove organic substances that would otherwise interfere with the chromatography. For anion analysis. Quite often the retained species are substances that would interfere with the chromatography ha d they not been previously removed.in columns coupled online with detection and quantification in a flow through detectorµ. the sample can be treated with a cation exchanger in the H+ form that removes divalent cations that can mask any fast eluting anions. Different Types of Sample Preparation Techniques Employed Dilution of the Sample Dilution of the sample is performed when the concentration of the analytes of interest either exceed the working capacity of the separa tion column chosen.45mm filters to ensure that any particulate material from the samples don·t make their way onto the injection valve or the analytical column where they can cause blockages and considerably reduce the lifetime of the column(s). Filtration of the Sample It is recommended to filter all samples prior to injection with 0. one can employ an anion exchanger in the OH form to remove any interfering anions present in the sample. Similarly for cation analysis. For anion chromatography these are quaternary ammonium groups and for cation chromatography sulphonic acid groups. The sample solution is constantly being renewed resulting in no depleti on of the sample. The 833 Dialysis Unit is strongly recommended for demanding applications that contain strongly loaded samples. Dialysis is the diffusion of ions from a sample solution into an acceptor solution to achieve a concentration equilibrium. Instrumental Sample Preparation Modules Often with more complex sample matrices. Different types of digestion includ e wet. The ions pass through the membrane without hindrance but larger sample particles ² the sample matrix .Digestion of the Sample If digestion techniques are to be employed then analyte content should be changed as little as possible and any organic matter present should ideally be destroyed completely. Metrohm has actually been a pion eer of inline sample preparation modules with the release of the 754 IC Dialysis Unit in 1997. one has to add additional dedicated sample preparation modules to the standard ion chromatography configuration.838. It consists of a dual channel peristaltic pump for conveying the sample and acceptor solutions and the actual dialysis cell in which the ions from the flowing sample solution are enriched in the resting acceptor solution. since then the technology has been optimised and considerably improved so that today the Metrohm IC portfolio contains many different sample preparation instruments. Calibration of the standards is easily done requiring no extra outlay.are transported past the membrane to waste reducing matrix effects to an absolute minimum.0040) The Metrohm 833 Dialysis Unit is a module for online sample preparation in ion chromatography permitting the use of automatic sample dialysis directly before sample injection. microwave and UV.0210) . 833 Advanced IC Liquid Handling Dialysis Unit (2. the suitability of each depends on both the sample matrix and the analytes of interest being determined.833. One can obtain analytical inaccuracies due to an exaggerated blank value as a result of the chemicals used during the digestion. 838 Advanced IC Ultra-Filtration Sample Processor (2. There are a number of different instrument options available within the Metrohm range depending on the type of sample treatment required prior to analysis. The reported dialysis recovery rates have been found to be in excess of 98% using the patented stopped flow method. 833. It is eminently suitable for those samples with a light to medium load such as surface waters and digestion solutions. Only a small fraction of the sample is removed as filtrate so the contaminants remain mainly in the sample stream preventing the regenerated cellulose membrane from becoming blocked too quickly. The 838 IC Ultra-Filtration sample processor combines inline filtration with automatic sample injection through the use of an ultra-filtration cell. 833 Advanced IC Liquid Handling Sample Preparation Unit (2. At the same time the filtrate is aspirated off from the rear of this membrane and transferred to the sample loop by the second channel of the pump. The sample is conveyed by one channel of the pump through the ultra filtration cell passing the membrane.The aim of sample filtration is to protect the separation columns from contamination and blockage from particulates that may be present in the sample.0030) . The samples are placed onto the sample carousel before being processed automatically. Sample filtration and introduction to the injection valve is achieved by means of an integrated double channel peristaltic pump meaning that it is possible to aspirate slightly viscous samples. The modules consists of a reactor block that houses the cation exchangers with a control unit that contains a two channel peristaltic pump that conveys the regenerant and rinse solutions. .S.M) for difficult ani on analyses such as those found in concentrated alkaline solutions.Picture of Metrohm 833 Advanced IC Liquid Handling Sample Preparation Unit Inline sample preparation i s rapidly becoming the method of choice for eliminating difficult sample matrices in ion chromatography and Metrohm has developed the 833 Advanced IC Liquid Handling Sample Preparation Unit based upon its Metrohm Suppressor Module (M. ultra -filtrate or even dilute the sample automatically inline. . The sample solution then passes onto the preconcentration column where the trace anions to be determined are retained and then eluted by the eluent flo wing in a counter flow direction. Example Chromatogram Showing Matrix Elimination upon a Sample of Caustic Soda for the Determination of Chloride. the capacity of the 833 is unlimited. water is formed by neutralisation. The sample cations are exchanged against protons (H+).Schematic Diagram of Inline Sample Preparation The matrix elimination occurs inline whilst the regeneration and rinsing of the packed bed suppressor occur simultaneously offline. Chlorate and Sulphate Metrohm Inline Sample Preparation (MISP) With Metrohm it is possible to perform the time consuming sample preparation inline using the 838 Advanced IC Sample Processor equipped with MISP technology. Located on the side of the tower is the relevant sample preparation technology for example an ultra -filtration cell or an injection valve which is utilised for dilution along with the proven Dosino liquid handling technology. A fresh suppressor channel is used for each new analysis and because the ri nse and regeneration occurs after each determination. The sample solution is transferred to the 833 module from an autosampler via a loop injection and rinsed with deionised water. If sodium hydroxide constitutes the sample matrix. The 838 comes in a number of different variants that mean it is possible to dialyse. The analyte anions are then separated on the analytical column before quantification using chemical suppression with conductivity detection. the oxo halides and dichloroacetic acid. EPA 300. EPA 317. which yields extremely high plate numbers and accordingly excellent separation and detection characteristics.The new Range of Anion Columns from Metrohm include: - The disinfection by-products generated in water processing plants are suspected to be not only a health hazard but could even be carcinogenic.0). the column's counter-pressure remains small. Even at these high flow rates. Metrohm now presents a high-performance separation column for the simultaneous determination of the standard anions.g.1 Part B. The Metrosep Dual 4columns contain an entirely novel carrier material. whose dead volume is very low. This contributes to the high capacity of the Metrosep Dual 4 column. A special sodium chloride eluent is used for these applications. With the Metrosep A Supp 7 these ions can be determined reliably and precisely down to lower ppb range. Compared to conventional materials.m polyalcohol polymer. bromide and nitrate in concentrated salt solutions. namely a functionalized monolith based on silica. have become the object of many investigations and standard methods (e. For this reason the oxo halides. This monolith allows flow rates of up to 5 mL/min. the monolith with its structure made up of macro. The high-capacity Metrosep A Supp 8 ² 150 allows the determination of nitrite. UV detection at 215 nm opens up the determination of concentrations in the on edigit ppb range.and mesopores has a much larger surface area. above all bromate that is generated from bromide during the ozone-treatment of drinking water. . The separations require a temperature of 45 °C. The outstanding detection sensitivity is obtained by applying the 5 .. 5 ppb perchlorate in the presence of a total of 3 g/L of chloride.The high-capacity Metrosep Dual 4 ² 100 column is the separation column of choice for the detection of very small amounts of toxic perchlorate. The Metrosep A Supp 10 ² 100 separation column is based on a high-capacity polystyrene/divinylbenzene copolymer having a particle size of only 4. Dialysis ² An Overview Dialysis is a successful method for the separation of low molecular substances from high molecular ones by means of a semi-permeable membrane and is used on patients with kidney deficiencies. Metrohm has optimized this time-tested column concept. As the ions pass through the membrane virtually unhindered they diffuse from the high molecular strength blood into the low ionic strength acceptor solution. Deliberate care is taken to ensure that no concentration equilibrium can be established between the two solutions. Metrohm has developed the patented stopped flow method for dialysis where the sample solution is continuously pumped past a semi -permeable membrane but the acceptor solution lies at rest and here lies the inherent ¡ . high selectivity and excellent separation performance. combined with moderate run times. As the kidneys can not functioning correctly. The A Supp 10 ² 100 is the column of choice for routine applications. By variation o f temperature. The concentrations of these ions must be reduced at frequent intervals and this is achieved by continuous flow dialysis. flow rate and eluent composition. The acceptor solution is permanently renewed in its continuous flow ensuring a steep concentration gradient with relatively large efficiency. excellent price-performance ratio and very good separation performance. the A Supp 10 ² 100 is a universally applicable anion separation column. An acceptor solution of a low ionic strength (usually deionised wat er) is pumped along the semi-permeable membrane with the blood flowing past on the other side. Low molecular substances in the blood refer to ions that disturb the electrolyte balance.6 m. The Dual 4 ² 100 also easily achieves the base-line separation of chloride and nitrite present at a concentration ratio of 1000 to 1. It allows to determine 0. which is characterized by its robustness. the column characteristics can be adapted to the application at hand. Thanks to its robustness. then the concentrations of these ions increases impairing the metabolic functions. carbonate and sulfate. 830 IC interface and 838 Advanced IC Dialysis Sample Processor. with dialysis these types of samples no longer represent a problem for the ion chromatography user. time consuming sample preparation steps such as digestion that can potentially destroy the analytes of interest. 820 Advanced IC Separation Center. Once the equilibrium has been set up the dialysed acceptor solution is transferred to the sample loop and injected onto the separation column. Diagram Illustrating Stopped Flow Dialysis Technique Applications of Dialysis The benefit of dialysis is that there are no complicated. cutting oil emulsions and inks. This is particularly applicable to foodstuffs and other complex matrices that carry high organic loads such as waste waters or soil eluates. In the food industry the ionic contents of milk and other diary products can easily be determined using an ion chromatography system incorporating a dialysis module. It is no longer necessary to separate the proteins from the milk using Carrez precipitation ensuring that the sample preparation is reduced to a simple dilution step. 833 IC Suppressor Module. This ensures that equilibrium is attained between the sample solution and the acceptor solution usually in less than 10 minutes.difference from continuous flow dialysis. Other difficult matrices i nclude the analysis of fruit juices that contain fruit pulp. Method for Analysis of Milk Samples The modular system used for the determination of anions present in samples of milk comprised the Metrohm modules 818 Advanced IC Pump. 819 Advanced IC Detector. . The subsequent dialysis of the sample and injection of the dialysed sample onto the separation column was fully automated and the response for the peaks recorded using a mobile phase eluent of sodium carbonate/sodium bicarbonate. ionic form then very little sample preparation is required and quantified results can be obtained within a matter of minutes. For a sample in a homogeneous. Chromatograph of Anions Found in Milk Samples after Dialysis Ion chromatography as an analytical technique has seen an enormous surge in popularity due partly to the simplicity of the method as well as other factors such as market forces driving down the expenditure costs of the initial instrumentation and improved reliability and power.3 against a previously prepared calibration plot. the consumer places ever more stringent demands and requirements upon the manufacturer and the world of ion chromatography .Picture Showing Modular System Configuration for Dialysis of Milk Samples The milk sample was diluted 1:5 with deionised water and placed in the sample vials upon the rack of the 838 Advanced IC Sample Processor. As in any industry. The calculation was carried out automatically using integration software IC Net 2. still only a relatively small volume of sample is required. Because of the ease of use at which io n chromatography as a method can be manipulated. G. Blackie Academic & Professional. Fundamentals of Analytical Chemistry . This coupled with the low running costs of ion chromatography using Metrohm instruments really does mean that ion chromatography is the method of choice for the analyst even with the most difficult and problematic sample matrices. the end user today wishes to analyse ions within increasingly complicated sample matrices which until recently would not have been possible.is no different. ISBN 0-471-97412-9. ISBN 0 -216-92920-2. Saunders College Publishing.metrohm. The Essential Guide to Analytical Chemistry. 3. such as mass spectroscopy Ion chromatography with Metrohm means High sensitivity Accurate and reproducible results Low acquisition cost All instruments backed by the COOL guarantee (Cost of Ownership is Lower) Very low service expenditure . John Wiley & Sons. Schwedt. Kealey.A Skoog. Metrohm has developed a range of instrumental sample preparation modules that can be added to standard ion chromatography configurations to allow quantification even with the most difficult sample matrices. 2. ISBN 0 -03-075397-X. (1992) 3rd edition. Product Application Manager (IC/VA Division) for Metrohm UK Ltd. F. References 1. (1999) 2nd edition. D. Coupling techniques So-called coupling techniques represent the link -up of a chromatography system with an independent analytical method. Even for those samples requiring sample preparation by dialysis or ultra filtration. the analyst is then free to carry out other functions within the laboratory affording an increase in efficiency of the employed manpower. Principles and Practice of Analytical Chemistry.W Fifield and D.M West and F. The analysis is fully automated so once the sample is loaded. The following internet site was also used extensively as a reference and can be used to obtain further information: www. D.com Article written by Jonathan Bruce.J Holler. (1992) 6th edition. And don't forget: Swiss quality from over 60 years as a pioneer of Ion Analysis Metrohm ² First Class Ion Chromatography .
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